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[ { "content": "You are a medical writer. Summarize the following article: evidence has emerged that inflammatory mechanisms contribute to the onset and progression of type-1 diabetes ( t1d ) . \n recent data further suggest that the initiation of autoimmunity is preceded by inflammation reflected by a proinflammatory metabolic serum profile . \n many autoimmune diseases are associated with increased histamine levels in serum and tissue fluids , such as rheumatoid arthritis , sjgren 's syndrome , multiple sclerosis , and diabetes mellitus [ 14 ] . \n similarly , it has been known for many years that histamine is elevated in tissue biopsies and secretions in the intestine of patients with crohn 's disease and ulcerative colitis [ 5 , 6 ] . during inflammatory responses , \n histamine , a hydrophilic vasoactive amine , increases capillary permeability and facilitates the immune response by enhancing leukocyte rolling , adhesion , and vascular extravasation of inflammatory cells to the site of inflammation . \n histamine is recognized not only as an inflammatory mediator but also as a regulator of immune responses , including the th1/th2 balance and hematopoiesis . \n patients with diabetes are more susceptible to vascular diseases , which led to the assumption that their increased circulating histamine levels promoted lipolysis and facilitated the transport of lipids into tissues . beside its physiological role in vascular health , excess histamine , \n a result of physical or emotional stress or a chronic disease / inflammatory state , appears to elicit atherogenic effects . \n whether histamine is a critical marker or target in the treatment of type-1 diabetes has not yet been established . \n it is still poorly understood how the expression of histidine decarboxylase ( hdc ) , the unique enzyme responsible for histamine generation , is controlled . \n hdc is induced in a variety of tissues in response to bacterial components ( lipopolysaccharides , peptidoglycan ) and to various cytokines such as il-1 , il-3 , il-12 , tnf , g - csf , and gm - csf . \n its upregulation has been associated with pathological conditions , including t1d . on the other hand , \n an impaired glucose tolerance has been reported in hdc mice , together with autoantibodies reactive to glutamic acid decarboxylase ( gad ) . \n histamine exerts important functions in allergy , inflammation , neurotransmission , and the gastrointestinal tract through g - protein - coupled specific histamine receptors , termed h1r - h4r . in the context of diabetic physiopathology , it plays a pivotal role in various physiological functions , such as feeding behaviour and energy homeostasis . \n mice with genes disrupted for hdc are prone to become obese on a high - fat diet or at advanced age . \n these metabolic changes presumably are due to the impaired regulatory loop involving hypothalamic histamine and h1r [ 12 , 13 ] . \n consequently , treatment with imetit , a h3r agonist , reduced plasma histamine , leptin , and insulin levels and has anorexigenic effects in diet - induced obesity . \n histamine has been shown to regulate t cells by enhancing th1-type responses through the h1r and downregulating both the th1 and th2 type responses through the h2r [ 15 , 16 ] . \n it has recently been described that anti - inflammatory and immunomodulatory functions of histamine are mediated through the h4r . \n thus , they can be reproduced using the h4r agonist clobenpropit , which has been shown to suppress il-12 production , modulate dendritic cell migration [ 18 , 19 ] , and regulate the growth of cancer cells through h4r . autoimmune diabetes in nod mice is characterized by a progressive mononuclear cell infiltration in the pancreatic islets of nod mice and leads to beta cell destruction and hyperglycemia . \n it has been recently demonstrated that leptin can reverse diabetes . assuming that histamine might be involved in the pathogenesis of diabetes \n , we investigated the relationship between endogenous histamine production and disease development in the nod mouse model , using histamine - deficient hdc mice backcrossed on a nod background . here , we show that the lack of endogenous histamine provides a notable protection against t1d . \n we observed an increase of ly6gcd11b immature myeloid cells in these mice that was not correlated with diabetes onset , while il-12 and ifn- levels in the serum were decreased , suggesting that endogenous histamine modulates the inflammatory pattern . \n paradoxically , we found that t1d onset was also delayed when exogenous histamine was given to nod . \n our study supports a metabolic role of histamine during the pathogenesis of autoimmune diabetes in accordance with previous findings supporting its immunomodulatory functions . \n conventional nod mice ( k , i - a , and d ) were bred in our animal facility at the hpital necker . \n hdc mice generated by ohtsu et al . received a histamine - low diet ( safe ( scientific animal food and engineering ) ) to avoid exogenous uptake . \n we generated hdc - deficient mice by speed backcrossing the hdc - deficient allele against the nod background . in order to confirm the allele maps of nod \n diabetes susceptibility genes , we analysed several representative microsatellite markers : we chose the alleles related to mhc and numerous insulin - dependent diabetes genes . \n then , mice were intercrossed to generate the hdc mice used in this study . we purchased histamine and 4-methylhistamine ( 4-mh ) from sigma - aldrich and clobenpropit dihydrobromide ( cb ) from tocris . \n l of saline once a week for 10 weeks starting at 4 weeks of age . \n h4r agonists , cb ( 10 g / kg ) , 4mh ( 10 g / kg ) , and vehicle ( control group for h4r - agonist ) were injected i.p . in 100 \n leptin ( millipore ) , ifn- ( r&d ) , il-6 ( r&d ) , and il-12 ( r&d ) were quantified in sera from mice , using elisa kits according to the manufacturer 's instructions . \n we treated nod mice with histamine , cb , or 4-mh and monitored weekly for clinical signs of diabetes using gluko - test reagent sticks , to detect glucose in urine samples ( boehringer mannheim , meylan , france ) . when needed , glycemia was also measured in a drop of blood collected from the tail vein using a reflolux s glucometer ( boehringer mannheim ) . \n incidence of diabetes was defined based on the discovery , upon serial monitoring , of glycosuria and hyperglycemia ( fasting glycemia > 2.5 g / l ) . to induce t1d by adoptive transfer , \n purified cd4 diabetogenic t cells from prediabetic nod mice were injected i.v . into eight - week - old nod rag recipient mice with a cd11b population ( 5 10 cells / mice ) \n bone marrow cells were stained with cd11b microbeads ( miltenyi biotec ) then sorted by positive magnetic separation on ls columns ( macs separator , miltenyi ) . \n a high incidence of t1d is elicited within 10 weeks after t cell transfer . bone marrow ( bm ) , spleen , and peripheral whole blood cells \n bm and spleen cells were prepared as previously reported [ 25 , 26 ] and adjusted to a final concentration of 1 10 per ml in culture medium ( pbs from gibco brl ) . \n cell suspensions were stained with the following appropriately labeled antibodies : cd11b , ly6g- ly6c , cd4 , cd25 , foxp3 , mpdca-1 , cd11c , cd115 , mhc - ii , cd80 , and cd86 ( bd biosciences , pont - de - claix , france ) . \n mononuclear cells were gated according to forward- and side - scatter properties and analysed using the facs diva software . \n diabetes incidence was plotted using the kaplan - meier method , that is , nonparametric cumulative survival plot . statistical comparison between curves \n was performed using the log rank ( mantel - cox ) test that provided the corresponding values . when needed \n to evaluate the role of histamine in autoimmune diabetes , we generated nod mice deficient for the gene encoding hdc in the nod genetic background . \n these genetically engineered mice were healthy and heavier than their wild - type controls after several months . \n as shown in figure 1 , the onset of t1d was delayed in nod hdc mice and the disease incidence remained lower than that among wild - type controls , suggesting a deleterious role of histamine during pathogenesis . it may be hypothesized that an increase in plasma and cellular histamine content contributes to disease progression by enhancing endothelial permeability , as suggested for the intimal damage in atherosclerosis . in favour of this hypothesis , \n histamine levels were effectively increased in diabetic nod mice ( data not shown ) , similarly to what has been reported for streptozotocin - diabetic rats . to assess potential changes in immunological and hormonal parameters in the absence of histamine , we analysed interleukin-12 ( il-12 ) , il-6 , and leptin . il-12 and \n ifn- levels were lower in hdc than in nod mice , but no differences were noted for il-6 or leptin ( figure 1(b ) ) . as histamine contributes to the differentiation and maturation of hematopoietic myeloid cells , we performed a comparative cytometry analysis of these populations in histamine - deficient and wild - type nod mice in various organs , such as bone marrow , spleen , and peripheral blood based on gr-1 , cd11b , ly6 g , and ly6c expression by leukocytes . \n we found an increased percentage of immature cd11bgr-1 cells in the peripheral blood and the spleen from nod hdc mice ( figure 2(a ) ) . \n a similar result was observed for the cd11bly6 g ly6c subset from peripheral blood and spleen of nod hdc mice ( figures 2(b ) and 2(c ) ) . \n these results confirmed the previously reported importance of histamine for the differentiation of cd11bgr-1 myeloid subsets in a different genetic background . \n we hypothesized that the increased proportion of immature myeloid cells ( imcs ) might be involved in the protection against type-1 diabetes in histamine - deficient mice . to test this assumption , we performed adoptive transfers of effector cd4cd62l t cells together with cd11b bone marrow cells from hdc - deficient or wt mice to nod rag recipients , which did not alter diabetes onset ( figure 2(d ) ) , ruling out the increase of imcs as a cause for diabetes progression . \n we also analysed the status of myeloid - derived suppressive cells ( mdscs ) , which can modulate diabetes onset . \n however , no difference was observed when comparing this subset in both strains ( figure 2(e ) ) . \n similarly , the proportion of cd4cd25foxp3 regulatory t cells remained unchanged ( figure 2(f ) ) . \n since the generation of endogenous histamine appears to enhance the pathogenesis of t1d , we attempted to reproduce this deleterious effect by adding exogenous histamine . \n injections of either vehicle ( 100 l pbs ) or histamine ( 4 mg / kg ) , respectively , once a week for 10 weeks . as shown in figures 3(a ) and 3(b ) \n , exogenous histamine not only failed to increase the incidence of t1d , but also delayed the onset of disease in both wild - type and hdc mice . \n as shown in figure 3(c ) , decreased expression of class ii major histocompatibility complex ( mhc - ii ) and cd86 molecules occurred only on pdcs from pancreatic lymph nodes ( lns ) in mice treated with histamine , suggesting that this mediator may modulate the activation pattern of apcs . \n recent evidence supports the notion that the inflammatory functions of histamine are mediated through the h4r [ 20 , 29 , 30 ] . for this reason , we examined whether compounds characterized as potent h4r agonists could reproduce the protective effect provided by exogenous histamine . \n we injected nod mice with clobenpropit ( cb ) , an isothiourea derivative of histamine , or 4mh for 10 weeks and found no significant protection as compared with mice having received vehicle alone ( figures 3(d ) and 3(e ) ) . \n in the present study , we provide evidence that histamine deficiency causes a delay in diabetes onset and decreases its incidence . \n this result fits with clinical observations in diabetic patients since increased histamine levels were associated with pathogenesis in these patients . \n furthermore , there is now a compelling body of evidence for the contribution of histamine to the development of autoimmune diseases from animal studies . \n it has also been reported that inhibition of histamine synthesis in experimental diabetes reduces disease complication . \n several arguments suggest that in the present study the phenotype results from vascular and/or metabolic functions of histamine . this conclusion has already been proposed in a rat diabetic model , since \n the authors have shown that the gastric histidine decarboxylase activity and plasma gastrin levels were increased in connection with the depletion of insulin . \n we can thus anticipate that decreasing histamine levels in diabetic patients can decrease the risk of vascular complications , which is one critical issue in the cure for patients . \n there is increasing evidence that diabetes is under the control of numerous cytokines and hormones . \n since leptin can influence diabetes [ 22 , 33 ] , we have measured the level of this mediator in serum of starved mice of both strains . however , we observed no change in its level . \n we observed that il-12 and ifn- levels were lower in nod hdc female mice compared with wt controls . \n these cytokines are known for promoting diabetes progression in the nod strain , possibly via induction of th1-type cytokines [ 34 , 35 ] . by contrast \n , circulating il-6 or leptin did not differ between wt and ko mice , ruling out a major involvement of these factors in the delayed disease onset in hdc - deficient mice . \n even though we detected hematopoietic anomalies in the myeloid population of hdc mice , these cells were not involved in the protective response , as demonstrated by adoptive transfer . \n although neutrophils express the same markers as imcs , the protection against diabetes in hdc - deficient mice can not be ascribed to an increase of this population among the cd11bgr-1 subset since neutrophil depletion has been shown to inhibit type-1 diabetes development in nod mice . furthermore , the expansion of imcs is not concomitant with mdscs since the two populations bear different markers and most mdscs do not express ly6 g \n . furthermore , we found that the proportion of regulatory myeloid cells was not modified comparing both strains . \n finally , the increased histamine levels associated with the pathogenesis of diabetes are expected to be a consequence rather than a cause of disease . \n therefore , adding exogenous histamine may act on the gastric mucosa to enhance the flow of the lipidic factors but will be unable to reverse the phenotype observed in hdc - deficient mice . indeed , we observed the opposite effect . \n we describe a new immunomodulatory function of histamine in our model since diabetes onset is delayed in both wild - type and hdc mice after treatment . \n for example , the role of the h3r on food intake has been well documented . \n since it has been previously established that h4r drives inflammatory responses in asthma , dermatitis , arthritis , and colitis [ 29 , 30 , 37 ] , we hypothesized that its agonists could reproduce the effects of histamine . \n this was not the case in this study , but even though we could not validate the h4r pathway using the agonists cb or 4-mh , we can not exclude that the dose , the route of administration , or the choice of the agonists tested was inadequate . \n alternatively , a mixture of h2r and h4r agonists may be required to obtain a protective effect similar to the one induced by histamine . \n histamine , which is increased in inflammatory and pathological situations , has also been shown to decrease , in vitro , the production of il-12p70 of monocyte - derived dendritic cells ( dcs ) via h2r and h4r [ 17 , 19 , 38 ] and to inhibit il-27 production in activated apcs . \n the regulation of cytokine production in maturing dcs causes an alteration of the t cell polarization that has been well described [ 15 , 40 , 41 ] . \n more recently , it has been reported that histamine decreases the migration of human pdcs and their cytokine production , while increasing regulatory t cell recruitment . in histamine - treated nod mice \n , we observed that pdcs from pancreatic lymph nodes expressed lower levels of costimulatory and mhc - ii and cd86 molecules than their counterpart from control mice , consistent with a decreased inflammatory state . \n contrary to our anticipations , injection of histamine did not promote diabetes pathogenesis but , paradoxically , decreased disease onset . \n the immunomodulatory function of histamine has been described in other inflammatory models and histamine is used therapeutically in patients suffering from acute myeloid leukemia . \n histamine release can be induced by different stimuli , such as cytokines or microbial stimulation . \n it has been previously shown that human basophils release histamine in response to a toll - like receptor 2 ( tlr2 ) agonist . \n interestingly , tlr2 agonists have also been implicated in the modulation of t1d in the nod mouse . \n similarly , parasites such as schistosoma mansoni protect nod mice and parasites stimulate basophils and induce histamine release . \n these results support our hypothesis that local histamine release upon stimulation of immune cells , such as basophils , may participate in immunoregulatory pathways in autoimmune diabetes .", "role": "user" }, { "content": "recent evidence has highlighted the role of histamine in inflammation . since \n this monoamine has also been strongly implicated in the pathogenesis of type-1 diabetes , we assessed its effect in the nonobese diabetic ( nod ) mouse model . to this end , we used mice ( inactivated ) knocked out for the gene encoding histidine decarboxylase , the unique histamine - forming enzyme , backcrossed on a nod genetic background . \n we found that the lack of endogenous histamine in nod hdc/ mice decreased the incidence of diabetes in relation to their wild - type counterpart . whereas the proportion of regulatory t and myeloid - derived suppressive cells was similar in both strains , \n histamine deficiency was associated with increased levels of immature macrophages , as compared with wild - type nod mice . \n concerning the cytokine pattern , we found a decrease in circulating il-12 and ifn- in hdc/ mice , while il-6 or leptin remained unchanged , suggesting that histamine primarily modulates the inflammatory environment . \n paradoxically , exogenous histamine given to nod hdc/ mice provided also protection against t1d . \n our study supports the notion that histamine is involved in the pathogenesis of diabetes , thus providing additional evidence for its role in the regulation of the immune response .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: the amyloid hypothesis states that accumulation \n of amyloid- \n ( a ) peptides in the brain is the primary driver of pathogenesis \n in alzheimer s disease ( ad ) , including synapse loss and neuronal \n cell death . the full - length a1 - 42 peptide and its a1742 fragment ( p3 ) \n are formed via cleavage of the amyloid precursor protein \n ( app ) by the action of three secretase enzymes . \n the \n ape3 - 42 fragment is post - translationally \n generated by cleavage of the first two n - terminal amino acids of a1 - 42 , leaving an exposed glutamate ( e ) residue in position \n 3 . \n the lactam ring in the pyroglutamate ( pe ) residue is subsequently \n generated by intramolecular dehydration catalyzed by the glutaminyl \n cyclase ( qc ) enzyme . \n pe - modified \n as represent the dominant fraction of a \n oligomers in brains of ad patients . \n autopsied \n brains of elderly patients with normal cognition also show accumulation \n of a1 - 40/42 , but the amount of accumulated \n ape3 - 42 is significantly lower . consequently , the ratio of ape3 - 42/a1 - 42 oligomers is higher in ad brains \n than in brains of normal elderly individuals . \n the \n larger accumulation of ape3 - 42 in ad brains \n has been attributed to its increased stability and higher aggregation \n propensity . \n these properties are attributed \n to the lactam ring in the pe - modified third residue as well as the \n loss of electrical charge in three residues during the conversion \n of a1 - 42 to ape3 - 42 , thus resulting in higher ape3 - 42 hydrophobicity \n and increased resistance to degradation by peptidases . \n significantly , it is also believed that the formation of \n ape3 - 42 plaques precedes a1 - 40/42 plaque formation . \n this is supported \n by the observation that ape3 - 42 plaques appear \n earlier than a1 - 40/42 plaques in down syndrome \n ( ds ) brains . \n the additional copy of chromosome \n 21 , characteristic of ds , is responsible for generating more app , \n and thus individuals with ds are more likely to develop ad earlier \n in life . increasing evidence suggests \n that following initial interactions \n on the cell membrane , a oligomers insert into the membrane \n and form pore structures . \n cell toxicity \n results from an abrupt change in cell ionic concentration , producing \n loss of cell homeostasis . \n pore activity has been observed for full - length \n as , a \n fragments , and point substitutions . in \n addition to pore formation , a-induced toxicity mechanisms include \n lipid extraction by peptides on the membrane surface . \n these mechanisms are not mutually exclusive , as a recent \n study suggested that pore formation precedes nonspecific fragmentation \n of the lipid membrane during amyloid fiber formation . to the best of our knowledge \n , there is currently \n no experimental \n data for amyloid pore formation in vivo . ape3 - 42 \n these ape3 - 42 mice displayed a significantly reduced survival \n rate compared with a1 - 42 transgenic mice . in vitro , \n optical patch - clamping was used to \n characterize the single - channel ca fluorescence transients \n induced by a1 - 42 pores in membranes of xenopus laevis oocytes . \n however , \n most experimental evidence of amyloid pore formation stems from model \n membrane studies . here \n we characterize the electrical properties \n of pe - modified a \n in lipid membranes . \n we discuss the activity and structure of ape3 - 42 pores in anionic model cell membranes . \n we used \n phosphoethanolamine ( pe ) and phosphoserine ( ps ) lipid headgroups to \n mimic the brains of elderly patients . \n phospholipids with ethanolamine \n ( pe ) head groups are one of the dominant components in the brains \n of the elderly , and these levels , as \n well as those of ps , have been found \n to change in ad brains . \n using planar \n lipid bilayer ( plb ) electrical recording , we show that ape3 - 42 induces pore activity in anionic membranes , producing \n increased membrane permeability with respect to a1 - 42 pores . using atomistic molecular dynamics ( md ) simulations , we model \n the architecture of ape3 - 42 pores and correlate \n them with the activity observed experimentally . \n ape3 - 42 was purchased \n from bachem ( torrance , ca ) , and a1 - 42 was \n purchased from bachem and anaspec ( fremont , ca ) . the phospholipids \n 1,2-dioleoyl - sn - glycero-3-phosphoserine ( dops ) and \n 1-palmitoyl-2-oleoyl - sn - glycero-3-phosphoethanolamine \n ( pope ) were purchased from avanti polar lipids ( alabaster , al ) . all \n other chemicals were purchased from sigma - aldrich ( st . louis , mo ) . \n ape3 - 42 and \n a1 - 42 peptides were dissolved in milli - q \n water to a concentration of 1 mg / ml prior to being aliquoted for storage . \n these 50 l aliquots were stored at 80 c for a \n maximum of 60 days before use . \n bilayers were formed from a 1:1 ( w / w ) mixture \n of dops and pope in heptane at a total lipid concentration of 20 mg / ml . \n spontaneous membrane formation occurs following the addition of lipid \n directly over a pore with a diameter of 250 m in a \n delrin septum ( warner instruments , delrin perfusion cup , volume 1 \n ml ) . in previous studies \n , this membrane composition was shown to be \n stable for long recording times ( 4 h ) . \n as the electrolyte , we use 150 mm kcl , 10 mm hepes ( ph \n 7.4 ) , and 1 mm mgcl2 . before performing electrical \n recordings \n , we verified that the bilayer was stable for several minutes \n with low conductance ( < 10 ps ) across 100 mv applied voltage \n and that the system capacitance was > 110 pf . \n when both criteria \n were \n met , peptide was added directly to the cis ( hot wire ) side and stirred \n for 5 min . \n bilayer stability was monitored by periodic capacitance \n measurements throughout the course of the experiment . \n all traces \n were recorded in voltage clamp mode using the 2 khz \n built - in filter cutoff of our bc-535 amplifier ( warner instruments , \n hamden , ct ) . \n we used a custom - made labview program to record the current \n and clampfit 10.2 ( molecular devices , sunnyvale , ca ) to analyze traces . \n for representation in figures , \n we have filtered the recorded current \n versus time traces with a digital gaussian low - pass filter with a \n cutoff frequency of 50 hz . \n two u - shaped \n a monomer conformations with the -strand - turn--strand \n motif were extracted from a1 - 42 fibrils , \n where the structure was defined by hydrogen / deuterium - exchange nmr \n data , side - chain packing constraints from pairwise mutagenesis , ssnmr \n and em ( pdb code : 2beg ) , and small a1 - 40 protofibrils ( pdb codes : 2lmn and 2lmo ) , where the structure was based on \n a ssnmr model . in both structures , the n - terminal coordinates , residues \n 116 for the former and 18 for the latter structure , \n are missing due to disorder . \n we used the a116 coordinates , in the absence of zn ( pdb code : 1ze7 ) , for the missing portions of the peptides . for each combination \n of the n - terminal structure with the u - shaped motifs , \n conformer 1 has a turn at ser26-ile31 , and \n conformer 2 at asp23-gly29 . in the latter conformer , two c - terminal \n residues , ile41 and ala42 , \n , we removed the first two residues , asp1 and ala2 , \n from each conformer and converted glu3 into pe3 , generating ape3 - 42 . \n because the standard charmm force field does not provide a force field for pe , we first \n created the pe molecular topology using the avogadro software . \n then , we calculated partial charges , bond lengths , \n angles , and torsional angles for the atoms in the pe residue using \n the gaussian09 program on a biowulf cluster \n at the nih . \n two ape3 - 42 conformers , each derived from the wild - type ( wt ) \n a1 - 42 conformers with different turns , still \n retain the u - shaped structure with the -strand - turn--strand \n motif . \n to construct ape3 - 42 barrel structures , \n we inclined ape3 - 42 monomers 37 \n relative to the pore axis ; then , an 18-fold \n rotational symmetry operation was performed with respect to the pore \n axis creating an 18-mer ape3 - 42 barrel ( figure 1 ) . \n we modeled a barrels with the -sheet \n structure by mimicking naturally occurring -barrels observed \n in transmembrane proteins that are found frequently in the outer membranes \n of bacteria , mitochondria , and chloroplasts . \n the -barrel motif \n is a large -sheet composed of an even number of -strands . \n some known structures of -barrel membrane proteins have -strands \n ranging in size from 8 to 22 . \n we modeled \n 18-mer a barrels , with 18 -strands enclosing the solvated \n pore . \n this number is also in the range of the number of -strands \n for natural -barrels ranging from 8 to 22 , which can form a \n -barrel motif . \n our previous simulations for a channels \n indicate that different numbers of a monomers could produce \n channels with different outer and pore dimensions . \n we found that a channels obtained a preferred size range \n of 1624 -strands lining the pores . this range was also found to hold for other toxic -sheet \n channels : k3 channels with 24 -strands , 18- and 24-mer hiapp channels , pg-1 channels with 1620 -strands , and max channels with 20 -strands . in agreement with afm data , \n these channels have outer and \n pore dimensions within the range found with afm . in this work , \n the \n outer / pore diameters of the 18-mer ape3 - 42 barrels are in good agreement with the experimental afm ranges and the computational 18-mer wild - type a \n barrels . \n the afm experiments provide images of channels \n with a wide variety of sizes and shapes , but simulated a barrels \n are limited to cover all ranges of channel sizes that are imaged by \n afm . \n monomer conformations of ( a ) conformer 1 ape3 - 42 with turn at ser26-ile31 and ( b ) conformer 2 ape3 - 42 with turn at asp23-gly29 , and ( c ) highlight of pyroglutamate at \n residue 3 ( pe3 ) . \n the initial barrel structures of md \n simulations in ribbon representation for the ( d ) conformer 1 and ( e ) \n conformer 2 ape3 - 42 barrels . in the peptide \n ribbon , \n hydrophobic , polar / gly , positively charged , and negatively \n charged residues are colored white , green , blue , and red , respectively . \n the pe3 n - terminal termini are highlighted as threads . to obtain \n a lipid bilayer , we constructed a unit cell containing \n two layers of lipids . \n in the middle of the unit cell , lipid molecules \n were randomly selected from the library of the preequilibrated state \n and replaced by pseudo vdw spheres at the positions of the lipid headgroups , \n constituting the lipid bilayer topology . for a given number of lipid molecules , the optimal value of lateral \n cell dimensions can be determined . \n an anionic lipid bilayer composed \n of dops and pope with a mole ratio 1:2 , containing a total of 420 \n lipids , constitutes the unit cell with tip3p waters added at both \n sides . \n updated charmm all - atom additive \n force field for lipids ( c36 ) and the \n modified tip3p water model were used \n to construct the set of starting points and to relax the systems to \n a production - ready stage . \n the system contains mg , k , ca , and zn at the same concentration \n of 25 mm to satisfy a total cation concentration near 100 mm . \n the \n bilayer system containing an ape barrel , lipids , \n salts , and water has almost 200 000 atoms . \n we generated at \n least 10 different initial configurations for each conformer ape3 - 42 barrel for the relaxation process to obtain the \n best initial configuration for a starting point . in the pre - equilibrium \n stages , a series of minimizations was performed for the initial configurations \n to remove overlaps of the alkane chains in the lipids and to gradually \n relax the solvents around the harmonically restrained peptides . \n the \n initial configurations were gradually relaxed through dynamic cycles \n with electrostatic cutoffs ( 12 ) . \n the harmonic restraints were \n gradually diminished with the full ewald electrostatics calculation \n and constant temperature ( nos hoover ) thermostat / barostat \n at 303 k. for t < 30 ns , our simulation employed \n the npat ( constant number of atoms , pressure , surface area , and temperature ) \n ensemble with a constant normal pressure applied in the direction \n perpendicular to the membrane . \n production runs of 100 ns for \n the starting points with the namd code were performed on a biowulf cluster at the nih . \n plb electrical recording data demonstrate \n that ape3 - 42 peptides induce spontaneous \n pore activity through lipid membranes presenting the general features \n observed for a1 - 42 pores ( figure 2 ) . at a concentration of 10 m and at constant \n voltage both peptides show stepwise changes in the current flowing \n through the membrane ( figure 2a ) , characteristic \n of the opening and closing of ion channels . \n however , unlike ion channels \n that have regulated activity and possess integer values of a unitary \n conductance , amyloid pores are not regulated and present multiple \n conductance values due to the different number of monomers composing \n the pore and membrane channel interaction dynamics . \n the multilevel conductance seen here in both \n ape3 - 42 and a1 - 42 is a hallmark of amyloid pores . \n ape3 - 42 and a1 - 42 produce pore \n activity in anionic lipid membranes . \n representative \n traces for the activity of ( a ) ape3 - 42 and \n ( b ) a1 - 42 pores in stable membrane recording . \n the activity \n seen in both membranes , namely step , spike , and burst behavior is \n characteristic of amyloid ion channels . \n bilayers were formed by the \n painted technique using 1:1 ( w / w ) mixture of dops / pope . \n both sides \n of the bilayer chamber contained as electrolyte 150 mm kcl , 1 mm mgcl2 , and 10 mm hepes ( ph 7.4 ) . \n the first , as seen in figure 2 , was characterized \n by stable , long recording times ( > 30 min ) following the first onset \n of activity . \n the second type of activity was characterized by ionic \n conduction through the membrane that grows rapidly in an exponential - like \n fashion ( figure 3a ) . \n phase began within 20 min of the first observed activity and typically \n led to current saturation of the amplifier within 10 min . \n this type \n of growth was seen in 50% ( 3/6 ) of ape3 - 42 and 33% ( 2/6 ) of a1 - 42 membranes ( table 1 ) . at first , only a few pores open ( figure 3b ) , and the membrane stays in the same conductive \n state for several 10s of seconds ( region inside green rectangle ) . \n however , after an abrupt increase in the membrane conductivity due \n to the opening of a large single pore or several pores simultaneously , \n the activity is characterized by several short small steps only a \n few seconds long , suggestive of the opening of several pores in a \n cascade - like fashion . despite this rapid growth , \n discrete steps indicative \n of pore activity are still clearly observed ( figure 3c ) . \n in addition , similar to a1 - 42 , ape3 - 42 pores are voltage - independent \n and can be blocked by zn ( figure 3a ) . \n this type of membrane activity \n was observed in 50% of ape3 - 42 membranes \n and 33% of a1 - 42 membranes . \n ( a ) the current \n across the membrane increased rapidly upon disruption of the membrane , \n exceeding the 1.1 na saturation current of our amplifier . \n an abrupt increase in the membrane conductivity \n due to the opening of a large single pore or several pores simultaneously \n leads to the opening of several pores in a cascade - like fashion . \n ( c ) \n clear step behavior is still observed during exponential current growth \n ( region indicated by the short blue line an a ) . \n a mixed mechanism \n involving both channel activity and nonspecific leakage is likely \n involved . \n although the overall characteristics of the \n membrane activity induced \n by ape3 - 42 and a1 - 42 pores are similar , distinct differences are present . \n table 1 displays the characteristic parameters observed \n for the electrical activity of both ape3 - 42 and a1 - 42 . \n the onset of a1 - 42 activity in plb can be observed as soon as several minutes following \n peptide addition into the chamber and was typically seen within 60 \n min post - addition with an average of 31.6 25.7 min . \n ape3 - 42 demonstrated a significantly longer ( p < 0.05 for a one - tail t test ) and \n more variable lag time , with an average of 98.2 68.5 min . \n we next examined whether there was a difference between ape3 - 42 and a1 - 42 channel conductances . \n figure 4 shows the histogram distribution of \n single - channel conductances calculated from both step and spike activity . \n the histograms show that the conductances can be sorted into three \n groups that demonstrate a shift toward higher conductances for ape3 - 42 pores ( figure 4a ) compared \n with a1 - 42 ( figure 4b ) . \n the first group of conductances , below 100 ps , consisted of 74% \n of ape3 - 42 and 91% of a1 - 42 events . \n the second group is between 100 and 200 ps and , significantly , \n shows a second peak of activity for ape3 - 42 that is absent for a1 - 42 . \n while 21% of \n the events fall in this range for ape3 - 42 , only 5% of a1 - 42 events are in this range . \n the third grouping shows similar rates of sparse activity greater \n than 200 ps ( 5 and 4% for ape3 - 42 and a1 - 42 , respectively ) . \n while the percentage of activity \n in the high conductivity range is similar , close inspection of the \n histogram shows higher conductances for ape3 - 42 in this region . \n table 1 shows that the average conductance in this third \n region is 20% higher for ape3 - 42 . \n these results show an overall trend for higher ape3 - 42 conductance , which could have relevance to the ape3 - 42 toxicity mechanism . \n histogram analysis of ( a ) ape3 - 42 and \n ( b ) a1 - 42 conductances shows that there is \n an increased propensity for higher conductance events for ape3 - 42 . \n both sets of conductances \n can be sorted into three groups ; < 100 ps , 100200 ps and \n > 200 ps , as shown in table 1 . \n ape3 - 42 presents significantly more events than a1 - 42 in the 100200 ps interval . \n data sample \n size was 1400 for ape3 - 42 and 1192 for a1 - 42 . \n data were collected in the 50 mv range , \n with peptide at a concentration of 10 m . \n the electrolyte used \n was 150 mm kcl , 1 mm mgcl2 , and 10 mm hepes ( ph 7.4 ) . \n bilayers \n were made by the painted technique with dops / pope lipids dissolved \n in heptane . \n we performed 100 ns all - atom md simulations \n on ape3 - 42 barrels embedded in an anionic \n lipid bilayer composed of dops / pope ( mole ratio 1:2 ) . \n the ape3 - 42 barrels comprising two different u - shaped conformers \n were initially preassembled as an annular shape . \n the initial annular \n conformation is gradually lost via relaxation of the lipid bilayer , \n and no immediate peptide dissociation in the barrels was observed \n ( figure s1 of the supporting information ) . \n the u - shaped portions of the ape barrels ( residues \n 1542 and 1142 for the conformer 1 and 2 ape3 - 42 barrels , respectively ; see peptide topologies \n in figure 1 ) , which mostly include membrane \n embedded portions , reach equilibration after the initial transient \n state , while the extramembranous n - termini of the peptides ( residues \n 314 and 310 for the conformer 1 and 2 ape3 - 42 barrels , respectively ) are disordered ( figure \n s2 of the supporting information ) . small \n fluctuations in the pore and c - terminal strands in the lipid bilayer \n retain the u - shaped peptide motif in the ape3 - 42 barrels ( figure s3 of the supporting information ) . \n in our simulations , the ape3 - 42 peptide \n also presents heterogeneity in barrel conformations ( figure 5 ) , as observed in the wt a1 - 42 and mutant a barrels . \n the \n outer diameters for the membrane embedded portion are 7.6 \n and 7.1 nm for the conformer 1 and 2 ape3 - 42 barrels , respectively . \n the averaged pore diameters are 2.2 \n and 1.9 nm for the conformer 1 and 2 ape3 - 42 barrels , respectively . \n both outer / pore diameters of ape3 - 42 barrel are in the range of the wt barrel . averaged pore structures calculated with \n hole embedded in the average barrel conformations \n during the \n simulations for the ( a ) conformer 1 and ( b ) conformer 2 ape3 - 42 barrels . in the barrel structures with the surface \n ( top view ) and the ribbon representations ( angle and lateral views ) , \n hydrophobic \n , polar / gly , positively charged , and negatively charged \n residues are colored white , green , blue , and red , respectively . for \n the pore structures , \n red denotes pore diameter of d < 1.4 nm , green denotes pore diameter in the range , 1.4 nm d 2.0 nm , and blue denotes pore diameter of d > 2.0 nm . unlike the wt a1 - 42 barrels , where the \n n - terminal strands normally reside in the bulk water area ( figure \n s4 of the supporting information ) , the \n pe3 n - termini of ape3 - 42 barrels tend to \n retreat to the lipid hydrophobic core due to the hydrophobicity of \n the lactam ring . to locate the pe3 residue across the bilayer \n , we \n calculated the probability distributions for pe3 as well as few selected \n charged groups in the barrels ( figure 6 ) . \n the \n distribution curves for pe3 spanning the interior of the lipid bilayer \n indicate that several pe3 termini interact with the lipid hydrophobic \n tails . \n the interaction energy of the pe3 residue with lipids shows \n strong attraction for several pe3 n - termini ( blue bars in figure s5 \n of the supporting information ) , while no \n strong lipid interactions of the standard n - termini were observed \n in the wt a1 - 42 and mutant barrels . \n relocating \n the pe3 n - termini into the lipid hydrophobic core significantly reduces \n the fluctuations of the n - terminal portions of barrels at the lower \n bilayer leaflet , further stabilizing the barrel conformation . with \n the pe3 anchoring in the membrane , \n the ape3 - 42 barrel would provide a clear channel mouth at the lower bilayer \n leaflet and hence yield a wide - open water pore for ion leakage , suggesting \n that ape3 - 42 pores may lead to high ion conductance . \n probability \n distribution functions for pe3 and selected charged \n residues , pe3 ( purple ) , glu11 ( red ) , lys16 ( orange ) , and glu22 ( yellow ) , \n and for the phosphate group of lipid head , po4 ( green ) , \n as a function of the distance along the pore center axis for the ( a ) \n conformer 1 and ( b ) conformer 2 ape3 - 42 barrels . \n initial locations \n of pe3 are marked by arrows . to observe ion activity \n in the ape3 - 42 pores , we calculated the probability \n distribution for ions across \n the bilayer ( figure s6 of the supporting information ) . \n peaks in the distribution curves reflect the highly populated \n ion binding sites in the pore . \n the locations of ion binding sites \n in each conformer ape3 - 42 barrel are similar \n to those of each corresponding conformer of wt a1 - 42 barrels because both barrels share the same u - shaped motif in the \n lipid bilayer . to observe ion fluctuation across the pore , we calculated \n the change in total charge in the pore as a function of the simulation \n time ( figure 7 ) . \n two selected pore lengths \n along the pore axis , 1.0 < z < 1.0 \n nm and 1.8 < z < 1.8 nm , were used \n in the calculation . \n these pore lengths ensure that the charge fluctuations \n exclude a contribution of ion interactions with the lipid head groups . \n for |z| < 1.0 nm , \n the pore of the ape3 - 42 barrel conformer 1 exhibits larger charge fluctuations \n than the conformer 2 barrel , because the glu22 cationic binding site \n is located at z = 0.6 nm , attracting more cations into the \n pore . for the conformer 2 ape3 - 42 barrel , \n the glu22 cationic binding site is located at z = \n 1.8 nm near the channel mouth ( as indicated by ca peak at z = 1.8 nm in figure s6b of the supporting information ) . to correlate these charge \n fluctuations with experimental ion conductance , we calculated the \n maximum conductance , gmax , representing the ion transport , which can be \n described as1where qe is the elementary charge \n , kb denotes \n the boltzmann s constant , t is the simulation \n temperature , and l represents the pore length of \n 36 . in the bracket , d(z ) \n and gpmf(z ) denote the \n 1-d diffusion coefficient and the 1-d potential of mean force for \n ions , respectively . for mg , \n k , ca , and zn , the maximum conductances are 667 , 338 , 115 , \n 155 ps and 137 , 271 , 92 , 89 ps in the pores of conformers 1 and 2 \n ape3 - 42 barrels , respectively . averaged maximum \n conductances for ions are relatively higher than those calculated \n for the wt a1 - 42 barrels . \n change in total charge in the pore as a function of the simulation \n time for the ( a ) conformer 1 and ( b ) conformer 2 ape3 - 42 barrels . \n the pore heights with cutoff along the pore axis , 1.0 \n < z < 1.0 nm and 1.8 < z < 1.8 nm were used . \n we have shown that the toxic pe - modified a peptides produce \n pore activity in dops / pope anionic membranes . \n these pores demonstrate \n similar activity characteristics ( heterogeneous step , spike and burst \n conductance , zn blockage ) as a1 - 42 pores as well as other previously studied pores of the a \n family . however , there are also some significant differences . \n the \n onset of ape3 - 42 pore activity is generally \n delayed , but once started , ape3 - 42 pores \n show increased propensity for larger conductance events compared with \n a1 - 42 pores , particularly in the 100200 \n ps range ( figure 4 ) . \n structurally , the lactam \n ring of ape peptides induces a change in the conformation \n of the n - terminal strands of the ape3 - 42 pores . \n while the n - terminal strands of a1 - 42 pores \n normally reside in the bulk water region , the n - termini of ape pores tend to reside in the hydrophobic lipid core , providing \n the ape3 - 42 pores with higher stability . \n the longer times required for the onset of ape3 - 42 pore activity are tentatively attributed to ( i ) the higher hydrophobicity \n of ape3 - 42 , which decreases the attractive \n charge - dipole interactions with the zwitterionic lipid heads , thus \n also decreasing the number of adsorbed ape3 - 42 oligomers available for subsequent membrane insertion , and ( ii ) \n the possible longer times required for pores to assemble in the membrane . \n our plb data demonstrate that the ionic current produced by ape3 - 42 pores can rapidly rise to levels potentially \n toxic for cells . following pore formation , \n according to early \n estimations , a single pore with a gigantic \n 4 ns conductance can produce a change of 10 m / s in internal \n na concentration , producing loss of cell homeostasis in \n seconds . \n while no single catastrophic event of this nature was observed \n ( figure 4 ) , the summation of smaller ionic \n conductance events can have a similar toxic effect . \n ape3 - 42 has been shown to be more cytotoxic than a1 - 42 in vitro . \n we showed that ape3 - 42 shows a greater propensity for higher conductance values when compared \n with a1 - 42 . \n this would lead to a more prevalent \n and rapid dysregulation of cellular ionic homeostasis for ape3 - 42 , thus suggesting increased cytotoxic properties \n of ape3 - 42 pores . \n the pore structures \n found for ape3 - 42 had \n characteristics and dimensions in accordance with the pores previously \n reported . \n pore structures have been characterized by afm and modeled \n using md simulations for several a peptides . \n our previous studies \n indicate that full - length a1 - 40/42 , certain a mutants , a fragments , as well as other amyloids form heterogeneous pore structures showing 46 subunits and \n shapes varying from rectangular to hexagonal . \n our md simulations suggest \n similar pore sizes for both peptides but increased stability of the \n ape3 - 42 pores inserted in dops / pope membranes . \n stability in the membrane is likely more important for the formation \n and longevity of the larger pores with conductances in the 100200 \n ps range . as a result , \n ape3 - 42 pores with \n conductances in this range have a higher population than a1 - 42 pores ( figure 4 ) . \n pores \n with conductances > 200 ps likely have similar and fast decay rates \n for ape3 - 42 and a1 - 42 , and thus both populations are similarly low , although still somewhat \n larger for ape3 - 42 . \n such a scenario would \n also correlate with the greater in vitro cytotoxicity observed for \n ape3 - 42 peptides . \n the energy of a pore \n according to the continuum model is2where r is the pore \n radius , \n is the line tension at the pore edge , and is the surface \n tension of the membrane . \n our md results indicate that ape3 - 42 and a1 - 42 pores have \n similar diameters and general characteristics , while the main difference \n is the retreat of the n - termini of ape3 - 42 into the hydrophobic lipid core . \n therefore , this reconfiguration \n of the n - termini likely decreases the line tension of ape3 - 42 pores , minimizing their pore energy and rendering \n them more stable . \n the exponential growth of the current , seen in figure 3 , would lead to rapid cell death and was observed \n in 50% ( 3/6 ) of the ape3 - 42 membranes in \n this study . while pore activity appears to be the only mechanism present \n during the early stages ( green rectangle in figure 3b ) , the activity trace suggests that two mechanisms of ionic \n conductance act simultaneously during exponential saturation : ( i ) \n stepwise pore behavior and ( ii ) nonspecific drift leakage . \n the nonspecific \n leakage could be caused by a number of alternative mechanisms leading \n to a-induced membrane permeability , including carpeting and \n detergent effects . \n the data suggest that \n a combined mechanism ( pore and nonspecific leakage ) may be at work \n during the rapid growth of the exponential phase , \n at least some of the time . \n it is important to note that similar behavior \n was also seen for some of the a1 - 42 membranes . \n although this behavior was observed for more membranes with ape3 - 42 pores , this difference ( table 1 ) was not statistically significant to reach conclusions regarding \n specific ape3 - 42 cytotoxity . \n in summary , using plbs and md simulations , we have shown that ape3 - 42 forms ionic pores in dops / pope lipid bilayers . \n the activity of these pores follows the same general pattern as a1 - 42 pores . \n significantly , however , ape3 - 42 pores display higher conductances than a1 - 42 pores , in particular , in the 100200 ps range . \n in addition , \n the onset of ape3 - 42 pore activity is delayed \n compared with a1 - 42 pores . \n structurally , \n ape3 - 42 and a1 - 42 pores have similar dimensions , but the pe3 residue induces a change \n in the conformation of the n - terminal strands of the ape3 - 42 pores . \n while the n - terminal strands of a1 - 42 pores normally reside in the bulk water region , \n the n - termini of ape pores tend to reside in the \n hydrophobic lipid core , providing the ape3 - 42 pores with higher stability compared with the a1 - 42 pores . \n these studies are a first step in understanding the role \n of pores in the enhanced toxicity attributed to ape peptides .", "role": "user" }, { "content": "among the family of a peptides , \n pyroglutamate - modified a \n ( ape ) peptides are particularly associated with \n cytotoxicity in alzheimer s disease ( ad ) . \n they represent the \n dominant fraction of a oligomers in the brains of ad patients , \n but their accumulation in the brains of elderly individuals with normal \n cognition is significantly lower . \n accumulation of ape plaques precedes the formation of plaques of full - length a \n ( a1 - 40/42 ) . \n most of these properties appear \n to be associated with the higher hydrophobicity of ape as well as an increased resistance to enzymatic degradation . however , \n the important question of whether ape peptides induce \n pore activity in lipid membranes and their potential toxicity compared \n with other a pores is still open . \n here we examine the activity \n of ape pores in anionic membranes using planar bilayer \n electrical recording and provide their structures using molecular \n dynamics simulations . \n we find that ape pores spontaneously \n induce ionic current across the membrane and have some similar properties \n to the other previously studied pores of the a family . \n however , \n there are also some significant differences . \n the onset of ape3 - 42 pore activity is generally delayed compared with \n a1 - 42 pores . however , once formed , ape3 - 42 pores produce increased ion permeability of the \n membrane , as indicated by a greater occurrence of higher conductance \n electrical events . \n structurally , the lactam ring of ape peptides induces a change in the conformation of the n - terminal \n strands of the ape3 - 42 pores . while the n - termini \n of wild - type a142 peptides normally reside \n in the bulk water region , the n - termini of ape3 - 42 peptides tend to reside in the hydrophobic lipid core . \n these studies \n provide a first step to an understanding of the enhanced toxicity \n attributed to ape peptides .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: fabry disease ( fd ) is a rare x - linked disorder resulting from a deficient activity of the lysosomal enzyme -glycosidase a ( -gal a ) that leads to progressive accumulation of glycosphingolipids in several tissues and organs . in the classical form of the disease , \n renal involvement commonly ensues in the second to third decades of life with overt proteinuria ( > 300 mg / day ) rapidly leading to chronic kidney disease ( ckd ) and end - stage renal disease ( esrd ) . \n cardiac and renal atypical variants of the disease with different degrees of organ impairment and the pattern of disease have been described in males . in females , \n the disease presentation may range from a completely asymptomatic status to the classic form of fd . \n nonetheless , to the best of our knowledge , isolated proteinuria as the first sign of the disease is rare and has been described only in males . \n enzyme replacement therapy ( ert ) with agalsidase has been demonstrated to be safe and effective in the improvement of several signs and symptoms of fd in both genders , and it is accepted that the early establishment of ert might prevent organ damage . thus , ert is recommended in the presence of symptoms or organ involvement . \n however , due to the variable presentation and the slower progression of the disease in female patients , the questions of how to detect organ involvement and when to start therapy remain open . \n vv , a 20-year - old female was referred to our clinic in 2006 for a family screening programme due to one uncle , the mother and sister being diagnosed with fd [ point mutation i354k ( c1061t > a ) of the exon 7 ] . at that time , vv was completely asymptomatic for fabry disease ; leucocyte -gal a activity was 19.5 nmol / mg prot / h ( normal values 1850 nmol / mg prot / h ) , while molecular analysis showed the presence of the pathologic genotype , confirming the heterozygous status of the patient . at this time , clinical evaluation and periodic laboratory tests were unremarkable and no sign of fd could be detected . \n two years later in the first evaluation , urine analysis showed evidence of isolated persistent microalbuminuria ( 303 mg / l ; normal value < 30 mg / l ) without overt proteinuria or abnormalities in the urinary electrolytes . \n all laboratory tests were in the normal range including serum creatinine and creatinine clearance ( 0.7 mg / dl and 92 ml / min , respectively ) . \n similarly , a thorough physical examination of the patient could not elicit any sign of fd or any other disease . specifically , body mass index , blood pressure and heart rate were 20.5 kg / m , 120/70 mmhg and 68 bpm , respectively . \n audiometric evaluation did not show any evidence of hearing loss . magnetic resonance imaging ( mri ) of \n the brain was negative for infarctual encephalopathy . finally , both sweat test and electromyography were normal , with no evidence of sensitive - motor abnormalities . due to the presence of isolated and persistent microalbuminuria \n light microscopy revealed numerous vacuolated cells in both glomerular tuft and tubular cells ; myelin osmiophilic bodies were also present in glomeruli or between tubules ( figure 1a ) . \n direct immunofluorescent study showed negative staining for iga , igm , igg , c1q , c3 and c4 . \n zebra bodies in podocytes , epithelial and endothelial cells of the glomerular tuft ( figure 1b ) . \n thus , the renal biopsy confirmed that the patient presented fd with the sole involvement of the kidney . \n considering the aggressive course of the disease in the family , the optimal blood pressure control and what was suggested by the international guidelines for fd diagnosis and treatment , we started ert with agalsidase beta ( fabrazyme - genzyme corp . ) at a standard dose of 1 mg / kg every 2 weeks , in the hope to provide vv with an aetiological rather than antiproteinuric treatment . \n after 6 months of treatment , the microalbuminuria returned to the normal range ( 27 mg / l ) , while no other signs or symptoms of fd could be detected . \n renal biopsy of an asymptomatic fabry female patient presenting with isolated microalbuminuria ( 303 g / l , 24 h urine collection ) . \n light microscopy ( panel a , left ) shows numerous vacuolated cells in glomerular tufts ( upper panel ) and tubuli ( lower panel ) . \n tem ( panel b , right ) shows osmiophilic zebra bodies in podocytes and epithelial cells ( upper panel ) and endothelial cells ( lower panel ) . \n several studies have demonstrated that heterozygous patients , due to the lyon phenomenon , may present different phenotypes ranging from the complete absence of symptoms of the classical form of fabry disease . to the best of our knowledge , \n this is the first case reported of an adult heterozygous female in whom isolated microalbuminuria is the first and only presenting sign of both fd and renal involvement . \n the presentation of fd with a prevalent renal impairment ( so - called renal variant ) is rare and documented in male patients only . \n nonetheless , microalbuminuria has been recently described as a preliminary sign of renal impairment in symptomatic young female patients . \n it is the authors opinion that the present case report should question the watch and wait approach and pose several questions on what should be looked at as early signs of fd and when to start ert . indeed , the early detection of renal damage appears of paramount importance since the presence of proteinuria and/or ckd at the time of the start of ert are independent predictors of poor outcome . \n however , our case should suggest that microalbuminuria might be regarded as an early sign of fd nephropathy . \n furthermore , the disappearance of microalbuminuria after 6 months of ert without antiproteinuric agents [ such as angiotensin - converting enzyme inhibitors ( ace - i ) or angiotensin receptor blockers ( arbs ) ] should deserve attention since it might suggest that early intervention might also reverse fd nephropathy . \n this finding might be of importance if we consider data from the fabry registry that seem to corroborate the notion that females with fd are undertreated , as only 33.8% of females currently included in the fabry registry are on ert . in summary \n , this case suggests that the kidney may be the first , and sometimes only , organ damaged by fd in heterozygous female patients and that isolated and persistent microalbuminuria instead of proteinuria should be regarded as an early marker of fd nephropathy and an indication for ert . \n renal variant, as well as how to detect organ damage and when to start an early treatment in female fd patients .", "role": "user" }, { "content": "fabry disease ( fd ) is a rare x - linked disorder characterized by low or absent activity of the lysosomal enzyme -glycosidase - a that leads to progressive accumulation of glycosphingolipids in different organs and tissues . \n clinical manifestations vary from classic to atypical forms characterized by one prevalent organ involvement , and a renal variant has been described in men but not in women . \n however , little is known about renal manifestation in females affected by fd . \n we herein report a case of a 22-year - old female with isolated and persistent microalbuminuria as the only sign of fd . in light of the importance of early recognition and treatment of fd organ damage , this case should call for future studies to determine how to assess organ damage , investigate the existence of a \n renal variant in fd female patients and determine when best to start enzyme replacement therapy ( ert ) .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: supplementary material is available for this article at 10.1007/s13659 - 012 - 0026-z and is accessible for authorized users . \n ", "role": "user" }, { "content": "three new sesquilignans , 13 , a new sesquiterpenoid , 4 , and three known compounds were isolated from the stem barks of illicium simonsii . \n the structures of new compounds ( 14 ) were elucidated by spectroscopic methods . \n a biosynthetic pathway was proposed for simonsienols a - c ( 13 ) . \n anti - ache activity and anti - buche activity were evaluated for all compounds except for -cadinol ethyl ether ( 4 ) . as a result , \n isodunnianol ( 7 ) exhibited anti - ache activity with an ic50 value of 13.0 m . \n electronic supplementary materialsupplementary material is available for this article at 10.1007/s13659 - 012 - 0026-z and is accessible for authorized users .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: it is usually associated with an increased efflux of cytotoxic drugs by atp - binding cassette ( abc ) transporters including abcg2 . \n abc transporters are energy - dependent transporters that normally function in the detoxification and protection of normal cells from xenobiotics . \n overexpression of abcg2 in cancer cell lines in vitro has been shown to confer mdr to a variety of anticancer drugs including mitoxantrone , irinotecan , methotrexate , flavopiridol , and anthracyclines . moreover \n , growing evidence suggests that abcg2 underlies the mdr of clinical samples from different cancers . \n it may also contribute to the maintenance of stem or progenitor cells as a survival factor , ultimately driving tumor growth . \n identification of factors that can influence abcg2 expression and activity might lead to the development of new strategies to modulate abcg2-mediated drug transport clinically . \n they can be thought of as inhabiting within a complex milieu of normal cells , blood vessels , endogenous small molecules , and secreted factors , which collectively comprise the tumor microenvironment . \n recent advances have indicated that the tumor microenvironment is critically important for cancer initiation , progression , metastasis , and drug resistance , thus providing opportunities for therapeutic intervention . \n this study aims to evaluate the regulation of abcg2 in response to a few characteristic growth conditions within the tumor microenvironment , including hypoxia , glucose deprivation , and low ph . \n cancer cells are often confronted with a remarkable reduction in oxygen supply inside solid tumors , leading to intratumoral hypoxia . \n hypoxia - inducible factor ( hif-1 ) , the master regulator of hypoxic response , is accumulated in order to adapt to the adverse growth condition . \n the production of vascular endothelial growth factor ( vegf ) and other hypoxia - induced angiogenic cytokines to promote increase tissue vascularization , and the metabolic switch from oxidative to glycolytic metabolism represent the two major adaptive responses to enhance cell survival under tumor hypoxic condition . \n glucose depletion is often observed in malignant tumors due to insufficient blood supply in the core of solid tumors and increased rate of glycolysis . on the other hand , \n the production of lactic acid under anaerobic conditions and the hydrolysis of atp in an energy - deficient environment contribute to the acidic microenvironment that has been shown in many types of tumor ( warburg 's effect ) . \n while the effect of tumor microenvironment on oncogenes and tumor suppressor genes have been extensively studied recently , less is known about how this may modulate the xenobiotic transporters thus mediating multidrug resistance . \n a better understanding of this complicated interaction may allow us to design novel therapeutic strategies to improve treatment outcome in chemotherapy . \n mitoxantrone and 2-deoxyglucose ( dg ) were purchased from sigma chemical ( st . louis , mo ) . \n pheophorbide a ( pha ) and fumitremorgin c ( ftc ) were kind gifts obtained from dr . \n susan bates ( national cancer institute , nih , bethesda , md , usa ) . \n human colon carcinoma hct-116 , and s1 and its abcg2-overexpressing resistant s1m1 - 80 cell lines were kindly provided by dr . \n the resistant s1m1 - 80 subline used in this study was derived from the s1 clone of the ls174 human colon cancer cell line by prolonged selection in mitoxantrone , which was routinely maintained at 80 m mitoxantrone . \n there is no amplification of the abcg2 gene in s1m1 - 80 . the abcg2-mediated drug resistance in s1m1 - 80 is believed to be contributed by a balanced t(4,7 ) translocation downstream of the abcg2 gene and the escape from mir-519c repression by a shortening of its 3untranslated region . \n at the time of investigation , it has been allowed to grow in drug - free medium for at least 3 weeks . \n the cell lines were maintained in rpmi 1640 medium supplemented with 10% fetal bovine serum , 100 units / ml streptomycin sulfate , and 100 units / ml penicillin g sulfate , and incubated at 37c in 5% co2 . \n hypoxic incubation was accomplished by using the bactron anaerobe system ( sheldon manufacturing inc . , \n prior to hypoxic incubation , the culture was replenished with fresh complete medium and the cells were then subjected to 3 cycles of evacuation refilling of anaerobic gas ( 90% nitrogen and 10% co2 ) . \n the hypoxic environment was proved to be radiobiologic hypoxic ( oer = 3.0 , data not shown ) . to mimic glucose depletion , \n cells were cultured in glucose - free rpmi 1640 medium for 16 h or in regular medium but in the presence of 20 mm 2-deoxyglucose ( 2-dg ) for 16 h. under these conditions , no significant cell death was observed . to test the effect of acid , \n the glucose - containing culture medium was titrated with 0.1 m hcl to ph 5 . in order to ensure that any observed change in the transporter expression was not attributed by a change in medium osmolality , \n control cells were also grown in glucose - containing rpmi-1640 with an added volume of distilled water to achieve an osmolality equivalent to the acid - treated medium ( ~20% v / v of added water ) . \n total rna was isolated using the trizol regent ( invitrogen , carlsbad , ca ) . \n rna ( 1 g ) was reverse transcribed using the transcriptor high fidelity cdna synthesis kit ( roche applied science , indianapolis , in ) . \n quantitative real - time pcr was performed to quantify the change of abcg2 or pgk1 transcript expressions using the kapa sybr fast qpcr kit ( kapabiosystems , woburn , ma ) in a lightcycler 480 instrument i ( roche applied science , indianapolis , in ) . the human gapdh rna was amplified in parallel as the internal control . \n the specific primers used are as follows : abcg2 ( forward ) 5-tttccaagcgttcattcaaaaa-3 ( reverse ) 5-tacgactgtgacaatgatctgagc-3 ; pgk1 ( forward ) 5-tttctaacaagctgacgctg-3 , ( reverse ) 5-ttcttcctccacatgaaagc-3 ; gapdh ( forward ) 5-agccacatcgctcagacac-3 ( reverse ) 5-gttcaaacttctgctcctga-3. pcrs were performed at 95c for 5 min , followed by 50 cycles of 95c for 10 s and 60c for 10 s. fluorescence signal was acquired at the end of the elongation step of every pcr cycle ( 72c for 10 s ) to monitor the increasing amount of amplified dna . \n ct was calculated by subtracting the ct of gapdh from the ct of the transcript under investigation . \n ct was then calculated by subtracting the ct of the untreated cells ( or parental cells ) from the ct of the treated cells ( or resistant cells ) . \n fold change of gene expression was calculated by the equation 2 . a small interference hairpin - loop ( sh ) silencing vector against the human abcg2 ( pu6-abcg2 ) and a negative control shrna vector targeting firefly luciferase ( pu6-luc ) were prepared as described in to et al . . \n s1 and s1m1 - 80 cells were transfected with pu6-luc or pu6-abcg2 using lipofectamine 2000 ( invitrogen , grand island , ny ) . \n a series of human abcg2 promoter constructs with progressive deletions at the 5-end has been described previously . \n the abcg2 promoter / firefly luciferase fusion genes ( 400 ng dna ) were transfected in hct-116 or s1 cells on 24-well plates using fugene 6 ( roche , indianapolis , in ) . \n the pgl3-basic ( promoterless ) plasmid , encoding firefly luciferase ( promega , madison , wi ) , was used to determine the basal levels . in each experiment , the phrg - basic plasmid ( 100 ng ) , encoding renilla luciferase ( promega ) , was cotransfected for normalization purposes . \n luminescence was measured 48 h after transfection using the dual - luciferase reporter assay system ( promega ) with the glomax 20/20 luminometer ( promega ) . in experiments involving 2-dg or hypoxic \n treatment , the cells were treated at 24 h after transfection for 16 h. reporter activity was normalized by calculating the ratio of firefly / renilla values . \n results were expressed as mean sd of duplicate measurements from three independent transfections . \n the abcg2-mediated efflux activity in hct-116 , s1 and s1m1 - 80 cells with or without pretreatment with 2-dg or hypoxia were determined by flow cytometric assays as described previously . \n trypsinized cells was incubated in 1 m pha with or without 10 m abcg2-specific inhibitor ftc in complete medium ( phenol red - free rpmi 1640 with 10 % fbs ) at 37c in 5% co2 for 30 min . \n subsequently , the cells were washed with cold complete medium and then incubated for 1 h at 37c in pha - free medium continuing with ftc to generate the ftc / efflux histogram , or without ftc to generate the efflux histogram . \n the ftc - inhibitable pha efflux was determined as the difference in mean fluorescence intensity ( mfi ) between the ftc / efflux and efflux histograms , which indicates the abcg2-mediated transport activity . \n cells were finally washed with cold dulbecco 's pbs and placed on ice in the dark until analysis by flow cytometry . \n to measure the cell surface abcg2 expression , trypsinized cells were incubated in 2% bovine serum albumin / dulbecco 's pbs with either phycoerythrin - conjugated anti - abcg2 antibody 5d3 ( ebioscience , san diego , ca ) or phycoerythrin - conjugated mouse \n igg2b negative control antibody ( ebioscience ) according to manufacturer 's instructions for 30 min at room temperature . \n surface expression of abcg2 was calculated as the difference in mean channel numbers between the 5d3 antibody histogram and the negative control antibody histograms . \n samples were analyzed on an lsrfortessa cell analyzer ( bd biosciences , san jose , ca ) . \n phycoerythrin fluorescence was detected with a 488 nm argon laser and a 585 nm bandpass - filter , whereas pha fluorescence was detected with a 488 nm argon laser and a 670 nm bandpass filter . \n cell debris was eliminated by gating on forward versus side scatter and dead cells were excluded based on propidium iodide staining . \n growth inhibitory effect of mitoxantrone and cisplatin ( representing a typical abcg2 substrate and nonsubstrate , resp . ) , with or without the pretreatment of 2-dg or hypoxia , on hct-116 , s1 and s1m1 - 80 cells , were evaluated by the sulforhodamine b assay . \n cells were seeded into 96-well microtitre plates in 100 l at a plating density of 5,000 cells / well and allowed to incubate overnight . \n the cells were then exposed to 2-dg ( 20 mm ) or hypoxia for 16 h , before being treated with mitoxantrone or cisplatin at a range of concentrations and allowed to incubate at 37c in 5% co2 for another 72 h. each drug concentration was tested in quadruplicate and controls were tested in replicates of eight . \n each experiment was carried out independently at least three times . to determine whether differences between ic50 values were significant , \n the student 's t - test was performed with p < 0.05 being considered significant . \n soft agar plates were prepared in six - well plates with a bottom layer of 0.6% noble agar in serum - free rpmi 1640 . \n 1,000 cells , with or without a 24 h hypoxia treatment , were suspended in 0.3% noble agar in rpmi 1640 supplemented with 10% fetal bovine serum and seeded onto the bottom layer . \n the number of colonies was counted after staining with 0.05% crystal violet for 1 h. \n the abcg2 mrna expression was evaluated in three human colon carcinoma cell lines ( hct-116 , s1 and its resistant subline s1m1 - 80 ) after the pretreatment with glucose depletion , 2-dg , acidic ph and hypoxia . in hct-116 and s1 , a small but significant upregulation of abcg2 ( 2 to 4-fold ) \n since the effect was more pronounced after 2-dg and hypoxia treatment , these conditions were selected for later functional and more detailed mechanistic studies . on the other hand , \n the expression of pgk1 , a hypoxia responsive gene , was also measured to verify the hypoxia treatment . \n our data indicates that pgk1 was remarkably induced by the hypoxia treatment in all three cell lines ( figure 1 , lower panel ) . \n the increase in abcg2 mrna after 2-dg or hypoxia treatment was blocked by actinomycin d ( 5 g / ml ; data not shown ) , suggesting that these tumor microenvironment conditions regulate abcg2 mrna levels through an effect on abcg2 transcription . a series of 5-deletion reporter gene constructs harboring the abcg2 promoter region with the 3 end terminating at + 396 bp were used to elucidate the promoter region responsible for glucose depletion or hypoxia - mediated abcg2 upregulation ( figure 2(b ) ) . \n after transiently transfected into hct-116 and s1 cells , all the abcg2 promoter - luciferase constructs demonstrated good activities above the promoter - less pgl3basic background ( data for hct-116 and s1 are shown in figure 2(b ) and supplementary figure 1 ( available online at doi:10.5402/2012/746025 ) , resp . ) . \n while reporter activity of the full length abcg2 promoter construct ( 1662/+396 ) and the longer ones ( 628 and 312/+396 ) were found to be activated in hct-116 and s1 pretreated with 2-dg or hypoxia , the activation was abolished in the shortest construct ( 105/+396 ) . \n the data implies that the region between 312 and 105 , harboring two reported hif-1 response elements ( figure 2(a ) ) , is responsible for the observed induction of abcg2 transcription . \n the upregulation of abcg2 observed above suggested that all of the tumor microenvironment conditions investigated may confer a more drug - resistant phenotype . \n next we sought to demonstrate the functional evidence with special regard to drug efflux activity and in vitro drug response . by flow cytometric analysis , both 2-dg and hypoxia treatment led to a small but statistically significant elevation of abcg2-mediated drug efflux ( mfi = signal between the efflux and ftc / efflux histograms ) in all cell lines tested ( figure 3 ) . since the basal abcg2 levels in s1 and hct-116 cells are very low , we did not proceed to measure the protein expression either by western blot or surface staining by flow cytometry . \n nonetheless and more importantly , 2-dg and hypoxia were found to significantly decrease the cytotoxic effect of the abcg2 substrate anticancer drug ( mitoxantrone ) ( table 1 ) . on the other hand , \n the cytotoxicity of cisplatin was not affected , indicating the specific effect of these tumor microenvironment conditions on abcg2-mediated mdr ( table 1 ) . in the abcg2-overexpressing resistant s1m1 - 80 cells , western blot and flow cytometry - based cell surface staining analysis \n interestingly , by both assays , abcg2 protein expression was also found to be induced slightly after 2-dg and hypoxia treatment ( figures 4(a ) and 4(b ) ) . in figure 4(b ) ( flow cytometry cell surface staining ) , the channel # between the specific anti - abcg2 staining and the isotype antibody control represents the cell surface abcg2 protein expression . \n importantly , this slight induction of abcg2 protein by 2-dg and hypoxia was also found to decrease the cytotoxic effect of mitoxantrone ( an abcg2 substrate anticancer drug ) ( table 1 ) , probably through enhancing the abcg2 efflux activity ( figure 3 ) . in an attempt to further understand the induction of abcg2 protein expression by the 2-dg and hypoxia treatment , cycloheximide chase assay \n was performed to evaluate the degradation rate of abcg2 protein ( data not shown ) . \n unfortunately , abcg2 protein was found to be fairly stable and no appreciable degradation was observed with or without the 2-dg or hypoxia treatment after up to 36 h. anchorage - independent growth is one of the hallmarks of cell transformation , which is considered a reliable in vitro assay for detecting malignant transformation of cells . since abcg2 upregulation was observed after 2-dg and hypoxia treatment and that abcg2-mediated mdr may lead to a more malignant phenotype , the colony formation capability of s1 and s1m180 cells with or without 2-dg and hypoxia treatment was assessed . in both cell lines , \n hypoxia was found to significantly increase the colony formation capability ( figure 5(b ) ) . in s1m1 - 80 cells , \n 2-dg treatment was also found to enhance the tendency to form colony in a statistical significant manner . \n interestingly , the colony formation capability could be abolished when abcg2 was knocked down in the cells ( figures 5(a ) and 5(b ) ) . \n the concept of tumor microenvironment is gaining a lot of attention in recent years because it allows complex interaction between a tumor and its surrounding normal cells , molecules , and blood vessels , which all participate in tumor progression . \n a tumor can change its microenvironment and the microenvironment can affect the way the tumor grows and spreads . \n hypoxia , glucose deprivation , and acidosis are generally considered as the hallmarks of the tumor microenvironment . \n while chemotherapy is the mainstay of treatment strategy for cancer , drug resistance is seriously hindering the clinical efficacy of most anticancer drugs . \n solid tumors exhibit distinct structural abnormalities such as the leaky vasculature and thus have poor tissue perfusion , subsequently leading to physiological resistance to anticancer drugs . on the other hand , \n resistant cancer cells often express high levels of the multidrug resistance ( mdr ) transporters as their biochemical mechanism of drug resistance . \n they make use of the energy derived from atp hydrolysis to regulate intracellular drug concentration , thereby determining cell sensitivity to chemotherapeutic agents . \n abcg2 is one of these transporters , conferring mdr to a broad spectrum of antitumor agents . with high normal tissue expression in the brain endothelium , gastrointestinal tract , and placenta \n , abcg2 is believed to be important in the protection from xenobiotics , regulating oral bioavailability , forming part of the blood - brain barrier , and the maternal - fetal barrier . \n importantly , abcg2 may also contribute to the maintenance of stem or progenitor cells as a survival factor . among the various conditions within the tumor microenvironment \n this study attempted to understand the regulation of abcg2 by the various adverse growth conditions within the tumor microenvironment in colon carcinoma cell lines . \n in fact , it has been postulated that the atp - dependent drug efflux may be reduced under hypoxic condition . \n cells are known to switch from oxidative to glycolytic metabolism under hypoxia and glycolytic pathway is less efficient in generating atp . to this end \n , it has been shown that a limitation in atp supply ( by sodium azide or 2-dg ) could inhibit daunorubicin efflux in a non - pgp - overexpressing resistant cell line . \n xu et al . demonstrated that the inhibition of mitochondrial respiration leads to an increase in the cytotoxic efficacy of various chemotherapeutic drugs as a result of a reduced p - gp activity which was attributed to a depletion of atp . on the other hand \n , it has been shown that acidosis ( especially together with hypoxia ) could increase p - gp - mediated drug efflux and lead to reduced cytotoxicity of chemotherapeutic drugs . \n the discrepancy between the studies may be caused by the different cell lines used and different extent of hypoxia treatment . \n the data revealed that hypoxia , glucose deprivation , and acidosis can all upregulate abcg2 in the two parental colon cancer cell lines ( figure 1 ) and apparently leading to multidrug resistance ( table 1 ) . \n the upregulation is likely mediated at the transcription level because it can be abolished by pretreatment with actinomycin d , which inhibits rna synthesis ( data not shown ) . \n the effect on an abcg2-overexpressing resistant s1m1 - 80 cell line was much less pronounced at the rna level ( figure 1 ) . \n however and interestingly , the resistance level was further enhanced in s1m1 - 80 cells after 2-dg and hypoxia treatment ( table 1 ) . \n since hypoxic condition will commit the cells to glycolytic metabolism , a downstream glycolytic gene ( pgk1 ) was also evaluated in our study ( figure 1 ) as a surrogate for the hypoxic treatment and to demonstrate the functional linkage with glucose metabolism . \n as expected , pgk1 level was elevated remarkably after the hypoxia treatment . to evaluate the effect of glucose depletion , \n cells were grown either in glucose - free medium or in glucose - containing medium supplemented with 2-dg . \n 2-dg is a glucose analogue , which acts as a competitive inhibitor of glucose metabolism , where 2-dg competes with d - glucose to be transferred to the cells as a target for the hexokinase in the first step of the glycolytic pathway . upon transport into the cells , glucose and 2-dg are phosphorylated to glucose - po4 and 2-dg - po4 , respectively , by the hexokinase . \n however , unlike glucose - po4 , the 2-dg - po4 can not further be metabolized by the glucose phosphate isomerase ( gpi ) . \n in addition , 2-dg could inhibit gpi , and then downregulate its downstream glycolysis - related genes ( such as pgk1 ) , which agree well with our findings ( figure 1 ) . regarding the effect of acidosis , the upregulation of abcg2 was only statistically significant in hct-116 cells . \n since abcg2 upregulation is more pronounced after 2-dg and hypoxia treatment , further mechanistic and functional studies were performed under these conditions . by reporter gene assay , the abcg2 promoter region ( 312 to 105 ) close to the reported hif-1 response element was found to be required for our observed gene upregulation ( figure 2(b ) ) . \n consistent with the reduced response of cells to abcg2 substrate anticancer drug ( table 1 ) after 2-dg and hypoxia treatment , abcg2 efflux activity was found to be increased in all cell lines tested by the flow cytometry ( figure 3 ) . since mdr \n is usually mediated by overexpression of the mdr transporters , we intentionally investigated how abcg2 regulation is affected by 2-dg and hypoxia in the abcg2-overexpressing resistant s1m1 - 80 cell line . \n s1m1 - 80 has massive abcg2 overexpression probably caused by a genetic translocation and its escape from mir-519c repression reported previously . \n abcg2 protein expression was detected by western blot analysis and cell surface staining . by both assays , 2-dg and hypoxia treatment \n were found to increase abcg2 level ( figure 4 ) , which is consistent with the elevated efflux activity ( figure 3 ) and reduced response to abcg2 substrate anticancer drug ( table 1 ) . \n note that the massively overexpressed abcg2 transcript level in s1m1 - 80 was not significantly affected by these treatments ( table 1 ) ; therefore , the observed functional effect may have been caused by modulation at the posttranscriptional and/or translational levels . \n this piece of data is important because it suggests that tumor microenvironment could coordinately regulate abcg2 with preexisting mdr mechanisms to attain a higher resistance level . \n the ability of cancer cells to form colonies in vitro in an anchorage - independent manner is commonly used as a measure of their malignancy . \n after 2-dg and hypoxia treatment , both s1 and s1m1 - 80 cells were found to form more colonies in a soft agar colony formation assay ( figure 5 ) . \n the upregulation of abcg2 may play a specific role in this enhanced colony formation capability because genetic knockdown of abcg2 was found to significantly reduce the number of colonies formed ( figure 5 ) . \n taken together , the three adverse growth conditions in the tumor microenvironment tested were found to upregulate abcg2 to mediate multidrug resistance , with the possible enhancing effect on preexisting mdr mechanism , and to give rise to a more malignant phenotype . \n novel clinical interventions may be devised to help circumvent this abcg2-mediated resistance by modulating the tumor microenvironment .", "role": "user" }, { "content": "abcg2 is an efflux transporter commonly found to overexpress in multidrug resistant ( mdr ) cancer cells . \n it is also believed to be a survival factor for cancer stem cells to drive tumor growth . \n tumor microenvironment represents an attractive new drug target because it allows complex interaction between a tumor and its surrounding normal cells , molecules , and blood vessels , which all participate in tumor progression . \n hypoxia , glucose deprivation and acidosis are the hallmarks of tumor microenvironment . \n this study investigated the upregulation of abcg2 by these adverse growth conditions within the tumor microenvironment . \n reporter gene assay revealed that a region within the abcg2 promoter close to the reported hif-1 response element is responsible for abcg2 upregulation . increased abcg2 efflux activity \n was observed under the same conditions , subsequently leading to reduced response to abcg2 substrate anticancer drug . \n importantly , glucose deprivation and hypoxia were also found to enhance the resistance level of abcg2-overexpressing resistant cells with pre - existing genetic and epigenetic mdr mechanisms . \n hypoxia was further demonstrated to cause a more malignant anchorage - independent growth phenotype in the resistant cells , which can be abolished by knocking down abcg2 . \n a better understanding of abcg2 regulation by the tumor microenvironment may help design novel strategies to improve treatment outcome .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: the in - vivo study on ophthalmic products is done on experimental animals , usually on rabbit 's eye . \n preparation volume 50 l is instilled topically in the center of the lower cul - de - sac . \n eyes of the animal are observed for signs of irritation and injury . in vitro methods \n diclofenac sodium solid lipid nanoparticles ( slns ) , prepared with goat fat and phospholipids evaluated using bio - engineered human cornea , produced from immortalized human corneal endothelial cells ( henc ) , stromal fibroblasts , and epithelial cells . \n sustained release and high permeation of diclofenac sodium through the bio - engineered cornea were achieved . \n the bovine corneal opacity and permeability ( bcop ) assay has been accepted as a valid in - vitro alternative method to the draize eye irritation test with the development of an improved laser light - based opacitometer ( llbo ) for the analysis of the complete corneal surface instead of dual beam op - kit opacitometer . \n confocal laser scanning microscopy study on the transport mechanism through the corneal epithelium of nanostructured lipid carrier was carried out in rabbits in vivo . \n the corneas were isolated and rinsed in physiological saline to remove the adhesive fluorescence fragment . \n after 30 minute administration in to eyes , the interaction with corneal epithelia to be investigated by observing the corneal samples using confocal laser scanning microscopy ( clsm ) . \n the epiocular tissue model is a new in - vitro test method for ocular irritation , produced by mattek corporation tissue production facilities . \n the epiocular tissue model shows in - vivo like morphological and growth characteristics that are uniform and highly reproducible . \n the tissue consists of highly organized basal cells , which progressively flatten out as the apical surface of the tissue similar to the normal in - vivo corneal epithelium . \n this test could be applied as an alternate to rabbit eye test in the toxicological assessment of ophthalmic preparations . \n the in - vivo study on ophthalmic products is done on experimental animals , usually on rabbit 's eye . \n preparation volume 50 l is instilled topically in the center of the lower cul - de - sac . \n eyes of the animal are observed for signs of irritation and injury . in vitro methods \n diclofenac sodium solid lipid nanoparticles ( slns ) , prepared with goat fat and phospholipids evaluated using bio - engineered human cornea , produced from immortalized human corneal endothelial cells ( henc ) , stromal fibroblasts , and epithelial cells . \n sustained release and high permeation of diclofenac sodium through the bio - engineered cornea were achieved . \n the bovine corneal opacity and permeability ( bcop ) assay has been accepted as a valid in - vitro alternative method to the draize eye irritation test with the development of an improved laser light - based opacitometer ( llbo ) for the analysis of the complete corneal surface instead of dual beam op - kit opacitometer . \n confocal laser scanning microscopy study on the transport mechanism through the corneal epithelium of nanostructured lipid carrier was carried out in rabbits in vivo . \n the corneas were isolated and rinsed in physiological saline to remove the adhesive fluorescence fragment . \n after 30 minute administration in to eyes , the interaction with corneal epithelia to be investigated by observing the corneal samples using confocal laser scanning microscopy ( clsm ) . \n the epiocular tissue model is a new in - vitro test method for ocular irritation , produced by mattek corporation tissue production facilities . \n the epiocular tissue model shows in - vivo like morphological and growth characteristics that are uniform and highly reproducible . \n the tissue consists of highly organized basal cells , which progressively flatten out as the apical surface of the tissue similar to the normal in - vivo corneal epithelium . \n this test could be applied as an alternate to rabbit eye test in the toxicological assessment of ophthalmic preparations . \n most of the formulation efforts aim at maximizing ocular drug absorption through prolongation of the drug residence time in the eye as well as to slow drug release from the delivery system and minimize pre - corneal drug loss . \n nanostructured platforms provide the effective ways for delivery of insoluble drugs to the target site . \n as these platforms for ophthalmic delivery are very recent and the research continues , formulation factors such as loading of drug , release rate , stability , and therapeutic effectiveness for these needs to be optimized .", "role": "user" }, { "content": "the topically applied drugs as drops are washed off from the eye in very short period , resulting in low ocular bioavailability of drugs . \n number of approaches have been attempted to increase the bioavailability and the duration of action of ocular drugs . \n this review provides an insight into various novel approaches ; hydrophilic nanogels , solid lipid nanoparticles , and nanosponges applied very recently in the delivery of insoluble drugs , prolonging the ocular residence time , minimize pre - corneal drug loss and , therefore , bioavailability and therapeutic efficacy of the drugs . despite various scientific approaches , efficient ocular drug delivery remains a challenge for pharmaceutical scientists .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: clearance of potential cervical spine ( c - spine ) injury in the awake and cooperative patient with no distracting injury is a standardized procedure . \n the process becomes more problematic in the unconscious patient , leading to delay in c - spine collar removal and consequent complications such as tissue necrosis , raised intracranial pressure , excessive sedation and so on . \n two recent reports addressed this issue . in the first of these a postal questionnaire \n was sent to 32 neurosurgery and spinal injury departments in the uk , with the aim of determining how they assessed the c - spine in unconscious , adult trauma patients , and at what point immobilization was discontinued . \n the majority of the units questioned had no formal protocol for either screening investigations or criteria for discontinuation of c - spine immobilization . \n out of 27 units , 12 used two x - ray views alone , and only 10 out of 27 units routinely used computed tomography ( ct ) scanning . \n one unit used magnetic resonance imaging routinely and two used dynamic fluoroscopy . following negative imaging of one variety or other , 12 units discontinued immobilization immediately , 10 continued until they were able to clear spines clinically , and the remaining five were prepared to discontinue if the patient 's condition required it . \n over half of the patients had immobilization discontinued on the basis of plain x - rays alone , despite evidence that plain x - rays have poor diagnostic sensitivity for c - spine fractures [ 3 - 5 ] and are inferior to ct . \n the results suggest that there is often suboptimal and inconsistent investigation , with a subsequent lack of rationale for discontinuation of immobilization . \n it is suggested that head injured patients receiving a ct scan of the brain should routinely undergo c - spine ct scanning at the same time , and that magnetic resonance imaging and dynamic fluoroscopy are not necessary in these patients . \n the second article , also employing a postal questionnaire ( 95% response ) , looked at the major differences between clinicians of differing specialities in the management of potential c - spine injuries in unconscious adult patients . \n the report also reviews the available literature and goes on to suggest a management protocol . \n consistent with the findings of jones and coworkers , morris and mullan demonstrated great variations in practice , attitudes and perceptions with regard to management and evaluation of the potentially injured c - spine in the unconscious patient . \n no consensus existed as to the minimum standard of investigations required to clear the c - spine in these circumstances . \n recognition of the complications of prolonged c - spine immobilization in the critically ill patient was also patchy , with some clinicians suggesting that immobilization should be indefinite until clinical examination could be carried out in the awake patient . based upon a literature review and available consensus , the working group devised a protocol for the investigation and subsequent clearing , or not , of the c - spine in an unconscious patient . \n essentially , the protocol requires three - view x - rays of the c - spine , an anteroposterior and a lateral thoracolumbar x - ray , and a high - resolution ct of the craniocervical junction . \n this must be a step in the right direction , providing an evidence - based approach to an all too common dilemma in the intensive care unit ( icu ) . \n a randomized controlled trial , recently reported in jama , aimed to resolve the tricky issue of whether prone positioning for acute hypoxaemic respiratory failure improves patient survival . \n the trial involved 21 icus with a total of 791 patients being assigned to one of two groups : continually supine or intermittently prone . \n patients were eligible if they were intubated ( or tracheostomized ) with an arterial oxygen tension ( pao2)/fractional inspired oxygen ( fio2 ) ratio < 300 mmhg and had an expected duration of ventilation in excess of 48 hours . \n patients were excluded if they were at risk for harm from the prone positioning ( e.g. raised intracranial pressure ) or had recently been ventilated in the prone position . \n patients in the supine group were managed entirely in that position with a 30 head up tilt . \n patients assigned to the prone group were placed in a complete prone position for at least 8 hours per day . however , \n if a patient developed severe hypoxia in the supine position , then they could crossover to the prone group . \n if a major complication attributable to prone position occurred , then the patient was reverted to the supine position . sadly 28-day mortality rates were not significantly different between the two groups ( 31.5% for supine versus 32.4% for prone ) . \n ninety day mortality was also similar ( 42.2% for supine versus 43.3% for prone ) . \n however , ventilator associated pneumonia was found to be significantly reduced in the prone group . \n the authors acknowledged several limitations in their trial , including significant treatment group crossover and a failure rate in excess of 25% in the prone positioning protocol . \n however , the trial failed to show a reduction in mortality after early prone positioning for acute hypoxaemic failure , despite an improvement in oxygenation and a reduction in ventilator associated pneumonia ; also , it did demonstrate an increased incidence of serious side effects from ventilation in the prone position . \n this report echoes the findings of gattinoni and coworkers and begs the question , is there any evidence to justify prone ventilation ? \n the november issue of critical care medicine included reports on two retrospective studies based on the prowess ( recombinant human activated protein c worldwide evaluation in severe sepsis ) study data base , and discussed new post-28-day data . in the first of these reports , outcome beyond 28 days and health care resource utilization , employing the simplified therapeutic intervention scoring system , were examined . \n previous investigators have concluded that the cost - effectiveness of activated protein c ( apc ) in the management of severe sepsis compares favourably with that of other health care interventions , despite its high up - front cost . \n however , these findings were largely based on 28-day data , and as such they do not take the entire hospital stay into account . \n laterre and coworkers point out that , at 28 days , more than 40% of the prowess survivors were still in hospital and little is known of their health care consumption from there on . \n survival from severe sepsis is significantly better at hospital discharge for those treated with apc than for those not treated with apc , and this remained the case for the majority of subgroups . \n furthermore , this increase in survival was not associated with an increase in resource consumption , as measured using the therapeutic intervention scoring system , or in terms of icu and hospital length of stay . \n in addition , a greater number of survivors in the apc group than in the placebo group were discharged directly home . not happy with \n follow up to hospital discharge , angus and coworkers collected long - term survival data up to 3.5 years after prowess . \n they aimed to determine the effect of apc on hospital survival ( as described above ) and then to investigate the ensuing long - term survival and to analyze these results for selected subgroups based on age , premorbid functional dependency , acute physiology and chronic health evaluation ( apache ) ii score and number of organ dysfunctions , all of which are known to have prognostic significance in severe sepsis . sadly , despite survival to hospital discharge being better in those treated with apc , rates of overall median survival and survival at 3 months , 6 months , 1 year and 2.5 years were not significantly different between the apc and placebo groups . \n this finding was consistent for subgroup analyses other than for apache ii scores . in post \n hoc analysis , patients with an initial apache ii score in excess of 25 had a survival benefit when treated with apc . \n retrospective , cross - sectional observations , use of post hoc analysis and small sample sizes that are not powered for the analysis in question must temper over interpretation . \n however , it is unlikely that a new long - term prospective trial studying the effect of apc in severe sepsis is now possible , and we must therefore tailor our clinical practice with these findings in mind . \n in the past few months the journals , as has become the norm , have been peppered with studies investigating noninvasive ventilation ( niv ) in the management of acute respiratory failure ( arf ) of varying aetiology . \n the immunocompromised patient presenting with arf has been thought to carry a very poor prognosis , and as such intensive care physicians have been reluctant to institute invasive respiratory support . \n however , recent studies have demonstrated the efficacy of niv in this patient population , providing us with an alternative to endotracheal intubation . carrying this concept forward , \n rocco and colleagues reported a case control study comparing niv delivered via traditional face mask versus the much discussed helmet interface for immunocompromised patients with hypoxaemic arf and fever . accepting \n that numbers in this study were small and that patients in the helmet group were matched with historical control individuals receiving facemask niv , both groups demonstrated a similar improvement in pao2/fio2 ratio , but importantly the helmet group demonstrated a more sustained improvement . \n this was probably due to a lower incidence of complications , leading to patient intolerance of the device . \n a further study appearing in chest compared niv with invasive intubation and ventilation for this patient group . \n this was a retrospective study using a pair - wise matching system to compare the two treatment modalities and logistic regression analysis to identify factors affecting in hospital mortality . \n conclusions need to be drawn carefully because the study population was quite heterogeneous with varying types of malignancy and differing aetiologies to the respiratory failure . \n despite this the authors identified increasing severity of illness and a diagnosis of acute myeloid leukaemia as markers of poor outcome , whereas being female , intubated in the first 24 hours and the presence of recent positive blood cultures were markers of good outcome . \n make of this what you will . to end , we return to critical care medicine , december issue . \n a prospective randomized trial suggested that continuous positive airway pressure ( cpap ) and/or bilevel positive airway pressure ( bipap ) is superior to oxygen therapy alone in acute cardiogenic pulmonary oedema . \n specifically , cpap and bipap resulted in much improved pao2/fio2 ratios and a lower incidence of endotracheal intubation . \n however , there was no significant difference in mortality at hospital discharge between the groups . \n apache = acute physiology and chronic health evaluation ; apc = activated protein c ; arf = acute respiratory failure ; bipap = bilevel positive airway pressure ; cpap = continuous positive airway pressure ; ct = computed tomography ; fio2 = fractional inspired oxygen ; icu = intensive care unit ; niv = noninvasive ventilation ; pao2 = arterial oxygen tension . \n ", "role": "user" }, { "content": "' every day ' clinical conundrums are all too infrequently addressed in the mainstream literature , but in the past few months two reports attempted to tackle the thorny problem of the occult cervical spine injury on the intensive care unit . \n are we approaching the death knell for prone ventilation , and how much more can we squeeze out of the prowess study ? \n also , we must of course mention noninvasive ventilation .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: nuclear receptors ( nrs ) constitute a unique family of ligand - modulated transcription factors . \n nrs mediate cellular response to small lipophilic endogenous and exogenous ligands [ 1 , 2 ] and are responsible for sensing a number of hormones , including steroid and thyroid hormones , and act as positive and negative regulators of the expression of specific genes [ 35 ] . \n therefore , nrs play a central role in many aspects of mammalian development , as well as lipid homeostasis , physiology , and metabolism . \n nrs make up one of the most abundant classes of transcriptional regulators in the body and have emerged as promising pharmaceutical targets . \n classically , nrs consist of several functional domains , that is , a variable n - terminal ligand - independent transactivation domain ( which often exhibits a constitutive transcription activation function ( af-1 ) ) , a highly conserved dna - binding domain ( dbd ) that contains two zinc fingers , a hinge domain ( a variable linker region ) , and a multifunctional c - terminal domain . \n furthermore , the c - terminal domain includes the ligand binding ( lbd ) , the dimerization interface , and the ligand - dependent transactivation domain af-2 [ 1 , 6 ] . small heterodimer partner ( shp ; nr0b2 for nuclear receptor subfamily 0 , group b , member 2 ; mim number 604630 , 601665 ) is a member of the mammalian nr superfamily , due to the presence of a putative ligand - binding domain ( lbd ) . \n shp functions as a corepressor through heterodimeric interaction with a wide array of nuclear receptors and repressing their transcriptional activity . \n shp achieves its goal via several members of the nr superfamily that are able to regulate shp expression . \n however , shp is also a unique and atypical nr because it lacks the classical dna - binding domain ( dbd ) , generally present in other nrs . \n the nr0b family of nrs consists of 2 orphan receptors : shp and dax-1 ( dosage - sensitive sex reversal adrenal hypoplasia congenita ( ahc ) critical region on the x chromosome , gene 1 ) . \n dax1 is a gene whose mutation causes the x - linked adrenal hypoplasia congenita and is the only family member that lacks a conventional dbd . \n dax-1 ( nr0b1 ) is therefore seen as the closest relative of shp in the nr superfamily [ 1012 ] . \n whereas shp is different from other conventional nrs both structurally and functionally , it acts as a ligand - regulated receptor in metabolic pathways . \n shp belongs to the orphan subfamily since there is no known ligand for this receptor , except for some retinoid - related molecules . \n shp inhibits transcriptional activation by working on several other nuclear receptors , that is , directly modulating the activities of conventional nuclear receptors by acting as an inducible and tissue - specific corepressor [ 12 , 15 ] . \n the discovery of shp dates back to 1996 ; since then , this orphan nr has been identified as a key transcriptional regulator of signaling pathways [ 8 , 16 ] involving fundamental biological functions and metabolic processes . \n such processes include cholesterol , bile acid and fatty acid metabolism , glucose and energy homeostasis , and reproductive biology . \n experiments performed by fluorescence in situ hybridization ( fish ) analysis of the human metaphase chromosome have shown that shp is found at a single locus on chromosome 1 at position 1p36.1 and consists of two exons and a single intron spanning approximately 1.8 kb with 257 amino acids in humans . in mice and rats , \n shp resides on chromosomes 4 and 5 , respectively , both consisting of 260 amino acids . \n shp expression is predominantly observed in the liver [ 10 , 18 ] , but it is also detected at lower levels in other tissues , including the pancreas , spleen , small intestine , colon , gallbladder , kidney , adrenal gland , ovary , lung , brainstem , cerebellum , heart , and thymus ( table 1 ) [ 1921 ] . \n shp is indeed able to repress the transcriptional activities of its target nrs and transcriptional regulators through two functional leu - xaa - xaa - leu - leu- ( lxxll- ) like motifs [ 2224 ] . \n such motifs appear to be essential for the interaction with the ( activation function 2 ) af-2 domains of several sets of nrs [ 22 , 23 ] . \n the human shp is enriched by another 12 amino acids [ 2536 ] , and this region between helix 6 and 7 is also involved in the repression of the transactivation of nrs . \n since shp lacks dna - binding domain , it exerts the inhibitory effects through protein - protein interaction . \n shp expression seems to follow a circadian rhythm in the liver , involving the clock - bmal1 pathway and suggesting that some of the regulatory functions of shp and deriving functions must be temporal [ 19 , 20 , 38 ] . \n gene expression of shp is regulated by several factors including nrs , transcription factors , and a number of additional conditions and substances , as extensively reported in table 2 . \n also , the central role of shp is clear since this nr is able to act as a coregulator for wide range of targets , namely , nrs / transcription factors / transcriptional coregulators and few different molecules , as depicted in table 3 . in general \n , shp acts as a repressor of the transcriptional activity of the specific interacting partner ( via lbd of the partner and nr boxes of shp ) [ 12 , 3943 ] . \n however , it is also demonstrated that shp is able to upregulate gene transcription , as in the case of ppar and ppar [ 4446 ] and nf-b . \n both n - terminal nr interaction domain and c - terminal domain of shp are important for repression [ 47 , 48 ] . \n overall , the shp - mediated repression of target transcription factors occurs by at least three distinct transcriptional repression mechanisms ( figure 2 ) . \n a first mechanism involves direct interference with the af-2 coactivator domain of nrs ( competition for coactivator binding , leading to the repression of nr - mediated transcriptional activity ) . \n this is the case for the inhibition of estrogen receptor ( er ) and estrogen receptor ( er ) . a second mechanism for the shp - mediated repression involves the recruitment of corepressors including direct interactions among mammalian homolog of the saccharomyces cerevisiae transcriptional corepressor sin3p ( msin3a ) , human brahma ( brm ) , switch / sucrose nonfermentable ( swi / snf ) complexes leading to the repression of cholesterol 7-hydroxylase ( cyp7a1 ) ) . \n a third mechanism of inhibition of shp involves the direct interaction with the surface of nr or transcription factor , resulting in the blockade of dna binding and the consequent inhibition of its transcriptional activity . \n this is the case for rar - rxr heterodimers , pxr - rxr binding to dna by shp , interaction with hepatocyte nuclear factor ( hnf4 ) , or jun family of the activator protein 1 ( ap-1 ) transcription factor complex ( jund ) [ 51 , 52 ] . \n all three mechanisms might occur sequentially or alternatively according to type of cells and promoters . \n clearly , information on factors that increase or decrease shp expression and that are regulated by shp is essential for understanding the regulatory effects of this orphan nr . \n interestingly , it is suggested that targeting posttranslational modifications of shp may be an effective therapeutic strategy . \n selected groups of genes could be controlled to cure a vast range of metabolic and shp - related diseases . \n overall , the huge amount of information on shp function is currently available , making this nr essential in a number of functions involving cholesterol and bile acid metabolism , lipogenesis , glucose metabolism , steroid hormone biosynthesis , xenobiotic homeostasis / metabolism , and cell cycle . in particular , the ability of shp in interacting with different metabolic signaling pathways including bile acids and lipid homeostasis , fat mass , adipocytes , and obesity will be reviewed here \n the wide ability of shp to target multiple genes in diverse signaling pathways points to the key role of shp in various biological processes , including the metabolism of bile salts , glucose , and fatty acids . \n studies suggest that loss of shp might positively affect cholesterol and bile acid homeostasis in pathophysiologically relevant conditions . \n bile acids ( bas ) are amphipatic cholesterol metabolites which are synthesized in the liver , secreted into bile , stored in the gallbladder , and secreted postprandially into the duodenum . \n bas are synthesized from cholesterol , and this pathway provides the elimination of excess cholesterol in the body . moreover , bas should be seen as physiological detergents which , in the small intestine , are essential for the absorption , transport , and distribution of lipophilic molecules , including dietary lipids , steroids , and lipid - soluble vitamins . in the intestine , bas undergo extensive metabolism by the intestinal microflora . \n a high efficient system is the enterohepatic circulation of bas [ 55 , 56 ] , where more than 9095% of bas are returned to the liver from the terminal ileum via the portal vein . \n thus , the concentration of bas in serum , liver , and intestine is tightly regulated to prevent damage to enterohepatic tissues due to their strong detergent moiety [ 5759 ] . \n the major rate - limiting step in biosynthetic pathway of bas in humans is initiated by cholesterol 7-hydroxylase ( cyp7a1 ) , the microsomal p450 liver enzyme , to produce two primary bas , cholic acid , and chenodeoxycholic acid , essential in the overall balance of cholesterol homeostasis . \n sterol 12 hydroxylase ( cyp8b1 ) catalyzes the synthesis of cholic acid , a step which determines the cholic acid to cdca ratio in the bile . \n secondary bile acids ( deoxycholic acid and lithocholic acid ) and tertiary bile acids ( ursodeoxycholic acid ) in humans are produced following intestinal dehydroxylation of primary bile acids by intestinal bacteria [ 58 , 61 ] . \n regulation of ba biosynthesis is highly coordinated and is mediated by key nrs including the orphan receptor , liver receptor homologue-1 ( lrh1 ; nr5a2 ) , the hepatocyte nuclear factor 4 ( hnf4 ) , shp , and the bile acid receptor farnesoid x receptor ( fxr ; nr1h4 ) . \n thus , the activation of fxr initiates a feedback regulatory loop via induction of shp , which suppresses lrh-1- and hnf4-dependent expression of the two major pathway enzymes cholesterol 7hydroxylase ( cyp7a1 ) and sterol 12 hydroxylase ( cyp8b1 ) . \n the ba feedback regulation primarily occurs since bas act as transcriptional regulators for the expression of the gene encoding cyp7a1 . both cholic acid and chenodeoxycholic acid function as endogenous ligands for the nuclear bile acid receptor fxr . \n fxr expression is high in the intestine and liver , the two sites where bas reach high concentrations to activate fxr . \n the transcription by fxr includes heterodimerization with retinoid x receptors ( rxrs ) in the cytoplasm , translocation into the nucleus , and binding to dna response elements in the regulatory regions of target genes . \n when the bind of bas to fxr , shp transcription is increased [ 60 , 64 , 65 ] , this alteration leads to the inhibition of lrh-1 activity or hnf4 on the ba response elements ( bares ) of cyp7a1 and cyp8b1 promoters [ 64 , 65 ] . in this scenario , ba synthesis is downregulated by a precise feedback regulatory mechanism , which represents the major pathway under normal physiological conditions [ 6466 ] ( figure 3 ) . \n lrh1 is also a well - known activator of shp gene transcription [ 64 , 65 ] , and this step leads to an autoregulatory loop of gene expression by shp . \n this step also includes the g protein pathway suppressor 2 ( gps2 ) interacting with fxr , lrh-1 , and hnf4 to regulate cyp7a1 and cyp8b1 expression in human hepatocytes ( table 3 ) . \n a critical role in maintaining cholesterol homeostasis for cyp7a1 has been recently advocated in a model of in cyp7a1-tg mice . \n the hepatocyte nuclear factor-1 ( hnf1 ) , which haploinsufficiency causes the maturity - onset diabetes of the young type 3 ( mody3 ) , also appears to modulate shp expression via the fxr pathway . in this respect , \n hnf1 ( / ) mice displayed a defect in bile acid transport , increased bile acid and liver cholesterol synthesis , and impaired hdl metabolism . a role for shp in mediating the recruitment of msin3a - swi / \n snf to the cyp7a1 promoter , with chromatin remodeling and gene repression , has been described . in hepg2 cells , \n kemper et al . have shown that bile acid treatment resulted in shp - mediated recruitment of transcriptional coregulators msin3a and swi / snf complex to the promoter , chromatin remodeling , and gene repression ( table 3 ) . \n this is an additional mechanism involving transformation of nucleosome conformation for the repression by shp of genes activated by various nrs . in line with such results , increased synthesis and accumulation of bas occurs in shp ( / ) mice , due to the loss of shp repression and consequent derepression of the rate - limiting cyp7a1 and cholesterol 12-hydroxylase ( cyp8b1 ) ( the rate - determining enzyme of the alternative but minor ba synthesis pathway ) in the biosynthetic pathway [ 7072 ] . \n mechanisms independent of the fxr / shp / lrh pathway might also exist , since bas feeding to shp ( / ) mice reduced the levels of cyp7a1 mrna to similar levels of control mice [ 70 , 71 ] . \n such shp - independent and alternative pathways include the protein kinase c / jun n - terminal kinase ( pkc / jnk ) pathway , the fxr / fgfr4 ( fgf receptor 4 ) pathway [ 57 , 74 ] , the cytokine / jnk pathway , the pregnane x receptor ( pxr ) mediated pathway , and the jnk / c - jun signaling pathway . \n another study demonstrated , in shp ( / ) mice on a background of 129 strain , the protection against hypercholesterolemia in three different models : an atherogenic diet , hypothyroidism , and shp ( / ) mice intercrossed with ldlr ( / ) mice ( to generate shp / ldlr double ( / ) mice in a mixed 129-c57bl/6 background ) . when fed an atherogenic diet , the latter strain was almost completely resistant to diet - mediated increases in triglyceride , very low - density lipoprotein ( vldl ) cholesterol , and low - density lipoprotein ( ldl ) cholesterol but had an increase in high - density lipoprotein ( hdl ) cholesterol as compared with ldlr ( / ) mice . \n such results point to the protection against dyslipidemia following the inhibition of hepatic shp expression , although no antagonist ligands have yet been identified for shp . \n we have recently examined biliary lipid secretion and cholesterol gallstone formation in male shp ( / ) and ( + /+ ) mice before and during the feeding of a lithogenic diet for 56 days . \n deletion of the shp gene significantly increased hepatic bile salt synthesis , and doubled the increase of biliary bile salt outputs in shp ( / ) mice than in ( + /+ ) mice . \n the intestinal bile acid pool size was significantly greater in shp ( / ) mice than in ( + /+ ) mice . \n these increased bas are efficacious ligands of fxr and can stimulate the expression of intestinal fibroblast growth factor 15 ( fgf15 ) in mice through the fxr signaling pathway , which is consistent with the expanded bile acid pool size in shp ( / ) mice . at 14 days on the lithogenic diet , \n fasting gallbladder volume was significantly larger in shp ( + /+ ) mice than in ( / ) mice . \n is another fxr gene target in the intestine and appears to contribute to the fine tuning of bile acid synthesis in the liver . \n thus , a model for fxr - mediated repression of bile acid synthesis should also take into account the bile acid - mediated activation of intestinal fxr and fgf15 in the small intestine ( while the fxr - shp pathway is activated in the liver ) . according to the most plausible view , fgf15 acts as a hormone to signal between intestine and liver . \n the secreted fgf15 by the intestine circulates to the liver , likely through the portal circulation or lymph flow , and induces the activation of fgfr4 in the liver . \n as shown in figure 3 , the fgf15/fgfr4 pathway synergizes with shp in vivo to repress cyp7a1 expression . in humans , \n of note , activation of fxr transcription in the intestine protected the liver from cholestasis in mice by inducing fgf15 expression and reducing the hepatic pool of ba . \n hepatic fatty acid homeostasis is also regulated by shp since regulating these genes involves in fatty acid uptake , synthesis , and export [ 8387 ] . in a study exploring global gene expression profiling combined with chromatin immunoprecipitation assays in transgenic mice constitutively expressing shp in the liver , overexpression of shp in the liver \n was associated with the depletion of the hepatic bile acid pool and a concomitant accumulation of triglycerides in the liver . by contrast , fat accumulation induced by a high - cholesterol or high - fat diet \n the pleiotropic role of shp can also be found in the case of nonalcoholic liver steatosis since ob / shp double ( / ) mice ( a model of severe obesity and insulin resistance ) became resistant to liver steatosis and showed improved insulin sensitivity . \n another interesting role for shp emerged after it was found that bas negatively regulate the human angiotensinogen ( ang ) gene . \n ang is the precursor of vasoactive octapeptide angiotensin ii , and bas act through the shp pathway by preventing hepatocyte nuclear factor-4 ( hnf4 ) from binding to the human ang promoter . \n human obesity is considered a polygenic disorder characterized by partly known abnormal molecular mechanisms resulting in increased fat mass , with an imbalance between the energy acquired from nutrients that dissipated as heat ( i.e. , thermogenesis ) . in this respect , \n weight stability requires a balance between calories consumed and calories expended . in adipose tissue depots , two main types of adipocytes \n exist , that is , brown adipocytes and white adipocytes . in several animal species , \n some adipose tissue sites mainly include brown adipocytes ( bats ) and the other contains mainly white adipocytes ( wats ) . \n bat dissipates chemical energy to produce heat either as a defense against cold or as energy expenditure to compensate food intake [ 93 , 94 ] . \n the unusual function of bat might be better understood by considering that they share a common origin with myocytes [ 95 , 96 ] , and bat was indeed considered something in between muscle and adipose tissue . \n bat is deemed as the major site for sympathetic ( adrenergic ) mediated adaptive thermogenesis ; this pathway involves the uncoupling protein-1 ( ucp1 ) . \n wat is mainly implicated in the regulation of lipid storage and catabolism but also in the synthesis and secretion of adipokines [ 97100 ] . while the percentage of young men with bat is high , the activity of bat is reduced in men who are overweight or obese . \n thermogenesis unequivocally exists in both humans and animals , and bat is the major site of thermogenesis which can be increased by environmental factors ( i.e. , adaptive thermogenesis ) . in both human and animal species , dietary composition , \n chronic cold exposure , and exercise may increase thermogenesis . as far as adipose tissue biology is concerned \n , shp seems to play a distinct regulatory function in wat , as compared with bat . \n a number of experiments have focused on animal models of obesity and subtle molecular changes . \n shp - deficient mice are protected against high - fat - diet - induced obesity . \n peroxisome proliferator - activated receptor ( ppar ) coactivator-1 ( pgc-1 ) family members are multifunctional transcriptional coregulators . \n in particular , pgc-1 and pgc-1 regulate mitochondrial biogenesis , adaptive thermogenesis , fatty acid and glucose metabolism , fiber - type switching in skeletal muscle , peripheral circadian clock , and development of the heart . \n in particular , shp functions as a negative regulator of energy production in bat because shp is a negative regulator of pgc-1 expression in bat . in turn , pcg-1 is a coactivator of uncoupling protein 1 ( ucp1 ) which plays a major role in energy dissipation as heat in multilocular bat of different animal species and humans [ 104106 ] . \n fat - specific ( bat ) shp - overexpressed transgenic mice had increased body weight and adiposity . energy metabolism , however , was increased , and bat cold exposure function was enhanced with activation of thermogenic genes and mitochondrial biogenesis ( enhanced 1-ar gene expression and pgc1 ) . compared with wild - type mice on a high - fat diet , \n shp overexpression was associated with enhanced diet - induced obesity phenotype with weight gain , increased adiposity , and severe glucose intolerance . \n an additional feature of shp transgenic mice was a decreased diet - induced adaptive thermogenesis , increased intake of food , and decreased physical activity . \n this leads to the conclusion that , although expressed at low levels in fat , activation of shp in adipocytes has a strong effect on weight gain and diet - induced obesity . moreover , \n if mechanisms linked to energy metabolism and the development of obesity are considered , shp has distinct roles in wat and bat . \n as previously mentioned , while shp deletion in obese leptin - deficient mice ( ob / ob ) prevented the development of nonalcoholic fatty liver and improved peripheral insulin sensitivity , shp deletion did not overcome the severe obesity caused by leptin deficiency . \n a significant protective effect from obesity by shp deficiency was likely associated with the low basal level of shp expressed in fat . \n adipogenesis appears to be influenced by shp : when shp was overexpressed in 3t3-l1 preadipocytes , cell differentiation was inhibited , as well as the accumulation of neutral lipids within the cells . \n in particular , shp appears to be a potent adipogenic suppressor , and preadipocytes are kept in an undifferentiated state through the inhibition of the adipogenic transcription factors and stimulators . \n further studies will address whether the loss of shp function results in inhibition of lipid accumulation in adipocytes , similar to what is observed in hepatocytes . in a future clinical setting , treatment of obesity might also include drugs able to mimic or stimulate the effects of shp . \n mutations in the shp gene have also been reported in patients with lipodystrophy carrying four different polymorphisms . \n a number of important clinical studies have examined this issue ( table 4 ) ; however , hung et al . in uk examined the relationships between genetic variation in shp and weight at birth , adiposity , and insulin levels in three different populations ( the genetics of obesity study ) goos , the avon longitudinal study of parents and children ( alspac ) , and the ely studies ) . in the 329 cases of severe early - onset obesity ( goos study ) , two novel and rare missense mutations ( r34 g and r36 g ) \n furthermore , two common polymorphisms , namely , g171a ( 12% of subjects with higher birth weight ) and 195ctgadel ( 16% of subjects with lower birth weight ) were found . in the alspac cohort of 1,079 children , \n the g171a variant was associated with increased body mass index and waist circumference together with higher insulin secretion 30 minutes after glucose load . thus , whereas mutations in the shp gene can not be seen as a common cause of severe human obesity , genetic variation in the shp gene locus may influence birth weight and have effects on body size . \n the effect might ultimately involve insulin secretion by the negative regulation between shp and the hepatocyte nuclear factor-4 ( hfn-4 ) , a transcription factor involved in differentiation and function of pancreatic -cells . \n a possibility is that decreased shp expression or function results in increased hfn-4 activity with a cascade of events , including fetal hyperinsulinemia , and increased birth weight . at a later stage , \n mutations in the shp gene were also associated with influence on birth weight , mild obesity , and insulin levels in the study by nishigori et al . \n mutations in several genes encoding transcription factors of the hepatocyte nuclear factor ( hnf ) cascade are associated with maturity - onset diabetes of the young ( mody ) . \n mody is a monogenic form of early - onset diabetes mellitus ( defective insulin secretion with normal body weight ) , and shp is deemed as a plausible candidate mody gene ; this is because shp is able to inhibit the transcriptional activity of the hepatocyte nuclear factor-4 ( hfn-4 ) , a key member of the mody regulatory network . \n thus , further studies have looked for segregation of shp mutations with mody in a cohort of japanese patients with early - onset diabetes . in this context \n , variants in shp appeared to cosegregate with increased body mass index in families , thus contributing to obesity among japanese subjects . also , increased risk of morbidity was observed in another study from japan , examining patients with type 2 diabetes and shp mutations . \n major differences , however , might exist in the prevalence and function of shp variants in different populations . of note \n , the results from other caucasian cohorts did not confirm the association between shp mutation and obesity [ 113 , 114 ] . \n conducted an elegant study on the prevalence of shp variants by single - strand conformational polymorphism and heteroduplex analysis among 750 danish obese men with early - onset obesity . \n as control , a cohort of 795 nonobese control subjects was genotyped using pcr - rflp . \n functional analyses of the identified coding region variants were performed in both min6-m9 and hepg2 cell lines . \n five novel variants were identified ( including 3 missense variants ( c.100c > g [ p.r34 g ] , c.278g > a [ p.g93d ] , and c.415c > a [ p.p139h ] ) and 2 silent variants ( c.65c > t [ p.y22y ] and c.339g > a [ p.p113p ] ) ) . the previously reported c.512g > c [ p.g171a ] common polymorphism was identified ; however , the prevalence of functional shp variants associated with obesity was considerably lower among danish subjects ( 1 out of 750 obese , none of control subjects ) , compared to the prevalence observed in japan by nishigori et al . . \n investigated shp variants in 1927 uk subjects according to type 2 diabetes , obesity , and birth weight . \n although reporting a raised body mass index among homozygous carriers of the 171a variant ( < 1% ) , this polymorphism was unlikely to be associated with all three conditions in caucasians . \n taken together , the above - mentioned studies suggest that the 171a variant might contribute only to subsets of polygenic obesity . \n the existence of multiple interactions of shp with nrs , transcription factors and transcriptional cofactors ( tables 2 and 3 ) points to the pleiotropic and central role of shp in the body . \n shp has been hypothesized to act in glucose homeostasis via complex pathways involving the inhibition of glucocorticoid receptors ( gr ) in mammalian cells and the inhibition of pgc-1 gene , a coactivator of nrs important for gluconeogenic gene expression and the pgc-1-regulated phospho(enol)pyruvate carboxykinase ( pepck ) promoter . \n such steps underscore a physiologically relevant role for shp in modulating hepatic glucocorticoid action . following \n the bile acid - induced induction , shp inhibited a number of other pathways , including the hnf4-mediated transactivation of the pepck and fructose biphosphate ( fbp ) promoters , as well as the transactivation of the glucose-6-phosphatase ( g6pase ) promoter mediated by foxo1 . \n the interaction between shp inhibitory function and the 3 isoforms ( , , and ) of the hepatocyte nuclear factor-3 ( hnf4 ) points to the regulatory role of shp on gluconeogenesis . \n mutations in hepatocyte nuclear factor 1 ( hnf-1 ) is associated with maturity - onset diabetes of the young type 3 . \n indeed , loss of hnf-1 function in hnf-1 ( / ) mice resulted in altered expression of genes involved in glucose - stimulated insulin secretion , but also insulin synthesis , and beta - cell differentiation . \n pancreatic islets of hnf-1 ( / ) mice showed a distinctive reduction of shp expression and a downregulation of the hnf4 gene expression . since shp appears to repress its own transcriptional activation following heterodimerization with hnf4 , a feedback autoregulatory loop between shp and hnf4 has been hypothesized . \n shp ( / ) mice were characterized by hypoinsulinemia , increased glucose - dependent response of islets , increased peripheral insulin sensitivity , and increased glycogen stores . \n the role played by shp in the regulation of hepatic gluconeogenesis has also emerged in a number of additional experiments . \n for example , the liver of shp ( / ) mice showed increased glycogen stores , while hepatic shp gene expression ( induced by the antidiabetic biguanide drug metformin ) was associated with inhibition of hepatic gluconeogenesis . \n induction of shp was achieved via amp - activated protein kinase ( ampk ) and associated with downregulation of essential gluconeogenic enzyme genes , that is , phosphoenolpyruvate carboxykinase ( pepck ) , glucose-6-phosphatase ( g6pase ) , and fructose-1,6-bisphosphatase ( fbp1 ) . \n pgc-1 gene is a coactivator of nrs , and this step is relevant for gluconeogenic gene expression . \n yamagata et al . showed that bile acid ( chenodeoxycholic acid ) was able to induce the downregulation of pgc-1 gene , and this mechanism involved forkhead transcription factors ( foxo1 , foxo3a , foxo4 ) via a shp - dependent manner . \n this is also the case for excess bas : the pregnane x receptor ( pxr ) induces cyp3a and inhibits cyp7 , both involved in biochemical pathways leading to the conversion of cholesterol into primary bas , whereas cyp3a is also involved in the detoxification of toxic secondary bile acid derivatives . \n shp acts as a potent repressor of pxr transactivation , and this finding suggests that pxr can act on both bile acid synthesis and elimination detoxification . \n additional mechanisms involved in the shp - dependent control of pathways of drug metabolism have been identified . \n the expression of genes involved with the metabolism of xenobiotics might be regulated by shp in the spleen acting on ( aryl hydrocarbon receptor ( ahr)/ahr nuclear translocator ( arnt ) ) ahr / arnt heterodimers which , in turn , bind to xenobiotic response elements ( xres ) at the level of specific dna sequences . \n a number of genes involved in hormone and drug metabolism would be expressed ( i.e. , ugt16 , aldh3 , cyp1a1 , cyp1a2 , cyp1b1 , etc . ) . \n shp also appears to downregulate the constitutive - androstane - receptor- ( car- ) mediated cyp2b1 gene expression , induced by phenobarbital to form the car / rxr heterodimer which , in turn , binds to 2 dr-4 sites to form the phenobarbital responsive unit in the cyp2b gene ( table 3 ) . \n one role of shp in steroidogenesis has been identified in the testes with influence on testosterone synthesis and germ cell differentiation and in the intestine for glucocorticoid synthesis . \n a role for shp in cell proliferation and apoptosis signaling is emerging . depending on the cell type \n however , the manipulation of shp through the synthetic ligands adamantyl - substituted retinoid - related ( arr ) compounds 6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphthalenecarboxylic acid ( cd437/ahpn ) and 4-[3-(1-adamantyl)-4-hydroxyphenyl]-3-chlorocinnamic acid ( 3-cl - ahpc ) induces apoptosis of a number of malignant cells ( i.e. , leukemia and breast carcinoma ) both in vitro and in vivo [ 123 , 124 ] . the complex mechanism implies binding of arr and 3-cl - ahpc to shp with formation of a corepressor complex containing sin3a and nuclear receptor corepressor ( n - cor ) which activate local control of mitochondrial function and apoptosis , with a limiting function on tumorigenesis [ 17 , 123 ] ( table 3 ) . \n shp appears to be also involved in dna methylation and acting as a tumor suppressor , at least in the human and mouse livers [ 125127 ] . \n whether manipulation of shp will be helpful in the treatment of hepatic and other gastrointestinal cancers is still a matter of research . \n the recent finding that shp negatively regulates tlr signaling to nf-b has raised the interest for the role of shp in mechanisms governing innate immunity . \n shp appears to negatively regulate the expression of genes encoding inflammatory molecules . of note , \n direct binding of nf-b seems to occur in resting cells , while binding of shp to traf6 occurs in lps - stimulated cells [ 128 , 129 ] . \n a remarkable number of metabolic functions in the body appear to be regulated by the orphan unique nr , small heterodimer partner shp , which targets a complex set of genes in multiple pathways as a transcriptional corepressor ( figure 4 ) . \n when looking at complex mechanisms leading to some important lipidopathies , that is , obesity and liver steatosis , enlightening data about the regulatory function of shp are provided by studies using shp - deleted and shp - overexpressed animal models . \n most likely , a condition of shp deficiency might counteract lipid accumulation and improve plasma lipoprotein profiles . \n further studies are urgently needed to confirm that such an important metabolic regulatory mechanism of shp is true and has high translational value . to date , however , no synthetic antagonists or agonists for shp are available , and one should keep in mind that rather divergent and somewhat elusive data have been observed regarding the loss of shp function in humans and rodents . thus , \n careful examination of subtle shp intrinsic functions is essential to dissect potential modulatory pathways of shp for a variety of metabolic abnormalities but also in tumorigenesis . \n moreover , identifying specific endogenous ligands and synthetic agonists of shp will pave the way to for therapeutic intervention . \n the effect of synthetic ligands on shp modulation in hepatocytes and adipocytes , for example , might represent therapeutic tools for the treatment of constituents of the metabolic syndrome , namely , hypercholesterolemia , overweight obesity , and liver steatosis .", "role": "user" }, { "content": "nuclear receptors ( nrs ) comprise one of the most abundant classes of transcriptional regulators of metabolic diseases and have emerged as promising pharmaceutical targets . \n small heterodimer partner ( shp ; nr0b2 ) is a unique orphan nr lacking a dna - binding domain but contains a putative ligand - binding domain . \n shp is a transcriptional regulator affecting multiple key biological functions and metabolic processes including cholesterol , bile acid , and fatty acid metabolism , as well as reproductive biology and glucose - energy homeostasis . \n about half of all mammalian nrs and several transcriptional coregulators can interact with shp . \n the shp - mediated repression of target transcription factors includes at least three mechanisms including direct interference with the c - terminal activation function 2 ( af2 ) coactivator domains of nrs , recruitment of corepressors , or direct interaction with the surface of nr / transcription factors . \n future research must focus on synthetic ligands acting on shp as a potential therapeutic target in a series of metabolic abnormalities . \n current understanding about the pleiotropic role of shp is examined in this paper , and principal metabolic aspects connected with shp function will be also discussed .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: among bilaterian animals , the deuterostomes include the hemichordates , the echinoderms and the chordates . \n the chordates can further be subdivided into urochordates ( = tunicates ) , cephalochordates and vertebrates ( fig . \n since human beings have often wondered about their own origins , it is not surprising that there have been many studies aimed at understanding the evolutionary history of chordates ( i.e. the origin of chordates and the diversification of vertebrates ) . \n the main characters that evolved at the base of the chordates are a notochord and a dorsal hollow nerve cord 1 , while definite neural crest cells and placodes arose at the base of the vertebrates 2 . \n however , the evolutionary origins of these structures have been controversial , mainly because of difficulties in determining homologies between chordate - specific characters and structures in non - chordate deuterostomes ( i.e. in hemichordates and echinoderms ) as well as between the vertebrate - specific characters and structures in invertebrate chordates . \n the development of many chordate- and vertebrate - specific characters is controlled , directly or indirectly , by retinoic acid ( ra ) , a vitamin a - derived morphogen . in chordate embryos , \n too much or too little ra during early embryogenesis causes malformations , which are mainly due to a mispatterning of the embryo along the anteroposterior body axis . \n ra signaling is mediated by ra binding to retinoic acid receptors ( rars ) , which form heterodimers with retinoid x receptors ( rxrs ) 3 . during chordate development , \n important functions of ra signaling are mediated by hox genes , at least some of which are direct targets of ra 4 - 6 . in many animals , including most deuterostomes , hox genes mediate anteroposterior positional patterning of the embryo 7 , 8 . moreover , in vertebrates , hox genes have been suggested to play important roles in the patterning of all embryonic tissue layers 7 , 9 . ever since it became clear that the genetic mechanisms controlling key features of embryonic development are highly conserved between organisms as diverse as fruit flies and mice , comparisons of the developmental mechanisms in different organisms have been used to address questions about animal evolution , particularly about the invertebrate - to - vertebrate transition in chordates 10 . using this so - called evo - devo \n approach , this review discusses the developmental roles of ra signaling among chordates in regard to chordate origins and vertebrate evolution . \n the focus is on the roles of ra- and hox - dependent patterning during development and their conservation during chordate evolution . \n we show that some of the control mechanisms governing ra - mediated patterning are quite conserved among vertebrates and between vertebrates and invertebrate chordates , such as the cephalochordate amphioxus . \n in contrast , evidence for roles of ra signaling outside the chordates and the evolutionary origin of ra signaling itself still remain elusive . \n retinoic acid ( ra ) is a natural morphogen synthesized from vitamin a ( retinol ) , and it has been known for over fifty years that either too much or too little ra during early development is teratogenic mainly due to anteroposterior patterning defects 3 . in general , excess ra posteriorizes , while ra deficiency anteriorizes chordate embryos 3 , 11 - 14 . \n endogenous ra is synthesized in two steps : the first is the reversible oxidation of retinol to retinal performed by alcohol dehydrogenases ( adhs or rdhs / sdrs ) and the second is the oxidation of retinal to retinoic acid , which is carried out by retinaldehyde dehydrogenases ( raldhs ) . \n ra signaling levels are also regulated by binding of retinol to cellular retinol binding proteins ( crbps ) and of ra to retinoic acid binding proteins ( crabps ) ( fig . \n the roles of these proteins in regulating ra signaling in vertebrates have been elucidated with gene knock - outs . \n for example , individual knock - outs of three of the four raldh genes ( i.e. the knock - outs of raldh1 , 2 and 3 ) have been described in the mouse , but only that of raldh2 exhibits clear developmental defects . in these mice , \n the anteroposterior axis is shortened and the embryos , which die before birth , have defects in the heart , limbs and head , which are reminiscent of vitamin a deprivation phenotypes 16 . in the ra degradation pathway , knock - out of cyp26a1 , one of the three mouse cyp26 genes , is also lethal before birth and affects anteroposterior patterning as well as hindbrain and tail development 17 , 18 . \n this cyp26a1 phenotype is comparable to the teratogenic effects of excess ra , suggesting that cyp26 genes in general may help restrict the distribution of endogenous ra to ra target tissues . \n moreover , analysis of cyp26a1 and raldh2 double knock - out mice has shown that ra and not one of its metabolic derivatives is the major ( if not the only ) active retinoid during development 19 . \n ra signaling is mediated by ra binding to retinoic acid receptors ( rars ) , which form heterodimers with retinoid x receptors ( rxrs ) . \n this complex in turn binds to retinoic acid response elements ( rares ) in the regulatory regions of target genes 13 , 15 . \n these include the rars themselves , hox genes , some other transcription factors , such as hnf-3 and cdx1 , plus genes involved in retinoic acid metabolism ( e.g. crabp1 and 2 ) 5 . in the mouse , there are 3 rars and 3 rxrs ( , and ) . \n although knock - out of any one of these has only minor , tissue - specific effects on morphogenesis due most likely to functional redundancy among them , compound mutants with two or three of the genes inactivated are much more severely affected with defects in anteroposterior patterning of pharyngeal endoderm , hindbrain and neural crest cells 3 . \n in general , rar and rxr proteins share a common organization of functional domains : an amino terminal a / b region containing a transcriptional activation domain ( af-1 ) , a centrally located c region corresponding to the dna binding domain ( dbd ) plus a weak dimerization domain and the e region , which includes the ligand binding domain ( lbd ) , a strong dimerization interface and a surface allowing binding of transcriptional co - regulators 3 , 20 , 21 . in the absence of ligand , the rar / rxr heterodimer is constitutively bound to dna on rares and associated with co - repressor complexes that induce transcriptional silencing by deacetylating histones associated with the target sequences thus increasing chromatin condensation ( fig . \n binding of ra to the rar ligand binding pocket induces a conformational change of the lbd that creates a surface allowing the association of co - activators and the release of co - repressors . \n the co - activators ( e.g. tif2 and src-1 of the p160 co - activator family ) subsequently mediate histone acetylation resulting in decondensation of the chromatin and activation of target gene expression ( fig . \n 2b ) 20 - 22 . among the known targets of the rar / rxr heterodimer \n hox genes encode transcription factors that contain a highly conserved dna binding domain of 60 amino acids , the homeodomain 7 , 8 . \n hox genes are usually linked in a cluster and their order on the chromosome correlates with both their temporal and spatial expression during embryogenesis 7 - 9 , 23 . \n this so - called collinear expression is crucial for conveying anteroposterior positional patterning information to the embryo during development and is conserved in many animals , including most deuterostomes 7 - 9 , 23 . \n however , when genomic hox clustering has been lost , as for example in tunicates , temporal collinearity is lacking and spatial collinearity is only approximate 24 , 25 . in vertebrates , hox genes are direct targets of ra signaling 5 and ra signaling is involved in regulation of collinear hox expression along the anteroposterior body axis of the developing embryo . for example , in mice , the deletion of a single rare from the hoxa1 cis - regulatory region disrupts the establishment of the anterior boundary of hoxa1 as well as the normal expression pattern of hoxa2 in the hindbrain 26 . \n moreover , the hox cluster also contains several other rares , which are largely conserved among vertebrates and at least to some extent between vertebrates and the cephalochordate amphioxus 4 , 6 . \n for example , both the amphioxus and vertebrate hox1 genes have a conserved rare in their cis - regulatory region 4 . \n in addition , although clustering of hox genes has been lost in tunicates , ascidian hox1 expression is strongly upregulated after ra treatment 27 suggesting that , as in amphioxus and vertebrates , hox genes in tunicates might also be direct targets of ra signaling . \n early during vertebrate diversification , the total number of genes has been markedly increased by two rounds of whole genome duplications 28 . \n thus , while vertebrates have 3 or more rars , there are single rars in invertebrate chordates , such as amphioxus and tunicates 14 , 29 . a comprehensive search of the sea urchin genome sequence ( www.ncbi.nlm.nih.gov/genome/guide/sea_urchin ) also reveals the presence of a single rar ( fig . \n although less is known about the ra signaling pathway in invertebrate deuterostomes than in vertebrates , it seems likely that the overall pathway is very much the same in amphioxus and vertebrates and somewhat different in tunicates ( for example , rar does not regulate its own expression in tunicates , but it does in both amphioxus and vertebrates 29 ) . \n raldh and cyp26 genes have been identified in the genomes of both amphioxus ( ftp.ncbi.nih.gov/pub/tracedb/branchiostoma_floridae ) and tunicates ( www.ensembl.org/ciona_intestinalis ) ( fig . \n too little is known about ra signaling in echinoderms , such as sea urchins , for a comparison with chordates 31 - 33 . moreover , for hemichordates , the sister group of echinoderms , it is not known if there is an rar , and roles for ra signaling during hemichordate development have yet to be established . \n even so , bioinformatic analyses of the sea urchin genome sequence ( www.ncbi.nlm.nih.gov/genome/guide/sea_urchin ) and of hemichordate est data ( ftp.ncbi.nih.gov/pub/tracedb/saccoglossus_kowalevskii ) suggest that both raldh and cyp26 genes are present in echinoderms and hemichordates ( fig . \n interestingly , these analyses also revealed a putative rare ( a so - called dr5 element ) about 3770 base pairs upstream ( 5 ' ) of the sea urchin hox1 gene . \n thus , at least some components of the ra signaling cascade were probably already present in the last common ancestor of deuterostomes . \n in addition , these results raise two very important questions about the evolution of the ra signaling network : ( 1 ) if molecular components of the ra pathway are present in non - chordate deuterostomes , what are their functional roles during development ? ( 2 ) if molecular traces of the ra pathway exist in all deuterostomes , when did ra signaling first evolve ? in the last common ancestor of deuterostomes , in the urbilaterian ancestor of deuterostomes and protostomes or even earlier during animal evolution ? \n a first tentative step towards answering the second question was recently taken by an extensive phylogenetic analysis of the nuclear hormone receptor superfamily , which includes the rar and rxr subfamilies 34 . \n the results obtained from this analysis suggest that a proto - rar gene was already present in the genome of the last common ancestor of protostomes and deuterostomes and that in protostomes this rar gene was lost at least in the lineages leading to nematode worms and insects 34 . \n in vertebrates , the roles of ra signaling in anteroposterior patterning of the central nervous system ( cns ) and pharynx have received particular attention 35 , 36 . in the cns , when vertebrate embryos are treated with ra , anterior neural structures ( like the forebrain ) are lost and posterior structures , such as the hindbrain and spinal cord , are expanded . \n in addition , expression of genes in the anterior nervous system ( like otx2 ) is lost and the hox genes , normally expressed in hindbrain and spinal cord , are upregulated and shifted anteriorly 35 . in lampreys , for example , \n conversely , decreasing ra signaling levels has the opposite effect : anterior neural structures , such as the forebrain and midbrain are expanded posteriorly , as is expression of otx2 35 . \n loss of at least two of the three rar genes in mice also leads to mispatterning of the hindbrain , such as defects in segmental ( rhombomeric ) hindbrain organization , which correlate with an induction of hox gene expression at more posterior levels of the hindbrain 38 . \n thus , in vertebrates , ra signaling influences collinear expression of hox genes , which in turn is required to set up an anteroposterior patterning code along the neural tube . not only is anteroposterior patterning of the cns affected by ra signaling , but , since hindbrain neural crest carries the hox code of the cns with it when it migrates , the hox code carried by neural crest migrating into the branchial arches is also altered when levels of ra signaling are changed . in mammals , \n excess ra causes fusion of the first two pharyngeal arches while in ra - deficient mice and rats , pharyngeal structures caudal to the first pharyngeal arch are absent 39 - 42 . \n similarly in vitamin a - deficient quail , the pharynx is extended caudally , the first pouch forms normally , the second one is abnormal and the third and fourth pharyngeal pouches never form 43 . in lampreys , \n the effects of excess ra are even more severe , with complete deletion of anterior pharyngeal structures 44 . \n thus , ra signaling in the pharynx specifies the anteroposterior position of pharyngeal structures 36 , 45 , 46 . \n it was initially believed that the effects of exogenous ra on the vertebrate pharynx were solely due to a mispatterning of neural crest cells migrating into the branchial arches 36 , 45 , 46 . however \n , this influence of neural crest cells is subordinate to regional patterning within the pharyngeal endoderm itself , since removal of neural crest does not prevent the proper patterning of pharyngeal arches and pouches 47 . \n conversely , in the zebrafish , tbx1 function in the pharyngeal endoderm is required for proper development of neural crest - derived structures in the pharyngeal arches 48 , 49 . \n interestingly , the defects obtained in the tbx1 mutant van gogh in zebrafish are similar to those obtained after loss of both rar and in mice 38 , 48 , 49 suggesting that ra signaling may be upstream of tbx1 in the pharyngeal endoderm . moreover , treatment of developing mice with an rar-specific agonistic ligand has conclusively shown that rar-dependent ra signaling in the endoderm is independent of neural crest cells 50 . \n thus , while neural crest cells migrating into the branchial arches give rise to several structures , such as the pharyngeal arch cartilage , the initial patterning of the pharynx requires ra signaling within the endoderm 36 , 45 , 46 . in the cephalochordate amphioxus \n , rar is expressed in the hindbrain and anterior spinal cord of the developing cns 14 . moreover , in the region of the amphioxus cns strongly expressing rar , hox1 , hox3 and hox4 are collinearly expressed 51 . \n ra strongly upregulates rar expression throughout the cns , whereas an ra antagonist almost completely downregulates rar 14 . \n in addition , ra pushes the anterior limits of hox1 expression anteriorly in the cns 12 , while ra antagonist shifts the hox1 domain posteriorly ( fig . \n these data suggest that in cephalochordates , as in vertebrates , ra signaling , probably acting via hox genes , controls anteroposterior patterning of the developing cns . \n similarly , in the amphioxus general ectoderm , treatments with ra and ra antagonist affect collinear expression of hox genes . \n whereas ra shifts expression of hox genes ( such as hox1 ) anteriorly in the general ectoderm , the ra antagonist completely downregulates ectodermal hox expression ( fig . \n 4b ) 53 . moreover , in amphioxus , which lacks definitive neural crest , exogenous ra shifts the posterior limit of the pharynx anteriorly , while the application of ra antagonist has the opposite effect 12 , 14 . \n the amphioxus pharyngeal endoderm strongly expresses pax1/9 and otx 54 , 55 and treatments with ra or an ra antagonist shift the posterior limit of pax1/9 and otx expression , respectively , anteriorly or posteriorly 52 . \n rar and hox1 are expressed just posterior to pax1/9 and otx in the midgut region of the endoderm suggesting that , as in vertebrates , ra signaling within the endoderm might control anteroposterior pharyngeal patterning in amphioxus 52 . \n in addition , loss of hox1 function mimics the effect of a decrease of ra signaling in the amphioxus endoderm 52 . \n taken together , these results suggest that in the amphioxus endoderm , ra signaling activates hox1 in the midgut and that hox1 in turn mediates the effects of ra signaling by limiting expression of genes that are required for specification of the pharynx ( such as pax1/9 and otx ) to the anterior endoderm 52 . \n a role for ra signaling and hox genes in neural patterning of tunicates is much less obvious than in amphioxus and vertebrates . in the ascidian ciona intestinalis , treatment with ra \n does not affect the expression of rar suggesting that , as mentioned above , rar does not regulate its own expression as it does in other chordate groups 29 . \n in addition , tunicates have lost several hox genes and both cluster organization and collinear expression have been at least partly lost . \n for example , the ascidian ciona intestinalis has lost hox7 , hox8 , hox9 and hox11 24 , while the appendicularian oikopleura dioica has lost hox3 , hox6 , hox7 and hox8 25 . \n expression of some of the remaining hox genes can not be detected at all in the cns and the domains of those that are expressed are only approximately collinear 24 , 25 . \n even so , excess ra affects morphology and even hox gene expression at least in ascidian tunicates . \n for example , treatment of embryos with exogenous ra leads to incomplete closure of the anterior neural tube of ciona intestinalis 29 and expression of the hox1 gene in the cns of the tunicate halocynthia roretzi is shifted anteriorly 11 . \n thus , a role for ra signaling via hox genes in neural patterning of tunicates remains a possibility . \n in contrast to its effects on the cns , the effects of ra on pharyngeal structures in tunicates is more similar to that in amphioxus and vertebrates . as in amphioxus , \n pax1/9 and otx genes are expressed in the tunicate pharynx during gill slit formation 56 , 57 . moreover , in the ascidian tunicate herdmania curvata , ra treatment of juveniles leads to a decrease of otx expression in pharyngeal tissues and to an eventual loss of the pharyngeal basket by respecification of anterior endoderm to a more posterior fate 57 , 58 . taken together , \n these data suggest that , in tunicates , as in amphioxus , ra signaling might play roles in patterning and development of the cns and pharyngeal endoderm . in addition \n , there is evidence that at least some of the genetic machinery mediating ra signaling in amphioxus and vertebrates ( like the hox genes ) might also be important for mediating ra signaling in tunicates . \n thus , embryonic patterning mechanisms mediated by ra signaling and hox genes are present in all chordates . in contrast to chordates , enteropneust hemichordates do not have a centralized nervous system , but instead a diffuse ectodermal nerve net ( i.e. a skin brain ) 59 , 60 . as in the cephalochordate amphioxus , \n hox genes are collinearly expressed in the hemichordate ectoderm 53 , 60 . whether ra signaling is involved in the control of hox gene expression in the ectoderm of hemichordates \n is unknown , but ra does not appear to affect anteroposterior patterning in echinoderms , the sister group of hemichordates 31 . \n these data suggest that collinear expression of hox genes was already used for anteroposterior patterning of ectodermal tissues in the last common ancestor of hemichordates and cephalochordates and that a role for ra signaling in controlling collinear expression of hox genes in the ectoderm evolved very early during chordate evolution . whether this ra - hox hierarchy evolved even earlier , in the last common ancestor of all deuterostomes , remains to be determined . \n thus , very early during chordate evolution or perhaps even very early during deuterostome diversification , ra signaling was co - opted for controlling hox - dependent anteroposterior patterning mechanisms during development . in one scenario , \n the putative deuterostome ancestor had a ventral cns that was patterned by hox genes like other tissue layers , such as the general ectoderm ( fig . \n 5 ) , but the cns was lost in hemichordates 59 . in the second scenario , \n the ancestral deuterostome lacked a cns , and the cns evolved independently from the ectoderm in protostomes and deuterostomes ( fig . \n an ancestral ectodermal patterning mechanism was carried over into the cns at the base of the chordates , leading to the creation of two independent ra - hox control hierarchies , one in general ectoderm and one in neural ectoderm ( i.e. in the cns ) ( fig . \n thus , in invertebrate chordates , ra signaling and hox genes are involved in anteroposterior patterning of the general ectoderm , the cns , the endoderm ( and possibly even of the mesoderm ) . during vertebrate evolution , with the functional elaboration of neural crest cells \n neural crest cells carry patterning information acquired at the dorsal neural tube into other tissue layers ( fig . \n 5 ) . in either case , ra signaling became linked to hox gene function at least at the base of the chordates . \n future research on hemichordates will show whether it evolved even earlier in the deuterostome lineage . \n in sum , ra signaling is a key regulator of anteroposterior patterning orchestrated by hox codes in at least general ectoderm , central nervous system and endoderm of chordates . \n in contrast to the relative conservation of developmental mechanisms within chordates , evidence for roles of ra outside the chordates and the evolutionary origin of ra signaling itself still remain elusive . \n in fact , it is unclear when ra - controlled patterning mechanisms first evolved in the animal kingdom and when the ra signaling cascade was co - opted for regulating hox gene expression . \n future research on the evolution of ra signaling should thus focus on the identification of ra signaling components , such as the ra receptor ( rar ) , in non - chordate deuterostomes ( such as hemichordates and echinoderms ) , protostomes ( such as insects and mollusks ) and maybe even cnidarians . \n furthermore , to fully understand the elaboration of this signaling pathway during evolution , the functional roles of the identified components during development will have to be assessed . \n strongylocentrotus purpuratus rar ( xm_774883 ) , ciona intestinalis rar ( ab210661 ) , branchiostoma floridae rar ( af378827 ) , takifugu rubripes rar1 ( frup00000151563 ) , takifugu rubripes rar2 ( frup00000165122 ) , takifugu rubripes rar ( frup00000079596 ) , takifugu rubripes rar1 ( frup00000158774 ) , takifugu rubripes rar2 ( frup00000143282 ) , homo sapiens rar ( nm_001033603 ) , homo sapiens rar ( nm_000965 ) , homo sapiens rar ( nm_000966 ) . \n echinoderms and hemichordates together establish the sister group of chordates . the urochordates ( = tunicates ) include ascidians and appendicularians . \n the vertebrates include agnathan groups ( hagfish and lampreys ) as well as the gnathostome chondrichthyans ( cartilaginous fish ) , actinopterygians ( ray - finned fish ) and sarcopterygians ( lobe - finned fish and tetrapods ) . within chordates , \n the phylogenetic relationships between cephalochordates and urochordates and between hagfish and lampreys are still disputed and their respective positions within the tree are thus shown as polytomies . \n two important events during deuterostome evolution are the origin of chordates and the origin of vertebrates . the two rounds of extensive gene duplications early during vertebrate diversification are highlighted with green boxes labeled ' r ' . \n deuterostome groups , for which a whole genome sequencing ( wgs ) project has already been finished are indicated with a turquoise + , those , for which a wgs project is underway , are marked with a turquoise ( + ) . \n if known , the exact number of rar , raldh and cyp26 genes in the genome of a specific deuterostome group is indicated , the certain presence of a gene is marked with + and the lack of data is highlighted by ? . \n ra is synthesized by oxidation of retinal by retinaldehyde dehydrogenases ( raldhs ) . in a reversible reaction , retinal \n is synthesized from retinol ( vitamin a ) by either aldehyde dehydrogenases ( adhs ) or short - chain dehydrogenase / reductases ( rdhs / sdrs ) . cellular retinol binding proteins ( crbps ) can bind retinol , whereas cellular retinoic acid binding proteins ( crabps ) can bind ra . \n b ) the rar / rxr heterodimer mediates the effects of ra . in the absence of ligand ( ra ) \n binding of the ligand ( ra ) induces conformational changes and the binding of co - activators leading to histone acetylation and activation of transcription . \n the tree shows the phylogenetic relationships between rars from the sea urchin strongylocentrotus purpuratus , the ascidian tunicate ciona intestinalis , the cephalochordate branchiostoma floridae , pufferfish ( takifugu rubripes ) and humans ( homo sapiens ) . \n the rar sequences were added to an alignment of nuclear hormone receptors 34 and conserved sites ( 335 positions ) were subsequently selected for phylogenetic reconstruction using phyml ( wag + 8 + i ) 61 . \n 100 bootstrap replicates were carried out to determine the robustness of the obtained phylogenetic tree . in the tree , \n the rar subfamily as a whole is strongly supported ( bootstrap support 95% ) and within the rars the sea urchin sequence is at the base ( with a moderate support of 70% ) . \n the respective branching of the cephalochordate and tunicate rars is not resolved ( bootstrap support of 49% ) . \n nonetheless , the invertebrate chordate sequences are positioned between the sea urchin rar and the duplicated rars of vertebrates , which form a single clade that is very strongly supported ( 100% ) . \n this analysis shows that a rar gene is present in the genome of echinoderms and suggests that , despite the lack of data from hemichordates , the presence of rar is an ancestral character of deuterostomes . \n ra signaling controls hox1 expression in central nervous system ( cns ) and general ectoderm of developing amphioxus . \n a ) the rostral limit of hox1 expression in the cns ( arrowheads ) is shifted , respectively , anteriorly and posteriorly by 1x10 m ra and 1x10 m ra antagonist ( bms009 ) . \n b ) in the general ectoderm , the rostral limit of hox1 ( arrowheads ) is shifted anteriorly by 1x10 m ra , whereas treatment with 1x10 m ra antagonist ( bms009 ) downregulates hox1 expression . \n evolution of ra- and hox - dependent patterning mechanisms in deuterostomes . in scenario 1 , \n the putative deuterostome ancestor had a central nervous system ( cns ) ( located ventrally ) and both cns and general ectoderm were patterned by hox genes , while a role for ra signaling remains elusive . in this scenario , chordates evolved by dorso - ventral axis inversion and the cns was secondarily lost in the hemichordate lineage . in scenario 2 , \n hox - dependent patterning codes were present in the ectoderm , but again a role for ra signaling remains elusive . \n early during chordate evolution , condensation of a central nervous system ( cns ) dorsally led to the creation of two ra - hox patterning hierarchies one in general ectoderm and one in neural ectoderm ( i.e. in the cns ) . in the vertebrate lineage , \n neural crest function was elaborated and neural crest cells contribute to patterning and development of the embryo by carrying positional information from the cns into other tissues , for example in the pharyngeal region .", "role": "user" }, { "content": "in chordates , which comprise urochordates , cephalochordates and vertebrates , the vitamin a - derived morphogen retinoic acid ( ra ) has a pivotal role during development . altering levels of endogenous \n ra signaling during early embryology leads to severe malformations , mainly due to incorrect positional codes specifying the embryonic anteroposterior body axis . in this review \n , we present our current understanding of the ra signaling pathway and its roles during chordate development . \n in particular , we focus on the conserved roles of ra and its downstream mediators , the hox genes , in conveying positional patterning information to different embryonic tissues , such as the endoderm and the central nervous system . \n we find that some of the control mechanisms governing ra - mediated patterning are well conserved between vertebrates and invertebrate chordates , such as the cephalochordate amphioxus . \n in contrast , outside the chordates , evidence for roles of ra signaling is scarce and the evolutionary origin of the ra pathway itself thus remains elusive . in sum , to fully understand the evolutionary history of the ra pathway , future research should focus on identification and study of components of the ra signaling cascade in non - chordate deuterostomes ( such as hemichordates and echinoderms ) and other invertebrates , such as insects , mollusks and cnidarians .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: the basic function of a tracheal tube is to provide a reliable connection between the patient 's lung and the bag or ventilator . \n ideally , this connection should be leak - proof without causing undue pressure to laryngeal or tracheal structures . \n traditional teaching was that in children under 8 - 10 years of age , this sealing should be obtained with an uncuffed tracheal tube . \n disadvantages of uncuffed tubes include air leak , difficulties in effectively ventilating patients with lung disease , environmental contamination of anaesthetic gases , risk of aspiration , and difficulty in accurate measurement of end - tidal co2 concentration . \n cuffed endotracheal tubes may be useful in special situations like laparoscopy and in surgical conditions at risk of aspiration . \n intubation depth marks and short , cylindrical cuff near tracheal tube tip allow adequate placement with a cuff - free subglottic zone , without the risk of endobronchial intubation . \n we studied the appropriateness of microcuff endotracheal tube size selection , efficacy of ventilation and monitoring and post - operative complications in children undergoing laparoscopic surgery . \n after institutional ethical committee approval and written informed consent from the parents , a prospective observational study was conducted . \n the study population included 100 patients of american society of anesthesiologists i and ii physical status , aged between 8 months and 10 years , posted for elective laparoscopic surgery . \n patients with known airway anomalies , known or suspected difficult intubation and planned post - operative ventilation in the intensive care unit were excluded from the study . all patients received injection glycopyrrolate 0.004 mg / kg , injection ketamine 0.5 mg / kg , injection midazolam 0.05 mg / kg , all intravenous ( iv ) as premedication . \n standard monitoring included electrocardiogram , pulse oximetry , non - invasive blood pressure , capnography and temperature . \n anaesthesia was induced with injection fentanyl 2 g / kg , injection propofol 2.5 mg / kg . \n air leak pressure after intubation was tested with the patient supine and head in a neutral position . in the absence of air leak at 20 cm h2o inflation pressure , \n the tube would be judged to be too large and replaced with the next smaller size ( 0.5 mm i d ) . \n tracheal tube with excessive air leak , not allowing adequate ventilation would be exchanged to the next larger size ( + 0.5 mm i d ) . \n cuff pressure was monitored continuously and limited to 20 cm h2o with a pressure release valve . \n minimal sealing pressure was assessed under steady - state ventilation conditions and maintained during the procedure . \n this was performed by reducing the cuff pressure until an audible leak appeared at the patient 's mouth and then pressure was increased until leak disappeared . \n anaesthesia was maintained with o2 : air ( 50:50% ) and isoflurane , using circle absorber with a mean flow of 1 l / min . \n patients were extubated after reversal with injection glycopyrrolate 0.008 mg / kg and injection neostigmine 0.05 mg / kg . \n parameters studied were size of microcuff tube used , tube passage characteristics , sealing pressure , capnography characteristics , effectiveness of ventilation , changes in cuff pressure during surgery , complications if any . \n coughing , laryngospasm or stridor at extubation was recorded . any change in voice or cry pitch was also noted . \n we studied 100 patients with a mean age of 5.44 years ( range 8 months 5 days9 years 11 months ) [ table 1 ] . \n leak on intermittent positive pressure ventilation ( ippv ) at airway pressure 20 cm h2o was present in all patients . \n mean sealing pressure was 11.72 ( 1.9 standard deviation [ sd ] ) cm h2o . with the creation of pneumoperitoneum , \n mean intracuff pressure increased to 12.48 ( 3.12 sd ) cm h2o [ table 3 ] . \n further , with head low positioning , mean cuff pressure recorded was 13.32 ( 2.92 sd ) . \n ventilation at low flow ( mean flow1 l / min ) , plateau - type capnography was noted in all patients . \n coughing at extubation was seen in 6 patients , which responded to gentle nasopharyngeal suctioning . \n partial laryngospasm occurred in 4 patients , which responded to continuous positive airway pressure ( cpap ) via face mask . \n age of study population , size of microcuff tube used and number of patients in each group type of laparoscopic surgery intracuff pressure variations \n cuffed tracheal tubes in smaller children are increasingly used because of the high probability of inserting a correctly sized tracheal tube at the first intubation attempt and to create an effective airway seal without the use of an oversized tracheal tube . \n the cuff of the microcuff tracheal tube differs from the conventional cuff in that it is made from ultra - thin ( 10 m ) polyurethane foil instead of the much thicker ( 5070 m ) polyvinyl chloride or polyethylene foils . \n the murphy eye has been eliminated , which has allowed the cuff to be moved more distally on the tracheal tube shaft . \n the cuff is short and when inflated , it expands below the sub - glottis , providing a seal with cuff pressure < 15 cm h2o . \n the airway is sealed at upper trachea where the posterior membranous wall can stretch and produce a complete seal , rather than at the cricoid level . \n thus , the problem that the cuff will cause airway mucosal injury , leading to sub - glottic stenosis is circumvented . \n creation of pneumoperitoneum and trendelenberg positioning during laparoscopy reduces pulmonary compliance , which may lead to ineffective ventilation or excessive peritubal air leak with an uncuffed tracheal tube . \n caudal displacement of the tracheal tube may be considerable during laparoscopy with pneumoperitoneum , especially with additional head - down tilt , increasing the risk of endobronchial migration of the tracheal tube . \n advantages of the microcuff endotracheal tube in this population , therefore , would be constant minute ventilation , precise respiratory monitoring and capnography , low fresh gas flow and lesser risk of pulmonary aspiration . \n selection of an appropriately - sized cuffed endotracheal tube is important to prevent airway mucosal injury . \n we were able to intubate the trachea without resistance in all patients , confirming the appropriateness of the size selection recommendations . \n air leak was observed with ippv at airway pressure 20 cm h2o in all patients . \n in a randomised controlled multicentre study of more than 2000 patients , weiss et al . concluded that microcuff tracheal tubes can be safely used in children if cuff pressure is controlled and size selection criteria followed . \n the difference in sealing pressure in our study could be due to the difference in age range or simply due to a difference in population characteristics . \n though the upper limit of safety for cuff pressure in adults is 2530 cm h2o , there is no data in children regarding perfusion pressures of the tracheal mucous membrane , and it is believed that a lower cuff pressure would possibly be safe . for the purpose of our study , effective ventilation meant a square wave capnogram , with etco2 values in the clinically acceptable range of 3545 mm hg and ability to ventilate with low flow ( < 2 l / min ) . \n pneumoperitoneum and head low positioning caused an increase in intracuff pressure ( mean increase 12.48 [ 3.12 sd ] cm h2o and 13.32 [ 2.92 sd ] respectively ) , but the pressure was always < 20 mm hg . in our study , \n creation of pneumoperitoneum and trendelenburg position did not affect ventilation or the effectiveness of the seal . \n the intra - abdominal pressures were limited to 10 mm hg in patients < 4 years and 12 mm hg in older patients . despite the head low positioning , we did not observe endobronchial intubation in any of the patients confirming the appropriateness of the depth marker . \n confirmation of appropriate tube position in our study was determined clinically , by auscultation in all patients as we did not have a paediatric flexible bronchoscope at our disposal . \n several studies have confirmed the safety of the cuffed endotracheal tube with respect to airway complications , as long as an appropriate size is chosen , and the cuff pressure is monitored . in a study of 204 patients , duracher et al . reported 6 cases of complications ( dysphonia , hoarse cough , and laryngeal dyspnea ) , and attributed those to the use of an incorrectly predicted larger tube size . \n incidence of stridor with microcuff tube in study by weiss et al . was 4.4% . in our study , at extubation \n partial laryngospasm was seen in 4 patients who were treated with cpap using a jackson - rees circuit , and humidified oxygen . \n history of recent upper respiratory tract infection ( within 4 weeks ) was present in 5 of these patients , which could have been a contributing factor for laryngospasm . \n our study confirms the appropriateness of size selection recommendations of microcuff tube in our patient population . \n advantages include better airway seal , economy of fresh gas flows , and effective ventilation .", "role": "user" }, { "content": "background and aims : traditionally , uncuffed endotracheal tubes have been used in children . \n cuffed tubes may be useful in special situations like laparoscopy . \n microcuff endotracheal tube is a specifically designed cuffed endotracheal tube for the paediatric airway . \n we studied the appropriateness of microcuff tube size selection , efficacy of ventilation , and complications , in children undergoing laparoscopy.methods:in a prospective , observational study , 100 children undergoing elective laparoscopy were intubated with microcuff tube as per recommended size . \n we studied appropriateness of size selection , sealing pressure , ability to ventilate with low flow , quality of capnography and post - extubation laryngospasm or stridor.results:mean age of the patients was 5.44 years ( range 8 months 5 days9 years 11 months ) . \n there was no resistance for tube passage during intubation in any patient . \n leak on intermittent positive pressure ventilation at airway pressure 20 cm h2o was present in all patients . \n mean sealing pressure was 11.72 ( 1.9 standard deviation [ sd ] ) cm h2 o. with the creation of pnemoperitoneum , mean intracuff pressure increased to 12.48 ( 3.12 sd ) cm h2 o. with head low positioning , mean cuff pressure recorded was 13.32 ( 2.92 sd ) . \n ventilation at low flow ( mean flow 1 l / min ) , plateau - type capnography was noted in all patients . \n mean duration of intubation was 83.50 min . \n coughing at extubation occurred in 6 patients . \n partial laryngospasm occurred in 4 patients , which responded to continuous positive airway pressure via face mask . \n severe laryngospasm or stridor was not seen in any patient.conclusion : microcuff tubes can be safely used in children if size selection recommendations are followed and cuff pressure is strictly monitored . \n advantages are better airway seal and effective ventilation , permitting use of low flows .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: the frontal bone is a singular , median , and symmetrical bone that occupies the most anterior part of the cranium . \n it has joints with the parietal , ethmoid , sphenoid , nasal , zygomatic , lacrimal , and maxillary bones , thereby contributing towards uniting the neurocranium and the viscerocranium . during the development stage , \n the frontal bone is a double bone , with right and left halves that grow together and unite along the median line at the metopic suture . \n this usually starts to undergo the fusion process at the age of two years and may have completely fused by the age of six years [ 2 , 5 ] , eight years [ 4 , 6 ] , or ten years . \n however , in approximately 8% of adults , the two halves of the frontal bone do not fuse , and the metopic suture persists . \n the suture may be incomplete or complete ( when it extends from the nasium to the bregma ) , and this condition is known as metopism [ 79 ] . \n this name originates from the greek word metopion , which means a space between the eyebrows . \n metopism has various degrees of incidence : from 7%-8% among europeans to 1% in africans and 4 - 5% in mongolians . \n overall , the range of incidence can go from 1% to 12% , and it is slightly more prevalent among males . \n the incidence of metopism and the difference in forms also vary according to geographical region . in lebanon \n it was 0.82% , in nigeria 3.14% , in india 2.66% , in southern india 6.25% , and in europe 7 to 10% [ 12 , 17 ] . prevalence of metopism is important , because , if not , this can be confused with a fracture of the frontal bone or even with a sagittal suture on radiological images [ 13 , 18 ] . \n our objective was to determine the rate of occurrence of metopism in dry crania of adult humans . \n 134 dry crania from adult humans ( 95 from males and 39 from females ) were analyzed . \n all of them belonged to anatomy laboratories in higher education institutions in macei , al , and had been obtained in accordance with law 8501 of november 30 , 1992 , which deals with use of unclaimed cadavers for teaching and research purposes . \n all the crania were macroscopically examined with regard to the presence or absence of the metopic suture . \n it was considered to be complete when it continued uninterruptedly from the nasium to the bregma and incomplete when it was not present over this entire length . \n the crania were also classified according to the cephalic index ( ci ) as follows : brachycephalic ( ci > 80 ) , dolichocephalic ( ci < 80 ) , or mesocephalic ( ci = 80 ) . \n this index was calculated by dividing the maximum width , that is , the distance between the euryons ( the widest cross - sectional distance from side to side ) , by the maximum length , that is , the distance between the glabella and the opisthocranion ( the glabella is located on the median plane , between the eyebrow arches , and the opisthocranion is the posterior point which is furthest from the longitudinal plane ) , multiplied by 100 . \n the measurements of the maximum length and width of the cranium were made using willis compasses . \n out of the 134 crania examined , the metopic suture was found in 9.7% ( 13/134 ) of the cases , of which 61.5% ( 8/13 ) were in males and 38.5% ( 5/13 ) in females . \n a complete metopic suture ( figure 1 ) was found in six cases ( 46.2% ) , while an incomplete suture ( figure 2 ) was observed in seven cases ( 53.8% ) ( table 1 ) . according to the anthropometric characteristics of the crania studied by means of the cephalic index , 47.76% ( 64 ) were of brachycephalic type , 44.03% ( 59 ) \n however , when we classified the types of cranium according to whether a metopic suture was present , 7.46% ( 10 ) were dolichocephalic , 1.49% ( 2 ) were brachycephalic , and 0.74% ( 1 ) were mesocephalic . \n there are divergences in the scientific literature in relation to the exact timing of closure of the metopic suture . \n fusion has been reported as occurring between the first year and the beginning of the second year of life , before the sixth year , in the eighth year [ 4 , 6 ] , or in the tenth year . according to sant'ana castilho et al . \n , there are several causes of failure of the metopic suture to fuse , but genetic influence is currently the factor most accepted by the scientific community . in the present study , metopism \n was found in 4.48% of the crania , which was similar to what was found in indian populations ( punjab , uttar pradesh , and southern india ) , mongolians , east asians , and nigerians ( 3.31 to 5.1% ) but was lower than what was found among scots , mongoloids , and europeans ( 7% to 10% ) ( table 2 ) . \n a study conducted in brazil by del sol et al . on 400 crania at the so paulo medical school ( escola paulista de medicina ) , federal university of so paulo , reported that the incidence of metopism was 2.75% . \n however , in an analysis on 71 crania at paranaense university , sant'ana castilho et al . found an incidence of 7% . in the present study , out of the 134 crania found in anatomy laboratories in higher education institutions in macei , al , the rate of occurrence of metopism was 4.48% and suture metopic 9.7% . \n this rate was higher than that found by either of the previous brazilian studies . among the 13 crania in which a metopic suture was seen to be present , \n the rate of occurrence was greater among males ( 61.54% ) , which corroborates the findings from studies by skrzat et al . \n , murlimanju et al . , and van der meulen , in which there was also greater occurrence among males . \n however , this was contrary to the findings of sant'ana castilho et al . , who observed that 80% of the occurrences of metopic suture were in females . according to baaten et al . \n however , when we correlated occurrences of metopic suture with the type of cranium , according to the cephalic index , we found that the occurrences in the dolichocephalic type were much greater than in the brachycephalic type . \n this differed from the findings of sant'ana castilho et al . , who reported greater occurrence in the brachycephalic type . \n on the other hand , according to bryce , what exists in reality are equal occurrences between the dolichocephalic and brachycephalic cranial types . \n the rate of occurrence of metopism in dry crania in the city of macei was 4.48% , without predominance in one sex over the other . \n incomplete metopic sutures were observed in 5.22% of the crania , with slight predominance among males . \n knowledge of the possible presence of metopism serves to alert imaging professionals to the fact that this is just a variation and does not imply any harm to the population 's health . \n moreover , this variation should not be confused with a line of fractures in the frontal bone , especially if observed close to the median line .", "role": "user" }, { "content": "the metopic suture is located between the tubercles of the frontal bone . \n there are divergences regarding the exact time at which it closes , which ranges from the first to the tenth year of life , although it may persist into adulthood . \n this study was conducted on 134 dry crania from adult brazilians , of which 95 were male and 39 were female . \n these were available in the anatomy laboratories of higher education institutions in macei , al , brazil . \n all the crania were examined macroscopically with regard to the presence ( metopism : m ) on absence of the metopic suture . \n m was considered to be complete ( mc ) when it continued uninterruptedly from the nasium to the bregma and incomplete ( mi ) when it was not present over its entire length . \n it was observed that mc was present in 4.48% ( 6/134 ) of the skull examined , of which 50% ( 3/134 ) were male and 50% ( 3/134 ) were female . \n an incomplete metopic suture was found in 5.22% ( 7/144 ) of the crania and more frequently among males ( 3.73% ; 5/134 ) . among the crania with a metopic suture , the dolichocephalic type predominated ( 7.46% ) in relation to brachycephalic crania ( 1.49% ) and mesocephalic \n crania ( 0.74% ) . \n there was no predominance of metopism between the sexes , while an incomplete metopic suture was slightly more common among males .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: jejunal diverticular disease is a rare clinical entity with an incidence of between 0.06 and 1.5% . \n the true incidence however may be higher as the majority of jejunal diverticula are asymptomatic , and thereby remain undiagnosed . in symptomatic cases , \n non - specific epigastric pain and bloating are the most common complaints [ 2 , 3 ] . \n however , life - threatening complications such as gastrointestinal ( gi ) bleeding , bowel perforation and obstruction have been reported in up to 18% of cases . \n it is therefore vital that the diagnosis of the condition can be made promptly to ensure optimal and timely management for the patient . \n we present a case of a 78-year - old male with massive gi bleeding secondary to jejunal diverticular disease who was surgically managed with good post - operative outcome . \n a 78-year - old male with a history of ischaemic heart disease , stage iv chronic kidney disease and chronic obstructive pulmonary disease , presented to the emergency department after a sudden collapse at home . \n whilst in the department , he experienced a further syncopal event with two episodes of large - volume melaena . \n he was pale , clammy and haemodynamically unstable with a systolic blood pressure of 75 mmhg . \n his blood results showed a haemoglobin ( hb ) of 66 g / l and a urea of 32.7 mmol / l . \n he was initially resuscitated with intravenous fluids , red blood cells ( rbcs ) , platelets , fresh frozen plasma ( ffp ) and cryoprecipitate . \n the patient proceeded to urgent upper gi endoscopy ( ogd ) but the procedure failed to identify any evidence of recent or active bleeding . \n colonoscopy showed large amounts of altered blood in the colon as well as the presence of sigmoid diverticula but without evidence of active bleeding . \n segmental ct - angiogram was contraindicated given patient 's poor renal function . despite vasopressor support and a proton - pump - inhibitor infusion , with on - going melaena , \n the patient 's hb and blood pressure remained low , requiring 19 units of rbcs in the first 48 h of admission . \n over the subsequent 36 h , the patient 's condition stabilized with an hb consistently over 90 g / l . \n however , on day 4 of admission , he produced five further episodes of melaena , and became acutely tachycardic and hypotensive . \n the decision was then made to proceed to exploratory laparotomy with a pre - operative plan of performing a subtotal colectomy . \n the presumptive diagnosis was that the bleeding was likely secondary to his known colonic diverticulosis . \n intra - operatively however , we found multiple diverticula clustered along the proximal and mid jejunum at the mesenteric edge . \n partially digested blood was seen in the portion of small bowel distal to these diverticula . \n we performed a small bowel resection of 80 cm length of diseased jejunum and an end - to - end anastomosis ( figs 1 and 2 ) . \n histology confirmed numerous true diverticula , with no evidence of malignancy . in total , since admission , the patient received 23 units of rbcs , 16 units of ffp , 4 units of platelets and 3 units of cryoprecipitate . \n note that the diverticula arise at the mesenteric border and are clustered around proximal jejunum . \n figure 2:intra - operative photography demonstrating jejunal diveritcula proximally and presence of altered blood in the small bowel distal to these diverticula . \n note that the diverticula arise at the mesenteric border and are clustered around proximal jejunum . \n intra - operative photography demonstrating jejunal diveritcula proximally and presence of altered blood in the small bowel distal to these diverticula . \n the patient was discharged on day 7 post - operation and followed - up in outpatients clinic 2 weeks later . \n there were no further reports of bleeding and he was discharged from surgical follow - up . \n jejunal diverticular disease is an uncommon condition that is usually asymptomatic but may rarely present with life - threatening complications including massive gi bleeding [ 2 , 3 ] . \n other complications such as chronic malabsorption , volvulus , diverticulitis with or without perforation or abscess occur in 1030% of patients [ 1 , 5 ] . \n the majority of patients who present with gi bleeding do not display previous gi symptoms and are reported to develop an acute onset haemorrhage per rectum . \n jejunal diverticulosis is often difficult to locate endoscopically and diagnosing jejunal diverticular bleeding remains problematic , with current imaging techniques continuing to be unreliable . whilst jejunal diverticulosis can be identified by abdominal ct and barium follow - through studies \n , enteroclysis remains the investigation of choice . although some reports demonstrate success with capsule endoscopy and double balloon endoscopy [ 9 , 10 ] such investigations are of limited use in emergency settings . in cases of jejunal diverticular bleeding , selective mesenteric angiography or ct angiogram \n can be used to localize active bleeding but may be contraindicated in patients ' with severe kidney impairment as with our patient . \n urgent laparotomy is indicated in the presence of acute complications of jejunal diverticula including diverticulitis , massive bleeding or bowel perforation both as a diagnostic and therapeutic measure [ 3 , 6 , 7 ] . \n several successful cases with complete small bowel resection with primary entero - entero anastomosis have been reported . \n due to low incidence , low clinical index of suspicion and unreliable diagnostic imaging in emergency situations , diagnosis of jejunal diverticular disease is challenging and therefore often delayed several days after initial presentation . \n this has major implications with regard to prompt and timely management , and can lead to significantly increased morbidity and mortality . \n whilst there are undoubtedly more common causes of gi bleeding , this case demonstrates that jejunal diverticular disease should remain on the differential diagnosis and investigations to confirm the diagnosis should be considered . \n however , despite investigations , the diagnosis may remain elusive and in patients with on - going bleeding , laparotomy and surgical resection is currently the treatment of choice [ 3 , 6 , 7 ] .", "role": "user" }, { "content": "jejunal diverticula are rare and the condition remains mostly asymptomatic . \n however , they can present with vague chronic abdominal symptoms and , in some cases , acute life - threatening complications , such as gastrointestinal ( gi ) bleeding , bowel obstruction and perforation . \n we present a case of an adult male who presented with life - threatening gi bleeding secondary to jejunal diverticular disease . \n whilst there are undoubtedly more common causes of gi bleeding , this case demonstrates that jejunal diverticular disease should remain on the differential diagnosis and investigations to confirm the diagnosis should be considered . however , despite investigations , the diagnosis may remain elusive and in patients with on - going bleeding , laparotomy and surgical resection is currently the treatment of choice .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: while noise can lead to serious hearing - related health problems , it can also cause general health problems such as attention impairment , anxiety , sleep disorder , and depression . \n however , the best known of these problems is noise - induced hearing loss which is commonly studied due to its irreversible character . \n approximately , 16% of adult cases of hearing loss are believed to be associated with workplace noise exposure . \n although noise is a common hazard in almost all occupations and in nearly all workplaces , some sectors such as mining , metalworking , textile , agriculture , or transportation involve a higher level of noise exposure . \n furthermore , certain occupations such as musicians , traffic polices , and soldiers are also deemed as high - risk groups with regard to noise exposure . in recent years \n , new high - risk groups have emerged due to the development of new sectors . among those , \n call centers are business units that provide product or service support to the clients , suppliers , agents , and other third persons via phone calls by way of giving instant help or referring to the related units . in 2010 , call center sector has reached a market value of 340 million dollars worldwide , including 11.5 million call center operators . in turkey , \n the most common occupational health consequences can be categorized as visual problems due to working with a video display units , voice deformation due to continuous talking , and auditory problems due to intense headset use and acoustic shocks . however , there is scarce research on noise - induced hearing loss among call center operators . in this case report \n , we presented a case of noise - induced hearing loss in a call center operator who was evaluated in the dokuz eylul university ( deu ) , occupational disease outpatient clinic . \n 30-year - old male patient was admitted to the department of ear nose throat in deu hospital with the main complaint of not hearing the safety belt alarm in his car . \n subsequent to his history taking , he was referred to occupational disease outpatient clinic in the hospital . \n he has been working as a home agent for a call center for the last 50 months . \n the system is accessed by a password and the incoming calls are directed by team leaders to the related operators . \n he was mainly serving clients from mechanical maintaining and repair sector clients , thus he described frequent high pitch noise and parasite particularly at the beginning of the conversations . \n i have to control the noise level manually since i do not have the amplificatory device . in his occupational history , he describes the time between high noise level and parasite and manual modification as 3 - 5 s. he also described ear pain after some of these exposures . \n a call is received every 3 - 5 min ; 17% of the work time is passed in passive mode , waiting for a call . \n he also mentioned other employees with hearing problems during occupational history , but we did not have a chance to examine any other employees yet . \n he reported no periodical health examinations and very limited occupational health and safety training was given to him only at the beginning of the job . \n although we requested personal noise exposure and environmental noise measurements from the company , no answer was received . \n there was no history of smoking or alcohol use , regular drug use , autotoxic drug use , chronic diseases , and exposure to high - pitched noise such as the explosion in his personal and family history , and no particular noise exposure during his military service . \n there was no history of diving , shooting , or listening to music at high volumes . \n his daily activities do not include any high - level exposure yet we tried to categorize his daily exposure according to the national regulations . \n he does describe a very sedentary everyday routine including limited time in traffic ( daily city traffic 70 db and automobile horns 80 db ) . \n i prefer to stay in a quite environment thus i do not listen high volume music or television ( radio music sound 70 - 80 db ) . the case was evaluated in consultation with the department of ear nose and throat . \n the physical examination revealed normal autoscopic results in both ears , the weber was lateralized to the right , and the rinne test was pathologically positive in both ears . \n a pure sound audiogram showed a mean pure sound of 27 db loss in the right and a mean pure sound of 35 db loss in the left ear , while there was a sensorineural hearing loss manifesting a sudden decline after 1000 hz [ figure 2 ] . \n in accordance to the history , physical examination , and audiometric results along with the initial audiometric test , other reasons were excluded , and the patient has been diagnosed as noise - induced occupational hearing loss . he was recommended to reduce the noise exposure and put under yearly follow - up by the committee of occupational diseases . \n call center activities ( nace code : 82.20.01 ) is regarded as hazardous according to the workplace hazard classes of the work health and safety regulation issued by the ministry of labor and social security in turkey . \n exposure limit and exposure threshold are set to 87 db ( a ) and to 85 db ( a ) , respectively , in the legislation . \n according to the widely recognized definition , a value higher than 25 db at hearing thresholds of 1 , 2 , 3 , and 4 khz is regarded as hearing loss . \n occupational hearing loss requires having worked at least 2 years in a noisy workplace or having worked in a workplace emitting a noise higher than 85 db ( a ) at least for 30 days . \n however , despite values stated in the laws , in 2010 , verbeek et al . reported that prolonged noise exposure to a noise of 80 db ( a ) may constitute a risk for hearing loss . \n they also reviewed the suggestions aiming to prevent noise - induced hearing loss and the efficacy of programs built to prevent hearing loss were found to have a very little effect , highlighting the need for more efficient legal regulations . \n the results of personal noise exposure and environmental noise measurements could not be obtained in our case . \n the company did not let us to measure but in a study performed among 150 call center operators in an office environment in britain , personal noise exposure level varied between 68 and 77 db . \n authors particularly highlighted the importance of in - house training and emphasized the necessity of accurate headset use along with taking the required precautions . \n trompette and chatillon evaluated 117 operators from 21 different call centers revealing that personal noise exposure level was 60 - 90 db ( a ) and the environmental noise level was 50 - 62 db ( a ) . \n however , these studies and similar ones in the literature have been performed on call center operators in an office environment . \n therefore , since noise sources such as fax machine , printer , and air conditioner are not present at home , background noise may be lower among home agents . \n however , there was no study focusing on home - office agents in the literature . \n another major risk other than the average noise exposure was acoustic shock defined as exposure to a sudden and disturbing loud sound . \n a study in australia evaluated acoustic shock in 103 call center operators and noted the presence of symptoms such as headache , dizziness , tinnitus , and hearing loss after work hours . \n similarly , trompette and chatillon underscored the significance of equipment selection , noting that many pathologies such as acoustic shock can be eliminated by using correct and safe headsets . \n they also reported the importance of employee training and recommended repeated trainings until the employees are completely adapted to the headset . \n our case had no amplificator in the headset , and he had not received adequate training on occupational health issues which were related as the possible causes of acoustic shock . \n there were no recruitment and follow - up examinations and yet there were neither engineering noise control nor sufficient training for hearing protection in our case . \n furthermore , the flexible and precarious nature of home - based jobs is still out of standard occupational health and safety inspection mechanisms . \n therefore , our case is provided a clear evidence for the need of comprehensive occupational health and safety services for call center workers . \n most of the home office workers are not willing to speak out about their work - related health problems since the sector has a high employee turnover rate . when they are out of a particular job , it is very hard to trace such workers for work - related health problems . \n regardless of the work environment , noise control programs should be developed and applied in order to prevent hearing loss at home and in call centers . \n moreover , it should be borne in mind that noise may lead to nihl and other health problems even at values not surpassing the predetermined limits . \n \n ", "role": "user" }, { "content": "noise is defined as unwelcome sound . it has been estimated that 16% of adult hearing loss in the world is due to noise exposure at the workplace . \n this report offers a case that diagnosed with hearing loss of whom working as a call center operator at home . \n home agent operators should be explored .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: they are also referred as congenital granular cell tumor , congenital granular epulis , congenital granular cell myoblastoma and congenital granular cell fibroblastoma . although these lesions are benign , they need immediate surgical intervention because they cause interference in feeding and have the potential to cause the death of the child from asphyxia during the perinatal and postnatal period . \n the most common site is the alveolar ridges of maxilla and mandible with a marked predilection of occurrence in females . \n the lesion is 3 times more frequently seen in maxilla than in mandible and the female : male ratio is 10:1 . \n the incidence rates were found to be 0.0006% at a center in wales and epulis accounted for 10.8% of all the oral lesions in a center in india . \n mostly , they are solitary but , in some cases , large and multiple tumors are shown . addressing the problem may need a multidisciplinary team approach at the time of birth ; herein , we report a case of bilobed congenital epulis of the newborn arising from the maxillary alveolar ridge . \n a 3-day - old female full - term infant weighing 2.45 kg at birth was admitted with complaints of a mass in the oral cavity measuring about 4.3 cm 3.2 cm . \n clinical examination of the newborn revealed the presence of a pink , bilobed , pedunculated , nontender smooth surfaced mass with a firm consistency arising from the right side of the maxillary alveolar ridge [ figure 1 ] . \n respiratory distress was not evident , but the child had sucking problems . in discussion with the pediatrician and anesthetist , \n after preoperative evaluation , surgical excision of the mass was done by the use of cautery after orotracheal intubation under general anesthesia [ figure 2 ] on the 5 day of life without any complications . \n the examination of the resected specimen showed a bilobular , encapsulated , smooth mass which was pedunculated . \n on histological examination , the tissue was seen to be composed of sheets of large polygonal granular cells with distinct borders , having abundant granular eosinophilic cytoplasm and predominantly eccentrically located vesicular nuclei with conspicuous nucleoli [ figures 3 and 4 ] . \n infant was able to commence feeding on the second postoperative day and was discharged on his 8 day of life with normal closure of the mouth and was able to suck normally . \n the postoperative period was uneventful with infant not showing any signs of recurrence after 4 months of follow - up . \n tumor mass attached to the alveolar ridge of maxilla postoperative clinical image demonstrating the site of the wound . \n microscopic examination of excised specimen revealing polygonal cells with granular cytoplasm ( h&e stain , x100 ) microscopic examination of excised specimen demonstrating polygonal cells with granular cytoplasm and vesiculated nuclei ( h&e stain , x400 ) \n congenital epulis also known as congenital granular cell tumor , congenital granular epulis , congenital granular cell myoblastoma and congenital granular cell fibroblastoma is a very rare condition among newborn and is predominant in females . \n the tumor is benign with no clear etiology but is reported to be hormone - related although other theories do exist . \n histological and electron microscope studies suggest a reactive theory in which the tumor results from stromal gingival cells such as fibroblasts or histiocytes . \n the cases of spontaneous regression in the literature and the absence of recurrence after incomplete tumor resection support this theory . \n they are mostly recognized at birth or just after birth except in cases where the size is very small and , therefore , absence of symptoms . \n prenatal diagnosis remains difficult because of the absence of specific signs and also as the tumor generally develops beyond the 22 week of gestation . \n fetal three - dimensional ultrasound and magnetic resonance imaging ( mri ) can provide the diagnosis by the 36 week of gestation , thereby helping to decide and plan in advance an early multidisciplinary management of the newborn with participation from the neonatologist , the maxillofacial or head and neck surgeon or the pediatric surgeon and the anesthesiologist . \n classically , there is a single firm tumor with a quite regular surface , sometimes multilobed , sessile or pedenculated , pink or red in color and nonpainful on palpation . \n the tumor size varies from a few millimeters to around 10 cm at its widest diameter . \n the maxillary location opposite the future canines or incisors is the most frequent , but the mandibular region can also be involved . \n multiple lesions or maxillary and mandibular locations have been described in 516% of the cases . \n although benign , immediate surgical intervention is required because of risk to patient 's life due to asphyxia . \n some other common clinical manifestations include dyspnea , cyanosis , cough , difficulty in sucking and swallowing and occasionally , vomiting . \n perinatal mri of the oral cavity helps in differentiating congenital epulis from other masses of oral cavity and in determining characteristics of the mass . \n literature shows no evidence of surgical excision affecting the growth of the bone or the eruption of teeth . \n surgery can be performed under local or general anesthesia and the choice between the two depends on the size , number and location . \n authors have shown preference for general anesthesia when tumor size is large or is present at multiple locations . with single and sessile tumor local anesthesia \n , we performed the excision under general anesthesia because of large and pedunculated size of tumor . \n histological examination of the tumor specimen demonstrated large polygonal granular cells with distinct borders , having abundant granular eosinophilic cytoplasm and predominantly eccentrically located vesicular nuclei with conspicuous nucleoli . \n congenital epulis may regress spontaneously but requires surgical removal due to problems in feeding and respiration . \n it is important that general dentists and pediatricians recognize this lesion and give relevant information regarding the treatment . \n \n ", "role": "user" }, { "content": "congenital epulis , a benign tumor of the oral cavity , is an extremely rare condition in newborn . \n it may lead to mechanical obstruction , therefore resulting in respiratory distress and difficulty in feeding . addressing the problem \n may need a multidisciplinary team approach at the time of birth . \n antenatal ultrasonography and perinatal magnetic resonance imaging are an adjunct to treatment planning . \n prenatal diagnosis remains difficult as the findings are nonspecific due to the late development of the tumor . \n surgical excision is , therefore , the treatment of choice . \n our report discusses this condition and the treatment thereafter on a newborn , with an epulis originating from the upper alveolar ridge discovered at birth . \n histological examination confirmed the diagnosis of large polygonal granular cells . \n the mass was excised under general anesthesia , and the outcome was good after surgery allowing regular feeds on the second postoperative day .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: minimally invasive adrenalectomy by the laparoscopic or the retroperitoneal approach has become the gold standard procedure in cases of functioning or nonfunctioning lesions of the adrenal gland since its introduction in 1992 . \n partial adrenalectomy was introduced by irvin et al in 1983 when the open technique was the only option . \n walz et al included a tissue - sparing endoscopic technique in select patients in his series in 1996 . \n bilateral synchronous surgical pathology to preserve vital adrenal volume is the absolute indication for partial adrenalectomy . \n however , cortical - sparing adrenalectomies have been described by several authors for small solitary tumors emphasizing the benefits of the procedure , which is less traumatic and potentially less hazardous to patients , especially in right - sided lesions ( avoiding a possible torsion of the right adrenal vein ) ; moreover , preserving adrenal tissue allows total adrenalectomy in cases of future pathology of the contralateral gland . \n the potential benefit of partial adrenalectomy is obvious not only in hereditary but also in sporadic diseases as well . \n indications for partial adrenalectomy have yet to be defined along with technical aspects , such as the extent of the excision and the preservation or not of the adrenal vein . \n preservation of the adrenal vein ensures that the adrenal remnant will drain to the circulation . \n however , some authors advocate division of the vein but careful preservation of the minor venus plexuses of the retroperitoneal space instead . in this article , we present a short series of 7 patients in whom partial adrenalectomy was successfully performed without division of the adrenal vein . \n we retrospectively reviewed patients who underwent a partial adrenalectomy in our department between january 2002 and january 2006 . \n the pathology of the patients reviewed included : peripherally located aldosterone - producing adenomas and peripherally located myelolipomas . \n there were 5 female and 2 male patients with a median age of 54 years . \n there were 5 right - sided ( all 4 myelolipomas and 1 adenoma ) and 3 left - sided ( all 3 adenomas ) lesions . \n the following data were recorded : patient 's age and sex , preoperative diagnosis , final tumor size , intra- and postoperative complications , operation time , final histological diagnosis , and length of postoperative stay . \n scans using 5-mm slices were used to assess the location and size of the tumor . \n the preferable surgical approach was the lateral transperitoneal laparoscopic procedure , placing the patient in the lateral decubitus position with the affected side up . \n this approach offers a large working space , easier removal of large tumors , and a facile conversion to open adrenalectomy if necessary . after meticulous examination of the adrenal gland and the visualization of the lesion , \n the proximity of the lesion to the adrenal vein was assessed and a partial adrenalectomy was decided upon , provided that a 5-mm margin of healthy tissue from the tumor edge would be kept and that the adrenal stump would be at least 20% of the normal gland . \n the dissection was initiated from the tumor side of the gland . full mobilization of the gland with a harmonic scalpel was necessary before dividing the gland with a vascular stapler . \n the main adrenal vein and artery were preserved in 6 patients and the accessory vein with its corresponding artery in one . \n the median preoperative estimation of the size of the lesion was 6.5 cm ( range , 3 to 12 ) for the myelolipomas and 11 mm ( range , 4 to 29 ) for the aldosterone - producing adenomas . \n operating time , blood loss , tumor size , and diagnosis are summarized in table 1 . \n median hospital stay was 2 days , and median follow - up was 3.6 years for the myelolipoma patients and 23 months for the patient with aldosterone - producing adenomas . \n three of the adenoma patients are normotensive 44 , 23 , and 20 months postoperatively , while the fourth one 's pressure is refractory and is controlled with antihypertensive drugs at present . \n partial laparoscopic adrenalectomy proved a safe operation in 7 patients , resulting in complete removal of the lesion , in cases of hormone - producing adenomas and in cases of nonfunctioning benign incidentalomas . despite having histological confirmation of removal of the aldosteronoma , one failure to achieve postoperative normotension \n is attributed to possible cortical hyperplasia , but the patient 's aldosterone levels are marginal for the time being . \n aldosterone - producing adenomas are ideal for partial adrenalectomy because the tumor is usually small , solitary , almost always benign , and frequently peripherally located in the outer surface of the cortex of the adrenal gland . during recent years , surgeons and urologists have presented series of patients suffering from aldosteronomas and bilateral pheochromocytomas treated by partial adrenalectomy . \n walz et al included 100 partial retroperitoneoscopic approaches with comparable results to total adrenalectomies regarding cure rate . \n a functioning stump of at least one third of the initial adrenal volume should be left in place . \n preservation of the adrenal vein has been used during partial adrenalectomy for adenomas , because adequate venous drainage is important to preserve function of the remaining adrenal stump . \n others propose division of the adrenal vein and rely on the retroperitoneal vein plexus for the maintenance of adrenal function . \n adrenal cortical function does not necessarily require an intact vein , but ligation of the vein requires expensive postoperative investigations to confirm that the retroperitoneal plexus is sufficient for drainage and if not , the adrenalectomy is anatomically partial but functionally total . furthermore , if the vascular stapler is utilized , mobilization of the gland , which is rather compulsory , carries a high risk of destroying the plexus . \n we emphasize that preservation of the vein in a partially resected gland results in less congestion compared with congestion in a partially resected gland with ligation of the vein in which drainage relies only on the retroperitoneal plexus . \n we state that partial adrenalectomy is time sparing and potentially less hazardous compared with total adrenalectomy but not compared with partial adrenalectomy with vein ligation . \n exact preoperative knowledge of the anatomical variations of the adrenal vein is of great importance but difficult to obtain , so meticulous dissection at the area of the inferior vena cava , for those that advocate vein ligation , is mandatory and not always fruitful . on the other hand \n , retroperitoneal plexus preservation can be extremely difficult to preserve , particularly in small glands . \n nonfunctioning , benign tumors of the adrenal gland if conveniently situated are not a contraindication to partial adrenalectomy . in select cases , such as peripherally located , benign and solitary tumors where total excision of the gland is not mandatory , a cortical - sparing technique with preservation of the vein is a promising alternative to total removal of the gland ( figure 1 ) . in our experience with more than 40 laparoscopic adrenalectomies \n partial adrenalectomy in select cases must be used by surgeons who are capable of performing a total adrenalectomy . \n we believe that the final decision for partial or total adrenalectomy should be made intraoperatively . \n preservation of the adrenal vein may not be compulsory , but potential loss of the contralateral gland in the future ( in case of a nonfunctioning stump ) makes it desirable . \n peripherally situated , nonfunctioning , benign tumors of the adrenal gland , where total excision of the gland is not mandatory , can be treated with partial adrenalectomy . \n aldosterone - producing adenoma patients are ideal candidates for cortical - sparing adrenalectomy , due to tumor location and size . \n the vein - preservation technique ensures a functioning stump , provides better hemostasis due to adequate drainage avoiding congestion of the gland , omits a time - consuming and potentially hazardous step of the operation , and finally gland mobilization allows application of a vascular stapler , reducing operation time . the final decision for partial or total adrenalectomy \n preservation of the adrenal vein may not be compulsory , but potential loss of the contralateral gland in the future ( in case of a nonfunctioning stump ) , makes it desirable .", "role": "user" }, { "content": "background and objective : adrenal tissue - sparing or partial adrenalectomy evolved initially for patients with bilateral synchronous adrenal surgical pathology to preserve vital adrenal volume . in the laparoscopic era , the exact criteria for performing such procedures laparoscopically have yet to be defined . \n controversy exists regarding the importance of preserving the adrenal vein , main or accessory . \n the aim of this retrospective study was to present our short series of laparoscopic tissue - sparing adrenalectomies with vein preservation . \n our main goal is not to support partial adrenalectomy as an alternative to total ( this is already advocated by many surgeons ) but to emphasize the vein - preserving technique.methods:seven patients with peripherally located either aldosterone - producing adenomas ( 4 cases ) or myelolipomas ( 4 cases ) underwent laparoscopic lateral partial adrenalectomy . \n one patient harbored an aldosterone - producing adenoma and a myelolipoma as well . \n the main adrenal vein was identified and preserved in 6 patients and the accessory vein in one.results:no conversion to open adrenalectomy was necessary , and no perioperative morbidity or mortality occurred . \n three adenoma patients are normotensive 44 , 23 , and 20 months postoperatively , while the fourth one 's pressure is refractory.conclusions:surprisingly , total adrenalectomies preceded the partial ones , which is controversial compared with other procedures . \n laparoscopic lateral partial adrenalectomy is a technically challenging tissue - sparing operation . \n meticulous dissection allows preservation of the middle artery and main or accessory vein resulting in a functioning adrenal stump .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: n - glycosylation \n sequons of proteins ( asn - xaa - ser / thr , where xaa \n is any amino acid except proline ) typically carry a diverse repertoire \n of structures , called glycan microheterogeneity . \n the diversity of protein glycosylation is influenced by \n the activity of many enzymes involved in the n - glycan biosynthetic \n machinery . \n the source and the location of the glycoproteins \n also strongly influence the final glycan structure and site specificity . \n microheterogeneity of protein n - glycoforms changes \n significantly in cancer diseases often in connection with changes \n in the activity of enzymes involved in n - glycoprotein synthesis . \n one of the best documented examples of aberrant glycosylation is \n the progression of liver disease to hepatocellular carcinoma ( hcc ) . in the case of liver disease , \n changes in protein n - glycosylation \n can be analyzed efficiently by serologic monitoring of liver - secreted \n glycoproteins . reported changes of n - glycan repertoire in cancer \n include the formation of bisecting glycans with one molecule of n - acetylglucosamine attached via 14 bond \n to the branched core mannose , asialo and agalacto glycans , truncated \n structures with terminal galactoses or n - acetylglucosamines , \n ( poly)-n - acetyllactosamines with beta 16 \n branching , and fucosylated glycans with fucose attached mainly at \n the c-3 position of outer arm n - acetylglucosamine \n or less frequently at the c-6 n - acetylglucosamine \n of the n - glycan core . \n we have previously detected multiply outer arm fucosylated \n glycoforms \n of haptoglobin ( hp ) in the context of hcc . \n we have now isolated three additional glycoproteins from serum of \n a patient that carries hyper - fucosylated hp ( up to six fucoses per \n glycan ) . \n these liver - secreted glycoproteins copurify on hemin agarose \n and include hemopexin ( hpx ) , kininogen-1 ( kng-1 ) , and complement factor \n h ( cfh ) . \n our assumption is that the same glycosylation machinery that \n hyper - fucosylates hp would lead to hyper - fucosylation of additional \n proteins . \n we report the analysis of the distribution of n - glycans \n between the four proteins and on specific glycopeptides using two \n complementary analytical methods . \n we have used sialidase - assisted \n tandem mass spectrometry of glycopeptides to study the site - specific \n protein glycoforms and further determined by esi - itmsn the analysis \n of permethylated glycans detached from each of the purified proteins . \n this is to our knowledge the first case report of detailed structural \n characterization of site - specific glycoforms of proteins isolated \n directly from patient serum . \n proteins were isolated from serum of \n a patient previously shown to carry multiply fucosylated glycoforms \n of hp . \n this patient was enrolled under \n protocols approved by the georgetown university institutional review \n board in collaboration with the georgetown university hospital , department \n of hepatology and liver transplantation , washington , dc . \n the patient \n is a 61 year old caucasian male , blood group o , diagnosed by the attending \n physician as a cirrhotic patient of hcv etiology with hcc stage t2n0m0 , \n according to the seventh edition of the american joint committee on \n cancer staging manual . \n hpx , \n kng-1 , and cfh were isolated by incubation of 500 l of human \n plasma with 400 l of heme - agarose beads ( sigma aldrich , st . \n the resin \n was washed with 10 ml of pbs buffer , and bound glycoproteins were \n released by the addition of 500 l of 0.2 m citric acid , ph \n 2 and immediately neutralized by 270 l of 1 m tris / cl buffer , \n ph 9 . \n proteins were further separated by reversed - phase chromatography \n using c18 macroporous mrp hi - recovery protein column ( 4.6 50 \n mm , agilent technologies , santa clara , ca ) under the following conditions : \n 0 min 1% b , 5 min 35% b , 30 min 45% b , 31 min 100% b , 32 min 100% \n b , 33 min 1% b , 45 min 1% b ( a : 2% acn ( fisher scientific , fair lawn , \n nj ) , 0.1% tfa in water ( fisher scientific , fair lawn , nj ) , b : 98% \n acn , 0.08% tfa in water ) , flow rate 0.75 ml / min , and column was heated \n to 40 c . \n the cfh fraction was collected in the time range 9.7 to 10 \n min , kng-1 fraction in the time range 13.515 min , and hpx \n fraction in the time range 17.518.5 min . \n isolated glycoproteins \n were resuspended in 20 l of 50 mm nh4hco3 ( sigma - aldrich ) at ph 7.8 with 0.05% rapigest ( waters , milford , \n ma ) . \n proteins were reduced with 5 mm dtt ( sigma - aldrich ) and alkylated \n with 15 mm iodoacetamide ( sigma - aldrich ) . \n trypsin ( promega , madison , \n wi ) digestion ( 2.5 ng/l ) was carried out at 37 c in barocycler \n nep2320 ( pressure biosciences , south easton , ma ) for 1 h. protein \n digests were loaded on a micro trap peptide cartridge ( michrom bioresources , \n auburn , ca ) and washed three times with 250 l of 0.1% aqueous \n tfa . \n peptides were eluted with 100 l of 60% acn with 0.1% tfa \n and the eluate was dried using a speedvac concentrator . \n for exoglycosidase \n treatment , peptides were resuspended in 20 l of reaction buffer \n containing 50 mm sodium citrate , ph 6.0 . \n two microliters ( 100 units ) \n of neuraminidase ( new england biolabs , ipswich , ma ) was added to peptide \n digests derived from 2 g of each glycoprotein and incubated \n at 37 c for 20 h. peptides were desalted as previously described . \n mixture of \n tryptic peptides was \n resuspended in 40 l of 2% acn , 0.1% formic acid and analyzed \n by nanolc \n ms / ms using a c18 column ( 1.7 m \n particles , 75 m i d 150 mm , waters ) coupled to a qstar \n elite mass spectrometer ( applied biosystems , foster city , ca ) . \n two \n picomoles of each glycoprotein digest were separated at a flow rate \n of 0.4 l / min , using the following gradient : 0 min 1% b , 35 \n min 35% b , 37 min 90% b , 42 min 90% b , 44 min 1% b , 55 min 1% b ( solvent \n a : 0.1% formic acid in 2% acn ; solvent b : 0.1% formic acid in 98% \n acn ) . \n the mass spectrometer was operated in data - dependent mode ; after \n the full - scan ( m / z 400 to m / z 1800 ) survey , the three most intense \n precursor ions were selected for collision - induced dissociation . \n collision \n energy and ms / ms accumulation time were set automatically , and the \n ms / ms spectra were recorded from m / z 150 to m / z 2000 . \n dynamic exclusion \n was set at 15 s and five counts for two repeated precursors . \n fragment \n intensity multiplier was set at 16 with maximum accumulation 2 s , \n which resulted in a total cycle time of 6.5 s. hp , \n hpx , and cfh purified as previously described ( 50 g each ) were \n enzymatically deglycosylated with n - glycanase at 37 c overnight \n ( prozyme , hayward , ca ) . \n released n - glycans were purified by c18 and \n porous graphitized carbon solid - phase extraction steps as described \n previously . \n briefly , n - glycans were reduced \n using borane - ammonia and permethylated using spin columns , as previously \n described . \n maldi - tof analysis was \n performed on a shimadzu kratos axima - cfr ( columbia , md ) using dhb \n as a matrix . \n detailed mass spectrometric analysis was performed \n on a thermo ltq ( san jose , ca ) mass analyzer equipped with an advion \n triversa nanomate ( ithaca , ny ) . \n samples were dissolved in 1:1 methanol / water \n for direct infusion into the ltq . \n all ions were sodium adducts , and \n select fucosylated compositions were disassembled to determine detailed \n structure as described . \n lc ms / ms of glycopeptides : \n analyst qs 2.0 software ( applied biosystems , foster city , ca ) was \n used to process ms data sets , and glycoproteins were identified by \n mascot search engine . \n ms / ms spectra were converted to mzxml format using msconvert \n from the proteowizard project . \n glycopeptidesearch ( gps ) engine was \n used for finding of glycopeptides forms for each glycoprotein . \n peak area counts from extracted ion chromatogram \n ( xic ) of the precursor ion of each identified glycoform were used \n for calculation of percentage distribution of glycopeptides and also \n for calculation of ratio between fucosylated and nonfucosylated glycoforms . \n our recent analysis of hp isolated from serum of a patient with \n hcc detected multiply fucosylated glycans with up to six fucoses . \n ms / ms to show the presence \n of lewis y type glycoforms at one glycopeptide of hp . in this study \n ms / ms \n methods and msn structural analysis of detached n - glycans to compare \n glycoforms of hp with glycoforms of hpx , kng-1 , and cfh isolated from \n plasma of the same patient . \n our goal is to compare site - specific microheterogeneity \n of proteins synthesized by the same enzymatic machinery and under \n the same pathophysiological conditions . \n the proteins were purified \n by reversed - phase chromatography after incubation of the patient s \n plasma with heme - agarose beads . \n isolation of these proteins as a complex \n on heme - agarose is expected because all three proteins have high affinity \n for either heme or each other . because \n n - glycans have minimal interaction with the reversed - phase resin , \n all glycoforms of the protein coelute and their microheterogeneity \n hpx and cfh were \n both purified to apparent homogeneity , but kng-1 coeluted from the \n reversed - phase column with histidine - rich glycoprotein and other minor \n contaminants ( figure 1 ) . \n we were able to analyze \n glycopeptides of kng-1 efficiently in the simple coeluting mixture \n of proteins , but we were not able to isolate sufficient amounts of \n purified kng-1 for msn structural analysis ; msn analysis was therefore \n completed on hp , cfh , and hpx . \n we have focused on lc ms analysis \n of tryptic digests because trypsin provides sufficient coverage of \n each protein s glycosites to allow comparison of microheterogeneity \n and because the tryptic digest is standardized sufficiently to allow \n semiquantitative comparison of precursor ion intensities of the microheterogeneous \n glycopeptides . \n we describe the first results of analysis of each protein \n and follow the description by comparison of the results across the \n proteins . \n uv detection ( 214 nm ) of a glycoprotein complex isolated by heme - agarose \n affinity chromatography and separated by reversed - phase chromatography . \n peaks containing the proteins of interest are numbered as follows : \n 1-cfh , 2-kng-1 , and 3-hpx . \n haptoglobin : hp is a liver - secreted glycoprotein with four \n n - glycosylation \n sites . its major function is capture of released hemoglobin during \n intravascular hemolysis . \n our recent data revealed that hp isolated \n from serum of hcc patients carries up to six fucoses per glycan . in the present study , we have used neuraminidase \n assisted lc \n ms / ms to confirm that all glycosites of hp carry \n complex glycans with variable proportion of bi- , tri- , and tetra - antennary \n structures and variable degree of fucosylation ( table 1 ) . \n tri - antennary structures on peptide v(218)k(233 ) \n are as abundant as bi - antennary structures . \n glycan abbreviations are adopted \n from the nibrt glycobase databases ( a2g2 , bi - antennary glycan terminated \n with two galactoses ; a3g3 , tri - antennary glycan terminated with three \n galactoses , etc . \n ; f , fucose ; we do not resolve core and outer arm ) . to further refine structure \n assignments , we have used msn analysis \n of detached permethylated n - glycans . our previous study used glycosidase - assisted \n lc ms / ms to show that hyper - fucosylated glycoforms of hp contain \n mostly lewis - x- and lewis - y - type glycoforms \n . detailed msn structural analysis of tetra - antennary doubly fucosylated \n glycan with monosaccharide composition ( hex)4 ( hexnac)4 ( deoxyhexose)2 \n ( neuac)2 + ( man)3(glcnac)2 supports the \n presence of lewis x , h2 , and lewis y glycoforms . \n dissociation ( ms ) of this doubly fucosylated permethylated glycan at m / z 1366 ( 3 + ) gives lewis type fragments \n at m / z 834 and 660 . \n both fragments \n were further dissociated ( ms ) and product ion at m / z 646 ( derived from 834 fragment ) was \n further collided ( ms ) . \n msn analysis shows at least three \n possible isomers at this composition , with both core and antennal \n fucosylation . \n figure 2 shows selected msn spectra \n for the reduced and permethylated tetra - antennary , disialylated , and \n doubly fucosylated composition ( m / z 1366 ( 3 + ) ) . \n figure 2a shows the ms2 spectrum \n of this composition , with insets displaying zoomed in views of the \n terminal ( b2 ) and internal ( b3/y ) lactosamine fragments as well as \n the y1 fragment ( m / z 490 ) indicative \n of the fucosylated reducing - end glcnac ; the second inset shows the \n terminal doubly fucosylated lactosamine ( m / z 834 ) and the sialylated lactosamine ( m / z 847 ) fragments . \n the ms2 spectrum also clearly indicates two different antennal fucosylation \n fragments : the terminal ( b - type ) monofucosylated fragment at m / z 660 and the terminal ( b - type ) difucosylated \n fragment at m / z 834 . because there \n are several structural isomeric possibilities for these fragments , \n these ions were subjected to further msn interrogation . \n the ms3 fragmentation \n of the m / z 660 ion ( figure 2b ) is indicative of a mixture of lewis x and h2 \n epitopes ( figure 2 ) . \n these isomeric glycans \n differ in fucose position and linkage and are particularly difficult \n to separate by lc ms analysis ; msn fragmentation is a complementary \n method that resolves the linkage efficiently . \n the specific diagnostic \n fragments are the m / z 259 , a c - type \n terminal hexose that can arise from a lewis x structure but not an \n h2 structure , m / z 329 , a a cross - ring cleavage fragment that can only arise from lewis x , \n not h2 or lewis a , and the m / z 503 , \n another a cross - ring cleavage fragment which can only \n arise from h2 , not h1 or lewis a or x. detailed structures with fragment \n identifications are in the supporting information . the low - intensity fragment at m / z 834 ( terminal difucosylated lactosamine ) in the ms / ms spectrum is \n consistent with the possible lewis y epitope . \n disassembly of this \n ion through ms3 , m / z 834 ( figure 2c ) and ms4 , m / z 834 646 ( figure 2d ) are consistent \n with the lewis y epitope . \n the ms3 and \n ms4 spectra both contain the m / z 503 fragment , a a cross - ring cleavage ion , of the \n same structure as that found in h2 , which distinguishes this from \n lewis b. because this fragment is of \n the same structure as that from h2 , the fragmentation pathway , through \n the m / z 834 ion , is of great importance \n in assigning this structure . \n structural analysis of permethylated tetra - antennary \n doubly fucosylated \n glycan ( m / z 1366 ( 3 + ) ) from hpt \n by msn fragmentation . spectrum a shows the ms2 spectrum with zoomed - in \n views of the core fuc - glcnac ( y1 , m / z 490 ) and lewis y ( m / z 834 ) fragments . \n panel b shows the ms3 spectrum of m / z 660 , showing a mixture of lewis x and h2 structures . \n panels c and \n d show ms3 and ms4 spectra , respectively , of the lewis y epitope disassembly . \n we were not able to detect any core - fucosylated \n glycoforms in hp \n by lc \n the low abundant core - fucosylated glycoform detected \n by msn ( see above ) is either a minor component below the detection \n limit of our lc \n we can not exclude that low percentage of core fucose exists \n especially on the doubly fucosylated site , but it is clear that core - fucosylated \n glycoforms represent at most a very minor component of the hp glycoforms . \n hemopexin : hpx is a heme - binding 60 kda plasma glycoprotein containing \n five n - glycosylation sites and one n - terminal o - glycosylation site . \n its major biological role is defense against hemoglobin - mediated oxidative \n damage during intravascular hemolysis . \n each glycopeptide is occupied by complex glycans , primarily \n bi- and tri - antennary glycoforms with up to two fucoses . \n bi - antennary \n glycans dominate , and their fucosylated counterparts represent a minor \n ( < 10% ) contribution ( table 1 ) . \n one tetra - antennary \n glycoform with three fucoses was observed at swpavgncssalr \n peptide , but multiply fucosylated and highly branched structures are \n rare on hpx , contrary to hp . \n although the ion intensities of most \n singly fucosylated bi - antennary glycopeptides were not sufficient \n for fragmentation , the measurable cid spectra contain low - intensity \n peptide - hexnac - fuc fragment consistent with core fucosylation . \n the \n presence of oxonium ions at m / z 512.2 \n indicates the presence of outer - arm fucose . \n the intensity of fragments , \n which could distinguish fucose linkage in doubly and triply fucosylated \n glycoforms , was under the detection limit of our instrument . \n fragmentation \n spectra of swpavgncssalr glycopeptide bearing a3g3f1 \n and a3g3f2 glycans contain both oxonium ion m / z 512.2 consistent with lewis x type linkage . \n the presence \n of oxonium ion at m / z 658.2 in fragmentation \n spectrum of the a3g3f2 glycopeptide could suggest the presence of \n lewis - y - type structure , but rearrangement of fucose at outer arms \n is a likely event ; independent methods \n would have to confirm such linkage assignment . \n we were not able to \n confirm lewis y linkage by msn analysis and conclude that rearrangement \n of the lewis x structures is more likely in this case . \n this fucose \n rearrangement , however , was not described to happen between core and \n outer arms in tri- and tetra - antennary glycans , and we can conclude \n that core fucosylation is minor and below the limit of our detection \n in the tri - antennary structures . \n these \n assignments are quite consistent with analysis of n - glycans detached \n from hpx in a study of cirrhotic and hcc patients . \n observed intense bi - antennary glycan with \n small contribution of fucosylation and minor tri- and tetra - antennary \n glycoforms compared with the dominant bi - antennary structure . \n the \n authors found that outer - arm fucosylated tri- and tetra - antennary \n glycans are increased in hcc compared with bi - antennary glycan without \n fucose and suggest that their ratio can be used as a diagnostic test \n for detection of hcc . \n maldi - tof spectrum of glycans released \n from isolated hpx is dominated \n by bi - antennary structures ( supplemental figure \n 1 in the supporting information ) . \n disassembly of a bi - antennary , \n monosialylated , monofucosylated composition shows a mixture of core \n fucosylation , lewis x , and sialylated lewis x - containing isomers ( supplemental figure 4 in the supporting information ) . \n kininogen-1 : kng-1 , a plasma glycoprotein of 644 amino acids , \n contains \n a heavy chain with four n - glycosylation sites . \n serine proteases cleave from kng-1 bradykinin , a nine amino acid \n peptide that regulates many biological processes including inflammation , \n angiogenesis , and cell migration . \n we \n have identified bi- and tri - antennary glycans and their fucosylated \n forms at three of four n - glycosylation sites of kng-1 . \n however , the intensity of this peptide is very low , \n and we can not exclude the presence of other glycoforms below the detection \n limit of our method . \n two glycopeptides carry tri - antennary glycoforms \n with up to two fucoses per glycan ; moreover , glycopeptide hgiqyfnnntqhsslfmlnevkr \n carries tetra - antennary glycans with up to five fucoses ( figure 3a ) . \n this is similar to the multiply fucosylated \n glycoforms of hp and in contrast with the limited fucosylation of \n glycoforms associated with hpx and cfh . \n figure 3b shows a representative cid fragmentation spectrum of triply fucosylated \n tetra - antennary glycoform of hgiqyfnnntqhsslfmlnevkr \n peptide with glycan fragments consistent with lewis x structure at m / z 512.2 . \n there is no evidence of core \n fucosylation in kng-1 glycopeptides based on their cid fragmentation . \n unfortunately , we were not able to complete msn structural analysis \n of kng-1 s glycans because the protein was not available in \n sufficient amount and purity . \n ( a ) ms profile of kng-1 peptide hgiqyfnnntqhsslfmlnevkr \n bearing tetra - antennary glycans with up to five fucoses . \n ( b ) cid spectrum \n of tetra - antennary triply fucosylated hgiqyfnnntqhsslfmlnevkr \n glycopeptide with oxonium ions corresponding to glcnac and glcnac - hex \n fragments ( m / z 204 and 366 ) and \n glcnac - gal - fuc fragment ( m / z 512 ) \n consistent with lewis x structure . \n complement factor h : cfh is a 140 kda ( without carbohydrate \n moiety ) \n plasma glycoprotein that regulates enzymatic activity of complement \n c3 . \n cfh has nine n - glycosylation sequons \n and is primarily secreted to plasma by the liver even though a membrane \n bound form exists as well . \n we have detected \n seven glycopeptides in the tryptic digest of cfh which cover eight \n sequons ( one peptide is doubly glycosylated ) . \n all peptides carry a \n limited variety of glycoforms of the complex type ; high mannose or \n hybrid glycoforms were not detected . \n we have observed dominant bi - antennary \n glycoforms on all sequons except peptide l(528)-r(567 ) , which carries \n more intense tri - antennary glycoforms . \n the bi - antennary glycoforms \n are singly fucosylated , but the intensity of the fucosylated glycoforms \n is somewhat higher ( up to 20% ) than for hpx ( < 10% ) . \n similar to \n hpx , the singly fucosylated tri - antennary glycoforms tend to have \n higher intensity than their nonfucosylated counterparts . \n we have detected \n the minimal amount of doubly fucosylated glycoforms ; tetra - antennary \n glycoforms and glycoforms with more than two fucoses were not detected \n ( table 2 ) . \n structural analysis of detached \n permethylated glycans uncovered the presence of core - fucosylated glycans \n represented by fuc - glcnac fragment at m / z 490 ( supplemental figure 3 in the supporting \n information ) . \n the intensity of this fragment is roughly similar \n to the fragment at m / z 660 corresponding \n to lex epitope . \n this is in contrast with other analyzed singly fucosylated \n glycoforms from hpx and hp , where the 660 fragment was dominant , and \n the 490 fragment was small but detectable . \n these findings demonstrate \n that the process of glycan branching and fucosylation is site - specific \n and differs significantly between different glycoproteins isolated \n from the same patient . \n our aim was to examine whether hyper - fucosylation \n of hp is associated \n with hyper - fucosylation of other proteins isolated from serum of the \n same hcc patient . \n our analysis focused on fucosylation because we \n observe only complex n - glycans on all isolated proteins examined in \n this study and because we have previously observed hyper - fucosylated \n structures on hp . \n comparison of the protein and \n site - specific heterogeneity of the glycoforms shows interesting general \n trends . \n distribution of fucosylated glycoforms is protein- and site - specific \n ( table 1 ) . \n singly fucosylated bi - antennary \n glycoforms are absent or low - intensity compared with the nonfucosylated \n bi - antennary forms in almost all studied glycoproteins . \n the highest \n intensity of bi - antennary glycoforms was found on cfh , where fucosylated \n glycoforms reach up to 20% ion intensity of the nonfucosylated glycoforms \n ( table 1 ) . \n singly fucosylated tri- and tetra - antennary \n glycoforms have higher intensity than their nonfucosylated forms ( table 2 ) . \n tri - antennary singly fucosylated glycans are \n the most intense glycoforms across virtually all identified tri - antennary \n glycopeptides . \n tetra - antennary glycans are , in general , less abundant \n and observed on select glycopeptides only . \n the distribution \n of multifucosylated tri- and tetra - antennary glycoforms \n differs by protein and peptide . hyper - fucosylated glycoforms with \n more fucoses \n than arms were observed only in hp ( up to six fucoses \n per glycopeptide ) and kng-1 ( five fucoses per glycopeptide ) . \n in contrast with hp and kng-1 , we have detected \n at most two fucoses on three of eight glycopeptides of cfh and only \n one glycopeptide with three fucoses in the case of hpx ( a4g4f3 on \n swpavgncssalr peptide ) . \n structural analysis \n of detached permethylated glycans shows differences in level of core \n fucosylation between the glycoproteins . \n for example , in both hp and \n hpx , core fucosylation is less abundant compared with cfh . \n furthermore , \n it was observed that doubly fucosylated glycoforms of hp contain both \n lewisx and lewisy epitopes . \n this shows that factors other than up - regulated \n glycosyltransferases define how and which glycosite is fucosylated . \n the determinants of site - specific microheterogeneity are not entirely \n known to our knowledge , but it is expected that the types of mature \n glycoforms detectable in disease context depend not only on the activity \n of glycosyltransferases but also on sequence of their action , distribution \n of the synthesized glycoproteins , their stability and turnover , and \n also on protein structure . \n n - glycosylation is a cotranslational \n process that originates in \n the er , but the n - glycan structures continue to mature on folded proteins \n in golgi apparatus . \n the protein s \n secondary and tertiary structure determines the interaction with binding \n partners and accessibility of glycosyltransferases responsible for \n fucosylation , sialylation , and branching . \n the authors \n suggest that protein structure ( e.g. , solvent accessibility , protein \n size , and hydropathy index ) affects glycan microheterogeneity . \n this \n is in agreement with our observation that glycoproteins isolated from \n serum of the same patient are differentially fucosylated , although \n we assume that the repertoire of glycan biosynthetic machinery is \n the same for all of the glycoproteins being characterized . \n we can not \n exclude the possibility that some glycoforms are selectively removed \n from circulation , but it is hard to believe that all tri- and tetra - antennary \n fucosylated glycoforms of cfh or hpx would be so efficiently eliminated . \n in addition , differences in site - specific distribution of glycoform \n on each protein support influence of other factors ( table 1 ) . \n kng-1 glycopeptide y(44)-r(58 ) carries \n only one bi - antennary glycan compared with the tetra - antennary multiply \n fucosylated glycans on the neighboring peptide . \n it is more likely \n that the selection process is defined by local structure - recognition , \n which regulates the activity of glycosyltransferases at specific sequons .", "role": "user" }, { "content": "chronic liver diseases are a serious \n health problem worldwide . \n one of the frequently reported glycan alterations in liver disease \n is aberrant fucosylation , which was suggested as a marker for noninvasive \n serologic monitoring . \n we present a case study that compares site specific \n glycoforms of four proteins including haptoglobin , complement factor \n h , kininogen-1 , and hemopexin isolated from the same patient . \n our \n exoglycosidase - assisted lc \n ms / ms analysis confirms the high \n degree of fucosylation of some of the proteins but shows that microheterogeneity \n is protein- and site - specific . \n msn analysis of permethylated detached \n glycans confirms the presence of ley glycoforms on haptoglobin , which \n can not be detected in hemopexin or complement factor h ; all three \n proteins carry lewis and h epitopes . \n core fucosylation is detectable \n in only trace amounts in haptoglobin but with confidence on hemopexin \n and complement factor h , where core fucosylation of the bi - antennary \n glycans on select glycopeptides reaches 1520% intensity . \n these \n protein - specific differences in fucosylation , observed in proteins \n isolated from the same patient source , suggest that factors other \n than up - regulation of enzymatic activity regulate the microheterogeneity \n of glycoforms . \n this has implications for selection of candidate proteins \n for disease monitoring and suggests that site - specific glycoforms \n have structural determinants , which could lead to functional consequences \n for specific subsets of proteins or their domains .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: nkt cells are distinguished type i nkt cell and type ii nkt cells according to t cell receptor ( tcr ) usage . \n type i nkt cells are called invariant nkt ( inkt ) cells because they express only restricted tcr chain ( v14 ) and tcr chain ( v2 , 7 and 8) , whereas type ii nkt cells express various tcr chains ( 2,3 ) . \n nkt cells can help activation of other lymphocytes such as , t , b , nk and dendritc cells ( 3 - 6 ) . \n it has been known that dedritic cells ( dcs ) play a pivotal role in antigen presentation to nkt cells ( 7 ) . \n in contrast , resting b cells are known poor immunogenic antigen presenting cells ( apcs ) to nkt cells ( 8) . however \n nkt cells provide direct and indirect help for lipid and protein - specific b cells , respectively ( 9,10 ) . direct help from inkt cells for lipid - specific b cells is dependent on il-21 secreted from inkt cells ( 11 ) . \n high - density -galcer loaded b cells stimulate inkt cells although the efficiency of stimulation is not comparable to dendritic cells ( 12 ) . \n -galcer / peptide loaded b cells can induce peptide - specific cytotoxic cd8 t cells and induced cytotoxic cd8 t cells eliminate various kinds of tumor ( 12,13 ) . in micro - environment of foreign antigen stimuli , dc and b cells cooperate to activate antigen - specific cytotoxic cd8 t cells ( 14 ) . \n nevertheless , it remains to be resolved unknown nkt cells response upon co - operative stimulation of b cells and dcs . in this study . \n we found that b cells induced th1-skewed response of nkt cells in the presence of -galcer loaded dcs , regardless of exogenous antigen loading to b cells . \n we also demonstrated that regulation of nkt cell response by b cells stimulation was dependent on cd1d expression of b cells . \n six- to eight - week - old c57bl/6 ( b6 ) , cd1d and cd1d mice ( b6 background ) were bred in facilities at korea university . \n the experimental protocols of this study were approved by the institutional animal care and use committee of korea university . \n monoclonal antibodies specific for mouse tcr ( h57 ) , ifn- ( xmg-1 ) , il-4 ( 11b11 ) and nk1.1 ( pk136 ) were purchased from bd biosciences . \n b cells were isolated from splenocytes via macs by using anti - b220-conjugated magnetic beads ( miltenyi biotec ) after dc had been depleted by using anti - cd11c microbeads . \n purified b cells and dcs were loaded 10 ng / ml -galcer for 4 hr and they were washed extensively , then were co - cultured with nkt cells for 8 hr or 48 hr . after 8 hr or 48 hr , the amount of cytokines was measured by intracellular cytokine staining and elisa . \n cells were washed and blocked with an anti - fcrii / iii mab ( 2.4g2 ) for 15 min and then labeled for 30 min on ice with the appropriate mabs . for intracellular cytokine staining , \n cells were fixed with a cytofix / cytoperm kit ( bd biosciences ) according to the manufacturer 's protocol . \n the stained cells were analyzed with a facscalibur flow cytometer using cellquest software ( bd biosciences ) . \n student 's t - tests were used to determine the statistical significance of differences between the comparison groups unless otherwise stated . \n six- to eight - week - old c57bl/6 ( b6 ) , cd1d and cd1d mice ( b6 background ) were bred in facilities at korea university . \n the experimental protocols of this study were approved by the institutional animal care and use committee of korea university . \n monoclonal antibodies specific for mouse tcr ( h57 ) , ifn- ( xmg-1 ) , il-4 ( 11b11 ) and nk1.1 ( pk136 ) were purchased from bd biosciences . \n b cells were isolated from splenocytes via macs by using anti - b220-conjugated magnetic beads ( miltenyi biotec ) after dc had been depleted by using anti - cd11c microbeads . \n purified b cells and dcs were loaded 10 ng / ml -galcer for 4 hr and they were washed extensively , then were co - cultured with nkt cells for 8 hr or 48 hr . after 8 hr or 48 hr , the amount of cytokines was measured by intracellular cytokine staining and elisa . \n cells were washed and blocked with an anti - fcrii / iii mab ( 2.4g2 ) for 15 min and then labeled for 30 min on ice with the appropriate mabs . for intracellular \n cytokine staining , cells were fixed with a cytofix / cytoperm kit ( bd biosciences ) according to the manufacturer 's protocol . \n the stained cells were analyzed with a facscalibur flow cytometer using cellquest software ( bd biosciences ) . \n student 's t - tests were used to determine the statistical significance of differences between the comparison groups unless otherwise stated . \n to test whether b cells can affect nkt cell stimulating activity of dcs , we co - cultured wt cd11c dc with nkt cells and b cells and then assessed the ability of b cells to activate nkt cells . \n elisa analysis and intracellular staining demonstrated that ifn- from nkt cells was significantly increased , in contrast il-4 was substantially reduced upon the addition of -galcer pulsed b cells compared to the response by dcs alone ( fig . \n the co - culture of nkt cells with -galcer un - pulsed b cells also led to a significantly higher production of ifn- and lower production of il-4 than that with -galcer pulsed dcs alone ( fig . 2a and 2b ) . \n taken together , these results show that b cells promote th1 response of nkt cells in the presence of antigen - loaded dcs , regardless of exogenous antigen pulse to b cells . \n although b cells promote th1 response of nkt cells in the presence of antigen - loaded dcs , it was not clear whether nkt cells and b cells form directly cell - cell contact by cd1d - tcr interaction . to test whether cd1d expression on b cells was required to promote th1 response of nkt cells in the presence of -galcer pulsed dcs , we isolated b cells from cd1d or cd1d mice and co - cultured them with dcs to activate nkt cells . \n co - culture of nkt cells with cd1d b cells led to a significantly higher production of ifn- and lower production of il-4 than that with cd1d b cells ( fig . \n this data show that th1-skewed response of nkt cells upon the addition of conventional b cells is dependent on cd1d expression of b cells . \n similarly , it had been reported that marginal zone ( mz ) b cells amplify dc mediated - nkt cell activation ( 15 ) . \n the cytokine patterns of nkt cells upon mz b cells and dcs stimulation ( 15 ) was similar to our results , except il-4 expression was also increased when -galcer loaded mzb cells was used . however , there was a contradicting report where conventional b cells could dampen inkt cells activation in the presence of dcs ( 8) . we are not sure at the moment what made these difference in nkt cell response but the density of b cells and concentration of -galcer might be causative factor since they used much higher number of b cells and higher concentration of -galcer to activate nkt cells compared to the experimental condition in this study . \n nkt cell - mediated cytokines milieu can be influenced by pattern of cellular composition of lymphocytes in specific inflammatory site . \n for example , nkt cells in tumor model promote th1-skewed cytokine milieu , in contrast nkt cells in asthma model promote th2-skewed cytokine milieu ( 4,6 ) . in asthma model , \n the administration of -galcer ameliorated or worsened the disease symptoms by how to inject -galcer ( 16 - 18 ) . \n a shift from th2 to th1-response by the administration of -galcer suppressed airway hyper - responsiveness ( ahr ) ( 17,18 ) . in this study \n , we demonstrated that b cells promote th1 response of nkt cells in the presence of antigen - loaded dcs . \n thus , the administration of -galcer pulsed b cells and dcs may be a possible approach for the treatment of ahr . \n eventually , the elucidation of optimal composition of apcs such as dcs and b cells to regulate finely the direction of nkt cell response will be the basis to apply nkt cell - based cell therapy .", "role": "user" }, { "content": "cd1d expressing dendritic cells ( dcs ) are good glyco - lipid antigen presenting cells for nkt cells . however , resting b cells are very weak stimulators for nkt cells . \n although -galactosylceramide ( -galcer ) loaded b cells can activate nkt cells , it is not well defined whether b cells interfere nkt cell stimulating activity of dcs . \n unexpectedly , we found in this study that b cells can promote th1-skewed nkt cell response , which means a increased level of ifn- by nkt cells , concomitant with a decreased level of il-4 , in the circumstance of co - culture of dcs and b cells . remarkably , the response promoted by b cells was dependent on cd1d expression of b cells .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: paraneoplastic neurological syndromes are a heterogeneous group of neurological disorders occurring in less than 1% of patients with malignancy . \n peripheral neuropathy is commonly accompanied by cancer but paraneoplastic neuropathies which show a pure demyelinating pattern on electrophysiological studies are not commonly reported . \n he had presented with anorexia , asthenia and repeated falls since one month and was admitted 2 weeks ago for these complaints in the setting of a biologic inflammatory syndrome . \n he had no significant past medical history , neither alcohol abuse nor toxic substance exposure nor a significant family history . \n motor force was otherwise normal . at this time , clinical examination revealed motor deficit extension in the proximal areas of the 4 limbs with complete reflex abolition . \n electromyography revealed a pure motor demyelinating polyneuropathy of the four limbs with several conduction blocks between 55 and 90% in the upper limbs ( fig . \n no motor response was detected in the lower limbs and neither tibial f - wave nor h - reflex were recordable ( table 1 ) . \n on needle electromyography , we observed denervation potentials in the distal muscles of the four limbs . \n biological tests showed a raised c - reactive protein level at 75 mg / l , with normal renal , hepatic and thyroid function tests . \n immunological tests showed anti - gd1a ( igm at 1/2,000 and igg > 1/10,000 ) , anti - gd1b igm at 1/5,000 , anti - gm1 igm at 1/5,000 and onconeural ( anti - gray matter and anti - cerebellar ) antibodies in the serum . \n as an immune - mediated neuropathy was suspected , the patient was treated by one course of ivig ( 2 g / kg ) and corticosteroid pulse without any response to therapy . \n , the patient did not have any metastases or any cancer - related complaints . following a multidisciplinary approach , \n the progressive pure motor symptoms in the setting of a demyelinating neuropathy associated with conduction blocks in electromyography and presence of a high anti - gm1 antibody titre favour the diagnosis of multiple motor neuropathy ( mmn ) . however , late - age onset and a 50% decrease in distal amplitude of the sensory ulnar nerve are not typical of this diagnosis [ 3 , 4 ] . \n initial clinical presentation may be concordant with a motor variant of guillain - barr syndrome ( gbs ) . \n moreover , in a large study of 147 patients with gbs , high titres of anti - gm1 antibodies were significantly more common in motor gbs patients . however , electrophysiological data of these patients revealed little or no evidence for demyelination . \n high titres of anti - gd1a antibodies have been found in patients with gbs but were also found in 18% of patients with mmn and 5% of those with chronic inflammatory demyelinating polyneuropathy ( cidp ) . \n a chronic ( > 2 months ) course and electrophysiologic findings can raise the possibility of cidp and there are patients eventually diagnosed with cidp who have an acute onset resembling gbs ; however , acute - onset cidp should be suspected in patients with prominent sensory symptoms and signs at presentation . \n conduction blocks are the electrophysiological hallmarks of lewis and sumner syndrome as well but the major distinguishing features between lewis and sumner syndrome and motor neuropathy with block are the clinical and electrophysiological sensory involvement and lack of anti - gm1 antibodies in lewis and sumner syndrome . \n another paraneoplastic neuropathy which is a classical one is a sensory ganglionopathy characterized by non - length - dependent abnormalities of sensory nerve action potentials . \n however , clinical and electrophysiological patterns of our patient did not correspond with this entity . \n we could not determine the subtype diagnosis of this demyelinating motor neuropathy ; based on initial clinical presentation , the diagnosis of gbs was likely but the electrophysiological findings are in favour of cidp or mmn with some atypical features . \n there are different mechanisms by which cancer affects the peripheral nervous system ( pns ) . \n these mechanisms can include compression or infiltration by the tumour , deleterious effects of treatments , metabolic and nutritional factors and infections . \n it has been proposed to classify paraneoplastic pns disorders as definite or possible according to the presence or absence of the following items : classical paraneoplastic pns disorder , onconeural antibodies and development of cancer [ 1 , 10 ] . \n our patient presented with a non - classical neurological syndrome but the presence of onconeural antibodies and cancer occurrence within less than 5 years following neurological symptoms confirm the diagnosis of definite paraneoplastic polyneuropathy . \n gbs was reported accompanying endometrial , cholecystic , bronchial , renal , hepatic and gastric carcinoma [ 11 , 12 , 13 , 14 , 15 , 16 ] . however , in many of these cases , it is difficult to determine whether gbs should all the time be considered paraneoplastic because the frequency of both cancer and gbs makes a coincidental association possible . \n there are also reports of cipd associated with solid carcinoma including melanoma , breast , gastric , pancreas , liver and colon carcinoma [ 17 , 18 , 19 , 20 , 21 , 22 , 23 ] . as for gbs , paraneoplastic syndromes are highly suggested in these cases but the frequent lack of antineuron antibodies makes the link between the two diseases uncertain . to our knowledge , there is no case report of mmn associated with cancer . \n however , in the last decade , there were some reports of mmn associated with haematological malignancies [ 24 , 25 ] . \n proposed that in the setting of demyelinating neuropathies , either a severe or rapid course , a poor response to therapy with an early axonal loss or the presence of systemic abnormalities should lead to suspect an associated lymphoma . \n the neuropathy had a rapid course with early axonal loss witnessed by inexcitability of both peroneal and tibial nerves and a poor response to treatment . \n unfortunately , in our case there was no indication for tumour resection , but the findings in the literature are generally consistent with clinical improvement of paraneoplastic demyelinating neuropathy following a solid tumour resection [ 21 , 27 ] . \n the progressive pure motor symptoms in the setting of a demyelinating neuropathy associated with conduction blocks in electromyography and presence of a high anti - gm1 antibody titre favour the diagnosis of multiple motor neuropathy ( mmn ) . however , late - age onset and a 50% decrease in distal amplitude of the sensory ulnar nerve are not typical of this diagnosis [ 3 , 4 ] . \n initial clinical presentation may be concordant with a motor variant of guillain - barr syndrome ( gbs ) . \n moreover , in a large study of 147 patients with gbs , high titres of anti - gm1 antibodies were significantly more common in motor gbs patients . however , electrophysiological data of these patients revealed little or no evidence for demyelination . \n high titres of anti - gd1a antibodies have been found in patients with gbs but were also found in 18% of patients with mmn and 5% of those with chronic inflammatory demyelinating polyneuropathy ( cidp ) . \n a chronic ( > 2 months ) course and electrophysiologic findings can raise the possibility of cidp and there are patients eventually diagnosed with cidp who have an acute onset resembling gbs ; however , acute - onset cidp should be suspected in patients with prominent sensory symptoms and signs at presentation . \n conduction blocks are the electrophysiological hallmarks of lewis and sumner syndrome as well but the major distinguishing features between lewis and sumner syndrome and motor neuropathy with block are the clinical and electrophysiological sensory involvement and lack of anti - gm1 antibodies in lewis and sumner syndrome . \n another paraneoplastic neuropathy which is a classical one is a sensory ganglionopathy characterized by non - length - dependent abnormalities of sensory nerve action potentials . \n however , clinical and electrophysiological patterns of our patient did not correspond with this entity . \n we could not determine the subtype diagnosis of this demyelinating motor neuropathy ; based on initial clinical presentation , the diagnosis of gbs was likely but the electrophysiological findings are in favour of cidp or mmn with some atypical features . \n . these mechanisms can include compression or infiltration by the tumour , deleterious effects of treatments , metabolic and nutritional factors and infections . \n it has been proposed to classify paraneoplastic pns disorders as definite or possible according to the presence or absence of the following items : classical paraneoplastic pns disorder , onconeural antibodies and development of cancer [ 1 , 10 ] . \n our patient presented with a non - classical neurological syndrome but the presence of onconeural antibodies and cancer occurrence within less than 5 years following neurological symptoms confirm the diagnosis of definite paraneoplastic polyneuropathy . \n gbs was reported accompanying endometrial , cholecystic , bronchial , renal , hepatic and gastric carcinoma [ 11 , 12 , 13 , 14 , 15 , 16 ] . however , in many of these cases , it is difficult to determine whether gbs should all the time be considered paraneoplastic because the frequency of both cancer and gbs makes a coincidental association possible . \n there are also reports of cipd associated with solid carcinoma including melanoma , breast , gastric , pancreas , liver and colon carcinoma [ 17 , 18 , 19 , 20 , 21 , 22 , 23 ] . as for gbs , \n paraneoplastic syndromes are highly suggested in these cases but the frequent lack of antineuron antibodies makes the link between the two diseases uncertain . to our knowledge \n however , in the last decade , there were some reports of mmn associated with haematological malignancies [ 24 , 25 ] . \n proposed that in the setting of demyelinating neuropathies , either a severe or rapid course , a poor response to therapy with an early axonal loss or the presence of systemic abnormalities should lead to suspect an associated lymphoma . \n the neuropathy had a rapid course with early axonal loss witnessed by inexcitability of both peroneal and tibial nerves and a poor response to treatment . \n unfortunately , in our case there was no indication for tumour resection , but the findings in the literature are generally consistent with clinical improvement of paraneoplastic demyelinating neuropathy following a solid tumour resection [ 21 , 27 ] . \n \n in conclusion , we suggest looking for a paraneoplastic origin in the setting of demyelinating neuropathy in older patients , or when the clinical course is progressing rapidly within a few weeks or months .", "role": "user" }, { "content": "peripheral neuropathy is commonly accompanied by cancer but demyelinating ones are not commonly reported . \n we report the clinical , neurophysiological , and biological characteristics of an 82-year - old patient who presented with a demyelinating motor neuropathy and high titre of anti - ganglioside antibodies associated with oesophageal cancer . \n the neurological course worsened rapidly despite immunotherapy , leading to a bedridden status . we propose to suspect a paraneoplastic origin in older patients or when the clinical course progresses rapidly within a few weeks or months .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: the dry mass of the plantago asiatica was purchased from an oriental pharmacy ( iksan , korea ) , was according to the standard as mentioned in korean pharmacopoeia and korean herbal pharmacopoeia , which are the official compendia of standard . the procedure for preparing pae was as follows . \n the air - dried mass of plantago asiatica ( 100 g ) was cut into pieces and extracted twice with 50% ( v / v ) ethanol ( six times as much as the weight of the dried plants ) for 3 hours at 80. after filtration through a 400-mesh filter cloth , the filtrate was refiltered through filter paper ( whatman , no . \n 5 ) and concentrated on a rotary evaporator ( eyela , tokyo , japan ) and the concentrated filtrate was evaporated to dryness under vacuum with freezing dryer ( labconco , usafinally , the solid residue was collected , placed in sealed bottles and stored at 20. the extract yield of plantago asiatica with 50% ethanol was 15.55% . plantamajoside , which was used for the standard material of pae composition , was purchased from the natural product bank , institute for korea traditional medical industry ( geong - san , korea ) . \n plantago asiatica is a member of the plantaginaceae family , is widely distributed in east asia , used in chinese medicine , where its leaf is called plantaginis herba and its seed is plantaginis semen , and has been commercialized as an extract and as a dietary supplement in various forms . \n a previous report discussed the isolations of plantamajoside , plantagin and aucubin from p. asiatica , as well as the various biological properties of these compounds , including antibacterial , antiallergic , antiinammatory , antioxidative and lipoxygenase inhibitory activity . \n figure 1 shows the chemical structures of phenylethanoid glycosides ( phgs ) including acteoside and plantamajoside . \n waters acquity lc system ( waters corp . , milford , usa ) was used for lc system . \n the column was c18 type acquity uplc beh ( 2.150 mm , 1.7 m , waters corp . , milford , usa ) . a waters nova pack c-18 column ( acquity uplc beh ( 2.150 mm , 1.7 m , waters corp . , \n the column temperature was set at 30 with a flow rate of mobile phase at 0.6 ml / min ( 0.1% h3po4/acetonnitrile ) . \n this study was conducted on the 3-day - old chickens ( n=30 ) in the animal facility of center for animal resources development , wonkwang university , korea . \n animals were acclimatized and kept in an animal facility room with regulated temperature ( 282 ) , humidity ( 505% ) and light / dark cycle ( 12/12 hours ) . \n the animals were fed commercial post - broiler diet without antibiotics and coccidiostat ( hanil feed co. , yongin , korea ) and tab water ad libitum . \n all studies were performed in accordance with the guide for animal experimentation by wonkwang university and approved by the institutional animal care and use committee of wonkwang university . \n those animals were divided with 3 groups ; pae 0.1% treated / infected ( n=10 ) , pae untreated / infected ( n=10 ) and non - infected control ( n=10 ) . \n chickens were fed a standard diet supplemented with or without pae for 1 week prior to infection with e. tenella ( 10,000 sporulated oocysts per chicken ) . \n the effects of pae on e. tenella infection were assessed by two parameters , fecal oocysts shedding and body weights gain . \n oocysts of e. tenella were cleaned by flotation on 5.25% sodium hypochlorite and washed three times with phosphate buffered saline . \n e. tenella was provided kindly by professor wongi min at gyeongsang national university in korea . \n chickens were treated orally by gavages using a 24 gauge , mouse stainless steel animal feeding tube ( popper & sons , inc . \n the oral infectious dose of has been approximated 10 oocysts of e. tenella in 1 ml of saline . \n the control chickens ( n=10 ) received saline through the same route . during the study period \n body weights were individually measured for 2 weeks before infection and for 10 days post - infection . \n the fecal samples were analyzed for the presence of coccidial oocysts using a standard fecal flotation technique . \n briefly , 5 ml from each sample was pelleted by centrifugation at 1500g for 5 min . \n the resulting pellet was resuspended in saturated sodium chloride ( aqueous ) , passed through a 1 mm mesh size sieve to remove coarse fecal debris . \n the resulting filtrate was used in a standard gravity vial fecal flotation using 22 mm22 mm coverslips . \n after flotation , the coverslip was mounted on a slide and examined in its entirety for the presence of coccidial oocysts . \n total number of oocysts was calculated using the following formula : [ total number of oocysts = oocyst countdilution factor(fecal sample volume / counting chamber volume)/number of birds per cage ] . \n differences in mean oocysts production and mean weight gain between the 4 groups were tested by using one - way analysis of variance ( anova ; graphpad instat ; graphpad software inc . , \n the effects of pae on e. tenella infection were assessed by two parameters , fecal oocysts shedding and body weight gain . \n pae - fed chickens produced significantly reduced fecal oocysts ( p<0.05 ) when compared to the e. tenella - infected group fed standard diet . \n also , pae - based diet , improved body weight loss caused by e. tenella infection . \n the results showed that , compared to untreated controls , chickens treated with pae had significantly decreased fecal oocysts shedding and showed strong anticoccidial activities ( p<0.01 ) . \n as shown in table 1 , oocyst shedding was significantly higher in the inoculated chickens than in the control chickens ( p<0.05 ) . \n the number of fecal oocysts shed was highest on day 7 post - inoculation ( table 1 ) . moreover , body weight gain was lesser in the animals in the inoculated group than in the animals in the control group ( table 2 ) . \n coccidiosis of domestic fowl is a worldwide disease caused by obligatory intracellular protozoa of the genus eimeria . \n e. tenella is important pathogen causing avian coccidiosis in laboratory avian animals and known to affect influencing experimental results obtained with contaminated animals . \n the disease is characterized by enteric lesions of variable extent and severity , reducing the absorptive function of the intestinal mucosa , thus leading to weight loss , diarrhea , poorer feed conversion and a higher mortality in the affected flocks . \n the results of this study showed that pae had a strong anticoccidial effect on e. tenella . \n the major bioactive constitute of p. asiatica are phenylethanoid glycosides ( phgs ) including acteoside and plantamajoside as shown in scheme 1 . \n phgs from many plants have been extensively studied for their various biological functions such as anti - hepatotoxic , anti - inflammatory , antinociceptive activities , improving sexual function and antioxidant activity . the plantaginin , plantamajoside , acetoside and 6-hydroxyluteolin 7-glucoside isolated from p. asiatica were the chief compounds and they had various biological activities . \n the plantamajoside , in particular , was reported to show antibacterial activity against staphylococcus aureus , inhibition activity against camp phosphodiesterase and 5-lipoxygenase , and antioxidant activity on adp+ nadph - induced lipid peroxidation in rat liver microsome . \n also , the plantamajoside is used as a marker compound in chemotaxonomic studies , as are other members of the dihydroxyphenylethyl glycoside family . \n theses phenylethanoid glycosides - included bioactive structures could be suggested a plausible explanation of the anticoccidial properties in this study . in this study , \n the pae - fed chickens produced significantly reduced fecal oocysts ( p<0.05 ) when compared to the e. tenella - infected group fed standard diet . \n also , pae - based diet , improved body weights loss caused by e. tenella infection .", "role": "user" }, { "content": "anticoccidial effects of the plantago asiatica extract ( pae ) were evaluated in chickens following oral infection with eimeria ( e. ) tenella . \n this study was conducted on the 3-day - old chickens ( n=30 ) . \n those animals were divided with 3 groups ; pae 0.1% treated / infected ( n=10 ) , pae untreated / infected ( n=10 ) and non - infected control ( n=10 ) . \n chickens were fed a standard diet supplemented with or without pae for 1 week prior to infection with e. tenella ( 10,000 sporulated oocysts per chicken ) . \n the effects of pae on e. tenella infection were assessed by two parameters ; fecal oocysts shedding and body weights gain . \n the pae - fed chickens produced significantly reduced fecal oocysts ( p<0.05 ) when compared to the e. tenella - infected group fed standard diet . \n also , pae - based diet , improved body weight loss caused by e. tenella infection . \n our data demonstrated that pae had remarkable anticoccidial activities against e. tenella . \n this finding might have implications for the development of anticoccidial drug . \n this study is the first to demonstrate anticoccidial effect of pae on eimeria parasites .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: anderson - hynes dismembered pyeloplasty was first introduced in 1949 for the treatment of obstructed retrocaval ureter . \n however , it is still the gold standard therapeutic approach to ureteropelvic junction obstruction ( upjo ) ( 1 ) . \n the success rate of this procedure is estimated as high as 90% ( 2 ) . \n the mainstay of this surgical intervention is to respect the stenotic part and the enlarged part of the ureter , while performing repositioning of ureter and ureteropelvic anastomosis ( 3 ) . \n use of a drainage method to protect the suture line from leakage is a matter of controversy ( 4 ) , as retrograde and antegrade double j ( dj ) stenting is used as a splint cross the anastomosis ( 5 ) . \n currently , dj internal drainage is widely used by those surgeons who advocate the drainage methods . \n nevertheless , several stent related symptoms are reported in these cases and readmission of patients is necessary for catheter removal , which may impose extra charges to these patients ( 6 ) . \n the objective of this study was to compare the surgical outcome of anderson - hynes dismembered pyeloplasty for upjo repair with or without internal stenting . \n eighty two patients with upjo were evaluated from 1996 to 2002 in the academic center of mashhad university of medical sciences , mashhad , iran . the study population comprised of 51 male and 31 female patients in the age range of 17 - 49 years . \n the upjo diagnosis was made on the basis of patient reported symptoms , sonographic findings and intravenous pyelography ( ivp ) and diethylene - triamine - pentaacetate ( dtpa ) scan results . \n those cases with renal failure , history of previous upjo repair or requiring emergent drainage because of renal stone or infection were all excluded . \n after completion of the posterior suture line , a dj catheter was paced in the internal drainage group ( randomly selected ) while in other stentless cases , a temporary feeding tube was inserted for secure anastomosis , which was later removed . \n anastomosis was performed using 4 - 0 chromic separate sutures in a water tight fashion . \n a drain was also placed in the retroperitoneal space and fallow - up of patients was planned weekly . \n drain output was assessed and it was extracted when drainage became less than 50 cc per day . \n urine analysis was done and oral antibiotics were continued for at least 5 days after discharge . the ivp was performed after 4 weeks in all patients and the renal function and probable extravasation and structural anomalies were evaluated . \n those patients with internal stent were admitted again after 1 month to remove the stent if no complication had occurred . \n a 3 day course of oral antibiotic therapy was also started at the time of stent removal . \n double stenting was used in 30 cases and pyeloplasty without internal drainage was performed in 52 patients . \n the most common symptom was severe flank pain , which was present in 69% of patients . \n history of urinary tract infection was detected in 47% and 51% reported associated gastrointestinal symptoms . \n pyuria was seen in 58% and only 36% of patients had positive urine culture , with escherichia coli ( e. coli ) as the most common pathogen ( 60% ) . the preoperative sonographic evaluation and ivp \n intraoperative assessments showed ureteropelvic junction segmental atresia in 45 cases and aberrant vessels in 25 patients . \n the postoperative course was compared in two groups of patients , with and without internal drainage ( table 2 ) . among patients with extravasation in dtpa scan in the stent free group , retrograde dj stent \n one patient had distal ureter obstruction because of distal ureter stone , that was treated by trans - ureteral lithotripsy ( tulp ) and stent support . \n three other cases had no anatomical abnormality , and they were followed and extravasation resolved spontaneously at control ivp after one month . \n however , eight cases missed the time and late stent removal was accompanied with several complications , such as stone formation on the tip of catheter in the bladder of a patient that needed tulp before stent removal . \n urine analysis was done in 27 patients of the stent group and 32 cases of the stent free group . \n dtpa scan was done in 20 patients of stent group and 32 cases of stent free group . \n the upjo is a common anomaly , especially encountered in pediatric urology , which is traditionally treated by open pyeloplasty and the most common method is the anderson - hynes dismembered pyeloplasty , performed via a retroperitoneal approach by a flank incision . according to the original approach , insertion of stent in not necessary in this procedure except for complicated cases or doubtful anastomosis ( 1 ) . in recent decades and with the introduction of new stents , such as the dj stents , \n internal drainage has become popular as a result of the means of anastomosis support , lowering the risk of extravasation and urinoma formation , ensuring urinary drainage , maintain ureteral caliber and preventing the obstruction due to anastomotic site edema ( 7 ) . \n recently , and especially by promotion of minimally invasive methods and laparoscopic repair of upjo , an ongoing debate is seen regarding the pros and cons of internal drainage in upjo repair . \n stents associated symptoms , such as dysuria , frequency , flank pain , hematuria , are commonly reported , even with short term placemat of urethral catheters , that may be so severe in selected cases that they even interfere with daily activities and impair the quality of life ( 8) . \n reported these bothering urinary complains in 78% of cases with ureteral stent ( 9 ) . among these patients , 80% experienced voiding or flank pain , 32% sexual impairment and 58% the negative effects on normal life ( 9 ) . \n potential catheter related complications include catheter dislocation or migration that may cause stent dysfunction and even obstructive uropathy , retained fragments , stone formation on the foreign body , exposure of upper system to high pressure during voiding and an artificial reflux uropathy , flank pain and an increased rate of catheter related urinary infections ( 7 , 10 , 11 ) . \n several attempts have been done to prevent readmission and repeated general anesthesia for catheter removal , and unfortunately they failed ( 12 , 13 ) . in our study comparing two techniques of pyeloplasty with or without internal drainage , there was no significant difference between groups , concerning extravasation and anastomosis complications , such as leakage , stenosis , urinoma formation , evidence of obstruction at postoperative ivp or dtpa scan . \n however , a significant incidence of catheter related symptoms , such as irritative urinary symptoms and flank pain was reported . \n kumar et al . also concluded that stentless pyeloplasty may be a feasible option in adults ( 8) . \n in conclusion , pyeloplasty with proper spatulation , hemostasis and a watertight anastomosis is the mainstay of a successful pyeloplasty and there may be no significant benefit for urethral stenting , especially in non - complicated cases .", "role": "user" }, { "content": "background : anderson - hynes dismembered pyeloplasty is the gold standard therapeutic approach to ureteropelvic junction obstruction ( upjo ) . \n use of a drainage method to protect the suture line from leakage is a matter of controversy.objectives:we have compared the surgical outcome of anderson - hynes dismembered pyeloplasty for upjo repair , with or without internal stenting.patients and methods : eighty - two patients with upjo were evaluated from 1996 to 2002 . complicated or emergent cases were excluded . \n classic standard dismembered pyeloplasty was performed . \n internal drainage , with a double j catheter , was performed in several patients , randomly . \n another drain was also placed in the retroperitoneal space . \n the follow - up of patients was planned weekly , with patient visits and urine analysis and intravenous pyelography ( ivp ) and diethylene - triamine - pentaacetate ( dtpa ) scan after one month.results:the study group consisted of 51 male and 31 female patients , who were mostly in the age range of 20 - 40 years . comparing the two techniques of pyeloplasty with or without internal drainage , there was no significant difference between groups regarding extravasation and anastomosis complications , such as leakage , stenosis , urinoma formation or evidence of obstruction on postoperative ivp or dtpa scan . \n however , a higher incidence of catheter related urinary symptoms and flank pain was reported among those with internal stent.conclusions:pyeloplasty , with adequate spatulation , hemostasis and a watertight anastomosis , represents the mainstay of successful pyeloplasty and there may be no significant benefit for urethral stenting , especially in non - complicated cases .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: the trigeminal neuralgia is defined as sudden , usually unilateral , severe , brief , stabbing , recurrent episodes of pain in the distribution of one or more branches of the trigeminal nerve ( fifth cranial nerve ) . in 80 - 90% of cases , \n it is thought to be caused by compression of the trigeminal nerve by a loop of artery or vein . \n the incidence increases with age and is rare in people younger than 40 years of age . \n most common adverse effects observed with carbamazepine are drowsiness , unsteadiness , constipation , nausea , and vomiting . \n it is also reported to cause hypertension ; however , the frequency of occurrence is rare ( 1/10,000 to < 1/1,000 ) . \n a 74-year - old female presented to a private practitioner with complaint of facial pain on the left side . on examination her blood pressure ( b.p ) was 117/80 mmhg . \n her dental examination was normal and based on the symptoms a provisional diagnosis of trigeminal neuralgia was made . \n after 2 days , she complained of headache , restlessness and on examination b.p was found to be 290/110 mmhg . \n as the pain was persisting , she presented to the neurology opd of the institute with complaint of left side facial pain and the diagnosis was confirmed as trigeminal neuralgia . \n the patient was started with carbamazepine 50 mg tablet bd and was gradually titrated to 200 mg bd . \n the mri findings confirmed the diagnosis of trigeminal neuralgia . on examination , her b.p . \n she was advised to get thyroid profile , random blood sugar and lipid profile done , all of which were within normal range . \n she presented after 4 days with severe nausea , vomiting and dizziness with persisting constipation . \n the dose of carbamazepine was reduced to 100 mg bd and rest all the medicines were continuing . \n the ecg and fundus examination were normal , biochemistry reports revealed low uric acid 2.3 ( 2.5 - 6.2 mg / dl ) ; low serum sodium 119 mg / dl ( 135 - 155 ) ; low chloride 96 mmol / l ( 98 - 107 ) ; high total protein 9.3 g / dl ( 6.3 - 8.2 ) ; and high serum globulin 4.3 g / dl ( 2.5 - 3.5 ) . \n next day the patient was admitted to a private hospital with complaints of severe headache , epigastric pain and vomiting . on examination b.p . \n records were normal and the patient is off all the antihypertensive treatment . the patient is stable now [ table 1 ] . the trigeminal neuralgia resolved after the carbamazepine was stopped . no other treatment modality was used . \n sequence of events summary meanwhile , based on literature search and detailed review of the patient 's medical and family history , carbamazepine was suspected to be the causal agent for this adverse reaction as there was no underlying cardiovascular disease . \n after oral carbamazepine high dose ; and non - resolution of hypertension despite anti - hypertensive therapy confirmed that hypertension was induced by oral carbamazepine therapy as it resolved only after carbamazepine discontinuation . \n a causal association between hypertension and carbamazepine was assessed by world health organization ( who ) probability method and naranjo 's adverse drug reaction probability scale . the who probability method and naranjo 's adverse drug reaction probability scale showed \n certain and definite ( naranjo 's score 10 ) association , respectively . as per modified hartwig and siegel scale , the severity of the reaction was moderate : level 4b . \n literature search was conducted which showed six reports on carabamazepine - induced hypertension from various countries . \n all the cases had patients who developed hypertension during carbamazepine treatment which resolved on discontinuation . to the best of our knowledge , \n discussed the possible role of anti - diuretic hormone in the production of this infrequent side effect . \n the possible mechanism as to why the antihypertensive therapy is unresponsive as explained by downey et al . \n is that antiepileptic agents could induce drug - metabolizing system and thus reduce the effects of antihypertensive medications . \n another author stated that carbamazepine induces the cytochrome p450 , which catalyze the metabolism of most of the antihypertensive used . \n the time sequence of start of the suspected drug and onset of hypertension are consistent with the diagnosis . \n the rechallenge by another physician led to raised blood pressure , not successfully managed by the appropriate treatment . \n the symptoms recovered after withdrawal of the suspected drug and the patient did not develop hypertension further owing to cessation of carbamazepine which is suggestive of possible association between carbamazepine and hypertension . \n carbamazepine - induced hypertension is rare and this report may act as alerting mechanism to the health care professionals . \n neurologists , doctors and experts in various fields , pharmacist and patients need to be aware of the potential of carbamazepine - induced hypertension . \n \n ", "role": "user" }, { "content": "a 74-year - old female with trigeminal neuralgia developed hypertension after the initiation of carbamazepine therapy . \n the time sequence of start of the suspected drug and onset of hypertension are consistent with the diagnosis . \n the hypertension did not resolve with antihypertensive therapy or dose reduction of carbamazepine . \n patient recovered after the carbamazepine therapy was discontinued . \n the positive rechallenge and positive dechallenge showed association of carbamazepine therapy with hypertension as its adverse effect . \n this is a rare case that we report of carbamazepine - induced hypertension and this report may act as alerting mechanism to the health care professionals especially neurologists .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: from june 2009 to october 2010 , 210 patients who had been diagnosed with rotator cuff tears and treated via arthroscopic rotator cuff repair were included as subjects . after obtaining institutional review board approval ( no . \n 12 - 005 ) by inje university busan paik hospital to review patients ' charts , a retrospective study was initiated . \n inclusion criteria were as follows : ( 1 ) posterior superior rotator cuff tear of the supraspinatus and infraspinatus tendons , but not the subscapularis tendon , ( 2 ) no concomitant biceps or acromioclavicular joint lesion , ( 3 ) 6 weeks , 3 months , 6 months , and 12 months postoperative follow - up was possible , ( 4 ) the patients had been treated with the same postoperative medical treatment and rehabilitation according to type of tear size , and ( 5 ) different preoperative and postoperative pain character . for patients with small to medium tears , we started a level 1 rehabilitation program for 4 weeks after surgery . \n one day after surgery , we started pendulum exercises and the passive forward flexion exercise . \n one week after surgery , we carried out active forward flexion exercises , extension exercises and active scapular exercises . on the third postoperative week , we conducted supportive active external rotation exercises from an abducted state . between the fourth and sixth \n postoperative weeks , we allowed multidirectional stretching exercises and started internal rotation exercises and cross body adduction exercises as level 2 rehabilitation . from the sixth week to the twelfth week after surgery , we started the phase i strengthening exercises , including resistive scapular strengthening exercises and manual resistive exercises of the rotator cuff and deltoid muscle . between the twelfth week and sixth month \n , we allowed sports and job activities . for patients with large to massive tears , the level 1 rehabilitation program lasted for the first six weeks . on the day after surgery , we started pendulum exercises . \n one week after surgery , we conducted passive forward flexion exercises , passive external rotation exercises , and active scapular exercises . \n six weeks after the operation , we allowed multidirectional stretching exercises and started internal rotation exercises , cross body adduction exercises and extension exercises as part of the level 2 rehabilitation . \n in addition , we carried out isometric exercises of rotator cuff , phase i strengthening exercises , including active scapular exercise . between the twelfth and sixteenth weeks \n , we started resistive scapular strengthening exercises , manual resistive exercise of the rotator cuff and deltoid muscle , which was level 3 rehabilitation . on the sixteenth week after surgery \n , we allowed the patient to resume sports activities and return to his or her job activities as part of the level 4 rehabilitation program . \n patients were excluded for the following reasons : ( 1 ) they had undergone operations on the same shoulder previously , ( 2 ) had degenerative arthritis and inflammatory arthritis , ( 3 ) had a cervical spine lesion , ( 4 ) had auto insurance or industrial accident compensation insurance , ( 5 ) were diagnosed with neuropsychiatric problems , or ( 6 ) an inconsistent change of pain pattern , for example , the patient developed preoperative pain characteristics during postoperative follow - up . using these criteria , 84 patients among the initial 210 patients \n the degree of pain was evaluated by a visual analog scales ( vas)14 ) score . \n however , pain is subjective in the sense that it could vary for each patient . \n sometimes it could be meaningless itself , so we consider that the degree of pain change in an individual is a more valuable measurement . \n the vas scores from the first outpatient clinic department visit before surgery were used as the preoperative degree of pain . \n the evaluation of postoperative pain was conducted by vas scores during the post - operative outpatient interviews at 6 weeks , 3 months , 6 months , and 12 months . \n outpatients were asked to report vas scores based on when they felt the most severe pain , one week prior to the outpatient interviews . \n range of motion ( rom ) was checked by a single examiner during outpatient visits both before the operation and then again at 3 , 6 , and 12 months after the operation . \n radiologic evaluation was performed at the postoperative time frames of 3 and 12 months using ultrasonography , and at 6 months using computed tomography ( ct ) arthrography . \n the factors that were thought to affect postoperative pain were evaluated after dividing the factors into three groups : preoperative , operative , and postoperative . \n age , gender , occupation , pain onset , trauma history , degree of pain , smoking history , synovitis of glenohumeral joint , and the size and degenerative change of the ruptured rotator cuff were evaluated as preoperative factors . the operational method and the number of anchors used \n we evaluated mean pain during the outpatient follow - up period , and compared changes between visits in addition to determining the pain pattern following the operation . \n first , we classified the patients that displayed a strictly decreasing pain pattern over our set time points as group i , and the rest as group ii . \n the differences of the preoperative , operative , and postoperative factors between each group were evaluated . \n second , we devised another classification system to elucidate the factors behind low postoperative pain intensity . \n we defined group a as the group of patients showing lower pain intensity than average for each follow - up period , and we labeled the rest as group b. as we did with group i and group ii , we examined the differences in the preoperative , operative , and postoperative factors between group a and group b. we then analyzed the factors behind the lower intensity pain pattern for each follow - up period . also , the patients were divided into acute onset and chronic onset pain groups . \n the acute onset pain group consisted of patients who had their operations within 3 months of first noticing pain , while the chronic onset pain group was defined as patients who were operated on at least 3 months after the onset of pain . \n statistical differences for categorical variables between groups were analyzed by a chi - square test . \n the relationship between variables and postoperative pain pattern was analyzed by a logistic regression test . \n the level of significance was set at 0.05 by use of sas ver . 9.0 ( sas institute , cary , nc , usa ) . \n the average pain scores at 6 weeks , 3 months , 6 months , and 12 months after the operation were 4.04 1.78 , 3.01 1.79 , 1.54 1.18 , and 0.85 0.84 , respectively . \n postoperative pain tended to decrease for every period , and the differences between each period were statistically significant ( p = 0.001 ) ( fig . \n the changes in the mean vas score between 1.5 months and 3 months postoperatively , between 3 months and 6 months postoperatively , and between 6 months and 12 months postoperatively in group i were 1.56 0.96 , 1.83 1.24 , and 0.7 0.66 , respectively . \n for group i , there were no statistically significant differences in the mean vas score between any two successive time points . \n for group ii , the changes in the mean vas score between 1.5 months and 3 months , 3 months and 6 months , and 6 months and 12 months were 1.19 1.10 , 2.23 2.08 , and 1.01 0.92 , respectively . \n unlike group i , group ii 's differences in mean vas scores between any two successive periods were statistically significant ( p = 0.001 and p = 0.001 , respectively ) . the mean vas score at postoperative 6 weeks , 3 months , 6 months , and 12 months was in 2.4 1.06 , 1.44 0.58 , 0.66 0.24 , and 0.36 0.26 , respectively , in group a and 4.79 1.53 , 3.72 1.7 , 1.93 1.23 , and 1.07 0.84 , respectively , in group b. there was a statistically significant difference between the groups in the follow - up period ( p < 0.05 ) ( fig \n , initial vas scores showed a significant difference between group i and group ii ( p = 0.0161 ) . \n other factors , such as age , tear size anterior to posterior ( ap ) , tear size medial to lateral ( ml ) , and number of anchors used were not found to have a statistically significant difference between groups ( p = 0.9332 , 0.6633 , 0.5652 , and 0.1561 , respectively ) ( table 1 ) . among categorical factors , combined medical disease , pain onset , and synovitis showed a statistically significant difference between group i and group ii ( p = 0.034 , 0.042 , and 0.003 , respectively ) . \n other factors showed no statistically significant difference between the two groups ( table 2 ) . with regard to continuous factors , \n tear size ml showed a significant difference between group a and group b ( p = 0.041 ) . \n other factors such as age , tear size ap , number of anchors used , and initial vas score did not show a statistically significant difference between the groups ( p = 0.678 , 0.147 , 0.174 , and 0.443 , respectively ) ( table 3 ) . among the categorical factors , \n stiffness of forward flexion , internal rotation and external rotation at 3 months postoperatively , and synovitis were shown to have statistically significant differences between group a and group b ( p = 0.006 , 0.013 , 0.012 , and 0.008 , respectively ) . \n the results of the continuous and catergorical variables in previous analysis were different from the results of the univariate analysis , so we conducted a multiple regression analysis using the results of univariate analysis . in single analysis and multiple regression tests for factors influencing the group with the strictly decreasing pain pattern , initial vas and onset of pain \n were shown to be statistically significant factors ( table 5 ) . with regard to the factors influencing the lower intensity pain pattern for each period , stiffness of internal rotation at 3 months postoperatively was shown to be a statistically significant factor in both single and multiple regression tests ( table 6 ) . \n with regard to continuous factors , initial vas scores showed a significant difference between group i and group ii ( p = 0.0161 ) . \n other factors , such as age , tear size anterior to posterior ( ap ) , tear size medial to lateral ( ml ) , and number of anchors used were not found to have a statistically significant difference between groups ( p = 0.9332 , 0.6633 , 0.5652 , and 0.1561 , respectively ) ( table 1 ) . among categorical factors , combined medical disease , pain onset , and synovitis showed a statistically significant difference between group i and group ii ( p = 0.034 , 0.042 , and 0.003 , respectively ) . \n other factors showed no statistically significant difference between the two groups ( table 2 ) . \n with regard to continuous factors , tear size ml showed a significant difference between group a and group b ( p = 0.041 ) . \n other factors such as age , tear size ap , number of anchors used , and initial vas score did not show a statistically significant difference between the groups ( p = 0.678 , 0.147 , 0.174 , and 0.443 , respectively ) ( table 3 ) . among the categorical factors , \n stiffness of forward flexion , internal rotation and external rotation at 3 months postoperatively , and synovitis were shown to have statistically significant differences between group a and group b ( p = 0.006 , 0.013 , 0.012 , and 0.008 , respectively ) . \n the results of the continuous and catergorical variables in previous analysis were different from the results of the univariate analysis , so we conducted a multiple regression analysis using the results of univariate analysis . in single analysis and multiple regression tests for factors influencing the group with the strictly decreasing pain pattern , initial vas and onset of pain \n were shown to be statistically significant factors ( table 5 ) . with regard to the factors influencing the lower intensity pain pattern for each period , stiffness of internal rotation at 3 months postoperatively was shown to be a statistically significant factor in both single and multiple regression tests ( table 6 ) . \n in the current literature , postoperative pain has been shown to have a strictly decreasing pattern at all given time intervals . \n in addition , the change in the decrease in pain was statistically significant at all the time intervals . \n stiglitz et al.15 ) also evaluated postoperative pain patterns after arthroscopic shoulder surgery during a 1-year postoperative period . \n they found that postoperative pain decreases by 7 days after the operation , as compared with the preoperative pain . \n however , the current literature did not evaluate postoperative pain changes compared with preoperative pain , only evaluating postoperative pain changes themselves . \n the follow - up time interval in the current study , different from the previous study , is thought to be more consistent for evaluating changes in postoperative pain . \n the reason for postoperative pain and the changing pattern at different time intervals could not be evaluated in this study . \n however , we selected subjects who had posterior superior rotator cuff tears and no combined biceps and acromioclavicular lesions among those who had a different character of pain after their operation . by adhering to these strict inclusion and exclusion criteria , \n the subjects in this study represent uncomplicated postoperative pain during healing periods after an operation . \n so , the strictly decreasing pain pattern may show an uncomplicated postoperative pain pattern during healing . \n initial vas and pain onset affected the pattern of pain change during the follow - up period . \n the higher initial vas score and acute pain onset affected the strictly decreasing postoperative pain pattern . on the contrary , \n the lower initial vas score and chronic pain onset had an irregular effect in terms of decreasing postoperative pain patterns . \n desai and cheung16 ) assessed the pain of 39 elbow and 38 shoulder surgery patients preoperatively and postoperatively using the short - form mcgill pain questionnaire . \n each patient scored their pain both preoperatively and then 3 days and 6 weeks postoperatively . \n the researchers found that preoperative pain and the anticipated postoperative pain were both independent predictors of increased postoperative pain . \n kuijpers et al.17 ) evaluated prognostic indicators of favorable outcomes and poor outcomes in narrative reviews from literature on shoulder disorders and suggested that an extended duration of pain is related to worse outcomes . in the current literature , preoperative pain intensity and onset \n these results are similar to those of desai and cheung16 ) and kuijpers et al.17 ) however , the current study examined patients who had arthroscopic rotator cuff repair and evaluated postoperative pain changes and the factors affecting postoperative pain patterns . \n this is different from desai and cheung 's study,16 ) which evaluated pain changes between the preoperative and postoperative period among heterogeneous subjects , such as those with fracture reduction and nerve decompression . \n in addition , kuijpers et al.17 ) studied prognosis factors for shoulder disorders after non - operative treatment , which is different from our study 's inclusion of only arthroscopic rotator cuff repair patients . further , our research evaluated the factors affecting postoperative pain patterns and is different from kuijpers et al.,17 ) as their research showed only the prognostic factors for shoulder disorders . in the current literature , as the initial vas increases , the pattern of postoperative pain decrease is regular . \n this study can not define the pathomechanism of preoperative pain and its effect on the strictly decreasing postoperative pain pattern . in our opinion \n , these findings may result from continued decreasing pain and better satisfaction among patients who had high levels of pain before their operations . \n in addition , as the duration of preoperative pain increased , the decrease of postoperative pain was irregular . that may come from synaptic plasticity in chronic pain , which amplifies pain in the spinal cord . \n improvement of the shoulder lesion could not be recognized as a pain - decreasing factor . in the current study \n , we found that postoperative internal rotation stiffness 3 months after surgery affects the pain intensity for each period . \n as stiffness becomes worse , postoperative pain remained high , in contrast to the lower levels of postoperative pain in patients who had mild stiffness . \n loss of motion is a well - recognized postoperative complication of rotator cuff repair.5,18,19,20,21 ) the risk factors of stiffness are known include calcific tendinitis , adhesive capsulitis , single tendon cuff repair , pasta repair , age younger than 50 , and those with workers ' compensation insurance.22 ) namdari and green23 ) studied 345 patients who underwent rotator cuff repair and researched correlations between preoperative and postoperative variables and postoperative limitations of motion . \n they evaluated active forward elevation , active external rotation , and passive internal rotation after rotator cuff repair , and found that limitation of motion causes poor operative outcome and worse postoperative pain statistically . \n this study reached the same result as the study by namdari and green,23 ) which found that rom loss at 3 months after surgery affects postoperative pain , especially in those with higher than average intensity pain during the follow - up period . \n however , unlike in this study , the study of namdari and green23 ) includes both open and arthroscopic surgery , and did not observe the pattern of change of postoperative pain during each period throughout follow - up . \n however , we could not define how the postoperative stiffness affects postoperative pain in our study . \n franceschi et al.24 ) reported that substance p may play a role in the pathogenesis of shoulder pain and found a 3-fold increase in levels of substance p in patients in whom postoperative stiffness developed . in our opinion , this cytokine release in stiffness may be related to postoperative pain intensity . \n even if the same degree of pain is felt , the vas score , the digital assessment for pain , could vary for each patient . in this study , therefore , we decided the differences between each patient 's vas score were a weak point , and used the degree of change for pain in a person instead because it is more valuable for comparisons . \n second , with regard to evaluating the rom in order to measure postoperative stiffness , the variables were not presented as a continuous variable , but rather as interval variables . in this way \n third , in this study , radiologic evaluations were not carried out in all patients . \n for instance , ct arthrography was conducted at 6-month postoperation in 80% of the study sample . \n fourth , in this study , subtle differences of pain control medication , including the type , duration , and other therapeutic means , such as injection or physical therapy during the follow - up period were not analyzed . in the future , \n a study with a stricter control of variables is needed . in conclusion , noncomplicated postoperative pain after arthroscopic rotator cuff repair surgery showed a strictly decreasing pain pattern at each time interval . as preoperative factors , higher initial vas scores and a more acute onset of pain affected the strictly decreasing postoperative pain pattern . as a postoperative factor , \n the stiffness of internal rotation at postoperative 3 months affected the high intensity pain pattern for each period . \n so , preoperative factors , such as initial vas score and pain onset , are not correctable and we consider that these factors could be used to predict the pattern of postoperative pain for patients with these conditions . \n in addition , the postoperative factors , such as stiffness of internal rotation at postoperative 3 months , are correctable , and we consider that the method of reducing postoperative stiffness could be helpful to reducing postoperative pain intensity .", "role": "user" }, { "content": "backgroundwe evaluated the factors that affect pain pattern after arthroscopic rotator cuff repair.methodsfrom june 2009 to october 2010 , 210 patients underwent arthroscopic rotator cuff repair operations . of them , \n 84 patients were enrolled as subjects of the present study . \n the evaluation of postoperative pain was conducted by visual analog scale ( vas ) scores during postoperative outpatient interviews at 6 weeks , 3 months , 6 months , and 12 months . \n the factors that were thought to affect postoperative pain were evaluated by dividing into three categories : preoperative , operative , and postoperative.resultspain after arthroscopic rotator cuff repair surgery showed a strictly decreasing pain pattern . in single analysis and multiple regression tests for factors influencing the strictly decreasing pain pattern , initial vas and pain onset \n were shown to be statistically significant factors ( p = 0.012 , 0.012 , 0.044 and 0.028 , respectively ) . with regard to the factors influencing lower than average intensity pain pattern for each period , \n the stiffness of internal rotation at 3 months postoperatively was shown to be a statistically significant factor in single and multiple regression tests ( p = 0.017 and p = 0.004 , respectively).conclusionshigh initial vas scores and the acute onset of pain affected the strictly decreasing postoperative pain pattern . \n additionally , stiffness of internal rotation at postoperative 3 months affected the higher than average intensity pain pattern for each period after arthroscopic rotator cuff repair .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: human oral cavity is inhabited by many different kinds of microorganisms whose composition , metabolic activity , and pathogenicity are affected by external and internal factors [ 1 , 2 ] . among fungi , yeasts belonging to the genus candida \n candida species are the major causes of mucocutaneous infections that are usually classified as oropharyngeal , esophageal , and vulvovaginal candidiasis . \n oropharyngeal candidiasis ( opc ) is the commonest mucocutaneous candidiasis amongst hiv - positive patients worldwide [ 1 , 3 , 4 ] , occurring in more than 95% of aids patients [ 1 , 5 ] . \n high viral load , low cd4 + t lymphocyte count , and disease progression have been incriminated as the greatest risk factors for the development of opc [ 46 ] . \n although opc infection is caused by many different species of yeast with the genus candida , c. albicans remains the most predominant reported species globally . \n opportunistic fungal infections resistant to antifungal agents have been increasingly documented and there are concerns that their frequency will likely continue to increase . \n demonstrated that emergence of resistance to antifungal agents in hiv / aids patients with opc is common . increased use of antifungal agents to treat fungal infections that occur in immune - compromised patients \n has been incriminated as a major factor for emergence of drug resistant yeast isolates , failure to respond to antifungal treatment with appropriate doses for a standard duration of time , and noticeable shift towards non - albicans candida species with relative resistance to azole antifungal agents and associated refractory and recurrent infections [ 11 , 12 ] . \n although the introduction of highly active antiretroviral therapy ( haart ) has dramatically reduced the incidence of opportunistic infections [ 1315 ] in hiv - positive individuals who have received antifungal drugs , opc with a shift in the spectrum of candida species remains the most frequent hiv associated oral lesion in developing countries including ethiopia . \n emergence of non - albicans candida species as etiologic agents of mucocutaneous candidiasis and development of drug resistance by c. albicans to the currently available antifungal principles have initiated further studies in isolation and identification of the causative fungal species and determining of their antifungal profile . in ethiopia , fungal culture and in vitro drug susceptibility pattern of yeast \n therefore , little is known regarding the distribution and the in vitro antifungal susceptibility profile of yeasts isolated from hiv infected patients with opc . to this end , \n determining species distribution of yeast isolates from ethiopian hiv infected patients with and without haart and evaluating their drug susceptibility profile to antifungal drugs is one of the highest priorities . \n the present study was a single institutional cross - sectional study carried out in zewditu memorial hospital , addis ababa , ethiopia , over ten moths ( september 2013 to june 2014 ) . \n the hospital is one of the dedicated centers in the city for the management of hiv / aids patients and runs both outpatient and in - patient services . the hiv clinic of the hospital consists of an integrated counseling and testing center in addition to cd4 + t cells monitoring laboratory and art pharmacy . \n two hundred and twenty - four consecutive hiv infected patients with opc , 112 under haart and 112 not under haart , were recruited when they were referred to the hospital for voluntary hiv counseling and testing ( vct ) center and/or for outpatient departments ( opd ) and/or referred from other health institution . \n inclusion criteria were a previous positive diagnosis of hiv , will to participate in the study , a presumptive diagnosis of opc following an appropriate complaint made at the clinic visit , and no history of antifungal therapy within two weeks prior to the attendance . upon admission to the study , oral examination of each patient \n was performed and then oropharyngeal swabs were taken from oral lesions , oral cavity mucosa , and dorsal surface of the tongue and the floor of the mouth of each study subject with sterile rayon tipped applicator stick swabs . \n all samples were then inoculated into brain - heart infusion broth ( oxoid , basingstoke , uk ) supplemented with chloramphenicol ( 50 mg \n tubes of inoculated brain heart infusion were then transported to the microbiology laboratory of the department of medical laboratory science , college of health sciences , and addis ababa university , and incubated at 3537c for 18 to 24 hours . \n the brain - heart infusion broth was then subcultured onto sabouraud dextrose agar ( sda ) supplemented with chloramphenicol ( oxoid , basingstoke , uk ) and incubated aerobically for 48 hours . \n yeasts were identified by employing conventional biochemical and assimilation test procedures , using chromagar candida culture medium ( becton dickinson ) as per the instruction of the manufacture , germ - tube formation in human serum , and production of blastoconidia , pseudohyphae , and chlamydospore on cornmeal agar ( oxoid , basingstoke , uk ) . \n the in vitro antifungal susceptibility pattern of yeast isolates to six antifungal agents , fluconazole , ketoconazole , miconazole , amphotericin b , clotrimazole , and nystatin ( usp , rockville , usa ) was determined by agar diffusion procedure according to antifungal disk potency of rosco diagnostica company ( neosensitabs , denmark ) . \n all antifungal drugs were generously provided as standard powder by ethiopian food medicine and healthcare administration and control authority ( efmhaca ) . \n the stock solutions were prepared by dissolving standard powders of each drug in their specific solvents ( dmso , water , and ethanol ) , after which they were impregnated into 6 mm sterile blank antibiotic assay disks ( sigma aldrich , germany ) at the following potencies : clotrimazole 10 g / disk , fluconazole 25 g / disk , ketoconazole 10 g / disk , miconazole 10 g / disk , amphotericin b 10 g / disk , and nystatin 50 g / disk according to antifungal disk potency of rosco diagnostica company ( neosensitabs , denmark ) . \n yeast inocula were prepared by suspending five colonies of each isolate from 24-hour old pure culture in sterile saline and suspensions were then adjusted to the turbidity of a 0.5 mcfarland standard to obtain 1 10 to 5 10 cells per ml . \n yeast suspensions were then streaked onto muller hinton agar ( oxoid , basingstoke , uk ) supplemented with 2% glucose and 0.5 g / ml methylene blue ( gmb ) . dried antibiotic assay disks impregnated with antifungal agents as described above \n were placed on muller hinton agar seeded with yeast isolates after the plates were dried at ambient temperature for 15 minutes . \n diameter zone of inhibition was measured by a metal caliper after 24 and 48 h of incubation at 35c . \n filter paper disks soaked in respective solvents without antifungal agent were placed on culture plate to serve as control . \n criteria of susceptibility and resistance of antifungal agents were determined according to zone diameter interpretation for fungi of rosco diagnostica company ( neosensitabs , denmark ) as shown in table 1 . \n data were captured in ms excel and analyzed using spss version 20 ( ibm , chicago , usa ) . \n comparisons of proportion between groups were carried out by chi - square test and the significant level set at p < 0.05 . \n all ethical considerations and obligations were duly addressed and the study was conducted after the approval of the department research and ethical review committee ( drerc ) of the department of medical laboratory sciences , college of health sciences , and addis ababa university and the addis ababa city health bureau . \n the respondent was given the right to refuse to take part in the study and to withdraw at any time during the study period . \n when the participants were found to be positive for fungal pathogen , they were informed by the hospital clinician and received proper treatment . \n assent form was completed and signed by family member and/or adult guardian for participants under the age of 18 years . \n in the present study , a total of 224 clinical samples were collected from hiv - sero - positive individuals with opc of which 155 ( 69.2% ) were from female patients and 69 ( 30.8% ) were from male patients . \n the ages of study subjects ranged from 3 to 78 years with a median age of 34.5 years . among the study participants , 112 ( 50% ) were under haart among which 76 ( 49% ) were females and 36 ( 52.2% ) were males . \n out of a total of 224 study subjects , 155 yeast isolates were recovered giving a prevalence of 69.2% colonization rate , of which 105 ( 67.7% ) were from female patients and 50 ( 33.3% ) were from male patients ( table 3 ) . \n differences in the isolation rate between gender were not statistically significant [ ( or = 0.750 , 95% ci , 0.4141.36 ) ( p = 0.342 ) ] . \n of one hundred and fifty - five ( 155 ) yeast isolates recovered from hiv patients with opc , ninety - one ( 91 ) isolates were from patients that were not under haart and 64 isolates were from patients that were under haart ( table 4 ) . \n difference in yeast isolation rate between patients under haart and those that were not under haart was statistically significant [ ( or = 2.433 , 95% ci = 1.5964.240 ) ( p = 0.002 ) ] . \n species distribution as presented in table 4 showed that c. albicans was the most frequently isolated species accounting 106 ( 68.4% ) of the total yeast isolates followed by c. glabrata , 24 ( 15.5% ) , c. tropicalis , 17 ( 11.0% ) , c. krusei , 4 ( 2.6% ) , and c. kefyr , 2 ( 1.3% ) . \n c. laurentii , 1 ( 0.65% ) , and rhodotorula species , 1 ( 0.65% ) , were the only isolated non - candida species from these patients . \n sixteen ( 16 ) patients had mixed infections and c. albicans appeared in all mixed infections . c. albicans appeared in seven patients with c. glabrata and in five patients with c. tropicalis , c. krusei , c. kefyr , and c. laurentii , and rhodotorula species appeared once with c. albicans . \n the susceptibility of yeast isolates to the six antifungal drugs is summarized in table 5 . as can be shown from the table , 83.9% ( 130/155 ) of the total yeast isolates \n were susceptible , 10.3% ( 16/155 ) were intermediate , and 5.8% ( 9/155 ) were resistant to amphotericin b. one hundred and forty - eight ( 95.5% ) , 16 ( 10.3% ) , and 9 ( 5.8% ) of yeast isolates were susceptible , intermediate , and resistant to clotrimazole , respectively . \n yeast isolates were more resistant to fluconazole in which 19 ( 12.3 ) were resistant , while 123 ( 79.4% ) and 13 ( 8.4% ) were susceptible and intermediate , respectively . \n one hundred and thirty - four ( 86.5% ) , 8 ( 5.2% ) , and 13 ( 8.4% ) of yeast isolates were susceptible , intermediate , and resistant to ketoconazole . \n more intermediate and least resistant yeasts were recorded against miconazole 21 ( 13.5% ) and 1 ( 0.6% ) was being intermediate and resistant against the drug , s : susceptible ; i : intermediate ; r : resistant ; am : amphotericin b ; clo : clotrimazole ; flu : fluconazole ; keto : ketoconazole ; mic : miconazole ; ny : nystatin , while 133 ( 85.8% ) were being susceptible to the drug . \n more yeast isolates were susceptible to nystatin in which 150 ( 96.8 ) were being susceptible , while 3 ( 1.9% ) and 2 ( 1.3% ) were intermediate and resistant to the drug , respectively . \n nystatin was the most effective drug against c. tropicalis and c. krusei with no resistant strains of both species to the drug being observed . \n nystatin also exhibited the greatest activity against c. albicans with no resistant strain found ; overall , only one out of 106 c. albicans isolates ( 0.9% ) was found out to be intermediate , while 105 ( 99.1% ) were susceptible to this agent . \n nystatin was also relatively active drug against c. glabrata to which 22 ( 91.6% ) out of 24 isolates were susceptible to the drug . among the three major isolates of the present study , c. tropicalis was less sensitive to amphotericin b ( 53.0% ) , while the sensitivity of c. albicans and c. glabrata to the drug was 92.4% and 87.5% , respectively . \n fluconazole was relatively the least effective drug against c. albicans , c. glabrata , and c. tropicalis which were the major isolates of this study . the sensitivity of c. albicans , c. glabrata , and c. tropicalis to the drug was 89.6% , 79.2% , and 58.8% , respectively . none of the isolates of c. kefyr and c. krusei were susceptible to ketoconazole , but the sensitivity of c. albicans , c. glabrata , and c. tropicalis to the drug was 89.6% , 83.3 , and 58.8% , respectively . \n the sensitivity of c. albicans , c. glabrata , and c. tropicalis to clotrimazole was 99.1% , 87.5% , and 882% , respectively . among the azole \n antifungal agents , more intermediate strains were produced against miconazole in which 100% of c. krusei and c. kefir , 35.5% of c. tropicalis , \n 20.8% of c. glabrata , and 3.8% of c. albicans were intermediate to the drug . \n little is known about the profile and the in vitro antifungal susceptibility pattern of oral yeasts in ethiopian hiv infected patients with opc . \n our finding of c. albicans as the predominant species was consistent with similar earlier [ 6 , 1925 ] and recent [ 26 , 27 ] studies conducted internationally . \n our finding of c. albicans as the predominant species was also consistent with similar study conducted locally in gondar , southwest ethiopia , but there were some striking differences in the species spectrum and percentage of yeast isolates . \n for instance , the culture positivity rate obtained in our study ( 69.2% ) was comparatively lower than the culture positivity rate ( 82.3% ) reported in the study carried out in gondar . \n differences in the recovery rate of different species of yeasts were also observed between these two studies . \n the recovery rate for c. albicans in our study was 106 ( 68.4% ) , 24 ( 15.5% ) for c. glabrata , 17 ( 11% ) for c. tropicalis , 4 ( 2.6% ) for c. krusei , and 2 ( 1.3% ) for c. kefyr . in the study conducted in gondar , the recovery rate of different species of yeasts was higher , 139 ( 78.5 ) for c. albicans , 40 ( 22.5% ) for c. glabrata , 17 ( 14.1% ) for c. tropicalis , and 10 ( 5.6 ) for c. krusei , respectively . \n similarly , their study apparently could not isolate c. kefyr , c. laurentii , and rhodotorula species that made 2.6% of yeast isolates in our study . c. kefyr and other yeasts \n have been isolated as one of the oral yeasts by other works [ 6 , 19 ] . \n disparity in species spectrum and percentage of isolation rates of different yeasts between the two studies could be resulting from differences in study subjects enrolled in the two studies since half of our study subjects were under haart . a marked decrease in the incidence of oral candidiasis in patients receiving haart \n although numerous studies on the prevalence of different candida species have led to the general consensus that c. albicans is the most commonly isolated species in the oral cavity of patients infected with hiv , there has been a growing trend of recovery of non - albicans candida species . \n this is evident by the present study in which the isolation rate of non - albicans candida species was 30.8% . \n comparatively , less recovery rates of non - albicans candida species of 15% in tanzania , 19% in ethiopia , 22% in texas , and 16.5% in mexico have been reported . \n similarly , comparatively higher recovery rates of 55% and 50% of non - albicans candida species have been reported in similar studies conducted in nigeria and brazil , respectively . \n a total of 155 isolates were obtained from 224 specimens during the 10-month period . of clinical specimens collected 7.1% ( 16/224 ) \n yielded more than one yeast ( mixed culture ) species in which c. albicans was isolated from all mixed cultures . \n four ( 1.5% ) and forty ( 18.6% ) mixed infections were reported by similar studies conducted in tanzania and ethiopia , respectively . \n fifteen percent ( 15% ) and 29% mixed infections of candida species among hiv patients were also reported by baumgartner et al . and schoofs et al . \n our finding regarding the prevalence of mixed culture may reflect the change from single to multiple candida species isolates , which have been responsible for epidemiological shift in opc . \n the most predominant species isolated together with c. albicans in the present study was c. glabrata ( 43.8% ) . \n the significance of this finding is that c. glabrata may replace c. albicans under selective pressure of fluconazole , resulting in infections refractory to the current fluconazole based treatment in ethiopia . like other african countries , \n the present guideline of the ethiopian ministry of health ( moh ) for the management of candidiasis includes fluconazole as a first choice drug and ketoconazole , miconazole ointment as alternative antifungal agents [ 33 , 34 ] . in this study , \n the occurrence of opc was higher in the non - haart group than in the haart group . \n of one hundred and fifty - five ( 155 ) yeast isolates recovered from hiv patients with opc , ninety - one ( 91 ) isolates were from patients that were not under haart and 64 isolates were from patients that were under haart . \n ceballos - salobrena et al . , in their study of 154 patients who were on haart for a minimum of 6 months , reported a 30% reduction in the prevalence of opc . \n ramirez - amador et al . , in their 12-year prospective study in mexico , observed a decrease in the prevalence of opc by half during the course of their defined study periods . in the present study , \n the number of female patient participants ( 155 , 69.2% ) was greater than male patient participants ( 69 , 30.8% ) . \n the significant difference in the number of female - male study participants could be due to the fact that female patients are more exposed to hiv infection than male patients . \n opportunistic fungal infections resistant to antifungal agents have been increasingly documented and their frequency will likely continue to increase . \n this phenomenon appears largely due to the extensive use of antifungal agents to treat fungal infections that typically occur in severely immune - compromised and/or critically ill patients . \n are among the leading fungi responsible for these invasive infections . while antifungal resistance has been described with each of these fungi , resistance among candida constitutes by far the most significant problem . \n this was evident by the present study in which 31% ( 48/155 ) yeast isolates were found out to be resistant to one or more antifungal drugs tested . \n this result was comparable with earlier study conducted in ethiopia but relatively higher than similar studies from africa [ 6 , 21 ] and the rest of the world . \n of the 48 yeast isolates which were resistant to one or more drugs more than 64.6% ( 31/48 ) of the isolates were non - albicans candida and this may reflect the intrinsic less susceptibility of some of the non - albicans candida species to some antifungal agents such as fluconazole and/or the reflection of the inappropriate use of antifungal in study area . \n amphotericin b is traditionally used in topical formats , although it may be administrated systemically for the treatment of systemic infections in hospitalized patients . \n our result on in vitro susceptibility of candida species to polyene antifungal agents ( amphotericin b and nystatin ) was consistent with that reported previously and showed that candida isolates recovered in the study were highly sensitive to these polyene antifungal agents . \n out of 155 yeast isolates , only 2 ( 1.3% ) and 9 ( 5.8% ) isolates were resistant to nystatin and amphotericin b , respectively . \n these results would support the effectiveness of topical amphotericin b and nystatin therapy for superficial candidosis . \n recurrent opc and repeatedly and/or prolonged exposure of patients to antifungal therapy induce reduced susceptibility of yeasts to azole antifungal agents [ 8 , 9 ] . \n selective pressure caused by antifungal agents and azole cumulative doses due to exposure to several courses of short- or long - term suppressive therapies in patients with opc has been incriminated as a cause of induction of reduced susceptibility of yeasts to azole antifungal agents [ 9 , 10 ] . \n similarly , the risk of developing opc with reduced susceptibility to azoles has been associated with greater duration of hiv infection and severe immunosuppression . \n isolation of yeasts more resistant to azole antifungal agents in the present study could be explained based on the above risk factors . \n regardless of yeast identity , 19 ( 12.3% ) , 13 ( 8.4% ) , and 4 ( 2.6% ) isolates were resistant to fluconazole , ketoconazole , and clotrimazole , respectively . \n fluconazole was also the least active drug against c. albicans which is the most frequently isolated yeast . though fluconazole has been widely used for the treatment of mucosal candidiasis because of its low toxicity and ease of administration local , \n many international studies have reported fluconazole resistance in candida strains isolated from hiv infected patients with opc [ 4042 ] . \n although we have no immediate reasons for a higher resistance of yeast isolates to fluconazole in the present study as indicated above , the accessibility of the drug being free of charge with the scale - up of haart since 2005 in the country and the frequent usage of the drugs as the drug of choice in the settings could be responsible . \n reported that 9.8% c. albicans isolated from patients with no previous treatment were resistant to fluconazole which increased to 44.7% after treatment with this drug . \n this data suggest that the continued use of fluconazole may lead to clinical treatment failure which is significantly correlated with reduced susceptibility to fluconazole and other azoles . in general \n , our study has shown that polyene antifungal ( amphotericin b and nystatin ) was more active than azole antifungal agents . \n therefore , it is advisable to use these drugs as a second - line therapy for the mucosal candidiasis which is resistant to fluconazole or for empiric therapy as reported in other studies . \n only one isolate out of the 155 yeast isolates was found out to be resistant to miconazole , but 21 ( 13.5% ) of the isolates were found out to be intermediate to the agent and why more isolates were found out to be intermediate to the agent is not clear and thus further study is recommended . \n this study demonstrated that although c. albicans is the most frequently isolated species from hiv patients that are under haart and not under haart , high prevalence of mixed culture in this study may reflect the change from single to multiple candida species isolates , which have been responsible for epidemiological shift in opc . \n infections with candida are almost endogenous ; therefore , identification of the species and corresponding susceptibility patterns to antifungal agents can be helpful for the management of these infections . \n as demonstrated in this study , resistance to azole drugs such as fluconazole , the most frequent antifungal used in the country , could be suggestive of the need for routine fungal culture and in vitro drug susceptibility pattern of yeast in medical centers in order to manage more efficiently the invasive candidiasis in the above - mentioned patients .", "role": "user" }, { "content": "background . in ethiopia , little is known regarding the distribution and the in vitro antifungal susceptibility profile of yeasts . objective . \n this study was undertaken to determine the spectrum and the in vitro antifungal susceptibility pattern of yeasts isolated from hiv infected patients with opc . method . \n oral pharyngeal swabs taken from oral lesions of study subjects were inoculated onto sabouraud dextrose agar . \n yeasts were identified by employing conventional test procedures and the susceptibility of yeasts to antifungal agents was evaluated by disk diffusion assay method . result . \n one hundred and fifty - five yeast isolates were recovered of which 91 isolates were from patients that were not under haart and 64 were from patients that were under haart . c. albicans was the most frequently isolated species followed by c. glabrata , c. tropicalis , c. krusei , c. kefyr , cryptococcus laurentii , and rhodotorula species . \n irrespective of yeasts isolated and identified , 5.8% , 5.8% , 12.3% , 8.4% , 0.6% , and 1.3% of the isolates were resistant to amphotericin b , clotrimazole , fluconazole , ketoconazole , miconazole , and nystatin , respectively . \n conclusion . yeast colonization rate of 69.2% and 31% resistance to six antifungal agents was documented . \n these highlight the need for nationwide study on the epidemiology of opc and resistance to antifungal drugs .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: positron emission tomography ( pet ) with f - fluorodeoxyglucose ( f - fdg ) has emerged as an increasingly common component of pretreatment cancer staging and response assessment . \n the use of concurrent computed tomography ( ct ) and pet ( pet - ct ) can improve detection sensitivity and is used in target delineation for radiation treatment planning . the municon phase ii trial included 110 patients with locally advanced gastroesophageal adenocarcinoma and suggested that a metabolic response ( decrease 35% in standardized uptake value [ suv ] ) 2 weeks after starting chemotherapy was strongly associated with improved survival . \n despite established sensitivity , f - fdg pet is not tumor specific , and there can be nontumor - related uptake of fdg in postoperative wound sites , normal physiological fdg uptake , and sites of postradiation inflammatory changes . \n these false - positive findings can be mistaken as residual disease or sites of new disease that may trigger unwarranted biopsies and exploratory surgeries . \n we reviewed 64 patients and identified those who received f - fdg pet before and after concurrent chemotherapy and radiation . \n we present two patients with her2-amplified gastroesophageal adenocarcinoma who were identified to have a focal area of intense f - fdg uptake in the left lobe of the liver in posttreatment pet / ct obtained after completion of concurrent crt . \n close radiographic monitoring observed waning fdg - uptake confirming nonmalignant etiology and avoidance of more invasive approaches . \n a 60-year - old man was diagnosed with clinical stage iv , her2-amplified metastatic gastroesophageal junction ( gej ) adenocarcinoma . \n pretreatment pet - ct revealed an intensely fdg - avid gej mass extending to fundus with fdg - avid subcarinal and retroperitoneal lymph nodes . \n the patient underwent laparoscopic staging demonstrating large bulky tumor along lesser curvature extending to and encasing the left gastric artery with adenopathy . \n the patient received five cycles of biweekly leucovorin , 5-flurouracil , and oxaliplatin ( folfox ) and trastuzumab 6 mg / kg every 3 weeks . \n a pet / ct showed decreased fdg avidity in the gej mass [ figure 1a and b ] . to maximize local control and swallowing function , \n he was treated with 5-fluorouracil and oxaliplatin with concurrent radiation to 37.5 gy at 2.5 gy per fraction . following radiation , he resumed full dose folfox and trastuzumab . \n a pet / ct scan was obtained 34 days after completion of radiation [ figure 1c and d ] . \n there was 109 mm 82 mm focal area of fdg uptake in the left lobe measuring suv 12.1 . \n hepatic function was within normal limits , and he tolerated crt with good clinical response . due to concerning pet - ct findings , a magnetic resonance imaging ( mri ) was performed which did not show any focal hepatic abnormalities . on retrospective imaging review , \n the fdg avid area of the liver was within the 37.5 gy isodose line [ figure 1d ] . \n due to clinical stability , a plan for repeat imaging was favored over more invasive tissue sampling , and a repeat pet / ct scan 3 months [ figure 1e and f ] and 6 months [ figure 1 g and h ] later showed resolution of the fdg avid area in the left hepatic lobe and ongoing systemic response . a 60-year - old man with gastroesophageal junction adenocarcinoma with the radiographic evolution of segment three hepatic uptake in a patient with advanced gastroesophageal adenocarcinoma . \n ( a ) axial images of pretreatment positron emission tomography - computed tomography confirms the absence of fluorodeoxyglucose uptake in the liver ( b ) positron emission tomography - computed tomography 30 , 90 , and 180 days after completion of chemoradiation revealed fluorodeoxyglucose uptake in the left lobe of liver ( arrow ) ( d , f , h ) positron emission tomography - computed tomography at 3 and 6 months after crt revealed no fluorodeoxyglucose uptake in the left lobe of the liver . respective coronal images are shown ( a , c , e , g ) . \n area within red , white , green , and blue isodose line encompasses 37.5 gy , 33.3 gy , 30 gy , and 26.3 gy , respectively . \n a 55-year - old man diagnosed with clinical t3n0m0 overall stage iib her2-amplified distal esophageal . \n a pretreatment staging pet / ct scan demonstrated a large 51 mm mass in the lower esophagus extending to the gej and to the gastric fundus measuring suv 8.4 with no suspicious nodal uptake [ figure 2a and b ] . \n he was enrolled in a neoadjuvant crt trial investigating the addition of trastuzumab to trimodality treatment in her2-overexpressing esophageal adenocarcinomas . \n he received 50.4 gy of radiation with concurrent carboplatin , taxol , and trastuzumab 2 mg / kg weekly . radiation was delivered using volume modulated arc therapy . \n a repeat pet - ct was performed 42 days from completion of radiotherapy and demonstrated fdg uptake ( suv 4.5 ) in the left lobe of the liver corresponding an area of low attenuation in the periphery of the left lobe of the liver measuring 28 mm 35 mm on ct [ figure 2c and d ] . similar to case 1 , the hepatic function was normal , and retrospective review confirmed the fdg avid area was within the 50.4 gy isodose line [ figure 2d ] . \n a follow - up pet scan 3 months later showed the resolution of fdg uptake [ figure 2e and f ] . a 55-year - old man with adenocarcinoma of the distal esophagus . \n ( a and b ) axial images of pretreatment positron emission tomography - computed tomography confirms the absence of fluorodeoxyglucose uptake in the liver , ( c and d ) positron emission tomography - computed tomography 45 days after completion of chemoradiation revealed fluorodeoxyglucose uptake in the left lobe of liver ( arrow ) , ( e and f ) positron emission tomography - computed tomography 3 months posttreatment revealed no fluorodeoxyglucose uptake in the left lobe of the liver . \n area within red , white , green , and blue isodose line encompasses 50.4 gy , 45.4 gy , 40.3 gy , and 35.3 gy , respectively . \n a 60-year - old man was diagnosed with clinical stage iv , her2-amplified metastatic gastroesophageal junction ( gej ) adenocarcinoma . \n pretreatment pet - ct revealed an intensely fdg - avid gej mass extending to fundus with fdg - avid subcarinal and retroperitoneal lymph nodes . \n the patient underwent laparoscopic staging demonstrating large bulky tumor along lesser curvature extending to and encasing the left gastric artery with adenopathy . \n the patient received five cycles of biweekly leucovorin , 5-flurouracil , and oxaliplatin ( folfox ) and trastuzumab 6 mg / kg every 3 weeks . \n a pet / ct showed decreased fdg avidity in the gej mass [ figure 1a and b ] . to maximize local control and swallowing function , \n he was treated with 5-fluorouracil and oxaliplatin with concurrent radiation to 37.5 gy at 2.5 gy per fraction . following radiation , he resumed full dose folfox and trastuzumab . \n a pet / ct scan was obtained 34 days after completion of radiation [ figure 1c and d ] . \n there was 109 mm 82 mm focal area of fdg uptake in the left lobe measuring suv 12.1 . \n hepatic function was within normal limits , and he tolerated crt with good clinical response . due to concerning pet - ct findings , a magnetic resonance imaging ( mri ) was performed which did not show any focal hepatic abnormalities . on retrospective imaging review , \n the fdg avid area of the liver was within the 37.5 gy isodose line [ figure 1d ] . \n due to clinical stability , a plan for repeat imaging was favored over more invasive tissue sampling , and a repeat pet / ct scan 3 months [ figure 1e and f ] and 6 months [ figure 1 g and h ] later showed resolution of the fdg avid area in the left hepatic lobe and ongoing systemic response . a 60-year - old man with gastroesophageal junction adenocarcinoma with the radiographic evolution of segment three hepatic uptake in a patient with advanced gastroesophageal adenocarcinoma . \n ( a ) axial images of pretreatment positron emission tomography - computed tomography confirms the absence of fluorodeoxyglucose uptake in the liver ( b ) positron emission tomography - computed tomography 30 , 90 , and 180 days after completion of chemoradiation revealed fluorodeoxyglucose uptake in the left lobe of liver ( arrow ) ( d , f , h ) positron emission tomography - computed tomography at 3 and 6 months after crt revealed no fluorodeoxyglucose uptake in the left lobe of the liver . respective coronal images are shown ( a , c , e , g ) . \n area within red , white , green , and blue isodose line encompasses 37.5 gy , 33.3 gy , 30 gy , and 26.3 gy , respectively . \n a 55-year - old man diagnosed with clinical t3n0m0 overall stage iib her2-amplified distal esophageal . \n a pretreatment staging pet / ct scan demonstrated a large 51 mm mass in the lower esophagus extending to the gej and to the gastric fundus measuring suv 8.4 with no suspicious nodal uptake [ figure 2a and b ] . \n he was enrolled in a neoadjuvant crt trial investigating the addition of trastuzumab to trimodality treatment in her2-overexpressing esophageal adenocarcinomas . \n he received 50.4 gy of radiation with concurrent carboplatin , taxol , and trastuzumab 2 mg / kg weekly . \n a repeat pet - ct was performed 42 days from completion of radiotherapy and demonstrated fdg uptake ( suv 4.5 ) in the left lobe of the liver corresponding an area of low attenuation in the periphery of the left lobe of the liver measuring 28 mm 35 mm on ct [ figure 2c and d ] . similar to case 1 , the hepatic function was normal , and retrospective review confirmed the fdg avid area was within the 50.4 gy isodose line [ figure 2d ] . \n a follow - up pet scan 3 months later showed the resolution of fdg uptake [ figure 2e and f ] . a 55-year - old man with adenocarcinoma of the distal esophagus . \n ( a and b ) axial images of pretreatment positron emission tomography - computed tomography confirms the absence of fluorodeoxyglucose uptake in the liver , ( c and d ) positron emission tomography - computed tomography 45 days after completion of chemoradiation revealed fluorodeoxyglucose uptake in the left lobe of liver ( arrow ) , ( e and f ) positron emission tomography - computed tomography 3 months posttreatment revealed no fluorodeoxyglucose uptake in the left lobe of the liver . \n area within red , white , green , and blue isodose line encompasses 50.4 gy , 45.4 gy , 40.3 gy , and 35.3 gy , respectively . \n here , we present the largest series to date highlighting the radiographic phenomenon of focal fdg - uptake in the liver mimicking residual or progressive disease following radiotherapy in gastroesophageal cancers . \n our cases suggest that hepatic uptake resolves within 12 weeks of radiotherapy and with careful clinical and radiographic communication invasive procedure may be avoidable . \n patients with gej cancers who undergo concurrent chemotherapy and radiation can develop radiation - induced hepatic injury . \n the symptomatology of radiation - induced livery disease ( rild ) typically includes fatigue , right upper quadrant pain , ascites , anicteric hepatomegaly , and elevated alkaline phosphatase . \n pathologically , the liver parenchyma displays veno - occlusive disease , marked central venous congestion , sparing of large veins , and atrophy of adjacent hepatocytes \n paudel et al . reported two patients with f - pet uptake in the left lobe of the liver following chemoradiation . \n there were no elevated liver enzymes during or after chemoradiation and personal communication with corresponding author . \n our findings are consistent paudel et al . with neither of our patients experiencing liver function abnormalities . \n although not well studied , both our patients were treated with intensity - modulated radiotherapy which optimizes dose to target while minimizing dose to surrounding normal organs such as the liver and kidney , possibly reducing rild risk . \n previous small studies have suggested that hepatic fdg - uptake is seen within 26 weeks after radiation therapy in distal esophageal cancers . in a series of 26 patients , \n abnormal left lobe hepatic fdg - uptake was seen in 8% of patients at 6 weeks , with fdg uptake increasing focally > 50% over baseline in two patients ( 54% and 133% ) . \n false - positive fdg - uptake after rt has led to potentially avoidable surgical intervention in some cases . \n reported a case of fdg uptake in the left lobe of liver 28 days after completion of radiation for esophageal cancer , with resolution on follow - up pet scan 3 months later , consistent with our cases . in our series , we found fdg uptake within 34 and 42 days postradiation . \n interestingly , among the remaining 62 patients in our database , hepatic fdg - uptake was not observed , suggesting that intrinsic biologic features in addition to treatment and imaging timing may play a role . \n neither of our patients was known to have preexisting hepatitis , steatohepatitis , or on concurrent hepatotoxins . \n we conducted a literature search in the pubmed database to identify similar cases . using the search terms , false - positive , fdg uptake , liver , and radiation we identified a total of four additional cases . where available clinic pathologic characteristics are presented in table 1 . \n previous publications of false - positive fluorodeoxyglucose uptake in the left lobe of liver following radiotherapy with or without concurrent chemotherapy for gastroesophageal cancers our case reinforces the importance of a multidisciplinary approach to the care of patients with esophageal and gastric cancers to diagnose and manage complex cases . \n radiologists who suspect false - fdg uptake should correlate pet - ct findings with radiation treatment fields . \n the radiation oncologist can perform image fusion of the ct portion of the pet - ct scan with the ct planning scan used for radiation treatment planning , thus allowing for co - registration of pet images with radiation isodose lines to determine dose to the area of pet uptake . if there is continued concern for malignancy , a follow - up liver mri with and without gadolinium contrast would be warranted . however , \n if the multidisciplinary team is still concerned for malignancy , then a ct - guided biopsy is warranted . \n notably , both our patients had her2-amplified disease , a feature not reported most prior series but shared with paudel et al . to our knowledge , there is no preclinical or clinical evidence that her2 amplification is associated with increased radiation risk . \n however , trastuzumab has been affiliated with radiation recall in breast cancer patients receiving whole breast radiotherapy . \n larger studies are needed to confirm the possible associated between her2-amplified gej tumors and hepatic fdg - uptake . \n the ongoing radiation therapy oncology group 1010 trial of neoadjuvant chemoradiation with trastuzumab may clarify our findings . \n determination of whether radiographic findings represent true disease progression or not depends importantly on clinical considerations . \n although metastatic disease to the liver is common in gej cancers , the clinical features supported false - positive hepatic fdg - uptake in both our patients . with other imaging modalities such as mri , the confidence in short interval imaging over invasive procedure can be further increased . \n the follow - up mri in our patient failed to demonstrate any focal increased t2-signal intensity as may be expected from a liver metastasis . if clinical suspicion remains high , our data will support repeat pet - ct to provide a functional assessment 3 months after the incident imaging . with the rising incidence of distal esophageal adenocarcinomas and increasing use of pet - ct and neoadjuvant therapy , \n an appreciation of false - positive hepatic uptake is important to both radiologists , radiation oncologists , and medical oncologists . \n \n ", "role": "user" }, { "content": "patients with locally advanced gastroesophageal cancers frequently undergo concurrent chemotherapy and radiation ( crt ) . 18-fluorodeoxyglucose - positron emission tomography ( 18fdg - pet ) in combination with computed tomography \n is used for disease staging and assessing response to therapy . \n 18fdg - pet interpretation is subject to confounding influences including infectious / inflammatory conditions , serum glucose , and concurrent medications . \n radiotherapy induces tissue damage , which may be associated with fdg - avidity ; however , few reports have described the focal areas of hepatic uptake following concurrent chemoradiation ( crt ) . \n distinguishing hepatic fdg uptake from disease progression represents an important clinical scenario . here , we present two cases of unexpected fdg uptake in the liver after crt and review the literature describing incidental liver uptake on fdg - pet .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: the currently accepted management of advanced stage ovarian cancer is primary debulking surgery ( pds ) followed by chemotherapy . \n many studies have shown that pds is associated with high mortality and morbidity rates . \n the morbidity is particularly higher in patients with extensive intraabdominal disease and poor general condition of the patients . \n it is noteworthy to mention that most patients who presents to our institution are poor , nutritionally deprived and frail and hence can not tolerate the stress of prolonged surgery . \n several studies have discussed the benefit of neoadjuvant chemotherapy ( nact ) in patients with advanced ovarian cancer . \n interval debulking surgery ( ids ) is the standard of care at our center for patients with advanced epithelial ovarian cancer . in the present study , we have presented the survival statistics of 104 patients who have been managed with ids from january 2007 to december 2009 . \n this was a retrospective study conducted in dr b borooah cancer institute , which caters to the entire cancer patient population of north eastern india . \n as reported in the population based cancer registries 2009 - 2011 , the incidence of ovarian cancer was highest in the kamrup urban district where this hospital is situated . from january 2007 \n to december 2009 , 104 patients who underwent ids for stage iiic or iv advanced ovarian epithelial cancer were selected for the study . \n patients with poor nutritional status were routinely advised nutritional supplementation in consultation with a dietician . \n ids was attempted after three to five courses of chemotherapy with paclitaxal ( 175 mg / m ) and carboplatin ( 5 - 6 of area under curve ) . \n optimal cytoreduction was taken as macroscopic disease < 1 cm . diagnosis was based on previous histopathology report of a biopsy specimen . if a biopsy specimen was not available , fine needle aspiration cytology of the ascitic fluid , metastatic disease or tru - cut biopsy of metastatic lesion was used . \n tumor marker included ca 125 level . to rule out primary gastrointestinal ( gi ) tumors carcinoembryonic antigen ( cea ) , \n ca 125 to cea ratio , ca 19.9 , upper gi endoscopy and colonoscopy was done . \n routine blood tests included complete blood count , liver function test , blood urea and serum creatinine . \n radiologically contrast enhanced computerized tomography ( cect ) abdomen and pelvis was used for staging advanced disease . the response to nact \n standard surgical procedure involved peritoneal washings , total abdominal hysterectomy with bilateral salphingoophorectomy , supracolic omentectomy . \n formal retroperitoneal lymph node dissection was not performed in the patients and only nodal sampling of suspicious nodes was done . \n cases with deposits in the porta hepatis , dense tumor deposits in the pouch of douglas with infiltration into the rectal wall , extensive mesenteric deposits underwent suboptimal surgery . at the completion of adjuvant chemotherapy patients \n patients were kept on regular follow - up with history and clinical examination , ca 125 every 3 months and cect scan every 6 months . \n the primary end point of the study was overall survival ( os ) and secondary end point was progression free survival ( pfs ) . \n survival was measured in months and estimated by means of the kaplan - meier method . \n the association between the explanatory variables ( age , menopausal status , stage at presentation , grade , size of tumor , degree of cytoreduction ) was assessed by cox - regression ( unifunctional ) and results were presented as hazard ratio and 95% confidence interval ( ci ) . \n they were statistically analyzed by ibm spss statistics 19 ( armonk , ny : ibm corp . ) . \n a total of 104 patients with advanced epithelial ovarian cancer who underwent ids between january 2007 and december 2009 were followed - up till december 2013 and pfs and os was determined . \n grade i hematological toxicity was observed in 28.84% of cases , grade ii in 5.7% of cases and grade iii in 1.92% of patients . \n grade i gi toxicity was noted in 19.23% patients and grade ii in 3.8% of cases . \n 19 patients had complete response ( 18.26% ) and 80 patients ( 76.92% ) had partial response to nact . \n ids was performed using a midline incision including : hysterectomy ( n = 102 ) , salpingo - oophorectomy ( n = 96 ) , appendicectomy ( n = 7 ) bowel resection anastomosis ( n = 6 ) , repair of bladder ( n = 4 ) , lymphadenectomy ( pelvic : n = 7 : paraaortic : n = 4 patients ) [ table 2 ] . baseline demography and clinical characteristics characteristics of debulking surgery postoperative complications were observed in 6 patients ( 5 had stitch line abscess requiring secondary suturing and 1 developed postoperative pneumonia . \n forty - three patients received blood transfusion due to preexisting anemia and not because of blood loss . \n the median time period between ids and adjuvant chemotherapy was 31.5 days ( range 21 - 45 days ) . \n on assessment of the survival the median os time was 26 months [ figure 1 ] ( ci : 24.47 - 27.52 ) and the median pfs was 18 months ( ci : 12.73 - 23.26 ) [ figure 2 ] . \n overall survival in months progression free survival in months in cox - regression analysis , it was found that both os and pfs were affected by the stage , and degree of debulking [ figures 3 and 4 ] . \n the median os for stage iiic cancer at the time of presentation was 26 months , whereas stage iv cancer it was 16 months ( p = 0.001 ) . \n the median pfs stage iiic cancer at the time of presentation was 18 months whereas stage iv cancer it was 4 months ( p = 0.002 ) . \n the os for patients undergoing optimal cytoreduction ( < 1 cm ) was 26 months , whereas it was 12 months for suboptimal cytoreduction ( p = 0.001 ) . \n the median pfs for optimal cytoreduction ( < 1 cm ) was 18 months , whereas it was 4 months for suboptimal cytoreduction ( p = 0.014 ) \n . overall survival in months ( stage ) overall survival in months ( degree of debulking ) \n ovarian cancer usually presents in advanced stages with an overall 5 years survival rate of < 40% even in developed countries . \n this adds up to the high prevalence of nutritional anemia prevalent in developing countries resulting in poor surgical performance status . the median hemoglobin level of our patients was 9.6 g / dl [ table 1 ] . \n even though pds has been considered the gold standard of advanced epithelial ovarian cancer , the serious morbidity associated with such surgeries as reported is as high as 43% . \n in addition , due to bulky disease in advanced ovarian cancer , optimal cytoreduction is possible in < 50% of the cases . \n it thus contributes to a decreased postoperative morbidity and hospital stay and improves the patient 's ability to tolerate postoperative adjuvant chemotherapy . out of 104 patients who had undergone interval debulking , \n this was consistent with the results of increased optimal cytoreduction rates in patients with interval debulking found by vergote et al . who found that optimal cytoreduction ( < 1 cm ) was higher ( 80.6% ) in patients undergoing interval cytoreduction as compared to primary surgery ( 41.6% ) . \n bowel resection and anastomosis was done in only 5.7% and repair of bladder in 3.84% . \n these findings were similar to the lower incidence of intraoperative morbidity in patients undergoing interval debulking in other studies . \n os and pfs in our study was 26 months and 18 months respectively similar to the survival in patients undergoing interval debulking in the study by vergote et al . \n os in our study was however low when it was compared to the study by kumar et al . who found an os of 41 months in patients undergoing ids . \n on the other hand pfs was found to be slightly better ( 18 vs. 15 months ) . \n this might be explained due to the initial response to nact , followed by greater degree of optimal cytoreduction , which reflected on the pfs . on the other hand , poor os \n as compared to the pfs might be due to the fact that most of the patients did not receive second line chemotherapy . \n this was either due to nonaffordability of the second line drugs or poor performance status of these patients at the time of relapse . \n the os and pfs in our study was found to be affected by degree of debulking , which was also found in other studies . \n we have also found that os and pfs was significantly less in patients who were stage iv at the time of presentation reflecting the poor long term prognosis of these patients in spite of initial response to chemotherapy . \n our study has shown that nact is a promising approach in indian patients with advanced ovarian cancer as a significant number of patients respond to it leading to increased optimal cytoreduction . \n this has resulted in comparable survival for patients without increased morbidity and greater percentage of patients being able to undergo optimal cytoreduction .", "role": "user" }, { "content": "context : patients with advanced ovarian cancer have a poor prognosis in spite of the best possible care . \n primary debulking surgery has been the standard of care in advanced ovarian cancer ; however , it is associated with high mortality and morbidity rates as shown in various studies . \n several studies have discussed the benefit of neoadjuvant chemotherapy in patients with advanced ovarian cancer.aims:this study aims to evaluate the survival statistics of the patients who have been managed with interval debulking surgery ( ids ) from january 2007 to december 2009.materials and methods : during the period from january 2007 to december 2009 , a retrospective analysis of 104 patients who underwent ids for stage iiic or iv advanced epithelial ovarian cancer at our institute were selected for the study . \n ids was attempted after three to five courses of chemotherapy with paclitaxal ( 175 mg / m2 ) and carboplatin ( 5 - 6 of area under curve ) . overall survival \n ( os ) and progression free survival ( pfs ) were compared with results of primary debulking study from existing literature . \n os and pfs rates were estimated by means of the kaplan - meier method . \n results were statistically analyzed by ibm spss statistics 19.results:the median os was 26 months and the median pfs was 18 months . in multivariate analysis it was found that both os and pfs was affected by the stage , and extent of debulking.conclusions:neoadjuvant chemotherapy , followed by surgical cytoreduction is a promising treatment strategy for the management of advanced epithelial ovarian cancers .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: cortisol samples from 13 pdicb , 15 pd+icb patients and 14 healthy controls were collected . \n all patients were recruited from the national hospital for neurology and neurosurgery , queen square , london , fulfilling the queen square brain bank criteria for pd18 and were taking l - dopa . \n pd+icb patients were diagnosed using proposed criteria.1921 most pd+icb patients had more than one icb . \n healthy controls were usually recruited from among the patient 's partners and were not taking any medication that could influence cortisol measurement . \n written informed consent was obtained from all subjects according to approved ethical protocols from the regional and local research ethics committee . \n we excluded participants who scored under 27/30 points on the mini - mental state examination and who had a current or past medical history of an anxiety disorder , and patients with current depression . \n patients without a known previous diagnosis of anxiety or depression were screened for these conditions in a semi - structured clinical assessment . \n furthermore , all participants were asked to complete the positive and negative affect schedule which has been recommended as a supplement to measure anxiety and depression.22 samples were obtained in a quiet environment , usually at the patient 's home , to control for and reduce the amount of stress . to control for the potential effects of food23 and l - dopa24 on cortisol levels , \n pd patients were asked not to take their usual anti - parkinson medication for at least 12 h and not to have breakfast on the day of testing . \n the morning samples were obtained together with the researcher and the other samples were collected by the participants themselves . \n participants were instructed to collect saliva samples by turning the cotton roll for 23 min in their mouth . in total , \n all patients were tested in their off condition in the morning , which was assessed by the unified parkinson 's disease rating scale ( updrs ) part 3 motor score . \n one was obtained immediately prior to the risk task , one just after the risk test and prior to medication , the next 5 min after medication , then 15 min after medication , 30 min after medication and 45 min after medication . \n sixty minutes after medication , between 9:15 and 9:45 and after the second test ( on condition in patients ) , another sample was taken , this time from both controls and patients ( figure 1 ) . schematic outline of the time course of saliva samples . \n ten samples were taken from parkinson 's disease ( pd ) patients prior to medication ( off ) and after medication ( on ) . \n all participants then collected samples between 13:00 and 14:00 , between 19:00 and 20:00 and between 22:00 and 23:00 on their own . \n these times were deliberately flexible for patients to ensure that the cortisol sampling was undertaken approximately 1 h after taking their usual dopamine replacement therapy . \n subjects were asked to avoid excessive physical activity , stress and heavy meals on the study day , and were provided with a collection diary where they entered the time of saliva collection and their activity during the hour prior to each cortisol sample . \n in addition , participants were instructed not to eat anything at least 30 min prior to collecting a sample . the therapeutic motor response to l - dopa \n was assessed by updrs scores ( part 3 ) during off and on states . \n all patients had an excellent l - dopa response and had a similar improvement in updrs scores . \n levodopa equivalent units ( leu ) was calculated as described previously21 ( see table 1 for demographic information ) . \n da , dopamine agonists ; icb , impulsive compulsive behaviour ; leu , l - dopa equivalent units ; pd , parkinson 's disease ; updrs , unified parkinson 's disease rating scale . \n all patients were tested on a validated computerised gambling task to assess risk behaviour15 once in their off medication state after their first salivary cortisol sampling , and once in their on medication state approximately 50 min after receiving their usual morning anti - parkinson medications . \n controls were tested in the same way but did not take dopaminergic medication between the tests . \n participants had to choose between two gambles with varying levels of risks either a low risk or a high risk gamble , where participants could win real money . \n details about this task are published elsewhere.17 risk aversion was assessed parametrically by fitting a model to each participant 's behaviour , separately for each test session.17 saliva samples for the determination of cortisol concentration were collected in salivettes ( sarstedt , leicester , uk ) and stored at 20c until analysed . \n details on the analysis were published recently elsewhere.25 data analyses were performed using spss v.18 . \n age , gender , age of onset , updrs scores , l - dopa and leu dose were used as dependent variables and group ( pdicb , pd+icb and control ) was modelled as a between subject factor . \n we used anova , the student 's t test or the test where appropriate . \n the data were positively skewed as cortisol levels show a diurnal variation with a peak during the morning ( figure 2 ) . as a result , we log transformed the data and residuals were checked and found to be normally distributed . \n we performed a linear mixed model anova with the log transformed scores as the dependent variable and group ( pdicb , pd+icb and control ) was modelled as a between subject factor . \n the model also included subject as a random factor , and the interactions between the two fixed factors ( time and group ) . \n all post hoc comparisons were corrected by the bonferroni method ; the level of significance was p<0.05 . \n for the diurnal cortisol measurement we excluded those extra samples that were obtained from the patients between the tests . \n ( b ) cortisol levels between parkinson 's disease patients without ( pdicb ) and with ( pd+icb ) impulsive compulsive behaviour . \n although not significantly different , baseline morning cortisol levels were lower in the pd+icb group . \n specifically , we calculated the correlation coefficient between : r and c , where r = r1r2 is the difference in risk aversion in the first and second test session ; c was defined accordingly for cortisol levels . \n one value for r and one for c was derived for each subject , and these values were then correlated using pearson 's correlation test . \n cortisol samples from 13 pdicb , 15 pd+icb patients and 14 healthy controls were collected . \n all patients were recruited from the national hospital for neurology and neurosurgery , queen square , london , fulfilling the queen square brain bank criteria for pd18 and were taking l - dopa . \n pd+icb patients were diagnosed using proposed criteria.1921 most pd+icb patients had more than one icb . \n healthy controls were usually recruited from among the patient 's partners and were not taking any medication that could influence cortisol measurement . \n written informed consent was obtained from all subjects according to approved ethical protocols from the regional and local research ethics committee . \n we excluded participants who scored under 27/30 points on the mini - mental state examination and who had a current or past medical history of an anxiety disorder , and patients with current depression . \n patients without a known previous diagnosis of anxiety or depression were screened for these conditions in a semi - structured clinical assessment . \n furthermore , all participants were asked to complete the positive and negative affect schedule which has been recommended as a supplement to measure anxiety and depression.22 samples were obtained in a quiet environment , usually at the patient 's home , to control for and reduce the amount of stress . to control for the potential effects of food23 and l - dopa24 on cortisol levels , \n pd patients were asked not to take their usual anti - parkinson medication for at least 12 h and not to have breakfast on the day of testing . \n the morning samples were obtained together with the researcher and the other samples were collected by the participants themselves . \n participants were instructed to collect saliva samples by turning the cotton roll for 23 min in their mouth . in total , \n all patients were tested in their off condition in the morning , which was assessed by the unified parkinson 's disease rating scale ( updrs ) part 3 motor score . \n one was obtained immediately prior to the risk task , one just after the risk test and prior to medication , the next 5 min after medication , then 15 min after medication , 30 min after medication and 45 min after medication . \n sixty minutes after medication , between 9:15 and 9:45 and after the second test ( on condition in patients ) , another sample was taken , this time from both controls and patients ( figure 1 ) . schematic outline of the time course of saliva samples . \n ten samples were taken from parkinson 's disease ( pd ) patients prior to medication ( off ) and after medication ( on ) . \n all participants then collected samples between 13:00 and 14:00 , between 19:00 and 20:00 and between 22:00 and 23:00 on their own . \n these times were deliberately flexible for patients to ensure that the cortisol sampling was undertaken approximately 1 h after taking their usual dopamine replacement therapy . \n subjects were asked to avoid excessive physical activity , stress and heavy meals on the study day , and were provided with a collection diary where they entered the time of saliva collection and their activity during the hour prior to each cortisol sample . \n in addition , participants were instructed not to eat anything at least 30 min prior to collecting a sample . the therapeutic motor response to l - dopa \n was assessed by updrs scores ( part 3 ) during off and on states . \n all patients had an excellent l - dopa response and had a similar improvement in updrs scores . \n levodopa equivalent units ( leu ) was calculated as described previously21 ( see table 1 for demographic information ) . \n da , dopamine agonists ; icb , impulsive compulsive behaviour ; leu , l - dopa equivalent units ; pd , parkinson 's disease ; updrs , unified parkinson 's disease rating scale . \n all patients were tested on a validated computerised gambling task to assess risk behaviour15 once in their off medication state after their first salivary cortisol sampling , and once in their on medication state approximately 50 min after receiving their usual morning anti - parkinson medications . \n controls were tested in the same way but did not take dopaminergic medication between the tests . \n participants had to choose between two gambles with varying levels of risks either a low risk or a high risk gamble , where participants could win real money . \n details about this task are published elsewhere.17 risk aversion was assessed parametrically by fitting a model to each participant 's behaviour , separately for each test session.17 \n saliva samples for the determination of cortisol concentration were collected in salivettes ( sarstedt , leicester , uk ) and stored at 20c until analysed . details on the analysis were published recently elsewhere.25 \n age , gender , age of onset , updrs scores , l - dopa and leu dose were used as dependent variables and group ( pdicb , pd+icb and control ) was modelled as a between subject factor . \n we used anova , the student 's t test or the test where appropriate . \n the data were positively skewed as cortisol levels show a diurnal variation with a peak during the morning ( figure 2 ) . as a result , we log transformed the data and residuals were checked and found to be normally distributed . \n we performed a linear mixed model anova with the log transformed scores as the dependent variable and group ( pdicb , pd+icb and control ) was modelled as a between subject factor . \n the model also included subject as a random factor , and the interactions between the two fixed factors ( time and group ) . \n all post hoc comparisons were corrected by the bonferroni method ; the level of significance was p<0.05 . for the diurnal cortisol measurement we excluded those extra samples that were obtained from the patients between the tests . \n ( b ) cortisol levels between parkinson 's disease patients without ( pdicb ) and with ( pd+icb ) impulsive compulsive behaviour . \n although not significantly different , baseline morning cortisol levels were lower in the pd+icb group . \n specifically , we calculated the correlation coefficient between : r and c , where r = r1r2 is the difference in risk aversion in the first and second test session ; c was defined accordingly for cortisol levels . \n one value for r and one for c was derived for each subject , and these values were then correlated using pearson 's correlation test . \n age , gender , age of onset , updrs scores , l - dopa and leu dose were used as dependent variables and group ( pdicb , pd+icb and control ) was modelled as a between subject factor . \n we used anova , the student 's t test or the test where appropriate . \n the data were positively skewed as cortisol levels show a diurnal variation with a peak during the morning ( figure 2 ) . as a result , we log transformed the data and residuals were checked and found to be normally distributed . \n we performed a linear mixed model anova with the log transformed scores as the dependent variable and group ( pdicb , pd+icb and control ) was modelled as a between subject factor . \n the model also included subject as a random factor , and the interactions between the two fixed factors ( time and group ) . \n all post hoc comparisons were corrected by the bonferroni method ; the level of significance was p<0.05 . \n for the diurnal cortisol measurement we excluded those extra samples that were obtained from the patients between the tests . \n ( b ) cortisol levels between parkinson 's disease patients without ( pdicb ) and with ( pd+icb ) impulsive compulsive behaviour . \n although not significantly different , baseline morning cortisol levels were lower in the pd+icb group . \n specifically , we calculated the correlation coefficient between : r and c , where r = r1r2 is the difference in risk aversion in the first and second test session ; c was defined accordingly for cortisol levels . \n one value for r and one for c was derived for each subject , and these values were then correlated using pearson 's correlation test . \n there was no significant age difference between the groups ( f(2,38)=2.5 , p=0.091 ) . however , pd+icb patients had an earlier disease onset ( t24=2.6 , p=0.014 ) . \n there was no difference in the daily l - dopa doses or the dose given in the morning , leu dose , updrs ( part 3 motor ) scores or disease duration ( see table 1 for further details ) . \n we found a main effect of group ( f(2,37)=4.6 , p=0.016 ) and a main effect of time ( f(4,144)=51.0 , p<0.001 ) , with the highest cortisol level being in the morning . \n post hoc analysis revealed that the pdicb group had significantly higher cortisol levels than the control group ( p=0.019 ) ( figure 3 ) . \n there was no difference between the control and pd+icb group ( p=0.1 ) and no difference between the patient groups ( p=1.0 ) . \n we next looked at just the morning cortisol levels between the patient groups by excluding the control group but found no significant difference ( t24=2.4 , p=0.2 ) . \n there was no correlation between updrs off score and baseline cortisol levels ( all p values > 0.4 ) . \n log scores of diurnal salivary cortisol levels in controls and in patients with parkinson 's disease without ( pdicb ) and with ( pd+icb ) impulsive compulsive behaviour . \n box plot showing the median ( horizontal line ) within a box containing the central 50% of the observations ( ie , the upper and lower limits of the box are the 75th and 25th percentiles ) . \n participants were tested twice on a behavioural task that assessed risk taking behaviour15 once off medication and once on medication . \n the task assessed the extent to which participants preferred large , low probability rewards to smaller more probable rewards . \n we have reported detailed analyses of the behavioural results previously , the main finding being that patients became more risk prone on medication , relative to controls.17 here we focus on the relationship between cortisol and behaviour in the task . in the pd+icb group \n , there was a significant correlation between change in risk from the first to the second test session ( from the off medication state to the on medication state ) and change in cortisol levels measured just before each test session ( r=0.617 , p=0.0144 , n=15 ) . \n specifically , increased risk preference was associated with increased cortisol levels . in the pdicb group , \n furthermore , the correlation coefficients were significantly different between the pdicb and the pd+icb groups ( z=1.99 , p=0.047 ) such that there was a significantly stronger correlation in the pd+icb group than in the pdicb group . \n there was no significant age difference between the groups ( f(2,38)=2.5 , p=0.091 ) . however , pd+icb patients had an earlier disease onset ( t24=2.6 , p=0.014 ) . \n there was no difference in the daily l - dopa doses or the dose given in the morning , leu dose , updrs ( part 3 motor ) scores or disease duration ( see table 1 for further details ) . \n we found a main effect of group ( f(2,37)=4.6 , p=0.016 ) and a main effect of time ( f(4,144)=51.0 , p<0.001 ) , with the highest cortisol level being in the morning . \n post hoc analysis revealed that the pdicb group had significantly higher cortisol levels than the control group ( p=0.019 ) ( figure 3 ) . \n there was no difference between the control and pd+icb group ( p=0.1 ) and no difference between the patient groups ( p=1.0 ) . \n we next looked at just the morning cortisol levels between the patient groups by excluding the control group but found no significant difference ( t24=2.4 , p=0.2 ) . \n there was no correlation between updrs off score and baseline cortisol levels ( all p values > 0.4 ) . \n log scores of diurnal salivary cortisol levels in controls and in patients with parkinson 's disease without ( pdicb ) and with ( pd+icb ) impulsive compulsive behaviour . \n box plot showing the median ( horizontal line ) within a box containing the central 50% of the observations ( ie , the upper and lower limits of the box are the 75th and 25th percentiles ) . \n participants were tested twice on a behavioural task that assessed risk taking behaviour15 once off medication and once on medication . \n the task assessed the extent to which participants preferred large , low probability rewards to smaller more probable rewards . \n we have reported detailed analyses of the behavioural results previously , the main finding being that patients became more risk prone on medication , relative to controls.17 here we focus on the relationship between cortisol and behaviour in the task . in the pd+icb group \n , there was a significant correlation between change in risk from the first to the second test session ( from the off medication state to the on medication state ) and change in cortisol levels measured just before each test session ( r=0.617 , p=0.0144 , n=15 ) . \n specifically , increased risk preference was associated with increased cortisol levels . in the pdicb group , \n furthermore , the correlation coefficients were significantly different between the pdicb and the pd+icb groups ( z=1.99 , p=0.047 ) such that there was a significantly stronger correlation in the pd+icb group than in the pdicb group . \n as expected , cortisol levels were found to be highest in the morning in all participants and decreased over the day ( figure 2 ) . \n furthermore , we found significantly higher daily salivary cortisol levels in pdicb patients compared with healthy controls but no difference between pd+icb patients and controls . increased irritability and lability , higher scores of disinhibition and novelty seeking and a previous history of addictive behaviours \n have been reported in pd+icb patients.12 26 furthermore , all pd+icb patients had higher schizotypy scores , which is a personality trait related to the risk for psychosis , than controls.27 in this study , it is not possible to determine whether changes in cortisol are a cause or effect of the impulsive compulsive behaviours . \n however , impulsiveness , carelessness and aggressive behaviour have been associated with attenuated cortisol levels in adolescents and adults.13 2830 impulsive adults with illicit drugs abuse,14 patients with antisocial behaviour31 32 and controls with reduced negative feedback learning33 also had lower cortisol levels . \n increased temporal discounting , the tendency to choose earlier , smaller over delayed , larger rewards has been found only in pd+icb patients27 and has been associated with attenuated cortisol levels in healthy volunteers.34 various studies have also shown impairment in feedback learning in pd+icb patients.17 35 36 however , we did not find any difference between the two patient groups and between the control and pd+icb groups . \n previous studies found sustained elevated cortisol levels in aborigines after receiving their wages which they planned to gamble with.37 therefore , the expectance of being tested and the subsequent modest monetary reward for completing the study could have led to an increase in baseline cortisol levels in pd+icb patients . \n another possible explanation is that pd itself is associated with increased cortisol levels5 while personality traits typical of pd+icb patients are linked with lower cortisol levels . \n this might explain why we did not find any difference between pd+icb patients and the two other groups . \n critically , we observed a change of direction of cortisol levels only in the pd+icb group during gambling . this change of direction in cortisol levels following a stressor has been linked with antisocial behaviour.30 we have reported previously that all pd patients were more risk prone on medication compared with controls , with a subgroup of pd+icb with pathological gambling taking the most risky decisions.17 when correlating changes in cortisol levels with risk taking behaviour , we found a significant interaction in the pd+icb group but not in the pdicb group , despite both groups showing similar performance on the risk task . \n our findings are consistent with previous studies in male non - pd gamblers showing a rise in salivary cortisol levels16 38 and blood cortisol levels39 during gambling . \n acutely raised cortisol has been linked with anticipation of increased chances of making money and can be euphorogenic.40 41 alcohol and nicotine induce an increase in cortisol levels8 42 and addicts have an increased activation of the hypothalamic pituitary \n axis during drug intake.8 several preclinical studies have also shown that cortisol acts as a positive reinforcer and causes addiction.43 self - administration of cocaine leads to elevated cortisol levels in rodents,9 non - human primates44 and humans.45 our findings , with a relative increase in cortisol in pd+icb during a risk task , further strengthen the links between biological and drug addictions . \n this finding in a pd population is especially interesting since l - dopa has a dual effect on cortisol and behaviour . \n it increases risk behaviour in all pd patients17 and high doses of l - dopa can reduce cortisol levels.24 the latter finding contrasts with our results but in this study by muller et al , a control group was not included , morning samples were obtained at a time of the day when cortisol levels decrease fastest and no information on circadian cortisol levels was provided . to our knowledge \n our results suggest that in general , cortisol levels are elevated in pdicb patients compared with controls but not in the pd+icb group . \n this is in keeping with the literature which links lower cortisol levels with antisocial behaviour , and further links icbs with substance addiction . \n additionally , we found a significant correlation between risk taking behaviour and cortisol levels in the pd+icb group with higher cortisol levels being associated with risk prone behaviour but no significant interaction in the pdicb group .", "role": "user" }, { "content": "objectiveto investigate salivary cortisol samples in patients with parkinson 's disease ( pd ) with and without impulsive compulsive behaviours ( icb ) during a risk task.methodssalivary cortisol levels were measured in 13 pd patients without icb ( pdicb ) and in 15 pd patients with icb ( pd+icb ) before , after medication and throughout the day , and were compared with results with 14 healthy controls . \n all participants also performed a gambling task to assess risk taking behaviour.resultssignificantly higher diurnal cortisol levels were found in the pdicb group compared with healthy controls but no differences were seen between the pd+icb and the control group . \n increased cortisol levels were significantly correlated with increased risk taking in pd+icb patients but no interaction was found in the pdicb group.conclusionsthe findings are in keeping with previous studies which have linked low cortisol levels with antisocial behaviour . \n the higher cortisol levels during the risk task in the pd+icb group are consistent with reports in pathological gamblers during gambling and addicts during drug abuse . \n the results support the hypothesis that cortisol plays an important role in risk taking in icbs .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: timing of initiation of renal replacement therapy ( rrt ) is currently one of the major unresolved topics in critical care nephrology and this is addressed through different approaches in a previous issue of critical care . \n thakar and colleagues performed an international survey predominantly among north american nephrologists consulting in intensive care . \n they questioned practice patterns for the initiation of rrt using three case scenarios representing patients with increased severity of disease and , by implication , a higher probability of mortality . \n the majority of the 172 respondents ( 70% usa ) expressed a reticence in commencing rrt early given the lack of evidence , preferring to base their decision on absolute levels of creatinine or blood urea nitrogen ( bun ) ( > 442 mol / l ( > 5 mg / dl ) , > \n 35.6 mmol / l ( > 100 mg / dl ) , respectively ) rather than any relative rise . despite this , 94% of physicians reported that they would be likely to start dialysis early in patients with the highest disease burden with early rrt described as a lower bun on commencing treatment . thus , the proportion of physicians starting dialysis at a bun < 75 mg / dl tripled in the more severe case . \n finally , given a selection of five parameters ( bun , creatinine , urine output , oxygenation and potassium ) for starting rrt , the latter two were given the highest priority with oxygen saturation appearing as the most frequent trigger in severe cases . \n however , as in most studies of this nature , cohort selection is of great importance . \n the responses were substantially influenced by current practice in the us underlined by the significantly higher inclination towards early initiation of rrt in physicians practicing outside the us . \n these results also differ to a recently published survey among 275 , mainly european intensivists . \n whereas the median thresholds with regard to serum creatinine ( 300 mol / l ) or urea ( 40 mmol / l ) were similar , there was a higher priority attached to hyperkalemia , metabolic acidosis and volume overload . \n additionally , they demonstrated a trend towards early initiation of rrt , with the majority favouring initiation when a diagnosis of acute kidney injury ( aki ) was made based on akin ( acute kidney injury network)/rifle ( risk , injury , failure , loss , end - stage renal disease ) criteria , particularly with regard to oliguria . \n thus , early treatment was defined by starting rrt within 6 hours after aki diagnosis according to the akin / rifle criteria . \n the perceived importance of oliguria as a trigger for commencing rrt is supported by other studies . \n for example , the international best kidney study group identified oliguria / anuria as the most frequent reason to start rrt and similar results were seen in the renal trial where oliguria was the indication to start in 60% of all patients . \n a completely different approach towards initiation of treatment was chosen by clec'h and co - workers investigating the effect of rrt on hospital survival in the french longitudinal prospective multicenter outcomerea database . \n they addressed the question as to whether rrt confers a survival benefit in critically ill patients through sophisticated propensity analyses . using two different propensity scores the answer was a resounding ' no ' . \n of course not : rrt definitely does one thing - it clears toxins and re - establishes electrolyte , acid - base and fluid homeostasis . \n therefore , the maximum one could expect from provision of rrt would be hospital mortality equal to patients not requiring rrt . \n however , the occurrence of aki reflects a higher severity of disease and is independently associated with increased mortality [ 6 - 8 ] . therefore , observed outcome will always be worse , regardless of how efficiently we replace the failing kidneys . \n this is clearly supported by the significantly higher modified severity of organ failure score ( msofa ) and simplified acute physiology score ( saps ) ii scores found for patients treated with rrt in both the crude comparison and the two propensity models . \n interestingly , one of the main conclusions was to commence rrt earlier based on the finding that delayed rrt , that is , starting 48 hours after reaching maximum rifle , was associated with a two - fold increased odds ratio for mortality compared to early or immediate rrt . \n however , only 12% of all patients treated with rrt fell into the ' late ' category and the median duration from onset of aki to start of rrt was only one day in this cohort . \n so , although the data suggest that ' very late rrt ' will be probably detrimental , no reliable conclusions can be drawn regarding the benefit of early or very early initiation of rrt from this study . lower stages of aki are associated with a lower mortality ; thus , it could be argued that one should not wait until the patient has reached a higher level of aki before commencing treatment . \n this is compounded by the fact that the maximum aki stage remains a retrospective diagnosis and can not be reliably predicted . nor can patients who will progress to a stage where rrt will be absolutely necessary or those who will recover without the need for rrt . \n in fact , three smaller randomized controlled trials showed either no benefit with early rrt or even an increased mortality compared to standard medical treatment in severe sepsis . in trying to determine which parameter might be the most relevant for starting rrt , clec'h and colleagues found only oliguria ( 0.4 l versus 1.3 l ) and fluid accumulation ( 4 l versus 2 l ) as significant for patients treated with rrt when performing a matched pair analysis . \n this again may reflect changes in practice in terms of avoiding a positive fluid balance . \n the syndrome of aki continues to gain importance ; the numbers of aki patients requiring rrt have roughly doubled over the past 10 years in the usa , which contribute to increased mortality and an increased burden on health care systems . \n several surveys and clinical studies reveal significant variations in treatment patterns as well as outcomes , largely dependent on regional or national habits as well as the primary medical speciality delivering rrt in critically ill patients . \n the practice of using fixed thresholds of creatinine or urea for timing of rrt may be considered a relic from haemodialysis programmes where creatinine is taken as a surrogate for residual function . in critically ill patients with aki , urea and creatinine are not reliable indicators of renal function given the lack of steady state in terms of production and the influence of catabolism , volume status and production rates , particularly in sepsis . \n consequently , physicians treating critically ill patients put an increasing emphasis on volume overload , oliguria , impaired oxygenation and acidosis as triggers for initiation of rrt with a general trend to commence rrt earlier in sicker patients . despite some progress achieved by recent recommendations , \n probably the conventional criteria discussed above are not sufficient to discriminate between early and late initiation of rrt and including additional biomarkers indicating renal damage may be required for that . \n this , however , can only be answered by a future sufficiently powered prospective randomized controlled trial . \n aki : acute kidney injury ; akin : acute kidney injury network ; bun : blood urea nitrogen ; rifle : risk , injury , failure , loss , end - stage renal disease ; rrt : renal replacement therapy . \n \n ", "role": "user" }, { "content": "when to commence renal replacement therapy ( rrt ) in the critically ill remains an unresolved issue . \n the study by thakar and colleagues sheds some light on current practice through an international survey , demonstrating physicians ' inclination to start rrt earlier when the severity of disease is higher . \n however , clec'h and co - workers investigated the effect of rrt on hospital survival by performing a propensity analysis on the large multicentre french outcomerea database . \n they demonstrate that rrt does not confer survival benefit , with a delay in initiation being proposed as a contributing factor .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: the study of cannabinoids in the central nervous system ( cns ) has been the primary focus of many investigations , not only due to their psychotropic effects , but also due to their involvement in neuroplasticity and their therapeutic use in conditions such as obesity , pain , and neurodegenerative diseases . \n fewer studies have focused on the role of cannabinoids in the retina , as well as other critical players of the visual system [ 1 , 2 ] . \n the functional significance of the cannabinoids in ocular physiology and disease was noted by hepler and frank who reported that subjects who smoked marijuana ( cannabis sativa ) developed lower intraocular pressure ( iop ) . \n this finding was subsequently reproduced and the results suggested that cannabinoids isolated from cannabis may be useful in the treatment of glaucoma [ 46 ] . \n the presence of a functional endocannabinoid system in the retina , which includes ( a ) endogenous cannabinoids , ( b ) enzymes involved in their synthesis and metabolism , and ( c ) cannabinoid receptors , supports a role for cannabinoids in retinal circuitry and vision . \n arachidonoyl ethanolamide ( anandamide , aea ) and 2-arachidonoylglycerol ( 2-ag ) have been found in retinas of many species [ 710 ] . \n two receptors have been cloned , cb1 and cb2 , that mediate the physiological and pharmacological actions of endocannabinoids , as well as the actions of the natural and synthetic cannabinoids [ 1115 ] . \n cb1 receptors are predominant in the cns and are expressed in brain areas that influence motor and cognitive functions [ 1 , 2 ] , as well as in areas that comprise the visual pathway of the brain , namely , the thalamus and visual cortex [ 1618 ] . \n cb2 receptors are also found in the cns , but their location and function are mainly in peripheral tissues and the immune system [ 19 , 20 ] . \n the aim of this review is to summarize the knowledge acquired to date on the function of endogenous and synthetic cannabinoids in the retina and to address the neuroprotective and therapeutic potential of cannabinoids in retinal disease . \n immunohistochemical studies described the presence and localization of the cb1 receptor in retinal neurons of several animal species . \n specifically , cb1 immunoreactivity was detected in the inner and outer plexiform layers ( and/or on cone pedicles and rod spherules ) of tiger salamander , gold fish , chick , mouse , rat , and rhesus monkey , in amacrine and retinal ganglion cells ( rgcs ) and ganglion cell axons of all species except gold fish , and sparsely labeled photoreceptors of monkey , mouse , rat , and chicken . \n cb1 immunoreactivity has been detected in rod bipolar cells , gaba - amacrine cells , and horizontal cells . in the mouse retina \n , a specific cannabinoid circuitry has been defined based on colocalization of the cb1 receptor and a repertoire of proteins that influence the endocannabinoid system . \n the expression of cb2 cannabinoid receptor mrna has been reported in adult rat retina , as well as in human retinal pigment epithelial cells ( mrna and protein ) [ 23 , 24 ] . \n immunohistochemical studies supported the presence of cb2 receptors in retinal tissue of rats and mice [ 25 , 26 ] . specifically , the cb2 receptor was localized in the retinal pigment epithelium , inner photoreceptor segments , horizontal and amacrine cells , ganglion cell layer ( gcl ) , and inner plexiform layer ( ipl ) of the rat retina and in all five neuronal types of the mouse retina . \n the presence of cb2 receptors in mller cells of monkeys has also been reported . however , different groups , employing radioligand binding studies in rat retinal membranes , suggested the absence or low undetectable levels of cb2 receptor in rat retina [ 28 , 29 ] . \n thus , further studies are needed in order to elucidate the presence and functional role of cb2 receptors in the rodent retina . \n two other receptor systems were shown to be activated by cannabinoids , namely , the transient receptor potential cation channel subfamily v member 1 ( trpv1 channel ) and the orphan receptor gpr55 [ 3034 ] . \n immunohistochemical studies have shown the expression of trpv1 channel in ipl , inl ( inner nuclear layer ) , and gcl of adult rat retina , while gpr55 has been detected in rod photoreceptors of adult vervet monkey retina [ 33 , 34 ] . \n the colocalization of gpr55 only with the inner segments of rod photoreceptors , with less immunoreactivity observed in peripheral retina regions , suggested that this asymmetric distribution of gpr55 in the monkey retina may underlie its function in phototransduction . \n the most well - known signal transduction response for both cb1 and cb2 receptors is the inhibition of adenylyl cyclase through interaction with gi / o proteins [ 12 , 35 ] . \n activation of cb1 receptors affects g - protein - coupled inwardly rectifying potassium channels ( girks ) [ 36 , 37 ] . \n cb1 receptors interact with a variety of ion channels including ca and k channels [ 3739 ] . \n aea and 2-ag , as well as the enzymes responsible for their synthesis ( n - acyl phosphatidylethanolamine phospholipase , d - nape - pld , diacylglycerol lipase , dagl ) and degradation [ fatty acid amide hydrolase ( faah ) , monoacylglycerol lipase ( mgl ) ] , have been found in the retina of rodents and other mammals [ 8 , 9 , 22 ] . \n immunohistochemical localization of dagl has been reported in retinal tissue , especially in the postsynaptic terminals of cone bipolar cells . yet despite the reported presence of nape - pld in rat retina little is known about its specific localization in rat retinal neurons . \n faah , the metabolic enzyme of aea , has been shown to be localized in photoreceptor inner segments , horizontal , dopaminergic , and cholinergic amacrine and ganglion cells , as well as mller cells , in the rodent retina [ 21 , 22 ] . \n mgl , the metabolic enzyme of 2-ag , has also been reported in retinal tissue and specifically in the ipl and opl ( outer plexiform layer ) . as mentioned above \n , cb1 receptors can interact with a variety of ion channels [ 3739 ] . in the retina \n cb1 receptor activation results in pertussis toxin sensitive actions ( gi / o - linked cb1 receptor ) , such as the modulation of ion channel function . \n this cannabinoid function may influence retinal circuitry , neurotransmitter release , and neuroprotection . in retina \n , cannabinoids have been reported to modulate ca and/or k channels in bipolar cells and photoreceptors [ 10 , 4446 ] . \n the presence of cb1 receptors in rod bipolar cells and the cb1-dependent reduction in the amplitude of voltage - gated l - type calcium channel currents present in these retinal neurons suggest that endocannabinoids and synthetic cannabinoids may play an important role in retinal circuitry and in scotopic vision . \n glutamate is the major neurotransmitter in the retina released from photoreceptors , rod bipolar and ganglion cells . \n the presence of the cb1 receptor in all retinal neurons suggests that endocannabinoids and synthetic cannabinoids play a neuromodulatory role in retinal circuitry by influencing the release of other neurotransmitters . \n cb1 receptor agonists inhibited k induced [ h]-d - aspartate release from bovine retina , and ca evoked [ h]-noradrenaline and [ h]-dopamine release in guinea pig retina [ 4749 ] . \n cb1 regulation of gaba release was reported by spontaneous mini frequencies involving gaba - a receptor - mediated inward currents , in cultured chick embryonic amacrine cells . \n the authors of this study concluded that the regulation of spontaneous transmitter release by endocannabinoids might be important in network maintenance in amacrine cells and other inhibitory interneurons . \n in rat retinal explants hu-210 attenuated the release of the inhibitory neuropeptide somatostatin in a dose - dependent bimodal manner via the activation of the cb1 receptor ( figure 1 ) . \n bimodal modulation was also reported in a study that showed that win55212 - 2 affected voltage - dependent currents of retinal cones in a biphasic manner . \n the gpr55 receptor and the trpv1 channel are also activated by cannabinoids , such as the endogenous cannabinoid aea , the synthetic cannabinoid cp55940 , and the nonpsychotropic cannabinoid cbd [ 3034 ] . \n recently , the presence of gpr55 receptor protein in rod photoreceptor cells of the vervet monkey was reported and the authors suggested that gpr55 activation in these cells may play a role in scotopic vision . \n however , more studies are needed in order to evaluate the role of gpr55 receptor in retinal circuitry . \n the trpv1 channel has been recently suggested to be involved in rgc function and survival in the retina . \n further studies regarding the neuroprotective actions of the trpv1 channel in retina , upon its activation by the cannabinoids , are presented in section 3.2 . \n glaucoma is the second cause of blindness worldwide and is characterized by elevation of iop . \n the main treatment of the disease to date involves the administration of drugs that reduce iop by affecting either aqueous humour synthesis or outflow ( cholinomimetics or cholinesterase inhibitors , alpha adrenergic receptor agonists , beta adrenergic receptor blockers , and others ) . \n the cb1 receptor was found to be localized in the anterior segment of the human and rat eye , including the ciliary body , epithelium , and the trabecular meshwork [ 53 , 54 ] . \n mcintosh et al . reported that endogenous cb1 receptors couple with both gq/11 and gi / o , present in the trabecular meshwork , and elicit different responses according to the cannabinoid agonist employed in the study . \n activation of cb1 receptors by win55,212 - 2 , but not by cp55,940 or methanandamide , led to cb1 receptor coupling with gq/11 and subsequent activation of plc - dependent increase of intracellular ca levels . \n this study suggested that the differential effects of cannabinoid agonists in human trabecular meshwork cells may be used to identify cannabinoids that affect aqueous outflow and iop . \n these pharmacological actions of cannabinoids render them useful therapeutic targets in lowering iop in glaucoma patients . however , the reduction of iop is not sufficient to prevent visual field loss . \n the pathophysiology of glaucoma is multifactorial and recently more and more studies define glaucoma as a neurodegenerative disease , characterized by ischemia - induced excitotoxicity and loss of rgcs [ 5659 ] . \n elevated glutamate levels have been detected in the vitreous of humans and monkeys with glaucoma and glutamate - induced excitotoxicity has been suggested to play a fundamental role in the rgcs loss observed in experimental glaucoma . \n while being defined by its microvascular characteristics ( neovascularization ) , more recent reports suggest that it possesses neurodegenerative and inflammatory components . \n in fact , it has been suggested that retinal function is compromised before the appearance of neovascularization in diabetic patients [ 6365 ] . \n oxidative stress has also been documented in both animals and diabetic patients and has been correlated with neuronal loss [ 6668 ] . \n many strategies have been used to develop therapeutic agents for the successful treatment of ischemia - induced retinopathies and the prevention of blindness . \n the proneovascular agent vascular endothelium growth factor ( vegf ) is considered to be the factor most responsible for the development of new vessels in diseases , such as proliferative dr . \n the discovery of anti - vegf therapy has given new hope to patients , yet this treatment targets only the neovascular component of ischemia - induced retinopathies and has serious ocular adverse effects [ 7072 ] . \n however , there are no therapeutics available to date to treat the neurodegenerative component of diabetic retinopathy or glaucoma ( for a review , see ) . in order to preserve vision \n , all three components of ischemia - induced retinopathies , neovascularization , neurodegeneration , and inflammation , must be addressed . \n different animal disease models have been employed to examine the neuroprotective effects of putative therapeutics to treat neurodegenerative retinal diseases . \n these include ex vivo and in vivo models of ischemia and excitotoxicity , such as the ex vivo chemical ischemia model , the ex vivo and in vivo nmda ( n - methyl - d - aspartate ) or ampa ( -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid ) models of excitotoxicity , and the in vivo animal models of iop - reperfusion ( glaucoma ) , and the streptozotocin ( stz ) model of diabetic retinopathy . \n the ex vivo model of chemical ischemia involves the blockade of oxidative phosphorylation and glycolysis and is believed to be useful for the understanding of the early events underlying the pathophysiology of ischemia . \n it was previously reported that chemical ischemia influences the viability of a variety of amacrine and rod bipolar cells , but not rgc or photoreceptor viability . \n hu210 afforded neuroprotection to cholinergic amacrine and rod bipolar cells via activation of the cb1 receptor ( figures 2 and 3(a ) ) . \n ischemic insults lead to activation of voltage gated calcium channels and increase in glutamate levels and activation of ionotropic glutamate ( nmda and ampa ) receptors leading to an excess of intracellular calcium ions . \n nmda has been the excitatory amino acid of choice as a model of ischemia - induced cell death in brain and retina . \n -tetrahydroxycannabinol ( -thc ; partly via a cb1 mechanism ) and the non - cb1 agonist cbd were shown to protect the retina from nmda excitotoxicity . \n both cannabinoids afforded neuroprotection to retinal neurons located in the inl and gcl via attenuation of lipid peroxidation and/or nitrotyrosine formation . \n intravitreal administration of ampa in rat retina was shown to affect the viability of horizontal and bnos , chat , and calbindin - expressing amacrine cells , but not photoreceptors , bipolar or ganglion cells . \n this model has been employed to investigate the neuroprotective properties of new pharmacological agents in the early events of retinal ischemia in vivo [ 7779 ] . in this paradigm of excitotoxicity , \n aea and the synthetic cannabinoids hu-210 and methaea ( nonhydrolysable analogue of aea ) , intravitreally coinjected with ampa , provided neuroprotection to bnos and chat expressing amacrine cells . \n this study suggested that the cb1 receptor is responsible for the neuroprotective actions of the cannabinoids . \n in addition , aea afforded neuroprotection to horizontal and gaba amacrine calbindin - immunoreactive cells . \n the involvement of cb1 receptor in the neuroprotective actions of the endocannabinoid 2-ag has also been investigated in the ampa excitotoxicity model . \n 2-ag was intravitreally coinjected with ampa in wild - type , cb1 and cb2 c57bl/6 knockout mice . \n 2-ag reversed the ampa induced reduction of bnos expressing amacrine cells , in both wild - type and cb2 mice . \n these results substantiated the involvement of the cb1 receptor in the neuroprotective actions of 2-ag against ampa excitotoxicity in mouse retina . \n cannabinoids are known to activate prosurvival downstream signaling pathways in different paradigms which involve , among others , the prosurvival pi3k / akt and the mek / erk1/2 signaling pathways [ 8185 ] . in rat retina , \n the pi3k / akt signaling pathway was shown to be involved in the neuroprotective actions of hu-210 and aea in the ampa excitotoxicity model , whereas the mek / erk1/2 signaling pathway seemed to be involved in the neuroprotective actions of aea , but not hu-210 . \n . also presented similar results reporting that hu-210 acted in a neuroprotective manner against ampa excitotoxicity in primary cortical neuronal cultures by leading to the phosphorylation of akt , but not erk1/2 kinases . \n moreover , methaea has also been shown to induce erk1/2 phosphorylation in the hippocampus via a cb1-dependent mechanism . in \n the in vivo iop - reperfusion model of glaucoma , retinal ischemia - reperfusion led to an attenuation of aea levels and this was shown to be due to the enhanced activity of its metabolic enzyme faah . in this paradigm , \n administration of the faah inhibitor urb597 diminished the retinal damage , while intravitreal injection of methaea rescued rgcs via activation of cb1 receptors and trpv1 channels . \n this study supports that the endocannabinoid system may be involved in the loss of rgcs and that cannabinergic agents ( cb1 or trpv1 agonists and inhibitors of endocannabinoid metabolic enzymes ) may be important therapeutics in glaucoma . in the stz animal model of diabetic retinopathy , \n cbd treatment provided neuroprotection , blood brain barrier preservation , and anti - inflammatory actions . \n specifically , administration of cbd led to attenuation of proinflammatory cytokines , tumor necrosis factor ( tnf ) and vegf , reduction of oxidative and nitrative stress , and neuroprotection to inner retinal neurons . \n further support for the neuroprotection afforded by the synthetic cannabinoid hu210 to retinal neurons was shown in a study by lax et al . . \n these investigators performed immunohistochemical and electrophysiological studies and showed that hu210 afforded neuroprotection to photoreceptor cells in a model of retinitis pigmentosa . \n intraperitoneal administration of hu-210 improved the disrupted connectivity of photoreceptor cells with horizontal and bipolar cells and preserved the morphology and function of photoreceptors . \n previous studies suggested that the trpv1 ligand capsaicin induces degeneration of rgcs in preweanling rats and apoptosis in isolated rgcs via a ca - dependent mechanism . \n however , capsaicin has been shown to protect rgcs in an in vivo model of nmda retinal excitotoxicity in rats , via trpv1 channel activation . \n intravitreal administration of methaea via activation of the trpv1 channel rescued rgcs from ischemia - reperfusion insults . \n the involvement of the trpv1 channel in the neuroprotective properties of aea , 2-ag , and hu-210 was also examined in the ampa excitotoxicity model . \n the trpv1 antagonist capsazepine intravitreally administered with the above agents attenuated their neuroprotective actions on the bnos - immunoreactive amacrine cells , suggesting a neuroprotective role for the trpv1 channel in this paradigm ( figures 4(a)4(c ) ) . \n however , functional studies employing the trpv1 agonist capsaicin did not show any neuroprotective effects at any of the doses used ( figure 4(d ) ) . \n these results suggested that the trpv1 channel is not localized in bnos amacrine cells and that capsazepine 's actions are independent of the trpv1 channel . \n the results presented in this part of the review provide strong evidence for the neuroprotective role of the endocannabinoids and the synthetic cannabinoids in different models of retinal disease via the activation primarily of the cb1 receptor and its signal transduction mechanisms . in section 3.3 the interaction of the endocannabinoid system with other neuromodulatory systems in the retina \n will be presented in order to assess the influence of this interaction on the cannabinoid induced neuroprotection . \n the cb1 receptor has been localized in photoreceptors , rod bipolar , a variety of amacrine , horizontal , and ganglion cells in retinas of many species . \n this was assessed by immunohistochemical studies [ 10 , 21 ] and also confirmed from the functional neuroprotective roles of the endogenous and synthetic cannabinoids in the different models of retinopathy presented in this review . \n the localization of the cb1 receptor in each retinal neuron provided in many cases information on the mechanism involved in the neuroprotection of the agents employed . \n the presence of the cb1 receptor in rod bipolar cells suggests that its activation may influence glutamate neurotransmission . \n win55,212 - 2 was shown to inhibit l - type calcium currents in salamander bipolar cells . \n these channels regulate glutamate release and their inhibition will modulate in a negative manner glutamatergic transmission between the bipolar , amacrine , and ganglion cells . while release studies have not been performed on isolated rod bipolar cells , cb1 agonists were shown to inhibit k and ischemia - induced [ h]-d - aspartate release from isolated bovine retinas , as mentioned above . \n a decrease in glutamate and intracellular calcium ion levels will lead to neuroprotection , since the underlying cause of excitotoxicity induced cell death is the rise in intracellular calcium levels . \n a possible mechanism via which hu-210 afforded neuroprotection to rod bipolar cells in the model of chemical ischemia ( figure 2 ) may involve the direct activation of cb1 receptors and the subsequent inhibition of calcium and glutamate levels . \n a similar conjecture can not be made as to the mechanisms via which hu-210 provided neuroprotection to chat - immunoreactive amacrine cells . \n while the cb1 receptor has been localized in subpopulations of amacrine cells , its presence in chat expressing neurons , to our knowledge , has not been reported to date [ 10 , 21 ] . \n one can not exclude the presence of the cb1 receptor in cholinergic neurons , yet neuroprotection may also be afforded by cannabinoids via indirect mechanisms orchestrated by the modulatory actions of cannabinoids and other neurotransmitters in the retina , as will be presented in the following . \n inhibitory neurotransmitters , such as gaba , could counteract the toxic influence of glutamate on retinal neurons during retinal ischemia and would be expected to provide neuroprotection . \n a reduction of the excitatory input or an increase of the inhibitory input in the retina can afford neuroprotection . \n dopamine ( da ) is a major neuromodulator in the retina that affects retinal circuitry by activating two major families of dopamine receptors , d1 and d2 [ 93 , 94 ] . \n cb1 and the dopamine inhibitory d2 receptors are found in cones , whereas cb1 and the dopamine stimulatory d1 receptors are found in on bipolar cells . \n a reciprocal inhibition of voltage - gated potassium currents ( ik(v ) ) by activation of cannabinoid cb1 and dopamine d1 receptors in on bipolar cells of goldfish retina was reported . \n these findings support an antagonistic interaction between the cannabinoid and dopamine signaling that may influence transmitter release . \n in addition , it was proposed that endocannabinoids function as a scotopic signal , interacting with dopamine to set retinal sensitivity . \n da also regulates the release of other retinal neuromodulators , such as no ( nitric oxide ) and the neuropeptide somatostatin . \n it enhances no release , while a reciprocal effect was also observed , namely , no attenuating da release [ 9597 ] . \n no , cgmp , and the no donor sin-1 stimulated the release of gaba in the retina . \n somatostatin was also shown to influence the release of both dopamine and no in the retina [ 99 , 100 ] . \n da , via d1 receptor activation , and no , via guanylyl cyclase and cgmp , influence somatostatin levels in the retina . \n these results suggested that the triad of neuromodulators , somatostatin - dopamine - no , have reciprocal interactions via which they regulate retinal circuitry . \n cb1 agonists were also shown to inhibit the release of retinal [ h ] dopamine . \n hu-210 regulated somatostatin release in a bimodal manner ( figure 1(a ) ) , in agreement with what was reported for cannabinoid effects on gaba release in the globus pallidus . \n therefore , it appears that the interactions amongst transmitter and neuromodulator systems in the retina not only regulate retinal circuitry , retinal sensitivity , and light adaptation but may also provide the neuroprotective mechanisms involved in reducing ischemia - dependent toxicity . \n it is possible that cannabinoid interactions with other retinal modulatory systems may lead to the indirect neuroprotection of retinal neurons . \n hu-210 was shown to provide neuroprotection and counteract the oxidative stress and cellular injuries observed in diabetic encephalopathy , a cns neuropathy and one of the most common complications of diabetes , through a cannabinoid receptor - independent mechanism . \n the aim of this review was to summarize the knowledge acquired to date on the function of endogenous and synthetic cannabinoids in the retina , the receptors they activate to mediate their actions , and the neuroprotective role of cannabinoids in retinal models of disease . \n the studies presented in this review support the involvement of the endocannabinoids , aea and 2-ag , and synthetic cannabinoids in the neuroprotection of the early and final events underlying the pathophysiology of retinal ischemia . \n it appears that these agents have the pharmacological profile to target the neurodegenerative / proapoptotic components of retinal disease . \n the available treatments for neurodegenerative retinal diseases have limitations and further research is needed in order to provide efficacious therapies . \n cannabinergic agents , cb1 or trpv1 agonists and inhibitors of endocannabinoid metabolic enzymes , may prove to be efficacious therapeutics in the treatment of glaucoma and diabetic retinopathy . in order to provide \n cannabinergic agents , further studies are essential ( a ) to study the pharmacokinetic properties of cannabinoids that would suggest their use as effective topical retinal therapies and ( b ) to ascertain their pharmacological efficacy in chronic use . these studies will assess whether cannabinergic agents can play an important role alone or in a multidrug treatment to provide efficacious therapy and improve the eye sight of millions of people worldwide who are afflicted with neurodegenerative retinal disease .", "role": "user" }, { "content": "the functional significance of cannabinoids in ocular physiology and disease has been reported some decades ago . in the early 1970s , subjects who smoked cannabis \n sativa developed lower intraocular pressure ( iop ) . \n this led to the isolation of phytocannabinoids from this plant and the study of their therapeutic effects in glaucoma . \n the main treatment of this disease to date involves the administration of drugs mediating either the decrease of aqueous humour synthesis or the increase of its outflow and thus reduces iop . \n however , the reduction of iop is not sufficient to prevent visual field loss . \n retinal diseases , such as glaucoma and diabetic retinopathy , have been defined as neurodegenerative diseases and characterized by ischemia - induced excitotoxicity and loss of retinal neurons . \n therefore , new therapeutic strategies must be applied in order to target retinal cell death , reduction of visual acuity , and blindness . \n the aim of the present review is to address the neuroprotective and therapeutic potential of cannabinoids in retinal disease .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: an immune stress inhibits reproductive functions in many animal species and humans [ 14 ] . \n most studies examined the impact of immune stress on reproductive system activity that used bacterial endotoxin lipopolysaccharide ( lps ) as a model of infection induced changes . \n lipopolysaccharide is a pathogenic membrane component of virtually all gram - negative bacteria and it is released from the surface of replicating and dying gram - negative bacteria into the circulation . \n bacterial endotoxin is thought to play a major role in the pathophysiology of septic shock . \n endotoxin stimulation of animal cells occurs through a signalling cascades with several proteins including cd14 protein , md-2 protein , and lps - binding protein ( lbp ) , a necessary component of corresponding lps receptor called toll - like receptor 4 ( tlr4 ) [ 6 , 7 ] . \n then , lps - lbp complex binds to the cd14 protein , which is necessary for the activation of tlr4 . \n cd14 , md-2 , and tlr4 as a whole make up the cellular lps specific receptor [ 8 , 9 ] . \n after activation by endotoxin , tlr4 transduces its inflammatory signal through complex intracellular pathways , leading to activation of transcription factors such as nuclear factor kappa - light - chain - enhancer of activated b cells ( nf-b ) , c - jun n - terminal kinase ( jnk ) , and protein kinases p38 or inducing cell apoptosis [ 10 , 11 ] . \n administration of lps inhibits tonic luteinizing hormone ( lh ) secretion in many species including rats , sheep , cattle , and nonhuman primates as well as delays or completely blocks the preovulatory lh surge . in our earlier studies carried out on anestrous ewes \n , we showed that suppressive effect of lps on gnrh / lh secretion occurs primarily at the level of hypothalamus , changing the gonadotropin - releasing hormone ( gnrh ) synthesis and release as well at the pituitary level by inhibiting release of lh from gonadotropic cells [ 15 , 16 ] . \n data from literature indicate the existence of many possible mechanisms mediating antireproductive action of immune stress . \n it was shown that the crucial role in the mediation of the inhibitory influence of inflammation on gnrh / lh secretion play the role of proinflammatory cytokines such as il-1 , il-6 , tnf- [ 17 , 18 ] . \n the results of our studies showed that il-1 is one of the most important factors in modulating the function of gnrh neurons in anestrous ewes during immune stress . \n however , cytokine dependent pathway is not only mechanism via an immune challenge that affects the reproduction processes in animals . the studies on ewes in anestrous period indicated the presence of tlr4 mrna in the hypothalamic structures such as the preoptic area ( poa ) , the anterior hypothalamic area ( aha ) , the medial basal hypothalamus ( mbh ) , the median eminence ( me ) , and in the anterior pituitary gland ( ap ) in control animals as well as after lps treatment . \n the expression of the gene encoding this receptor in ewes treated with lps was significantly higher than that determined in control animals . \n it is worth to note that tlr4 expression was found in the central nervous system ( cns ) not only in microglia cells , but even in neurons . \n this suggests that tlr4 and its ligand lps could be involved in inhibition of the reproductive function at the cns and directly influence gnrh / lh secretion at the hypothalamic or pituitary level . \n the question of the possible penetration of endotoxin from blood to the cerebrospinal fluid ( csf ) during immune stress and thus direct lps action at the brain level is still open because the results of in vivo studies are not equivocal . \n the study performed on rats showed that peripherally injected lps labelled with iodine 125 crossed the blood - brain barrier ( bbb ) . although the brain uptake of circulating lps was found to be low , it was measurable . on the other hand , \n singh and jiang suggested that lps modulates the functioning and permeability of the bbb but does not cross it . \n the research hypothesis assumes that lps given peripherally ( intravenously iv ) can directly affect the hypothalamic - pituitary - gonadal axis ( hpg ) at the cns level that was verified . \n the aim of our experiments was to study the influence of the intracerebroventricular ( icv ) injection of anti - lps antibody or blockade of tlr4 receptor during immune stress induced by iv lps administration on the gnrh / lh secretion and tlr4 gene expression in hypothalamus and pituitary of anestrous ewes . \n the studies were conducted on adult , 3-year - old polish longwool ewes in the anestrous season ( april - may ) . \n all animals were in good condition , and their body condition score was estimated at 3 points ( on a five - point scale ) . \n the ewes were well adapted to the experimental conditions ; they always had visual contact with their neighbours , even during the experimental period , to prevent the stress of social isolation . \n the animals were fed a constant diet of commercial concentrates with hay and water available ad libitum . \n one month before starting of the experiment all groups of ewes were cannulated with stainless steel guide cannulas ( 1.2 mm o.d . ) into the third ventricle under stereotaxic control . \n the guide cannula was fixed to the skull with stainless steel screws and dental cement . \n the correct placement of the guide cannula into the third ventricle was established by the efflux of the cerebrospinal fluid from cannula during the surgery . \n additionally , the placement of the cannula was checked by inspection of the brain after decapitation . \n all animals had a venous catheters implanted into jugular vein the day before the experiment . \n all experimental procedures were performed in accordance with the polish guide for the care and use of animals ( 1997 ) and were approved by the local ethics committee of the warsaw university of life sciences . \n immune stress was induced in treated animals by intravenous ( iv ) bolus injection of an appropriate volume of lps ( 400 ng / kg body weight ) from e. coli 055:b5 ( sigma - aldrich , st . \n louis , mo , usa ) dissolved in saline ( 0.9% w / v nacl ) ( baxter , deerfield , il , usa ) at a concentration of 10 mg / l into the jugular vein . \n the maximum volume of lps solution ( 10 mg / l ) administered to any animal was 2.5 ml . \n the control group received an equivalent volume of saline . the animals ( n = 20 ) \n were randomly assigned to four experimental groups : ( 1 ) nacl control group \n ( n = 5 ) , received intracerebroventricular ( icv ) injection of ringer - locke 's solution ( rls ) into the third ventricle of the brain and 15 min later iv injection of nacl ; ( 2 ) lps control group \n ( n = 5 ) , received rls ( icv ) and 15 min later lps ( iv ) ; ( 3 ) anti - lps group ( n = 5 ) , received monoclonal anti - lps antibody ( abcam , cambridge , uk ) ( icv ) in dose of 10 g / animal and 15 min later lps ( iv ) ; ( 4 ) anti - tlr4 group , received ( icv ) antibodies binding tlr4 receptor complex components , anti - lbp ( abcam , cambridge , uk ) in dose of 20 g / animal , and anti - md2 ( abcam , cambridge , uk ) in dose of 10 g / animal and 15 min later lps ( iv ) . \n jugular blood samples from each ewe were taken for lh and cortisol measurement at 15 min intervals , beginning 2 h before endotoxin or saline injection ( iv ) preceded by the injection of appropriate substances ( icv ) , and continuing 4 h after lps or saline treatment . \n effect of immune stress on the gene expression in hypothalamic structures and in the ap . \n after 2 weeks of convalescence , the same animals ( n = 20 ) were used . \n mbh , and the median eminence me ) and the anterior pituitary glands ( ap ) were collected 2 h after iv injection of lps or saline preceded by the icv injection of corresponding substances as described above . \n the animals were slaughtered by decapitation , the brains were rapidly removed from the skulls , and then chosen hypothalamic structures and aps were dissected . \n all tissues were frozen immediately after collection in liquid nitrogen and were stored in 80c until assay . \n the concentration of lh in plasma was assayed by the radioimmunoassay ( ria ) double - antibody method using anti - ovine - lh and anti - rabbit--globulin antisera and ovine standard ( nih - lh - so18 ) as described by stupnicki and madej . \n the sensitivity was 0.3 ng / ml ; intra - assay and interassay coefficients of variation were 8.3% and 12.5% , respectively . \n the cortisol concentrations were determined by the ria method according to kokot and stupnicki , using rabbit anticortisol antisera ( r/75 ) and hplc grade cortisol standard ( sigma ) . \n the assay sensitivity was 0.95 ng / ml and the intra - assay and interassay coefficients of variation were 10% and 12% , respectively . \n total rna from hypothalamic and pituitary tissues was isolated using nucleospin rna ii kit ( macherey - nagel gmbh & co , dren , germany ) according to manufacturer 's protocol . \n the purity and concentration of isolated rna were quantified spectrophotometrically by measuring the optical density at 260 and 280 nm in a nanodrop 1000 instrument ( thermo fisher scientific inc . , \n the rna integrity was verified by electrophoresis using 1% agarose gel stained with ethidium bromide . \n dynamo sybr green 2-step qrt - pcr kit ( finnzymes , espoo , finland ) was used to prepare cdna synthesis . as a starting material for this pcr synthesis 800 ng of total rna \n real - time rt - pcr was carried out using sybr green 2-step qrt - pcr kit ( finnzymes , espoo , finland ) components and hplc - grade oligonucleotide primers synthesized by genomed ( poland ) . \n specific primers for determining the expression of housekeeping genes and the genes of interest ( table 1 ) were designed using primer 3 software . \n one tube contained 10 l pcr master mix ( 2x ) , 7 l rnase - free water , 2 l primers ( 1 l each , working concentration was 0.5 m ) , and 1 l cdna template . \n the tubes were run on the rotor - gene 6000 ( qiagen , duesseldorf , germany ) . \n the following protocol was used : 95c in 15 min for activating hot start dna polymerase and finally the pcr including 30 cycles at 94c in 5 sec for denaturation , 56c in 20 sec for annealing , and 72c in 15 sec for extension . after the cycles , a final melting curve analysis under continuous fluorescence measurements \n the results of lps treatments on the concentrations of plasma lh and cortisol were examined by two - way analysis of variance anova ( statistica ; stat - soft , inc . , \n tulsa , ok , usa ) to identify treatment effects and significant interactions between the control and experimental groups . \n all experiments consisted of a baseline period when no treatment was given ( 2 to 0 h before ) and period when treatments were applied ( + 1 to + 4 h after lps or saline injection ) . \n anova for hormone parameters excluded data during the first hour after lps or saline treatment to allow time for treatments to take effect . \n when a significant treatment by time interaction was observed , post hoc analysis was conducted to identify treatment effects . \n relative gene expression was calculated using the comparative quantification option of rotor gene 6000 software 1.7 ( qiagen , duesseldorf , germany ) . \n the second differential maximum method was used in this analysis to calculate reaction efficiencies and a set percentage of the maximum fluorescence value to calculate the beginning of the exponential phase . to compensate a variation in cdna concentrations and the pcr efficiency between tubes , \n initially , three housekeeping genes glyceraldehyde-3-phosphate dehydrogenase ( gapdh ) , -actin ( actb ) , and cyclophilin c ( ppic ) were tested . \n the bestkeeper was used to determine the most stable housekeeping gene , for normalizing genes of interest expression . \n the bestkeeper was based on the pair - wise correlation analysis of all pairs of candidate genes and calculates variations of all reference genes ( sd ( ct ) ) . \n it had the lowest sd ( ct ) value and a good correlation coefficient with the remaining analysed housekeeping genes . \n the results are presented as relative gene expression of the target gene versus housekeeping gene , relative expression value and mean sem . \n the significance of differences between the experimental groups was assessed by the mann - whitney u test . \n lps administration significantly ( p < 0.01 ) increased plasma cortisol level in all lps - treated groups ( lps control , anti - lps , anti - tlr4 ) compared with nacl control group ( figure 1 ) . \n intravenous injection of lps significantly reduced plasma lh release in all lps - treated groups ( lps control : p < 0.01 ; anti - lps : p < 0.05 ; anti - tlr4 : p < 0.05 ) compared with the saline control . \n the central administration of anti - lps as well as icv injection of anti - lbp and anti - md2 did not influence on lowered lh secretion ( figure 2 ) . \n peripheral administration of endotoxin significantly ( p < 0.05 ) decreased the gene expression of lh- in the ap in lps control and anti - lps groups compared with nacl control group . on the other hand , injection of antibodies binding tlr4 receptor complex components anti - lbp and anti - md2 ( anti - tlr4 group ) released lh-gene expression from the suppressive effect of lps administration ( figure 3 ) . \n injection of lps significantly ( p < 0.01 ) decreased gnrh gene expression in lps control group from hypothalamic structures such as the poa ( by 54% ) and the me ( by 50% ) compared with nacl control group . in the me , the central administration of anti - lps antibody ( anti - lps group ) and antibodies binding tlr4 receptor complex components anti - lbp and anti - md2 ( anti - tlr4 group ) significantly ( p < 0.01 ) reduced lps dependent suppression of gnrh gene expression compared with lps control group . \n no effects of iv and icv treatments on gnrh expression were found in the mbh . \n the amount of gnrh mrna determined in the aha was too low and did not enable the quantitative analysis in this hypothalamic structure ( figure 4 ) . \n gene expression of receptor for gnrh - r significantly ( p < 0.01 ) decreased in lps control groups in the me and in the ap compared with nacl control group . \n central administration of anti - lps or antibodies binding tlr4 receptor complex components anti - lbp and anti - md2 did not affect gnrh - r gene expression compared to lps control group ( figure 5 ) . \n concomitant administration of lps ( iv ) and anti - lps ( icv ) increased ( p < 0.05 ) mrna tlr4 level in the mbh but decreased ( p < 0.01 ) it in the ap compared with nacl control group . \n it has been shown that the administration of lps ( iv ) together with antibodies binding tlr4 receptor complex components anti - lbp and anti - md2 ( icv ) increased tlr4 gene expression in the me compared with the other groups ( nacl control : p < 0.01 ; lps control : p < 0.05 ; anti - lps : p < 0.05 ) . \n an increase ( p < 0.05 ; p < 0.05 ) of tlr4 gene expression in anti - tlr4 group was also demonstrated in the mbh compared with nacl control and lps control groups , respectively , whereas a decrease ( p < 0.01 ) in the ap compared with nacl control group . in the poa and \n our study shows the inhibitory effect of lps - induced immune stress on lh secretion in sheep which is consistent with previous in vivo studies conducted on anestrous ewes [ 15 , 16 ] . \n other researchers also showed that lps affects lh secretion and even disturbs the preovulatory lh surge in ewes [ 29 , 30 ] . \n in castrated rams , lps significantly reduced plasma lh level and the number of lh pulses . likewise , refojo et al . demonstrated that endotoxin lowered lh concentrations by inhibiting several pulsatility parameters such as frequency , amplitude , and maximum values in male rats . \n the most important regulator of lh secretion is gnrh , which affects reproduction processes at level of the cns by stimulation of the gonadotrophs in the ap to secrete lh . \n functional regulation of lh secretion is mediated by the pulsatile secretion of gnrh into the hypophyseal portal vasculature . \n it has been demonstrated that pulsatile pattern of lh secretion is a direct reflection of gnrh secretion from hypothalamus in ovariectomized ewes . \n changes in lh secretion observed after peripheral injection of lps suggest that immune stress acts on the reproductive functions at the level of the hypothalamus through alterations of gnrh secretion . \n fergani et al . showed that peripheral endotoxin administration caused disorders in the gnrh / lh surge . \n it could be caused by less gnrh release or that pituitary responsiveness to gnrh may have been comprised . \n previous studies on the ovine model clearly showed that administration of bacterial endotoxin induced an immune / inflammatory stress and reduced pulsatile gnrh secretion [ 2 , 13 ] . \n in presented study , bacterial endotoxin lowered gnrh mrna level in the poa , where more than half of all gnrh perikarya are located . \n it was determined that gnrh mrna level in the poa was the highest among other analysed hypothalamic structures , which confirms earlier reports that neurons located in the poa synthesized most of gnrh transcripts in anestrous phase ewes . \n this fact further supports the assumption that activity of gnrhergic neurons in the hypothalamus is modulated by immune stress . \n however , it has not been proven that immune challenge affects the gnrh synthesis at the transcriptional or posttranscriptional levels . observed changes in gnrh mrna content in the hypothalamic area may not result from decreased gnrh gene transcription which is fairly stable but from lowered accumulation or increased degradation cytoplasmic gnrh mrna . \n this data suggests that the suppressive effect of immune stress on gnrh release to the hypophyseal portal blood previously described in sheep could result from reduced gnrh gene expression in the poa . \n it is worth to mention that in the present study there were no significant changes in gnrh gene expression in the mbh and the amount of gnrh mrna in the aha was below the limit of detection , which is consistent with previously obtained results and suggests that in anestrous ewes gnrh neurons located in these hypothalamic structures do not play pivotal role in communication between neuroendocrine and immune systems . \n it was also found that lps significantly decreased gnrh gene expression in the me , where gnrh neurons terminals are located . \n this phenomenon has been described in detail in our previous study and it has been suggested that the selective transport of gnrh transcript to the distal part of neurons occurred in the gnrhergic neurons . decreased content of gnrh in the me after lps treatment \n could result from decreased transport of gnrh mrna to the nerves terminals as well as increased degradation of gnrh transcript in this structure . \n it was previously found that a gradual reduction of the poly(a ) tail of mrna occurs during its translocation from the perykaryon to the nerve terminal [ 3739 ] . \n therefore , it may be assumed that mrna stored in nerves terminals is more sensitive to all factors affecting the stability of these transcripts than mrna that occurs in the region of the neuronal body . in the present study , decreased gnrh - r gene expression after lps administration was determined in the ap and me . \n it supports the previous studies carried out on ovariectomized ewes that immune stress lowered gnrh - r gene expression in the pituitary gland . \n similar results were also obtained in rats where the administration of lps affected the gnrh - r expression both in hypothalamus and pituitary . \n the decrease in the level of gnrh - r mrna in the ap may be due to lower gonadotrophs stimulation by gnrh , which is the main factor controlling the amount of its receptors . in turn , the reduction of gnrh - r gene expression may lead to a decrease gnrh - r expression in gonadotrophs and lower sensitivity of these cells on gnrh stimulation . \n this may lead to lower lh- synthesis in the ap . in the presented study , \n it fully supports the previous studies reported about stimulatory effect of immune stress on cortisol release in various animal species including sheep [ 2 , 16 , 20 , 43 ] . \n the elevation of the cortisol release suggests the activation of the hypothalamic - pituitary - adrenal ( hpa ) axis , which may result in inhibition of the hpg axis [ 2 , 43 ] . \n immune challenge stimulates the synthesis of hpa axis components , such as arginine vasopressin , corticotropin - releasing hormone ( crh ) , adrenocorticotrophic hormone , and corticosterone / cortisone from adrenal cortex [ 44 , 45 ] . \n all these factors have an inhibitory effect on the hpg axis [ 46 , 47 ] \n . however , the role of cortisol and other hpa axis components in the suppression of the gnrh / lh secretion during immune stress seems to be ambiguous . \n rivest and rivier demonstrated in rats that reproductive system inhibited by lps injection has not been released from its suppressive action by crh antibodies administration , although this administration prevented the increase in the hpa axis activity . in study carried out on sheep , debus et al \n . demonstrated that the cortisol secretion blockage did not lower the suppression of gnrh / lh release caused by lps treatment . \n one of the mechanisms through endotoxin that may modulate the neuroendocrine system is induction of pro - inflammatory cytokines [ 23 , 49 ] . \n however , the main source of centrally acting cytokines seems to be their local synthesis in the brain parenchyma . \n these cytokines can be also secreted by the bbb cells activated by endotoxin [ 23 , 51 , 52 ] as well as the choroid plexus cells . \n it is worth mentioning that some amounts of the central cytokines could have a peripheral origin and cross the bbb due to the existence of saturated transport mechanism . \n another possible pathway of endotoxin penetration to the brain is through the organum vasculosum laminae terminalis ( ovlt ) , which is one of the sensory circumventricular organs , forming the anterior wall of the third ventricle . \n this structure is devoid of the bbb , so ovlt could be a potential location for lps bypassing into the brain parenchyma . a direct response of ovlt cells to exposure to endotoxin or cytokines \n their study showed that the ovlt cells secrete proinflammatory cytokines ( e.g. , tnf- , il-1 , and il-6 ) . \n it has been previously reported that lps acting indirectly via stimulation of central cytokines synthesis affects gnrh secretion in the hypothalamus and can disturb lh secretion from the ap [ 13 , 18 , 19 ] . on the other hand , one of the mechanisms by which peripherally administered \n endotoxin affects central response is the activation of the afferent vagal nerves by prostaglandins ( pgs ) , other important regulatory factors of gnrh / lh levels suppression during immune stress . \n . showed that inhibition of prostaglandin e2 ( pge2 ) suppressed the release of gnrh / lh . \n peripheral administration of lps induces synthesis of endogenous cytokines ( e.g. , il-1 ) and activates the projection area of the vagal nerves in the brain . \n presence of receptors for il-1 was demonstrated in study of ek et al . which suggested that il-1 stimulates vagal sensory activity . \n this activation of afferent nerve fibers by peripherally released cytokines could be a fast pathway of immune signals transfer from the periphery to the brain . \n however , in response to circulating cytokines , a slow humoral pathway of transmission is activated . \n immune challenge could act as well in this pg - dependent manner represented by pgs synthesis by cyclooxygenase-2 ( cox-2 ) around blood vessels . \n these observations suggest that pgs play a role in mediating between the immune and neuroendocrine systems [ 57 , 63 ] . in our study , tlr4 gene expression was determined in the hypothalamic structures such as the poa , aha , mbh , and me and in the ap . however , no effect of lps administration on tlr4 transcription in all these structures was observed . \n these results are partially contrary to our previous study performed on anestrous ewes where significant increase of the tlr4 gene expression was determined . \n the existence of tlr4 receptor in the hypothalamus may suggest the possible direct action of lps in the cns . although experiments carried out on cats seem to exclude the penetration of endotoxin from the blood to the brain , the results of experiments conducted on rats are inconclusive . \n singh and jiang suggest that lps modulates the permeability of the bbb but does not exceed it . \n however , in vivo research performed on mice and rats have shown that iodine - radiolabelled lps penetrated the bbb in measured quantities . \n the study performed on rats showed that central administration of endotoxin suppressed the secretion of lh in rats . \n this proves the potential of centrally acting lps to suppress the hpg activity at the hypothalamic level . \n in present study , it was determined that the blockade of tlr4 receptor in the hypothalamus as well as administration of anti - lps antibody into the region of hypothalamus reverses decreasing effect of lps treatment on gnrh mrna level in the me . \n however , no effects of these treatments were observed in the structures where gnrh neurons perikarya are located . \n the fact that the blockade of tlr4 receptor as well as administration of anti - lps antibody into the third ventricle restored gnrh mrna content only in the me suggests that these treatments prevented the inflammation , dependent decreasing of the gnrh mrna stability rather than decreasing gnrh gene transcription . \n the inhibition of tlr4 receptor as well as decreasing the number of its interacting ligand could result in decreased proinflammatory cytokines synthesis in the hypothalamus . \n it was previously suggested that acting in the region of hypothalamus proinflammatory il-1 could be responsible for decreasing the stability of gnrh mrna and reduction of its translation . in our study , \n restoration of lh- mrna content to the control level was observed only in the anti - tlr4 group . \n however , this change in the lh- gene expression was not accompanied by the elevation of the circulating lh concentration . \n the lack of parallelism between the increased lh transcription and the peripheral level of lh in the anti - tlr4 group suggests that lh release was still inhibited by the peripheral immune / inflammatory challenges affecting the hpg axis at the pituitary level . \n this suppression may result from the action of proinflammatory cytokines whose receptors are widespread in the pituitary gland . \n the results of our ex vivo study showed that il-1 is a potent downregulator of lh secretion directly from the pituitary and suggested that this direct action of interleukin could have a profound effect on the suppression of lh release occurring during an inflammatory state . \n the in vitro study performed on the mouse att-20 pituitary tumor cells showed that direct lps treatment increases the number of il-1r1 in a dose - dependent manner . \n the studies carried out on mice and sheep also reported the stimulating effect of lps on il-1r1 mrna . \n another inflammatory cytokine involved in direct modulation of the secretory activity of the pituitary is il-6 . \n the in vitro study showed that il-6 significantly suppressed gnrh - stimulated lh release from male rats dispersed pituitaries throughout the dose range but did not influence basal lh release . \n it is worth mentioning that stress caused by lps injection may increase the number of cytokines receptors expressed in the ap . \n the factor suppressing the lh secretion at the level of pituitary could be also lps itself . \n our previous ex vivo study showed that lps directly decreases lh secretion from the ovine ap explants . \n it was suggested that the secretion of lh from the pituitary could be affected directly by lps and/or could result from autocrine action of proinflammatory cytokines secrete by the folliculostellate cells . \n the study suggests that the blockade of tlr4 receptor in the hypothalamus during lps - induced immune stress restores the lh- transcription in the pituitary gland \n . however , this treatment is not sufficient to unblock the release of lh suppressed by the peripheral immune / inflammatory challenges .", "role": "user" }, { "content": "the present study was designed to examine the effect of intracerebroventricular ( icv ) administration of antilipopolysaccharide ( lps ) antibody and blockade of toll - like receptor 4 ( tlr4 ) during immune stress induced by intravenous ( iv ) lps injection on the gonadotropin - releasing hormone / luteinizing hormone ( gnrh / lh ) secretion in anestrous ewes . \n injection of anti - lps antibody and tlr4 blockade significantly ( p < 0.01 ) reduced the lps dependent lowering amount of gnrh mrna in the median eminence ( me ) . \n moreover , blockade of tlr4 caused restoration of lh- transcription in the anterior pituitary decreased by the immune stress . however , there was no effect of this treatment on reduced lh release . \n the results of our study showed that the blockade of tlr4 receptor in the hypothalamus is not sufficient to unblock the release of lh suppressed by the immune / inflammatory challenges . \n this suggests that during inflammation the lh secretion could be inhibited directly at the pituitary level by peripheral factors such as proinflammatory cytokines and circulating endotoxin as well .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: family planning is a conscious activity of individuals and couples of reproductive age that incline , not only to regulate the number and timing of birth , but also to have a healthy child and to fulfill all of its role throughout the life cycle ( 1 ) . \n constitution of montenegro provides to its citizens the inalienable ( indefeasible ) right to health care and healthy living by items that ensure protection and promotion of reproductive health . \n hence , family planning comprises various aspects biological , medical , demographic , sociological , psychological , economic , ethical , political , and from the standpoint of carrier , there are three levels : as individual practice , as a movement of social groups and as a family planning program that is brought by government ( 2 ) . \n family planning is a complex process which is related to other aspects of reproductive health . \n the average age of engaging into the first sexual intercourse has decreasing tendency and it is also associated with a multiple sexual partners during active sex life , and thus an increased risk of acquiring sexually transmitted diseases ( 3 ) . it is estimated that each year , sexually transmitted diseases ( stds ) are diagnosed in more than 400 million adults \n . out of that number , approximately 60% of stds and infections occur in people younger than 25 years . \n past experience with young people shows that they get knowledge about sexuality mostly from unreliable sources , such as television , magazines and experience exchange with peers and older adolescents ( 4 ) . \n information about delicate topics at home is scarce and insufficient , as well as school curricula ( 5 ) . \n first sexual experience are rarely planned by adolescents , which is the reason for not using protection , either from ignorance or passivity and passing the responsibility to partner . \n the sexual behavior of adolescents is complex , and the facts show that even when they are well and accurately informed , they often behave inconsistently in protection of pregnancy and stds \n . growing number of young with different forms of risky behavior demands faster and better managing in early detection and eradication of risk factors , appropriate approach , prevention through contemporary and continuous methods of health education and promotion , counseling and regular preventive and gynecological follow up ( 6 ) . \n effective birth control could allow young couples to achieve individual goals and to plan family , when they want . \n objective of this survey is , primarily , to study knowledge , attitude and behavior of young people regarding family planning , as well as their awareness of sexuality and contraception , degree of motivation for using contraceptives and pattern of sexual behavior . \n the survey was conducted from may to december 2009 and it was cross - sectional study , created and coordinated by experts of the institute of public health . \n the schedule of conducting the survey on faculties was made by random selection and it was agreed by the university of montenegro . \n the interviewers were trained for conducting the survey . the general plan was made and precise survey day for each faculty was defined . according to the activity plan , all students , who were present at the faculty on the planned day , were surveyed . \n some faculties refused to participate in the survey , while some others were unable to participate because of earlier completion of school year due to new influenza pandemic . \n the rate of faculties response was 90% , whereas the rate of students response ( their consent to survey ) was 87% . \n it was planned to involve 10% of all students in montenegro , and considering that there are 19325 students at the university of montenegro , number of students included in the study had to be 1933 with the rate of students response 90% . \n regarding aforesaid , plan of number of students to be interviewed was prepared on each respective faculty . \n but since the number of students , who refused to participate in the study , differed from faculty to faculty , the planned number of interviewed students was not reached . \n namely , the survey included 1804 students of various generations , which is approximately 9.3% of all students of the university of montenegro . \n for the purpose of results interpretation , only students aged up to 26 years have been taken into account . \n furthermore , some questions from the survey were intended only for sexually active female students . \n the experts of the institute of public health had defined the questionnaire used for the survey and its adequacy was tested on the fifth year students of the medical faculty of the university of montenegro . \n the questionnaire was consisted of 25 closed type questions but only answers to questions , which were related to the topics of this work , were analyzed . \n the following methods for statistical data analyses were used : methods of descriptive statistics ( frequencies distribution of qualitative characteristics and relative numbers ) and nonparametric methods ( the 2 test ) for analyzing significance of correlation between responses and knowledge of students about family planning , as well as attitude and knowledge about family planning and sexual behavior of students , at the significance p<0.05 . \n the study comprised 53.9% of students aged up to 20 years and 46.1% of students aged from 21 to 26 years . \n it is observed that more than a third of participants were first year students , more than a quarter second year , around 20% third year , 15% fourth year , 2% fifth year and 0.7% sixth year students . \n there is not statistically significant difference among students regarding sources of knowledge on family planning by gender ( 2 = 7.66 , p>0.05 ) , but it appears by age ( 2 = 14.54 , p<0.05 ) . \n the students have got information on family planning from parents ( 46.7% ) , mass media ( 27.5% ) , books and school ( 23.6% ) and at least from doctors ( 2.2% ) ( graph 1 ) . out of total number of interviewed students , who stated that abortion is good method of contraception , 12.8% claimed doctors as the source of information about family planning , then books ( 12% ) , mass media ( 9% ) and parents ( 5.5% ) . out of total number of male students 11.2% considers abortion as good method of contraception , \n there is statistically significant difference among interviewed students by gender ( 2 = 16.02 , p<0.01 ) . \n attitude that abortion is a good method of family planning was commonly found in sexually active female students who reported that only their partners make decision on contraceptive methods ( 9.5% ) , then those who reported that they choose contraceptive methods themselves ( 9.3% ) . \n there were 5.3% of female students who consider abortion as a good family planning method among them who do not use any contraception , while this attitude was stated by 4.9% of female students who make decision on contraceptive methods with their partners ( graph 2 ) . \n more than a quarter of female students ( 28.6% ) , who had more induced abortions and 14.3% of female students , who had one induced abortion , consider abortion as a good method of contraception . among female students , who consider abortion as a good method of contraception , majority received information about family planning from doctors , followed by those whose source of information were books . \n mass media as the source of information was on the third place , and then information received in the school , and at the last place there were information received from parents ( table 1 ) . \n there was statistically significant difference in students responses ( 2 = 15.230 , p<0.05 ) . \n the most of interviewed students find condom as the best contraceptive method , then pill , contraceptive coil , calendar - based method , and interrupted sexual intercourse , and at the end local contraceptives . \n thereby , there is statistically significant difference regarding the source of information about family planning ( c2 test = 39.74 , p<0.05 ) . \n there is statistically significant difference among students in estimating their own knowledge about contraception ( 2 = 32.25 , p<0.05 ) . \n the most of students estimate their knowledge as sufficient , then as very good , excellent , insufficient and at the end only 3.4% of students state that they have no knowledge about contraception . \n the students , who have estimated their knowledge as excellent , stated the sources of knowledge by following order : books ( 21.1% ) , mass media ( 18.3% ) , school curricula ( 18.1% ) , doctors ( 17.9% ) and parents ( 13.4% ) . \n the contraceptive method , that is being used , is considered by the largest number of students as partially safe , then as completely safe , followed by those , that use no protection at all and at the end students who find their method unsafe . \n the largest number of students , who use protection occasionally , considers their contraceptives as partially safe , while the most of students , who use protection regularly , find their method completely safe . \n students that use protection occasionally prevail in group , which finds their method unsafe ( table 2 ) . \n there is statistically significant difference in students responses ( 2 = 257.62 , p<0.01 ) . \n the early engaging in sexual intercourse , with other forms of risky behavior such as sexual intercourse under influence of alcohol or other abuse substances , leads to multiple sexual partners and danger from sexual abuse , prostitution and crime ( 8) . on the other hand , society became more tolerant toward behavior and attitude of individuals and groups . \n there are less forbiddances and opinions which constrain young people and their behavior , so this tolerance could be seen in their sexual life . \n but , source of information for young about family planning and reproductive health , in general , is considerable problem ( 9 ) . \n more than a half of young people in serbia have said that they do not have enough information about new contraception , although two - thirds of them consider sex as very important in their relationship , according to survey conducted by the association for reproductive health among young people in belgrade last year ( 10 ) . \n the results of other surveys confirmed that there was trend of engaging in sexual activities at very young age with incomplete and insufficient knowledge about it . \n one of the characteristics of modern societies is the increase of sexual activities among young people , especially the first sexual experience in early adolescence period . \n consequently , there are increased risks related to sexuality of young people , proved by epidemiological researches in that area . at the same time \n the young people are getting informed about sexuality through mass media or from peers and through school curricula on a biology class in elementary and secondary school . nowadays , parents are too busy , many of them are uninformed , and often it is taboo in family ( 11 ) . \n the research in other countries confirmed the same experience about awareness and knowledge on reproductive health of young people from sources of various values ( 12 ) . \n however , results obtained in our study show that respondents dominantly received information by competent sources . \n the particular research shows changing in family relationships and that the young people talk more often to their parents on this topic . \n they are entitled to open access to verified and accurate information concerning their sexuality , reproductive health and rights . \n the results of this survey show that all sexually active students do not use contraceptives . \n it is very similar with results of surveys on young people behavior conducted in serbia . \n these surveys show that the most of them stated that they use contraceptives always , while a quarter of participants stated that they use contraception often and one in five stated rarely or never . \n these surveys also showed that condom is the commonly used contraceptive , and contraceptive pills were on the second place . \n the most of interviewed makes decision about using contraception together with partner whereas almost half of them stated they make decision alone ( 13 ) . \n the results of survey conducted among students in serbia showed that two - thirds of female students and more than a third of male have sexual intercourse . \n the most used contraceptive methods are the interrupted sexual intercourse and calendar - based method , then pill , local contraceptives , diaphragm and contraceptive coil ( 14 ) . \n the goal of the family planning is having healthy and wanted child , and responsible and conscientious parenthood .", "role": "user" }, { "content": "conflict of interest : none declared.family planning is a conscious activity of reproductive age that inclines , not only to regulate the number and timing of birth , but also to have a healthy child and to fulfill all of its role throughout the life cycle . the survey included 1804 students of various generations , which is approximately 9.3% of all students of the university of montenegro . \n instrument of survey was questionnaire with 25 closed type questions . \n methods used for statistical data analyses were methods of descriptive statistics and nonparametric methods to analyze significance of differences at the significance p>0.05 . \n the study comprised 53.9% of students aged up to 20 years and 46.1% of students aged from 21 to 26 years . \n there is statistically significant difference among students regarding sources of knowledge on family planning by age ( chi 2 = 14.54 , p<0.05 ) . \n the students have got information on family planning mostly from parents ( 46.7% ) . out of total number of interviewed students , who stated that abortion is good method of contraception , \n 12.8% claimed doctors as the source of information , then books , mass media and parents . \n the most of interviewed students find condom as the best contraceptive method . \n students used to get information on family planning mostly often from their parents , and most of them consider that they are well informed . \n one third of students do not protect themselves from undesired pregnancy , but most of them consider abortion as non adequate method of family planning .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: presbyopia affects an estimated one billion people worldwide.1 current surgical treatment options for presbyopia depend on the presence of a cataract at initial assessment . \n if present , a cataract extraction with the implantation of a multifocal or accommodative intraocular lens implantation may be chosen.2,3 if a clear lens is present , a non - lens - based treatment , eg , presbyopic laser - assisted in situ keratomileusis ( lasik ) , monovision with a contact lens or lasik , conductive keratoplasty or femtosecond laser intrastromal presbyopia treatment ( intracor ; technolas perfect vision gmbh , munich , germany ) may be the preferred choice . \n one emerging non - lens - based treatment modality for presbyopia is the insertion of intrastromal corneal inlays that work by a variety of different mechanisms . \n the vue+ ( revision optics , lake forest , ca , usa ) inlay consists of a space - occupying lenticule that changes corneal curvature , creating a hyperprolate cornea for both near and distance vision.4 the flexivue microlens ( presbia , amsterdam , netherlands ) inlay is itself a bifocal inlay , with a plano central zone for distance vision and a peripheral annulus with refractive add power for near vision.5 both the vue+ and flexivue microlens inlays have received conformit europenne ( ce ) marking , but are not yet us food and drug administration ( fda)-approved . the kamra small aperture inlay ( acufocus , irvine , ca , usa ) , previously known as the aci 7000 , has a 3.8 mm outer diameter and an inner 1.6 mm central aperture that selects for central light rays and creates a pinhole effect , thus increasing the depth of focus . \n this is inserted in the nondominant eye , and has been shown to improve uncorrected near visual acuity ( unva ) , without affecting uncorrected distance visual acuity ( udva).6 the kamra inlay was the first corneal inlay for presbyopia to receive ce marking in 2005 , and is currently pending approval from the fda for use in the usa outside of clinical trials . \n presbyopia typically affects patients from about 40 years of age , and phakic patients who undergo corneal inlay implantation may subsequently develop cataracts as they grow older . as corneal inlay implantation for presbyopia \n is relatively new , there are not many patients who have yet undergone cataract surgery after corneal inlay implantation . \n potential concerns with this procedure include technical difficulty performing cataract surgery with the corneal inlay in place , accuracy of biometry readings , and visual outcome after cataract surgery . \n this paper presents the experiences and results of two patients who underwent phacoemulsification and intraocular lens implantation for cataracts that developed after implantation of the kamra corneal inlay for presbyopia . \n the two patients in this case series , aged 53 and 62 years of age , were first seen in january 2010 and may 2007 , respectively , at the singapore national eye centre , singapore , requesting for correction of presbyopia without the use of corrective lenses . \n slit - lamp examination of the anterior and posterior segments revealed no significant abnormalities , and both patients had lenses graded as nc1 according to the lens opacities classification system iii ( locs iii)7 in their nondominant left eyes . \n various surgical options were presented to the patients , and each opted for implantation of the kamra corneal inlay . \n one patient underwent implantation of the kamra corneal inlay with a microkeratome - created lamellar flap and the other had implantation via a femtosecond laser - created lamellar flap . \n case 1 , a 53-year - old chinese male , underwent implantation of the kamra corneal inlay in the nondominant left eye on january 6 , 2010 . \n a 7.80 mm - diameter lamellar flap was created at a depth of 190 microns with a visumax femtosecond laser ( carl zeiss meditec , jena , germany ) . \n the patient s pre- and 1-month postoperative visual acuity results can be found in table 1 . \n a slit - lamp photograph of the kamra corneal inlay in place is shown in figure 1 . \n twelve months after implantation , even though reading well , the patient noted ghosting and slight monocular diplopia in the left eye . \n the patient underwent surgical repositioning of the implant , resulting in resolution of his symptoms . \n figure 2 shows acutarget ( acufocus ) photographs of the kamra implant positioning before and after surgical repositioning . \n seven months after repositioning of the implant , the patient had an unva of j10 and was noted to have developed cataract of grade nc3p3 ( locs iii ) in the left eye . \n various options were discussed with the patient , including cataract surgery with the implant in place , removal of the implant followed by cataract surgery and subsequent re - implantation , and implant removal followed by cataract surgery with a multifocal or accommodative intraocular lens implant . \n centration of the kamra implant was confirmed with acutarget photographs ( figure 2 ) . due to the excellent centration ( inlay vs purkinje : 95 m x , 27 m y ) , the patient opted for cataract surgery , leaving the kamra implant in place . \n corneal topography remained stable after implantation of the kamra inlay and before cataract surgery ( figure 3 ) . \n the patient underwent uncomplicated phacoemulsification and intraocular lens implantation of the left eye under local anesthesia . \n a 2.60 mm temporal clear corneal incision was made , and a + 22.0 d monofocal tecnis zcb00 intraocular lens ( abbott medical optics , santa ana , ca , usa ) was implanted . \n no modifications were made to the surgical procedure , and the surgeon reported no difference in the ease of surgery with the kamra implant in place , apart from a few extra ocular rotations during capsulorhexis and phacoemulsification to improve visualization . \n one month after cataract surgery , posterior capsular opacification was noted in the left eye , and the patient underwent nd : yag laser capsulotomy 6 weeks after cataract surgery ( figure 1 ) . \n the patient reported good near , intermediate , and distance vision without the use of spectacles , and remains spectacle - independent . \n case 2 , a 62-year - old chinese male , underwent implantation of the kamra corneal inlay in the nondominant left eye on july 4 , 2007 . \n a 9.00 mm - diameter lamellar flap was created at a depth of 151 microns with a schwind microkeratome ( schwind , kleinostheim , germany ) . \n the patient s pre- and 1-month postoperative visual acuity results can be found in table 1 . \n four years after kamra implantation , cataract of grade nc2 ( locs iii ) was noted in the implanted left eye . \n nevertheless , vision in this eye remained good ; udva was 20/20 and unva was j2 . \n however , 6 years after kamra implantation , the patient felt that his distance vision in the left eye had deteriorated significantly ( table 1 ) and , on assessment , cataract of grade nc4 ( locs iii ) was noted . \n acutarget photographs confirmed that the implant was well centered ( inlay vs purkinje : 146 m x , 78 m y ) . \n various treatment options were presented to the patient , who opted for cataract surgery , leaving the kamra implant in place . \n the patient underwent phacoemulsification and intraocular lens implantation of the left eye under local anesthesia . \n a 2.60 mm temporal clear corneal incision was made , and a + 20.0 d monofocal tecnis zcb00 intraocular lens was implanted . \n no modifications were made to the surgical technique , and the surgeon reported no difference in ease of the surgery with the kamra implant in place , except that some extra ocular rotations during capsulorhexis and phacoemulsification to optimize the surgical view were required . \n surgical time was 15 minutes , and there were no intraoperative or early postoperative complications . \n the patient reported good near , intermediate and distance vision without the use of spectacles , and remains spectacle - independent . \n case 1 , a 53-year - old chinese male , underwent implantation of the kamra corneal inlay in the nondominant left eye on january 6 , 2010 . \n a 7.80 mm - diameter lamellar flap was created at a depth of 190 microns with a visumax femtosecond laser ( carl zeiss meditec , jena , germany ) . \n the patient s pre- and 1-month postoperative visual acuity results can be found in table 1 . \n a slit - lamp photograph of the kamra corneal inlay in place is shown in figure 1 . \n twelve months after implantation , even though reading well , the patient noted ghosting and slight monocular diplopia in the left eye . \n the patient underwent surgical repositioning of the implant , resulting in resolution of his symptoms . \n figure 2 shows acutarget ( acufocus ) photographs of the kamra implant positioning before and after surgical repositioning . \n seven months after repositioning of the implant , the patient had an unva of j10 and was noted to have developed cataract of grade nc3p3 ( locs iii ) in the left eye . \n various options were discussed with the patient , including cataract surgery with the implant in place , removal of the implant followed by cataract surgery and subsequent re - implantation , and implant removal followed by cataract surgery with a multifocal or accommodative intraocular lens implant . \n centration of the kamra implant was confirmed with acutarget photographs ( figure 2 ) . due to the excellent centration ( inlay vs purkinje : 95 m x , 27 m y ) , the patient opted for cataract surgery , leaving the kamra implant in place . \n corneal topography remained stable after implantation of the kamra inlay and before cataract surgery ( figure 3 ) . \n the patient underwent uncomplicated phacoemulsification and intraocular lens implantation of the left eye under local anesthesia . \n a 2.60 mm temporal clear corneal incision was made , and a + 22.0 d monofocal tecnis zcb00 intraocular lens ( abbott medical optics , santa ana , ca , usa ) was implanted . \n no modifications were made to the surgical procedure , and the surgeon reported no difference in the ease of surgery with the kamra implant in place , apart from a few extra ocular rotations during capsulorhexis and phacoemulsification to improve visualization . \n one month after cataract surgery , posterior capsular opacification was noted in the left eye , and the patient underwent nd : yag laser capsulotomy 6 weeks after cataract surgery ( figure 1 ) . \n the patient reported good near , intermediate , and distance vision without the use of spectacles , and remains spectacle - independent . \n case 2 , a 62-year - old chinese male , underwent implantation of the kamra corneal inlay in the nondominant left eye on july 4 , 2007 . \n a 9.00 mm - diameter lamellar flap was created at a depth of 151 microns with a schwind microkeratome ( schwind , kleinostheim , germany ) . \n the patient s pre- and 1-month postoperative visual acuity results can be found in table 1 . \n four years after kamra implantation , cataract of grade nc2 ( locs iii ) was noted in the implanted left eye . \n nevertheless , vision in this eye remained good ; udva was 20/20 and unva was j2 . \n however , 6 years after kamra implantation , the patient felt that his distance vision in the left eye had deteriorated significantly ( table 1 ) and , on assessment , cataract of grade nc4 ( locs iii ) was noted . \n acutarget photographs confirmed that the implant was well centered ( inlay vs purkinje : 146 m x , 78 m y ) . \n various treatment options were presented to the patient , who opted for cataract surgery , leaving the kamra implant in place . \n the patient underwent phacoemulsification and intraocular lens implantation of the left eye under local anesthesia . \n a 2.60 mm temporal clear corneal incision was made , and a + 20.0 d monofocal tecnis zcb00 intraocular lens was implanted . \n no modifications were made to the surgical technique , and the surgeon reported no difference in ease of the surgery with the kamra implant in place , except that some extra ocular rotations during capsulorhexis and phacoemulsification to optimize the surgical view were required . \n surgical time was 15 minutes , and there were no intraoperative or early postoperative complications . \n the patient reported good near , intermediate and distance vision without the use of spectacles , and remains spectacle - independent . \n implantation of the kamra corneal inlay is an effective nonlens - based treatment option for presbyopia in patients with a clear lens at the time of presentation.6 however , some of these patients may eventually develop cataracts requiring surgical treatment . at this point \n , there are a multitude of available options , including cataract surgery with the kamra implant in place , implant removal followed by cataract surgery and subsequent re - implantation , and implant removal followed by cataract surgery with implantation of a multifocal or accommodative intraocular lens . \n cataract surgery with the kamra implant in place poses a few potential concerns , such as technical difficulty performing cataract surgery with the implant in place , the accuracy of biometry readings with various intraocular lens power formulae , and the visual outcome following cataract surgery . \n regarding the technical aspects of cataract surgery , no significant modifications to the surgical technique were required in either case , and the surgeon reported no significant difference in the ease of surgery ; surgical time was 15 minutes in both cases . \n the only slight difference was the need for some additional ocular rotations during capsulorhexis and phacoemulsification in order to improve surgical visualization . \n since both patients underwent local anesthesia , this was easily performed by asking them to voluntarily rotate the eye during the surgery . \n the authors feel that such rotations do not significantly increase the risk of intraoperative complications , such as radial tears or opening of the posterior capsule , as the surgical view through the 1.6 mm central aperture after rotation was very good and , as such , a 5 mm capsulorhexis and phacoemulsification were performed without difficulty \n . however , we acknowledge the experience of the surgeon performing the cataract operations in the patients in this series . therefore , cataract surgery with the kamra implant in place was not significantly more technically difficult to perform . \n the unva was j2 for both patients . corrected and unaided distance visual acuities varied from 20/20 to 20/40 ( table 1 ) . \n both patients were satisfied with their near , intermediate , and distance vision and remained spectacle - independent . \n the biometry readings and calculations were satisfactory and accurately predicted the achieved power ( table 2 ) . in this series , \n the surgeon used only one formula , srk / t , to calculate the power of the intraocular lenses . since there were no prior publications on this topic , an attempt was made , retrospectively , to determine if this choice of formula was reasonable . \n a comparison was made between four commonly used formulae for the calculation of intraocular lens power , to see which would have most accurately predicted the achieved power after surgery . \n the biometry readings for both cases were used for this purpose ( table 2 ) . in case 1 , using srk / t , a + 22.0 d monofocal tecnis zcb00 intraocular lens was implanted , aiming for a target power of 0.33 d. the achieved spherical equivalent was 0.38 d , ie , a difference of 0.05 d. the same biometry data were entered into three other commonly used formulae ( haigis , holladay 1 , and hoffer q ) , and the same + 22.0 d tecnis zcb00 lens was chosen . for haigis , \n the target power was 0.76 d , with a difference of 0.38 d from the achieved power of 0.38 d. for holladay 1 , the target power was 0.51 d ( a 0.13 d difference ) . \n hoffer q gave a target of 0.60 d , with a difference of 0.22 d from the achieved power . in case 1 \n , it appears that srk / t was the formula that most accurately predicted the achieved spherical equivalent . for case 2 , using srk / t , \n a + 20.0 d monofocal tecnis zcb00 intraocular lens was implanted , aiming for a target power of 0.72 d. the achieved spherical equivalent was 0.38 d , giving a difference of 0.34 d. using the same biometry parameters and the same + 20.0 d tecnis zcb00 lens , haigis gave a target of 0.56 d ( a difference of 0.18 d ) ; holladay 1 gave a target of 0.87 d , with a difference of 0.49 d ; and hoffer q gave a target of 1.00 d , with a difference of 0.62 d. in case 2 , srk / t was the second most accurate formula . \n therefore , based on these retrospective calculations , srk / t appears to be a reasonable choice of formula for choice of intraocular lens . however , this case series is too small to definitively establish the validity of srk / t in kamra implant patients . \n biometry readings and intraocular lens power calculations were not done prior to kamra implantation . however , in order to compare the biometry calculations before and after kamra implantation , the authors retrospectively calculated biometry for both cases before kamra implantation . \n the assumption was made that there was no change in axial length in either case . for case 1 , the pre - kamra k readings were 47.14 and 45.99 ; hence , using the same + 22.0 d lens would have given a target outcome of 0.46 d instead of 0.33 d. for case 2 , the pre - kamra k readings were 44.30 and 43.50 , and so using a + 20.0 d lens , as was done in case 2 , would have given a target outcome of 0.33 d instead of 0.72 d. therefore , implantation of the kamra inlay does not appear to have had a major effect on the biometry or intraocular lens calculations in these two patients . \n cataract surgery with the kamra implant in place is technically feasible with slight modifications to surgical technique . \n visual outcomes for both near and distance vision were satisfactory . additionally , the biometry readings were accurate , and srk / t appears to be an accurate formula for the prediction of eventual power . \n however , this case series is too small for drawing definitive conclusions , and larger studies with longer follow - up will be needed to properly evaluate the safety and visual outcome of cataract surgery with the kamra implant in place , as well as to determine the appropriate formula for calculation of intraocular lens power . \n nevertheless , cataract surgery with the kamra implant in place appears to be an attractive option for patients presenting with a cataract after previous kamra implantation for presbyopia correction .", "role": "user" }, { "content": "intrastromal corneal inlays are an emerging treatment for presbyopic patients . \n the kamra small aperture inlay was the first such inlay to receive conformit europenne ( ce ) marking in 2005 . \n it has been shown to improve uncorrected near and intermediate visual acuity without adversely affecting uncorrected distance visual acuity . due to the age of presbyopic patients \n , they may eventually develop cataracts . in two such cases \n , we found that cataract surgery with the kamra implant left in place was not technically more difficult , and that the surgical procedure could be improved by additional ocular rotations to improve visualization . \n biometry readings were reliable , and it appeared that the srk / t formula was accurate for calculation of intraocular lens power . \n cataract surgery with the kamra implant left in situ is a viable option for patients .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: although , iron is a vital element for the normal functioning of body organs ; excessive amounts in the body are very toxic , especially for liver and heart . \n iron - overload can result from the genetic disorder named hereditary hemochromatosis ( hh ) or secondary causes . \n hh is an autosomal recessive disorder that affects one in two hundred in caucasian people and higher prevalence in irish population . \n secondary iron overload is an evitable response to repeated transfusions as in thalassemia and sickle cell anemia or overdose of iron intake . \n iron chelators are mainstays of treatment for secondary hemochromatosis . since 1970 until now , the gold standard iron chelator is deferoxamine ( dfo ) that improve life expectancy of patients ; it must be administered for long - term by parenteral infusion for 5 - 7 days / week . \n regardless cumbersome administration , high cost and side - effects leading to poor patient compliance . \n more recently literature has emerged to discover more safe , efficient and affordable alternative chelator to treat patients with iron overload . \n diethylenetriaminepentaacetic acid ( dtpa ) is a member of a family of polyaminocarboxylic acid that is similar to ethylenediaminetetraacetic acid ( edta ) . \n this molecule widely used as a chelating agent to treat internal contamination of multivalent cations like : plutonium and americium [ figure 1 ] . \n the penta - anion dtpa is potentially an octadentate ligand , usually can form less than eight coordination bonds with divalent metal cations . \n currently , dtpa has been approved by the food and drug administration for chelating of transuranic elements including : plutonium , americium and curium . \n furthermore , dtpa can form complex with gd , in and tc in order to label chemical compounds to be used in nuclear medicine . \n structural formula of puchel , lipophilic derivative of diethylenetriaminepentaacetic acid used for plutonium decorporation the main drawback of this compound is impermeability to the cell membrane so that is classified as an extracellular fluid complexing agent . to overcome this handicap , \n for example , puchel is a lipophilic derivative of dtpa is used in removing intracellular plutonium or dtpa - conjugate compounds ( dtpa - folate [ figure 2 ] , dtpa - monoclonal antibody ) that increase cellular uptake of chelator through receptor mediated endocytosis . \n diethylenetriaminepentaacetic acid - folate structure , conjugated form of dtpa for tumor targeting - delivery recently , we reported that glucosamine - dtpa conjugate diethylenetriaminepentaacetic acid d - deoxy - glucosamine ( dtpa - dg ) [ figure 3 ] could be penetrated to inside the cancer cells easily . with this capability , \n chemical structure of diethylenetriaminepentaacetic acid - d - deoxy - glucosamine the aim of this study is to report that dtpa - dg complex can enhance clearance of intracellular free iron . \n the iron chelator dtpa - dg was synthesized in large scale using recently published method . \n various concentrations of dtpa - dg ( 20 , 40 , 60 , 80,100 mm ) were used in this investigation to evaluate the iron chelating capacity . \n dfo ( dfo ; novartis - pharma ) at 10 and 20 m concentration was used in parallel cultures for comparative purposes . \n the human hepatocellular carcinoma ( hepg2 ) obtained from american type culture collection ( atcc no : hb-8065 ) . \n hepg2 cells were grown in high glucose dulbecco 's modification of eagle 's medium ; gibco supplemented with 10% heat - inactivated fetal bovine serum , 1% glutamine , 10% penicillin / streptomycin and nahco3 ( mg ) . \n the hepatocytes were cultured in 75 cm sterile polystyrene flasks ( seromed / biochrom , germany ) in an incubator with saturating humidified atmosphere of 5% co2 in the air at 37c . for cell harvesting 0.05% \n the viability and count of cells were determined by 0.4% trypan blue exclusion using hemocytometer . \n iron overload was induced by adding 1 mm ferrous sulfate ( feso4 ) solution to the media for 72 h. cells were harvested and washed in pbs . to ensure removal of non - internalized iron that present in the media , cells were washed once in pbs , then in washing buffer including 500 m dfo ( dfo ; desferal ; novartis ) and 1.5 mm edta in pbs and in pbs buffer again . \n this method is one of the more practical ways to determine cytotoxicity of test agent . \n the principle of mtt assay is that yellow tetrazolium salt converted to blue formazan crystals by mitochondrial dehydrogenases of living cells . \n cells ( 3 10 ) were placed in a 96-well plate ( seromed / biochrom , germany ) and incubated for routinely 24 h at 37c prior to use . \n after 24 h , they were treated with various concentrations of dtpa - dg for 24 h. then remained media into wells were removed carefully and 100 l of 0.5 mg / ml mtt reagent was added to each well . \n the plate covered with aluminum foil and incubated for 2 h until formazan crystal is formed . \n finally , the formazan product in each well was dissolved in 100 l of dimethyl sulfoxide completely . \n the optical density of the 96-well plate was measured at 570 nm by microplate reader and the results were compared with untreated cell group . \n the total amount of free iron in the presence or absence of chelator was determined by perkin elmer flame atomic absorption spectrophotometer ( varian spectra \n iron loaded hepatocyte cultures incubated for 24 - 48 h in the presence of 20 , 40 , 60 , 80 , 100 nm dtpa - dg or 10 , 20 m dfo , then washed with isotonic buffer and subsequently lysed with glass beads , the lysate was centrifuged at 7000 rpm for 5 min . \n finally , iron chelating capacity of dtpa - dg was compared with the effect of dfo at several concentrations . \n data were expressed as mean standard deviation of three independent measurements ( n = 3 ) \n . statistical analysis of in - vitro data was performed using the spss ( version 19 , spss inc . , \n chicago , illinois , usa ) software package and prism ( version 5 , graphpad software inc . , san diego , california , usa ) software for determination of 50% inhibitory concentration value ( ic50 ) . \n measurement data were analyzed by student 's t - test and one - way anova followed by tukey 's post - hoc test . a value of p < 0.05 was considered as statistically significant . \n the iron chelator dtpa - dg was synthesized in large scale using recently published method . \n various concentrations of dtpa - dg ( 20 , 40 , 60 , 80,100 mm ) were used in this investigation to evaluate the iron chelating capacity . \n dfo ( dfo ; novartis - pharma ) at 10 and 20 m concentration was used in parallel cultures for comparative purposes . \n the human hepatocellular carcinoma ( hepg2 ) obtained from american type culture collection ( atcc no : hb-8065 ) . \n hepg2 cells were grown in high glucose dulbecco 's modification of eagle 's medium ; gibco supplemented with 10% heat - inactivated fetal bovine serum , 1% glutamine , 10% penicillin / streptomycin and nahco3 ( mg ) . \n the hepatocytes were cultured in 75 cm sterile polystyrene flasks ( seromed / biochrom , germany ) in an incubator with saturating humidified atmosphere of 5% co2 in the air at 37c . for cell harvesting 0.05% \n the viability and count of cells were determined by 0.4% trypan blue exclusion using hemocytometer . \n iron overload was induced by adding 1 mm ferrous sulfate ( feso4 ) solution to the media for 72 h. cells were harvested and washed in pbs . to ensure removal of non - internalized iron \n that present in the media , cells were washed once in pbs , then in washing buffer including 500 m dfo ( dfo ; desferal ; novartis ) and 1.5 mm edta in pbs and in pbs buffer again . \n this method is one of the more practical ways to determine cytotoxicity of test agent . \n the principle of mtt assay is that yellow tetrazolium salt converted to blue formazan crystals by mitochondrial dehydrogenases of living cells . \n cells ( 3 10 ) were placed in a 96-well plate ( seromed / biochrom , germany ) and incubated for routinely 24 h at 37c prior to use . \n after 24 h , they were treated with various concentrations of dtpa - dg for 24 h. then remained media into wells were removed carefully and 100 l of 0.5 mg / ml mtt reagent was added to each well . \n the plate covered with aluminum foil and incubated for 2 h until formazan crystal is formed . \n finally , the formazan product in each well was dissolved in 100 l of dimethyl sulfoxide completely . \n the optical density of the 96-well plate was measured at 570 nm by microplate reader and the results were compared with untreated cell group . \n the total amount of free iron in the presence or absence of chelator was determined by perkin elmer flame atomic absorption spectrophotometer ( varian spectra aa-220fs / australia ) . in practice \n , iron loaded hepatocyte cultures incubated for 24 - 48 h in the presence of 20 , 40 , 60 , 80 , 100 nm dtpa - dg or 10 , 20 m dfo , then washed with isotonic buffer and subsequently lysed with glass beads , the lysate was centrifuged at 7000 rpm for 5 min . \n finally , iron chelating capacity of dtpa - dg was compared with the effect of dfo at several concentrations . \n data were expressed as mean standard deviation of three independent measurements ( n = 3 ) \n . statistical analysis of in - vitro data was performed using the spss ( version 19 , spss inc . , \n chicago , illinois , usa ) software package and prism ( version 5 , graphpad software inc . , san diego , california , usa ) software for determination of 50% inhibitory concentration value ( ic50 ) . \n measurement data were analyzed by student 's t - test and one - way anova followed by tukey 's post - hoc test . a value of p < 0.05 was considered as statistically significant . \n the experimental model obtained by the addition of 1 mm feso4 to the culture media for 72 h. this procedure is in agreement with perak 's ( 2006 ) findings , which showed this iron amount can be tolerable for hepg2 cells in short - term of cultivation . following 72 h iron exposure duration , \n total intracellular iron was increased from 16 g fe / g of wet cell pellet to 98 g fe / g . \n in fact , there is increased 6-fold in iron level [ figure 4 ] . cellular iron level in control and iron - overload human hepatocellular carcinoma cell model as measurement by atomic absorption spectrometry . \n measurements were performed with sample triplicates ( n = 3 ) mtt assay of cytotoxicity associated with various concentrations of dtpa - dg after 48 h demonstrated a negligible ( p > 0.05 ) decrease of cell viability observed by mtt test [ figure 5 ] . \n mtt assay of cytotoxicity associated with various concentrations of diethylenetriaminepentaacetic acid - d - deoxy - glucosamine after 48 h. data are presented as the percentage of cell viability relative to the untreated normal human hepatocellular carcinoma ( mean standard deviation , n = 3 ) . as a result , a negligible ( p > 0.05 ) decrease of cell viability observed by mtt test in order to demonstrate the ability of chelator to decrease intracellular free iron content , the amount of free iron in the presence of several concentrations of dtpa - dg was quantified by using atomic absorption spectrophotometer . \n exposure of iron overload hepg2 cell model for about 24 h to different concentrations of dtpa - dg ( 20 - 100 nm ) dramatically decreases free iron level [ figure 6 ] . \n the lowest cellular iron content was observed with the concentration 100 nm that approximately reached as much as control cell group ( p < 0.05 ) . for concentration higher than 40 \n nm dtpa - dg significant differences ( p < 0.05 , p = 0.022 ) versus untreated cell group were observed ; can prove successful dtpa - dg 's function as intracellular iron chelators in the low concentration range . \n the comparison of chelator efficacy with dfo exhibited that the highest rate of decrease in the iron content from hepg2 cells occurred during 24 h treatment with dtpa - dg . \n the rate of decrease was significantly ( p < 0.05 , p = 0.011 ) higher in cell groups in which 60 nm dtpa - dg were injected than those of 20 m dfo [ figure 7 ] . \n a dose - dependent decrease of free iron content after 24 h incubation of human hepatocellular carcinoma cells at 37c which treated with 20 - 100 nm of diethylenetriaminepentaacetic acid - d - deoxy - glucosamine . \n the decrease was significant ( p < 0.05 ) at concentration equal to or higher than 40 nm of dtpa - dg in comparison with untreated cells ( n = 3 ) . \n results are the mean standard deviation of three separate experiments the comparison of two chelators diethylenetriaminepentaacetic acid - d - deoxy - glucosamine and deferoxamine efficacy in similar conditions after 24 h indicated statistically ( p < 0.05 ) a higher reducing of cellular iron content with dtpa - dg than with dfo at much lower doses . \n results are the mean standard deviation of three separate experiments ( n = 3 ) as a result of copper intracellular assay experiments , reducing in free iron level was significantly correlated ( p < 0.05 , p = 0.001 , r = 0.994 ) with iron chelator concentration [ figure 8 ] . \n dtpa - dg was the most effective at decreasing free iron content in hepg2 cell line with an ic50 value near 65.77 nm . \n a significant ( p < 0.05 , r = 0.994 ) correlation with iron chelator dose and its success in reducing cellular iron content is shown in this figure . \n diethylenetriaminepentaacetic acid d - deoxy - glucosamine at concentration of 65.77 nm has been able to reduce 50% of iron deposits \n the experimental model obtained by the addition of 1 mm feso4 to the culture media for 72 h. this procedure is in agreement with perak 's ( 2006 ) findings , which showed this iron amount can be tolerable for hepg2 cells in short - term of cultivation . following 72 h iron exposure duration , \n total intracellular iron was increased from 16 g fe / g of wet cell pellet to 98 g fe / g . \n in fact , there is increased 6-fold in iron level [ figure 4 ] . cellular iron level in control and iron - overload human hepatocellular carcinoma cell model as measurement by atomic absorption spectrometry . \n mtt assay of cytotoxicity associated with various concentrations of dtpa - dg after 48 h demonstrated a negligible ( p > 0.05 ) decrease of cell viability observed by mtt test [ figure 5 ] . \n mtt assay of cytotoxicity associated with various concentrations of diethylenetriaminepentaacetic acid - d - deoxy - glucosamine after 48 h. data are presented as the percentage of cell viability relative to the untreated normal human hepatocellular carcinoma ( mean standard deviation , n = 3 ) . as a result , a negligible ( p > 0.05 ) decrease of cell viability observed by mtt test \n in order to demonstrate the ability of chelator to decrease intracellular free iron content , the amount of free iron in the presence of several concentrations of dtpa - dg was quantified by using atomic absorption spectrophotometer . \n exposure of iron overload hepg2 cell model for about 24 h to different concentrations of dtpa - dg ( 20 - 100 nm ) dramatically decreases free iron level [ figure 6 ] . \n the lowest cellular iron content was observed with the concentration 100 nm that approximately reached as much as control cell group ( p < 0.05 ) . for concentration higher than 40 \n nm dtpa - dg significant differences ( p < 0.05 , p = 0.022 ) versus untreated cell group were observed ; can prove successful dtpa - dg 's function as intracellular iron chelators in the low concentration range . \n the comparison of chelator efficacy with dfo exhibited that the highest rate of decrease in the iron content from hepg2 cells occurred during 24 h treatment with dtpa - dg . \n the rate of decrease was significantly ( p < 0.05 , p = 0.011 ) higher in cell groups in which 60 nm dtpa - dg were injected than those of 20 m dfo [ figure 7 ] \n . a dose - dependent decrease of free iron content after 24 h incubation of human hepatocellular carcinoma cells at 37c which treated with 20 - 100 nm of diethylenetriaminepentaacetic acid - d - deoxy - glucosamine . \n the decrease was significant ( p < 0.05 ) at concentration equal to or higher than 40 nm of dtpa - dg in comparison with untreated cells ( n = 3 ) . \n results are the mean standard deviation of three separate experiments the comparison of two chelators diethylenetriaminepentaacetic acid - d - deoxy - glucosamine and deferoxamine efficacy in similar conditions after 24 h indicated statistically ( p < 0.05 ) a higher reducing of cellular iron content with dtpa - dg than with dfo at much lower doses . \n results are the mean standard deviation of three separate experiments ( n = 3 ) as a result of copper intracellular assay experiments , reducing in free iron level was significantly correlated ( p < 0.05 , p = 0.001 , r = 0.994 ) with iron chelator concentration [ figure 8 ] . \n dtpa - dg was the most effective at decreasing free iron content in hepg2 cell line with an ic50 value near 65.77 nm . \n a significant ( p < 0.05 , r = 0.994 ) correlation with iron chelator dose and its success in reducing cellular iron content is shown in this figure . \n diethylenetriaminepentaacetic acid d - deoxy - glucosamine at concentration of 65.77 nm has been able to reduce 50% of iron deposits \n in this study , iron - clearing efficiency and safety profile of dtpa - dg was assessed using iron overload hepg2 cell culture model . \n this model was chosen because the liver is primary iron storage site and hepg2 are well - established in - vitro model for human hepatocytes . \n recently , we have reported that linking of glucosamine through its 2-amino group to dtpa increased remaining of dtpa - dg into cells , which is mechanistically when dtpa - dg enters the cell after 6-phosphorilation by the hexokinase enzyme , is not metabolized further ; then dtpa more efficiently reduces intracellular iron in the iron loaded hepg2 cells . \n dtpa - dg was found to be much more efficient at iron decorporation than dfo in hepg2 cell line ; scilicet , the chelator has shown a significant effect at lower doses . because glucose receptor mediated endocytosis pathway \n have shown that when glucose is linked to a molecule through c2 position , its affinity for cell surface receptor does nt alter essentially . \n so , dtpa - dg can be transported through glucose channels with similar manner as unconjugated glucose . \n another experiment was mtt colorimetric assay that is a suitable technique for determining cell viability after drug treatment . \n our finding about hepg2 cell cultures were subjected to the mtt assay after 48 h indicated no significant detrimental effect of dtpa - dg on cell viability at concentration 20 - 100 nm . in conclusion , \n actually , the concentration of compound that is effective to reduce up to 50% of iron deposits was calculated by prism software . in 1963 \n , in which they claimed that long - term use of dtpa could be a possible method for eliminating excessive iron in the urine . \n the current study defines that glucose - targeted delivery of dtpa will be more successful in minimizing accumulation of iron than dtpa by itself . while future study with more focus on pharmacodynamics and pharmacokinetic of dtpa - dg will be required . \n based on a pressing need to design a new iron - chelating agent , this project was undertaken to evaluate the efficacy of a new dtpa - dg iron chelator in iron overload hepg2 cell model . \n the stark decrease of iron deposits showed the high efficiency of dtpa - dg in iron decorporation from the main target of iron overload . \n we found that the glycosyl moiety facilitates entrance of dtpa into cells without any iron interaction capabilities . \n it was also shown dtpa - dg is a non - toxic compound that can be rescue cells from deleterious effects of free iron . \n finally , this new cell - permeable iron chelator can be decreased tissue iron deposits more effectively than the known chelator dfo . \n these promising results encourage us to continue more experiments on dtpa - dg complex for treatment of iron overload disease and animal experiments that will be published in near future .", "role": "user" }, { "content": "objective : to find out whether dtpa - dg complex can enhance clearance of intracellular free iron.materials and methods : diethylenetriaminepentaacetic acid - d - deoxy - glucosamine ( dtpa - dg ) was synthesized and examined for its activity as a cell - permeable iron chelator in human hepatocellular carcinoma ( hepg2 ) cell line exposed to high concentration of iron sulfate and compared with deferoxamine ( dfo ) , a prototype iron chelator . \n the effect of dtpa - dg on cell viability was monitored using the 3-(4,5-dimethythiazol-2-yl)-2,5-diphenyl tetrazolium bromide mtt assay as well.results:there was a significant increase of iron level after iron overload induction in hepg2 cell culture . \n dtpa - dg presented a remarkable capacity to iron burden reducing with estimated 50% inhibitory concentration value of 65.77 nm . \n in fact , glycosyl moiety was gained access of dtpa to intracellular iron deposits through glucose transporter systems.conclusion:dtpa-dg , more potent than dfo to sequester deposits of free iron with no profound toxic effect . \n the results suggest the potential of dtpa - dg in chelating iron and permitting its excretion from primary organ storage .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: biochemical and structural studies conducted in the past two decades have established the general mechanism by which type-2a topoisomerases transport , in an atp - dependent manner , one dna duplex through a transient double - strand break produced in another ( 14 ) . \n all the enzymes of this class ( bacterial dna gyrase , bacterial topoisomerase iv and eukaryotic topoisomerase ii ) are functional dimers of three structural domains : an atp - ase or n - terminal domain , and the b and a domains that constitute the dna cleavage - rejoining core ( 3 ) . before interacting with dna , \n type-2a topoisomerases have the shape of an open clamp , in which the n - terminal domains are located at the tips and the a domains dimerize at the hinge ( 5,6 ) . the dna double helix \n to be gated ( named g - segment ) first enters the open clamp and binds to the cleavage - rejoining core ( 7 ) . \n binding of atp causes the n - terminal domains to interact and the clamp closes . \n if this closure leads to the capture of a second dna double helix inside the clamp , a cascade of conformational changes ensues ( 8) . the captured dna ( named t - segment ) is transported through a gate transiently opened in the bound g - segment . for this process , the g - segment is cleaved by two trans - estherification reactions that let the 5 ends of the severed strands covalently linked with a pair of tyrosine residues , one in each half of the topoisomerase ( 9,10 ) . after crossing the g - segment , \n the t - segment reaches the cavity delimited by the a domains and it is then expelled from the topoisomerase through the side opposite the one it entered , by the aa dimer interface at the hinge of the closed clamp ( 8,11 ) . \n atp hydrolysis begins during t - segment transport and release of the hydrolytic products allows the n - terminal domains to come apart leaving the enzyme ready for a new reaction cycle . \n one of them is the fine coupling between inter - domainal movements and atp usage during dna transport ( 1,4 ) . \n the translocation of the t - segment through the complete dimer interface of type-2 topoisomerases demands energetically well - balanced and robust conformational states during the critical dna cleavage , dna gating and dna rejoining steps . \n backtracking of the t - segment would cause unnecessary breakages of the g - segment and no net dna transport . \n dissociation of the enzyme halves linked to cleaved dna would cause chromosomal breaks and rearrangements . \n the other intriguing aspect of type-2a topoisomerases are the dna transport preferences that distinguish some enzymes . on the one side , \n dna gyrase is specialized to carry out intra - molecular transport . after binding the g - segment , the gyrase \n dna complex enforces the juxtaposition of a contiguous t - segment such that it configures a positive dna crossing ( 12,13 ) . as long as this configuration can be settled , dna gyrase removes positive supercoils and introduce negative ones into circular dna ( 14 ) . on the other side , bacterial topoisomerase iv and eukaryotic topoisomerase ii can carry out both intra- and inter - molecular dna transport . \n decatenate , knot unknot and remove supercoils . these enzymes , however , can not supercoil dna alike gyrase . instead , topoisomerase iv and topoisomerase ii reduce the topological complexity of dna ( catenation , knotting and supercoiling ) below the levels expected at thermal equilibrium ( 15 ) . \n several models have been proposed to explain how they assess the global topology of a dna molecule by their local interactions with dna segments ( 16,17 ) . in parallel to biochemical and structural studies on type-2a topoisomerases , \n the biological functions of this abundant and essential class of enzymes had also been the focus of attention for long time ( 18 ) . in eubacteria , dna gyrase and topoisomerase iv complete different tasks according to their distinct dna transport preferences . in most eukaryotes , however , a single form of topoisomerase ii appears to carry out diverse biological roles ( 18,19 ) . \n the major known and essential function of the enzyme is the decatenation of replicated dna duplexes after the s phase . \n when two replication forks converge , the last unreplicated segment becomes too short to be unlinked by topoisomerase i or ii . \n the residual intertwines between the parental dna are then converted to catenane links between the newly replicated daughter duplexes . \n removal of such catenanes by topoisomerase ii is essential for proper chromosome segregation ( recent studies on this topic are refs . \n another major known cellular activity of topoisomerase ii is the relaxation of positive supercoils generated ahead of the replication forks , as well as of oppositely supercoiled domains generated during dna transcription ( 23,24 ) . \n although eukaryotic topoisomerase i can also fulfil this function , topoisomerase ii is likely to be the main relaxase of dna supercoils inside the cell ( 25 ) . \n in addition to decatenating replicated dna and relaxing polymerase - driven supercoils , eukaryotic topoisomerase ii has been implicated in many other chromosomal processes . \n a common trait in all these processes is that the topological transformation ( if any ) catalysed by the enzyme is uncertain . \n for instance , topoisomerase ii is well known to be involved in chromosome condensation , but its precise role in the many steps for assembling morphologically distinct interphase chromosomes and highly condensed metaphase chromosomes remain unclear . at each of these steps , whether a continual dna transport activity of topoisomerase ii is required for chromatin condensation - decondensation or whether the enzyme mainly has a structural role are still controversial questions ( 26 ) . \n other examples of unknown topology conversion are the implications of topoisomerase ii in the local regulation of chromatin architecture and in gene expression . such biological roles became evident after discovering that mammals harbour two topoisomerase ii isozymes , ii and ii ( 27 ) . \n both isozymes carry out similar strand - passing activities in vitro ( 28 ) , though ii is found to relax positively supercoiled plasmids faster than negatively supercoiled ones ( 29 ) . \n both isozymes are differentially regulated during cell growth and can not compensate for each other in vivo ( 3032 ) . \n on one hand , ii is only found in proliferating cells , with an expression peak at late s and g2/m phases of the cell cycle ( 33,34 ) , and it is most likely involved in dna replication , chromosome condensation decondensation and sister chromatid segregation ( 3537 ) . \n these functions are quite analogous to those known for the single topoisomerase ii form found in budding yeast , fission yeast and drosophila ( 3841 ) . on the other hand , \n ii is found in all mammalian cell types but with prominent expression in terminally differentiated cells ( 4247 ) . \n the enzyme binds to the 5 region of a number of such ii-sensitive genes ( 48 ) . \n topoisomerase ii has been also found catalysing a site - specific transient double - stranded dna break in some gene promoters , which lead to local changes of chromatin architecture and activation of transcription ( 49 ) . \n these findings suggest that topoisomerase ii , and perhaps the single topoisomerase ii form found in yeasts and drosophila , produce critical structural changes in localized chromatin regions to regulate genome transactions . to find out the range of dna manipulations that topoisomerase ii might catalyse in vivo , it is essential to sum up the capabilities of the enzyme inferred from in vitro studies . \n some topoisomerase ii activities learned with dna in free solution that might be relevant for the performance of the enzyme in the context of chromatin architecture and dynamics are surveyed below . \n most studies on topoisomerase ii mechanism had focussed on the dna cleavage and gating steps , the dna transport preferences , and the coupling of inter - domainal movements to atp usage . however , a less - attended but critical aspect of the enzyme operation that needs to be absolutely effective is the rejoining of the gated dna . \n consider , for example , yeast cells deficient in topoisomerase i , in which topoisomerase ii alone relaxes the supercoils raised during dna transcription and reduces to zero the linkage between complementary dna strands during their replication . \n any single dna gating event not followed by the proper rejoining step will put at risk genome integrity and cell survival . \n it is reasonable to assume then that most mechanic traits of the enzyme are tuned to ensure rejoining rather than dna cleavage and dna transport . \n not surprisingly , those compounds affecting the dna - rejoining step readily produce cell death and have , therefore , prominent pharmacological interest . \n but apart from biochemical targeting , many physiological processes might also threaten the dna rejoining step . \n for instance , in contrast to dna in free solution , intracellular dna is subjected to considerable twisting , bending and pulling forces generated by dna tracking ensembles and the conformational dynamics of chromatin fibers . in this environment \n yet , the mechanism of the type-2 topoisomerases implies that none of its dimer interfaces is a permanent interaction . \n the path of the t - segment involves the sequential crossing of the entrance or n - gate , the dna gate and the exit or c - gate . \n coordination among these three gates is therefore critical to preclude the risk of the two halves of the enzyme coming apart while gating the dna . in that regard , \n biochemical studies with yeast topoisomerase ii indicate that gate opening and closure follow a double - lock rule : a given gate can open only if the other two are closed ( 50 ) . \n although the n - gate domain is not required for g - segment cleavage , the dna gate per se is not able to widen unless atp binds and closes the n - gate ( 50 ) . \n since a captured t - segment can not be held in the inter - domainal region between the n - gate and the dna gate , closure of the n - gate and consequent capture of a t - segment is likely to enforce the opening of the dna gate ( 50 ) . \n once a t - segment has reached the central cavity of the enzyme and before the c - gate opens , the gated g - segment dna can be rejoined ( 11,50 ) . \n hence , opening of the c - gate is likely to be triggered by the entrapment of the t - segment after being squeezed by the sequential closure of the n - gate and the dna gate . \n these inter - domainal couplings maximize the dimer stability , ensure a very transient dna gating state and enforce unidirectional dna transport by precluding the backtracking of the t - segment ( figure 1a ) . \n figure 1.mechanical couplings for gate opening and closure in type-2 topoisomerases . \n ( a ) in type-2a enzymes ( bacterial gyrase , topoisomerase iv , topoisomerase ii ) , the n - gate , dna gate and c - gate appear to be mechanically coupled with a double - lock rule , such that a given gate can open only if the other two are closed . \n this coordination minimizes the risk of the two enzyme halves from coming apart while gating the dna . \n ( a ) when the entrance n - gate is open , dna might be cleaved but the dna gate is not able to widen . \n ( b ) upon atp binding and closure of the n - gate , a captured t - segment can not be held in the inter - domainal region between the n - gate and dna gate . \n the t - segment quickly crosses the dna - gate and reaches the central cavity of the enzyme . \n ( d ) the consequent entrapment of the t - segment enforces its exit by a transient opening of the c - gate . \n ( b ) in type-2b enzymes ( topoisomerase vi ) there is no c - gate . \n ( a ) when the entrance n - gate is open ( no atp bound ) the dna gate is locked . \n ( b ) upon atp binding and closure of the n - gate , a captured t - segment is held in the central cavity of the enzyme prior to the aperture of the dna gate . \n ( c ) the captured t - segment crosses then the dna - gate and exits the complex . \n figure 2.topoisomerase ii might discern whether or not a dna domain is suitable for successful rejoining by bending the g - segment before the gating step . ( a ) proper bending of the interacting duplex will be possible as long as the stretching tension along the dna does not exceed a threshold value ( o ) . \n this threshold is determined by the tension ( p ) that the enzyme is able to counteract to close the dna gate . \n ( b ) the active deformation of dna , coupled to atp binding and hydrolysis , could operate then as a checkpoint to unlock the dna - gate , as well as a gain of elastic energy to be delivered in the rejoining step . \n ( a ) in type-2a enzymes ( bacterial gyrase , topoisomerase iv , topoisomerase ii ) , the n - gate , dna gate and c - gate appear to be mechanically coupled with a double - lock rule , such that a given gate can open only if the other two are closed . \n this coordination minimizes the risk of the two enzyme halves from coming apart while gating the dna . \n ( a ) when the entrance n - gate is open , dna might be cleaved but the dna gate is not able to widen . \n ( b ) upon atp binding and closure of the n - gate , a captured t - segment can not be held in the inter - domainal region between the n - gate and dna gate . \n the t - segment quickly crosses the dna - gate and reaches the central cavity of the enzyme . \n ( d ) the consequent entrapment of the t - segment enforces its exit by a transient opening of the c - gate . \n ( b ) in type-2b enzymes ( topoisomerase vi ) there is no c - gate . \n ( a ) when the entrance n - gate is open ( no atp bound ) the dna gate is locked . \n ( b ) upon atp binding and closure of the n - gate , a captured t - segment is held in the central cavity of the enzyme prior to the aperture of the dna gate . \n ( c ) the captured t - segment crosses then the dna - gate and exits the complex . \n topoisomerase ii might discern whether or not a dna domain is suitable for successful rejoining by bending the g - segment before the gating step . \n ( a ) proper bending of the interacting duplex will be possible as long as the stretching tension along the dna does not exceed a threshold value ( o ) . \n this threshold is determined by the tension ( p ) that the enzyme is able to counteract to close the dna gate . \n ( b ) the active deformation of dna , coupled to atp binding and hydrolysis , could operate then as a checkpoint to unlock the dna - gate , as well as a gain of elastic energy to be delivered in the rejoining step . \n the quaternary couplings during dna transport in topoisomerase ii must be alike in topoisomerase iv and dna gyrase . \n however , they might differ from those in the less - extensively studied type-2b topoisomerases ( archea topoisomerase vi and plant topoisomerase vi ) ( 51,52 ) . \n these enzymes share several mechanistic features with the type-2a topoisomerases , but there are significant structural differences between the two families ( 53 ) . for t - segment capture , the atpase domains of type-2b dimerize upon atp binding alike the n - gate in type-2a enzymes ( 54 ) . \n the dna cleavage - rejoining core of type-2b also contains counterparts of the structural motifs of the dna - gate of the type-2a enzymes . \n however , type-2b enzymes do not have an exit- or c - gate ( 55 ) . \n the transported duplex is outside the complex right after crossing the dna gate ( figure 1b ) . \n therefore , dimer stability during the dna gating step relies mostly in the closure of the n - gate . type-2a and \n type-2b topoisomerases should have then distintc couplings between their atpase and dna cleavage - rejoining modules . \n remarkably , contrary to type-2a topoisomerases ( 50 ) , the atpase domains of type-2b enzymes are required for dna cleavage ( 56 ) and the cavity between the two gates of these enzymes can accommodate a captured t - segment before widening the dna gate ( 55 ) . \n the absence of a c - gate in type-2b topoisomerases facilitates perhaps the dissociation of the enzyme halves for some biological tasks . \n for instance , reduced dimer stability could be operational in spo11 , a type-2b related enzyme that mediates dna double - strand breaks during recombination ( 51,57 ) . \n aside of ensuring dimer stability during the dna cleavage and gating steps , another way to guarantee genome integrity is to not cleave the dna in the first place . \n it is conceivable that topoisomerase ii discerns whether or not a duplex is suitable for successful rejoining before gating it . for example , if dna is undergoing extreme tension , its conformation might not be appropriate for catalytic cleavage . in that regard , \n the recently solved crystal structure of the cleavage - rejoining core of yeast topoisomerase ii bound to dna , reveals a complex producing a 150 dna bend in the g - segment ( 58 ) . a sharp bending of the bound g - segment \n had previously been inferred for bacterial topoisomerase iv and it is postulated to contribute to the capacity of type-2a topoisomerases to simplify dna equilibrium configurations ( 59 ) . \n bending of the g - segment might be a prerequisite for dna cleavage , thus protein conformational changes that accompany this deformation create a bipartite catalytic site , in which each dna strand gets close to a reactive tyrosine and a coordinated magnesium ion ( 58 ) . \n another possible function of such dna bending is that it would prevent topoisomerase ii from gating the duplex in regions undergoing severe stretching forces ( figure 2a ) . \n recently , oligonucleotides with a pair of fluorophores straddling the dna cleavage site have been used to observe the dynamics of the dna gate by fret ( 60 ) . \n the experiment revealed that during steady - state atp hydrolysis fluorophores repeatedly move over a distance of 20 with comparable back and forth rate constants . whether these displacements reflected opening and \n first , the putative open and closed states are nearly equally probable in a steady state population , whereas all previous biochemical data indicate that in this condition only a small fraction of complexes is in the open dna - gate state . \n second , as mentioned above , the capture of a t - segment is likely required to enforce the opening of the dna gate . in the fret experiment , however , no free dna was available to provide a t - segment . \n thus , the observed displacements could reflect mostly bending or twisting distortions of the g - segment coupled to atp binding and hydrolysis but with no significant widening of the dna gate . \n such active deformation of dna could operate as a checkpoint to unlock the dna - gate , as well as a stock of elastic energy to deliver at the rejoining step . by these means , \n topoisomerase ii could discern whether a duplex is suitable for proper rejoining before gating it ( figure 2b ) . \n identification of topoisomerase ii as one of the main components of the nuclear matrix and metaphase chromosome scaffold ( 6163 ) , as well as its preferential interaction to sar and mar sequences ( 6466 ) , had lead to the proposal of an structural role for the enzyme . \n the possibility that topoisomerase ii could be anchoraging dna chromosomal loops fit well with the enzyme capabilities to hold dna through covalent bonds and to interact simultaneously with several dna segments ( 67,68 ) . \n this putative structural role was further supported by the discovery of that , upon binding of a non - hydrolysable atp analogue , the topoisomerase constituted in vitro a high - salt resistant complex with circular dna molecules ( 7,69,70 ) . \n this observation was essential for unravelling the mechanism of type-2 topoisomerases , reflecting the closure of the n - gate and the subsequent impossibility of the protein ring detaching from a bound g - segment in the absence of a dna free - end . \n the topoisomerase dna complex was resistant to salt concentrations up to 5 m nacl or cscl during several hours ( 71 ) . \n therefore , such a topological interaction was an attractive mechanism to fasten large chromosomal dna loops , and it seemed more convenient than holding the duplex with covalent links that implied a permanent cleavage of dna . in the last decade , however , support for the proposed structural role of topoisomerase ii had faded since some observations disputed the requirement of the enzyme to maintain the shape of mitotic chromosomes . although topoisomerase ii is required for mitotic chromosome assembly in vitro , the enzyme could be extracted from in vitro condensed chromosomes with no significant changes in the chromosome conformation ( 72 ) . \n other in vivo studies , which used fluorescently tagged human dna topoisomerases ii and ii , indicated that neither enzyme is an immobile structural component of the chromosomes ( 73 ) . \n moreover , several studies reported significant mitotic chromosome condensation even after genetic knockdown or knockout of topoisomerase ii ( 30,7476 ) . yet , other demonstrations indicated that topoisomerase ii remains stably bound in chromosomes assembled in vitro after a round of dna replication ( 77 ) . \n more recent analyses in several species including mammals , chicken , c. elegans and drosophila , came to agree in that topoisomerase ii is not required for initial chromatin compaction and formation of a longitudinal chromosome axis , but that an axial distribution of the enzyme in late prophase to metaphase is real , being dependent on the presence of condensins ( 76,78,79 ) . on the other side , \n the existence of a rigid chromosome scaffold , in which dna loops could be anchored , is also no longer sustained after some morphological and biophysical observations . \n for instance , depletion of histone h1 results in longer and thinner compacted mitotic chromosomes , and so arguing against a stiff scaffolding structure driving chromosome condensation ( 80 ) . also , the elastic response of native metaphase chromosomes and its alteration by endonucleases rule out a contiguous internal non - dna ( e.g. protein ) scaffold . \n the elastic behaviour of mitotic chromosomes suggests instead that they are shaped by isolated dna - crosslinking elements spaced by few kilo base pairs ( 81,82 ) . from the above , the possibility that a highly stable topoisomerase ii dna complex plays a structural role in chromosome architecture can not be excluded . \n the closure of the enzyme around dna is driven by atp binding and this conformation could be easily stabilized by some cellular factor . such factor could interact across the closed n - gate domains , thus behaving like a molecular padlock . essentially , this is the case with bisdioxopiperazines , such as icrf-193 , that bind to the interface of the dimerized atp - ase domains of topoisomerase ii and preclude n - gate reopening ( 83,84 ) . \n the hypothetic existence of gate padlocks could be also considered for the c - gate of topoisomerase ii . in this case , stabilisation of the exit gate would be analogous to locking it by means of engineered disulfide bonds ( 11,50 ) . \n in such a case , a t - segment would be captured , transported across the gated g - segment and end up entrapped in the central chamber of the topoisomerase between the rejoined dna - gate and the locked exit - gate . \n if padlocks close both the c - gate and the n - gate simultaneously , the topoisomerase would become a tight toroidal structure , inside which two dna segments could be entrapped . \n this extreme can be observed in vitro by closing the n - gate with a non - hydrolysable atp analogue in the yeast topoisomerase ii mutant having the c - gate locked by disulfide bonds ( 50 ) . in this condition , \n analogously to bisdioxopiperazines , such as icrf-193 that inhibit topoisomerase ii activity , closure of the n - gate of topoisomerase ii upon atp binding can be stabilized also by some cellular factor . \n ( a ) such gate padlocks could operate as inhibitors of enzyme binding to chromosomal dna . \n ( b ) if dna is already bound to the topoisomerase , an n - gate padlock would produce a high salt resistant complex , which might serve to regulate the enzyme activity or operate as a structural element for dna organization . \n if both the n - gate and the c - gate were stabilized in the closed conformation , the topoisomerase could clamp two dna duplexes , the g - segment and passed t - segment . \n analogously to bisdioxopiperazines , such as icrf-193 that inhibit topoisomerase ii activity , closure of the n - gate of topoisomerase ii upon atp binding can be stabilized also by some cellular factor . ( a ) such gate padlocks could operate as inhibitors of enzyme binding to chromosomal dna . \n ( b ) if dna is already bound to the topoisomerase , an n - gate padlock would produce a high salt resistant complex , which might serve to regulate the enzyme activity or operate as a structural element for dna organization . \n if both the n - gate and the c - gate were stabilized in the closed conformation , the topoisomerase could clamp two dna duplexes , the g - segment and passed t - segment . \n dna complexes for structural purposes , another plausible function of gate padlocks could be the regulation of the enzyme activity , a subject that remains poorly understood . \n topoisomerase ii can be phosphorylated in vitro on multiple serine and threonine residues , the majority of which are located in its non - conserved c - terminal region . \n the plausible effects of phosphorylation in vivo remain controversial , since most studies report that these modifications do not produce significant changes in the dna transport activity of the enzyme in vitro ( 85 ) . \n topoisomerase ii can also undergo poly(adp - ribosyl)ation in vitro . whether this modification occurs in vivo and has physiological significance \n it is suggested that poly(adp - ribosyl)ation may play a role in the cellular distribution and degradation the enzyme ( 87 ) . \n more recently , topoisomerase ii has been shown to undergo sumo - lation during mitosis ( 8890 ) . \n this modification seems to be a signal for targeting the enzyme to pericentromeric regions of mitotic chromosomes and promotes the decatenation of sister chromatids at anaphase ( 8890 ) . \n the question then is what might prevent intracellular topoisomerase ii to hydrolyse atp , to cleave dna or to transport dna when it is not necessary . \n gate padlocks could be an effective way to turn - off these enzyme activities . as observed with the topoisomerase inhibitor icrf-193 , locking of the n - gate upon atp binding \n can occur in the absence of dna ( 83 ) . in this closed conformation , \n gate padlocks could therefore be used to transport and stock topoisomerase ii inside the cell and let it operate only where needed . \n so far , no gate padlock has been found in cell extracts or described among the several proteins that might physically interact with topoisomerase ii . \n another reason , however , is that proper assays to find them have not been conducted . \n once topoisomerase ii is bound to a g - segment , dna transport depends solely on the probability of finding a potential t - segment properly positioned in the vicinity of the n - gate of the enzyme ( 91 ) . for naked dna molecules in free solution , such juxtaposition probability will be determined by the global configuration of the duplex , and also by constrains that the topoisomerase might produce by its local interactions with dna . \n this last aspect should account for the enzyme capacity to drive dna topology below thermal equilibrium configurations and for its chiral preferences ( 92,93 ) . \n the question now is whether our current understanding of juxtaposition probability of dna segments on topoisomerase ii , as inferred from in vitro studies , is applicable to intracellular dna . \n although the topological problems inside the cell are equivalent to those portrayed in vitro , the conformational dynamics of dna is determined by a complex set of variables involving multiple molecular interactions , in addition to the topology of the duplex . \n for example , although in living yeast cells both dna topoisomerases i and ii are capable to relax plasmid minichromosomes ( 94 ) , in vitro studies revealed that topoisomerase ii relaxes nucleosomal dna much faster than topoisomerase i ( 25 ) . \n apparently , the dna cross - inversion mechanism of topoisomerase ii is facilitated in nucleosome fibres that might favour the juxtaposition of dna segments , whereas the dna strand - rotation mechanism of topoisomerase i might be stalled by twist diffusion barriers imposed by nucleosome organization . \n how chromatin affects dna segment juxtaposition and the consequent topoisomerase ii activity is complex to envisage . on one hand \n , juxtaposition can be favoured as dna is intensely and extensively more folded than in free solution . \n on the other hand , the number of dna segments suitable for topoisomerase ii manipulation is more reduced since many proteins cover most of the dna length . to further dissect this problem , \n three types of dna juxtaposition should be considered to occur in the intracellular landscape , each one producing a different outcome upon dna transport by topoisomerase ii . \n this is the case of the pre - catenanes and catenane links generated between newly replicated duplexes and the supercoils generated ahead and behind dna tracking ensembles . \n one question here is whether such juxtapositions are configured inside the cell in the same way as in naked dna in vitro , and how such configurations might enhance the dna transport efficiency of topoisomerase ii . \n for instance , dna decatenation by yeast topoisomerase ii is greatly enhanced by dna supercoiling in vitro , being catenane links removed much faster than coexisting supercoil crossings ( 91 ) . moreover \n , dna decatenation is more efficient in negative than in positively supercoiled circles ( 93 ) . \n these preferences , however , might not apply to intracellular dna unless newly replicated duplexes were supercoiled . \n dna configurations behind these in vitro preferences are perhaps reproduced in a different way by the chromatin architecture of post - replicative catenanes in vivo . \n another dissimilarity between in vitro and in vivo configurations is that intracellular catenane links might produce very tight dna juxtapositions when chromosome condensation and segregation forces pull apart replicated dna molecules . in that case \n , their configuration likely facilitates its removal by topoisomerase ii . in the case of dna relaxation \n , yeast topoisomerase ii does not have significant preference to invert positive or negative crossings in supercoiled dna in vitro ( 91 ) , though this is not the case in relaxed dna ( 95 ) . \n inside the cell , the enzyme must remove also positive and negative supercoils with comparable efficiency . \n otherwise , dna supercoils of a particular sign would accumulate in yeast mutants deficient in topoisomerase i ( 96 ) \n . this situation might be slightly different in mammalian cells , since topoisomerase ii appears to relax in vitro positive supercoils faster than negative ones , while ii displays no such preference ( 29 ) . \n to the question of how helical tension configures dna juxtapositions in vivo , one can assume that dna is not adopting the same plectoneme folds seen in naked dna . \n the nucleosome structure has its intrinsic mode of constraining and adapting to dna helical stress . yet \n , topoisomerase ii is found to relax positive helical tension in native nucleosome arrays nearly at the same rate as in naked dna ( 25 ) . without displacement of the histones \n , recent studies indicated that positive helical tension drives a chiral transition of nucleosomes to a meta - stable conformation ( reversome ) in which h2a h2b dimers and the h3h4 tetramer refold to shape a right - handed path for dna ( 97 ) . \n interestingly , the entry and exit dna segments of the reversome might configure a positive dna crossing , which could be inverted by topoisomerase ii . \n strand passage would readily revert the chiral transition of the nucleosome while relaxing the dna ( figure 4a ) . \n ( a ) nucleosomes accommodate to positive helical tension ( + ) by adopting a meta - stable conformation in which h2a \n h2b dimers and the h3h4 tetramer reorganize and shape a right - handed path for dna . \n the entry and exit dna segments of this nucleosomal conformation might configure an ideal positive dna crossing to be targeted by topoisomerase ii . \n dna transport would relax the helical tension and revert the chiral transition of the nucleosome . \n this scenery might explain why topoisomerase ii relaxes helical tension in nucleosome arrays as efficiently as in naked dna . \n ( b ) in absence of dna helical tension , dna juxtaposition could be tailored by neighbouring dna protein interactions . \n the only difference is that protein dna interactions enforcing the juxtaposition of a t - segment are established outside rather than inside the topoisomerase \n ( a ) nucleosomes accommodate to positive helical tension ( + ) by adopting a meta - stable conformation in which h2a \n h2b dimers and the h3h4 tetramer reorganize and shape a right - handed path for dna . \n the entry and exit dna segments of this nucleosomal conformation might configure an ideal positive dna crossing to be targeted by topoisomerase ii . \n dna transport would relax the helical tension and revert the chiral transition of the nucleosome . \n this scenery might explain why topoisomerase ii relaxes helical tension in nucleosome arrays as efficiently as in naked dna . \n ( b ) in absence of dna helical tension , dna juxtaposition could be tailored by neighbouring dna protein interactions . \n the only difference is that protein dna interactions enforcing the juxtaposition of a t - segment are established outside rather than inside the topoisomerase \n the second type of dna juxtaposition is that enforced by the molecular crowding of intracellular dna . \n computer simulations of dna chains under tough condensation or volume confinement predict that the duplex configuration would equilibrate with many knot and catenane links ( 98100 ) . \n such equilibrium would be achieved upon random transport of dna segments , but this extreme is unlikely to occur . \n first , chromatin architecture might prevent random dna juxtapositions leading to undesired knots and catenane links . \n second , topoisomerase ii might keep random knot and catenate links below the equilibrium values , as it does with dna in free solution . third , undesired links provoked by molecular crowding will be cleared at the time of chromosome replication and segregation . \n the fact is , however , than the extent of knots and catenanes driven by molecular crowding of intracellular dna is unknown . \n circular minichromosomes ( < 10 kb ) extracted from yeast cells show negligible catenation and knotting probability ( 101 ) , but these figures could become significant in larger chromatin domains . in such a case , the question is whether such links might have some biological relevance . for example , knotting of dna driven by the molecular crowding could contribute to stabilize chromosome condensation . \n the third type of dna juxtaposition is that enforced by the architecture of dna protein interactions . \n wrapping of dna around histones brings linker dna segments in close proximity ; and many other dna - interacting proteins bring together nearby dna regions by looping or kinking the duplex . \n the question here is what portion of such juxtapositions not driven by the topology state of dna can be effective pairs of g- and t - segments . \n it is conceivable that a juxtaposition is specifically tailored to promote dna transport by topoisomerase ii , so creating a situation analogous to that of dna gyrase . \n the only difference would be that protein dna interactions enforcing the juxtaposition of a t - segment to configure a positive ( or negative ) crossing would be established outside rather than inside the topoisomerase - dna complex ( figure 4b ) . \n dna transport could result then in supercoiling energy that , unless constrained by other interactions , could drive critical structural changes in chromatin micro - domains . \n such supercoiling activity could be behind the implication of topoisomerase ii in local chromatin condensation - decondensation and gene regulation . \n most studies on topoisomerase ii mechanism had focussed on the dna cleavage and gating steps , the dna transport preferences , and the coupling of inter - domainal movements to atp usage . however , a less - attended but critical aspect of the enzyme operation that needs to be absolutely effective is the rejoining of the gated dna . \n consider , for example , yeast cells deficient in topoisomerase i , in which topoisomerase ii alone relaxes the supercoils raised during dna transcription and reduces to zero the linkage between complementary dna strands during their replication . any single dna gating event not followed by the proper rejoining step will put at risk genome integrity and cell survival . \n it is reasonable to assume then that most mechanic traits of the enzyme are tuned to ensure rejoining rather than dna cleavage and dna transport . \n not surprisingly , those compounds affecting the dna - rejoining step readily produce cell death and have , therefore , prominent pharmacological interest . \n but apart from biochemical targeting , many physiological processes might also threaten the dna rejoining step . \n for instance , in contrast to dna in free solution , intracellular dna is subjected to considerable twisting , bending and pulling forces generated by dna tracking ensembles and the conformational dynamics of chromatin fibers . in this environment \n yet , the mechanism of the type-2 topoisomerases implies that none of its dimer interfaces is a permanent interaction . \n the path of the t - segment involves the sequential crossing of the entrance or n - gate , the dna gate and the exit or c - gate . \n coordination among these three gates is therefore critical to preclude the risk of the two halves of the enzyme coming apart while gating the dna . in that regard , \n biochemical studies with yeast topoisomerase ii indicate that gate opening and closure follow a double - lock rule : a given gate can open only if the other two are closed ( 50 ) . \n although the n - gate domain is not required for g - segment cleavage , the dna gate per se is not able to widen unless atp binds and closes the n - gate ( 50 ) . \n since a captured t - segment can not be held in the inter - domainal region between the n - gate and the dna gate , closure of the n - gate and consequent capture of a t - segment is likely to enforce the opening of the dna gate ( 50 ) . once a t - segment has reached the central cavity of the enzyme and before the c - gate opens , the gated g - segment dna can be rejoined ( 11,50 ) . \n hence , opening of the c - gate is likely to be triggered by the entrapment of the t - segment after being squeezed by the sequential closure of the n - gate and the dna gate . \n these inter - domainal couplings maximize the dimer stability , ensure a very transient dna gating state and enforce unidirectional dna transport by precluding the backtracking of the t - segment ( figure 1a ) . \n figure 1.mechanical couplings for gate opening and closure in type-2 topoisomerases . \n ( a ) in type-2a enzymes ( bacterial gyrase , topoisomerase iv , topoisomerase ii ) , the n - gate , dna gate and c - gate appear to be mechanically coupled with a double - lock rule , such that a given gate can open only if the other two are closed . \n this coordination minimizes the risk of the two enzyme halves from coming apart while gating the dna . \n ( a ) when the entrance n - gate is open , dna might be cleaved but the dna gate is not able to widen . \n ( b ) upon atp binding and closure of the n - gate , a captured t - segment can not be held in the inter - domainal region between the n - gate and dna gate . \n the t - segment quickly crosses the dna - gate and reaches the central cavity of the enzyme . \n ( d ) the consequent entrapment of the t - segment enforces its exit by a transient opening of the c - gate . \n ( b ) in type-2b enzymes ( topoisomerase vi ) there is no c - gate . \n ( a ) when the entrance n - gate is open ( no atp bound ) the dna gate is locked . \n ( b ) upon atp binding and closure of the n - gate , a captured t - segment is held in the central cavity of the enzyme prior to the aperture of the dna gate . \n ( c ) the captured t - segment crosses then the dna - gate and exits the complex . \n figure 2.topoisomerase ii might discern whether or not a dna domain is suitable for successful rejoining by bending the g - segment before the gating step . \n ( a ) proper bending of the interacting duplex will be possible as long as the stretching tension along the dna does not exceed a threshold value ( o ) . \n this threshold is determined by the tension ( p ) that the enzyme is able to counteract to close the dna gate . \n ( b ) the active deformation of dna , coupled to atp binding and hydrolysis , could operate then as a checkpoint to unlock the dna - gate , as well as a gain of elastic energy to be delivered in the rejoining step . \n ( a ) in type-2a enzymes ( bacterial gyrase , topoisomerase iv , topoisomerase ii ) , the n - gate , dna gate and c - gate appear to be mechanically coupled with a double - lock rule , such that a given gate can open only if the other two are closed . \n this coordination minimizes the risk of the two enzyme halves from coming apart while gating the dna . \n ( a ) when the entrance n - gate is open , dna might be cleaved but the dna gate is not able to widen . \n ( b ) upon atp binding and closure of the n - gate , a captured t - segment can not be held in the inter - domainal region between the n - gate and dna gate . \n the t - segment quickly crosses the dna - gate and reaches the central cavity of the enzyme . \n ( d ) the consequent entrapment of the t - segment enforces its exit by a transient opening of the c - gate . \n ( b ) in type-2b enzymes ( topoisomerase vi ) there is no c - gate . \n ( a ) when the entrance n - gate is open ( no atp bound ) the dna gate is locked . \n ( b ) upon atp binding and closure of the n - gate , a captured t - segment is held in the central cavity of the enzyme prior to the aperture of the dna gate . \n ( c ) the captured t - segment crosses then the dna - gate and exits the complex . \n topoisomerase ii might discern whether or not a dna domain is suitable for successful rejoining by bending the g - segment before the gating step . \n ( a ) proper bending of the interacting duplex will be possible as long as the stretching tension along the dna does not exceed a threshold value ( o ) . \n this threshold is determined by the tension ( p ) that the enzyme is able to counteract to close the dna gate . \n ( b ) the active deformation of dna , coupled to atp binding and hydrolysis , could operate then as a checkpoint to unlock the dna - gate , as well as a gain of elastic energy to be delivered in the rejoining step . \n the quaternary couplings during dna transport in topoisomerase ii must be alike in topoisomerase iv and dna gyrase . however \n , they might differ from those in the less - extensively studied type-2b topoisomerases ( archea topoisomerase vi and plant topoisomerase vi ) ( 51,52 ) . \n these enzymes share several mechanistic features with the type-2a topoisomerases , but there are significant structural differences between the two families ( 53 ) . for t - segment capture , the atpase domains of type-2b dimerize upon atp binding alike the n - gate in type-2a enzymes ( 54 ) . \n the dna cleavage - rejoining core of type-2b also contains counterparts of the structural motifs of the dna - gate of the type-2a enzymes . \n however , type-2b enzymes do not have an exit- or c - gate ( 55 ) . \n the transported duplex is outside the complex right after crossing the dna gate ( figure 1b ) . \n therefore , dimer stability during the dna gating step relies mostly in the closure of the n - gate . type-2a and \n type-2b topoisomerases should have then distintc couplings between their atpase and dna cleavage - rejoining modules . \n remarkably , contrary to type-2a topoisomerases ( 50 ) , the atpase domains of type-2b enzymes are required for dna cleavage ( 56 ) and the cavity between the two gates of these enzymes can accommodate a captured t - segment before widening the dna gate ( 55 ) . \n the absence of a c - gate in type-2b topoisomerases facilitates perhaps the dissociation of the enzyme halves for some biological tasks . \n for instance , reduced dimer stability could be operational in spo11 , a type-2b related enzyme that mediates dna double - strand breaks during recombination ( 51,57 ) . \n aside of ensuring dimer stability during the dna cleavage and gating steps , another way to guarantee genome integrity is to not cleave the dna in the first place . \n it is conceivable that topoisomerase ii discerns whether or not a duplex is suitable for successful rejoining before gating it . for example \n , if dna is undergoing extreme tension , its conformation might not be appropriate for catalytic cleavage . in that regard \n , the recently solved crystal structure of the cleavage - rejoining core of yeast topoisomerase ii bound to dna , reveals a complex producing a 150 dna bend in the g - segment ( 58 ) . \n a sharp bending of the bound g - segment had previously been inferred for bacterial topoisomerase iv and it is postulated to contribute to the capacity of type-2a topoisomerases to simplify dna equilibrium configurations ( 59 ) . \n bending of the g - segment might be a prerequisite for dna cleavage , thus protein conformational changes that accompany this deformation create a bipartite catalytic site , in which each dna strand gets close to a reactive tyrosine and a coordinated magnesium ion ( 58 ) . \n another possible function of such dna bending is that it would prevent topoisomerase ii from gating the duplex in regions undergoing severe stretching forces ( figure 2a ) . \n recently , oligonucleotides with a pair of fluorophores straddling the dna cleavage site have been used to observe the dynamics of the dna gate by fret ( 60 ) . \n the experiment revealed that during steady - state atp hydrolysis fluorophores repeatedly move over a distance of 20 with comparable back and forth rate constants . whether these displacements reflected opening and \n first , the putative open and closed states are nearly equally probable in a steady state population , whereas all previous biochemical data indicate that in this condition only a small fraction of complexes is in the open dna - gate state . \n second , as mentioned above , the capture of a t - segment is likely required to enforce the opening of the dna gate . in the fret experiment , however , no free dna was available to provide a t - segment . \n thus , the observed displacements could reflect mostly bending or twisting distortions of the g - segment coupled to atp binding and hydrolysis but with no significant widening of the dna gate . \n such active deformation of dna could operate as a checkpoint to unlock the dna - gate , as well as a stock of elastic energy to deliver at the rejoining step . by these means , \n topoisomerase ii could discern whether a duplex is suitable for proper rejoining before gating it ( figure 2b ) . \n identification of topoisomerase ii as one of the main components of the nuclear matrix and metaphase chromosome scaffold ( 6163 ) , as well as its preferential interaction to sar and mar sequences ( 6466 ) , had lead to the proposal of an structural role for the enzyme . \n the possibility that topoisomerase ii could be anchoraging dna chromosomal loops fit well with the enzyme capabilities to hold dna through covalent bonds and to interact simultaneously with several dna segments ( 67,68 ) . \n this putative structural role was further supported by the discovery of that , upon binding of a non - hydrolysable atp analogue , the topoisomerase constituted in vitro a high - salt resistant complex with circular dna molecules ( 7,69,70 ) . \n this observation was essential for unravelling the mechanism of type-2 topoisomerases , reflecting the closure of the n - gate and the subsequent impossibility of the protein ring detaching from a bound g - segment in the absence of a dna free - end . \n the topoisomerase dna complex was resistant to salt concentrations up to 5 m nacl or cscl during several hours ( 71 ) . \n therefore , such a topological interaction was an attractive mechanism to fasten large chromosomal dna loops , and it seemed more convenient than holding the duplex with covalent links that implied a permanent cleavage of dna . in the last decade , however , support for the proposed structural role of topoisomerase ii had faded since some observations disputed the requirement of the enzyme to maintain the shape of mitotic chromosomes . although topoisomerase ii is required for mitotic chromosome assembly in vitro , the enzyme could be extracted from in vitro condensed chromosomes with no significant changes in the chromosome conformation ( 72 ) . \n other in vivo studies , which used fluorescently tagged human dna topoisomerases ii and ii , indicated that neither enzyme is an immobile structural component of the chromosomes ( 73 ) . \n moreover , several studies reported significant mitotic chromosome condensation even after genetic knockdown or knockout of topoisomerase ii ( 30,7476 ) . yet , other demonstrations indicated that topoisomerase ii remains stably bound in chromosomes assembled in vitro after a round of dna replication ( 77 ) . \n more recent analyses in several species including mammals , chicken , c. elegans and drosophila , came to agree in that topoisomerase ii is not required for initial chromatin compaction and formation of a longitudinal chromosome axis , but that an axial distribution of the enzyme in late prophase to metaphase is real , being dependent on the presence of condensins ( 76,78,79 ) . on the other side , \n the existence of a rigid chromosome scaffold , in which dna loops could be anchored , is also no longer sustained after some morphological and biophysical observations . \n for instance , depletion of histone h1 results in longer and thinner compacted mitotic chromosomes , and so arguing against a stiff scaffolding structure driving chromosome condensation ( 80 ) . also , the elastic response of native metaphase chromosomes and its alteration by endonucleases rule out a contiguous internal non - dna ( e.g. protein ) scaffold . \n the elastic behaviour of mitotic chromosomes suggests instead that they are shaped by isolated dna - crosslinking elements spaced by few kilo base pairs ( 81,82 ) . from the above , the possibility that a highly stable topoisomerase ii dna complex plays a structural role in chromosome architecture can not be excluded \n the closure of the enzyme around dna is driven by atp binding and this conformation could be easily stabilized by some cellular factor . such factor could interact across the closed n - gate domains , thus behaving like a molecular padlock . essentially , this is the case with bisdioxopiperazines , such as icrf-193 , that bind to the interface of the dimerized atp - ase domains of topoisomerase ii and preclude n - gate reopening ( 83,84 ) . \n the hypothetic existence of gate padlocks could be also considered for the c - gate of topoisomerase ii . in this case , stabilisation of the exit gate would be analogous to locking it by means of engineered disulfide bonds ( 11,50 ) . \n in such a case , a t - segment would be captured , transported across the gated g - segment and end up entrapped in the central chamber of the topoisomerase between the rejoined dna - gate and the locked exit - gate . \n if padlocks close both the c - gate and the n - gate simultaneously , the topoisomerase would become a tight toroidal structure , inside which two dna segments could be entrapped . \n this extreme can be observed in vitro by closing the n - gate with a non - hydrolysable atp analogue in the yeast topoisomerase ii mutant having the c - gate locked by disulfide bonds ( 50 ) . in this condition , \n analogously to bisdioxopiperazines , such as icrf-193 that inhibit topoisomerase ii activity , closure of the n - gate of topoisomerase ii upon atp binding can be stabilized also by some cellular factor . \n ( a ) such gate padlocks could operate as inhibitors of enzyme binding to chromosomal dna . \n ( b ) if dna is already bound to the topoisomerase , an n - gate padlock would produce a high salt resistant complex , which might serve to regulate the enzyme activity or operate as a structural element for dna organization . \n if both the n - gate and the c - gate were stabilized in the closed conformation , the topoisomerase could clamp two dna duplexes , the g - segment and passed t - segment . \n analogously to bisdioxopiperazines , such as icrf-193 that inhibit topoisomerase ii activity , closure of the n - gate of topoisomerase ii upon atp binding can be stabilized also by some cellular factor . \n ( a ) such gate padlocks could operate as inhibitors of enzyme binding to chromosomal dna . \n ( b ) if dna is already bound to the topoisomerase , an n - gate padlock would produce a high salt resistant complex , which might serve to regulate the enzyme activity or operate as a structural element for dna organization . \n if both the n - gate and the c - gate were stabilized in the closed conformation , the topoisomerase could clamp two dna duplexes , the g - segment and passed t - segment . \n dna complexes for structural purposes , another plausible function of gate padlocks could be the regulation of the enzyme activity , a subject that remains poorly understood . \n topoisomerase ii can be phosphorylated in vitro on multiple serine and threonine residues , the majority of which are located in its non - conserved c - terminal region . \n the plausible effects of phosphorylation in vivo remain controversial , since most studies report that these modifications do not produce significant changes in the dna transport activity of the enzyme in vitro ( 85 ) . \n whether this modification occurs in vivo and has physiological significance is unclear ( 86 ) . \n it is suggested that poly(adp - ribosyl)ation may play a role in the cellular distribution and degradation the enzyme ( 87 ) . \n more recently , topoisomerase ii has been shown to undergo sumo - lation during mitosis ( 8890 ) . \n this modification seems to be a signal for targeting the enzyme to pericentromeric regions of mitotic chromosomes and promotes the decatenation of sister chromatids at anaphase ( 8890 ) . \n the question then is what might prevent intracellular topoisomerase ii to hydrolyse atp , to cleave dna or to transport dna when it is not necessary . \n as observed with the topoisomerase inhibitor icrf-193 , locking of the n - gate upon atp binding can occur in the absence of dna ( 83 ) . in this closed conformation , \n gate padlocks could therefore be used to transport and stock topoisomerase ii inside the cell and let it operate only where needed . \n so far , no gate padlock has been found in cell extracts or described among the several proteins that might physically interact with topoisomerase ii . \n another reason , however , is that proper assays to find them have not been conducted . \n once topoisomerase ii is bound to a g - segment , dna transport depends solely on the probability of finding a potential t - segment properly positioned in the vicinity of the n - gate of the enzyme ( 91 ) . for naked dna molecules in free solution , such juxtaposition probability will be determined by the global configuration of the duplex , and also by constrains that the topoisomerase might produce by its local interactions with dna . \n this last aspect should account for the enzyme capacity to drive dna topology below thermal equilibrium configurations and for its chiral preferences ( 92,93 ) . \n the question now is whether our current understanding of juxtaposition probability of dna segments on topoisomerase ii , as inferred from in vitro studies , is applicable to intracellular dna . \n although the topological problems inside the cell are equivalent to those portrayed in vitro , the conformational dynamics of dna is determined by a complex set of variables involving multiple molecular interactions , in addition to the topology of the duplex . \n for example , although in living yeast cells both dna topoisomerases i and ii are capable to relax plasmid minichromosomes ( 94 ) , in vitro studies revealed that topoisomerase ii relaxes nucleosomal dna much faster than topoisomerase i ( 25 ) . \n apparently , the dna cross - inversion mechanism of topoisomerase ii is facilitated in nucleosome fibres that might favour the juxtaposition of dna segments , whereas the dna strand - rotation mechanism of topoisomerase i might be stalled by twist diffusion barriers imposed by nucleosome organization . \n how chromatin affects dna segment juxtaposition and the consequent topoisomerase ii activity is complex to envisage . on one hand \n , juxtaposition can be favoured as dna is intensely and extensively more folded than in free solution . on the other hand , \n the number of dna segments suitable for topoisomerase ii manipulation is more reduced since many proteins cover most of the dna length . to further dissect this problem , \n three types of dna juxtaposition should be considered to occur in the intracellular landscape , each one producing a different outcome upon dna transport by topoisomerase ii . \n this is the case of the pre - catenanes and catenane links generated between newly replicated duplexes and the supercoils generated ahead and behind dna tracking ensembles . \n one question here is whether such juxtapositions are configured inside the cell in the same way as in naked dna in vitro , and how such configurations might enhance the dna transport efficiency of topoisomerase ii . \n for instance , dna decatenation by yeast topoisomerase ii is greatly enhanced by dna supercoiling in vitro , being catenane links removed much faster than coexisting supercoil crossings ( 91 ) . moreover , dna decatenation is more efficient in negative than in positively supercoiled circles ( 93 ) . \n these preferences , however , might not apply to intracellular dna unless newly replicated duplexes were supercoiled . \n dna configurations behind these in vitro preferences are perhaps reproduced in a different way by the chromatin architecture of post - replicative catenanes in vivo . \n another dissimilarity between in vitro and in vivo configurations is that intracellular catenane links might produce very tight dna juxtapositions when chromosome condensation and segregation forces pull apart replicated dna molecules . in that case \n , their configuration likely facilitates its removal by topoisomerase ii . in the case of dna relaxation \n , yeast topoisomerase ii does not have significant preference to invert positive or negative crossings in supercoiled dna in vitro ( 91 ) , though this is not the case in relaxed dna ( 95 ) . \n inside the cell , the enzyme must remove also positive and negative supercoils with comparable efficiency . \n otherwise , dna supercoils of a particular sign would accumulate in yeast mutants deficient in topoisomerase i ( 96 ) \n . this situation might be slightly different in mammalian cells , since topoisomerase ii appears to relax in vitro positive supercoils faster than negative ones , while ii displays no such preference ( 29 ) . \n to the question of how helical tension configures dna juxtapositions in vivo , one can assume that dna is not adopting the same plectoneme folds seen in naked dna . \n the nucleosome structure has its intrinsic mode of constraining and adapting to dna helical stress . yet \n , topoisomerase ii is found to relax positive helical tension in native nucleosome arrays nearly at the same rate as in naked dna ( 25 ) . without displacement of the histones \n , recent studies indicated that positive helical tension drives a chiral transition of nucleosomes to a meta - stable conformation ( reversome ) in which h2a \n h2b dimers and the h3h4 tetramer refold to shape a right - handed path for dna ( 97 ) . \n interestingly , the entry and exit dna segments of the reversome might configure a positive dna crossing , which could be inverted by topoisomerase ii . \n strand passage would readily revert the chiral transition of the nucleosome while relaxing the dna ( figure 4a ) . \n ( a ) nucleosomes accommodate to positive helical tension ( + ) by adopting a meta - stable conformation in which h2a \n dimers and the h3h4 tetramer reorganize and shape a right - handed path for dna . \n the entry and exit dna segments of this nucleosomal conformation might configure an ideal positive dna crossing to be targeted by topoisomerase ii . \n dna transport would relax the helical tension and revert the chiral transition of the nucleosome . \n this scenery might explain why topoisomerase ii relaxes helical tension in nucleosome arrays as efficiently as in naked dna . \n ( b ) in absence of dna helical tension , dna juxtaposition could be tailored by neighbouring dna protein interactions . \n the only difference is that protein dna interactions enforcing the juxtaposition of a t - segment are established outside rather than inside the topoisomerase \n ( a ) nucleosomes accommodate to positive helical tension ( + ) by adopting a meta - stable conformation in which h2a \n h2b dimers and the h3h4 tetramer reorganize and shape a right - handed path for dna . \n the entry and exit dna segments of this nucleosomal conformation might configure an ideal positive dna crossing to be targeted by topoisomerase ii . \n dna transport would relax the helical tension and revert the chiral transition of the nucleosome . \n this scenery might explain why topoisomerase ii relaxes helical tension in nucleosome arrays as efficiently as in naked dna . \n ( b ) in absence of dna helical tension , dna juxtaposition could be tailored by neighbouring dna protein interactions . \n the only difference is that protein dna interactions enforcing the juxtaposition of a t - segment are established outside rather than inside the topoisomerase \n the second type of dna juxtaposition is that enforced by the molecular crowding of intracellular dna . \n computer simulations of dna chains under tough condensation or volume confinement predict that the duplex configuration would equilibrate with many knot and catenane links ( 98100 ) . \n such equilibrium would be achieved upon random transport of dna segments , but this extreme is unlikely to occur . \n first , chromatin architecture might prevent random dna juxtapositions leading to undesired knots and catenane links . \n second , topoisomerase ii might keep random knot and catenate links below the equilibrium values , as it does with dna in free solution . \n third , undesired links provoked by molecular crowding will be cleared at the time of chromosome replication and segregation . \n the fact is , however , than the extent of knots and catenanes driven by molecular crowding of intracellular dna is unknown . \n circular minichromosomes ( < 10 kb ) extracted from yeast cells show negligible catenation and knotting probability ( 101 ) , but these figures could become significant in larger chromatin domains . in such a case , the question is whether such links might have some biological relevance . for example , knotting of dna driven by the molecular crowding could contribute to stabilize chromosome condensation . \n the third type of dna juxtaposition is that enforced by the architecture of dna protein interactions . \n wrapping of dna around histones brings linker dna segments in close proximity ; and many other dna - interacting proteins bring together nearby dna regions by looping or kinking the duplex . \n the question here is what portion of such juxtapositions not driven by the topology state of dna can be effective pairs of g- and t - segments . \n it is conceivable that a juxtaposition is specifically tailored to promote dna transport by topoisomerase ii , so creating a situation analogous to that of dna gyrase . \n the only difference would be that protein dna interactions enforcing the juxtaposition of a t - segment to configure a positive ( or negative ) crossing would be established outside rather than inside the topoisomerase - dna complex ( figure 4b ) . \n dna transport could result then in supercoiling energy that , unless constrained by other interactions , could drive critical structural changes in chromatin micro - domains . \n such supercoiling activity could be behind the implication of topoisomerase ii in local chromatin condensation - decondensation and gene regulation . \n in view of our substantial knowledge on topoisomerase ii structure and mechanism , as inferred from in vitro studies , many questions need to be now addressed pertaining to how the enzyme performs inside the cell . to the question of how dimer stability is ensured during the dna gating step , one answer is the double - lock rule that allows any of its three gates to open only if the other two are closed . \n an additional answer is a checkpoint to unlock the dna - gate , which might allow topoisomerase ii to discern whether a duplex is suitable for rejoining before gating it . to the question of what prevents atp hydrolysis , dna cleavage or dna transport by topoisomerase \n gate padlocks might also operate the transition from the catalytic to the putative structural functions of the enzyme . finally , to the question of what directs dna transport probability in the chromatin milieu , three kinds of dna juxtaposition can configure potential pairs of g- and t - segments to be inverted by intracellular topoisomerase ii : those enforced by dna topology configuration , those resulting from molecular crowding , and those customized by chromatin architecture . \n dna transport produces a different outcome in each case , so accounting for the multiple biological roles reserved for topoisomerase ii . \n more questions will arise , and the initial answers will be imperfect or quite speculative , as some in this survey . yet , this exercise is essential to design crucial experiments en route for new discoveries . \n funding for open access charge : ministerio de educacin y ciencia de espaa ( plan nacional i+d+i ) and generalitat de catalunya ( dursi ) .", "role": "user" }, { "content": "the mechanism by which type-2a topoisomerases transport one dna duplex through a transient double - strand break produced in another exhibits fascinating traits . \n one of them is the fine coupling between inter - domainal movements and atp usage ; another is their preference to transport dna in particular directions . \n these capabilities have been inferred from in vitro studies but we ignore their significance inside the cell , where dna configurations markedly differ from those of dna in free solution . the eukaryotic type-2a enzyme , topoisomerase ii , is the second most abundant chromatin protein after histones and its biological roles include the decatenation of newly replicated dna and the relaxation of polymerase - driven supercoils . yet , topoisomerase ii is also implicated in other cellular processes such as chromatin folding and gene expression , in which the topological transformations catalysed by the enzyme are uncertain . here , some capabilities of topoisomerase ii that might be relevant to infer the enzyme performance in the context of chromatin architecture are discussed . \n some aspects addressed are the importance of the dna rejoining step to ensure genome stability , the regulation of the enzyme activity and of its putative structural role , and the selectively of dna transport in the chromatin milieu .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: small bowel ischemia following laparoscopy was described recently as a rare fatal complication of the co2 pneumoperitoneum . of the 8 cases reported in the surgical literature , \n a 34-year - old woman who underwent laparoscopy with extensive adhesiolysis and myolysis was re - admitted with an acute abdomen on postoperative day 4 . \n immediate laparotomy revealed acute peritonitis , extensive adhesions , and a 3-cm defect in the small bowel . \n she was discharged after a 2-month hospital stay in the intensive care unit for rehabilitation . \n some abdominal pain occurred in all , nausea and vomiting in 4 , diarrhea in 2 , abdominal distention in 1 , fever in none . \n quick reintervention laparotomy was performed in 2 and delayed in 5 ( up to 4 days ) . \n the co2 pneumoperitoneum is a predisposing factor for intestinal ischemia as it reduces cardiac output and splanchnic blood flow . \n patient selection , maintaining intraabdominal pressure at 15 mm hg or less , and intermittent decompression of the gas represent the best options for preventing this complication . \n laparoscopic surgery has become the standard of care for many gynecologic conditions , and the laparoscopic technique is recognized as being safe and effective . \n however , recent case reports in the surgical literature describe small bowel ischemia as a rare fatal complication of the co2 pneumoperitoneum . \n andrei et al reviewed the literature on small bowel ischemia in 1999 and added 1 case that occurred following laparoscopic cholecystectomy . \n of the 8 cases in the literature , 7 were fatal , 1 was not . \n the co2 pneumoperitoneum appears to be a predisposing factor in the development of intestinal ischemia because it compromises the mesenteric circulation through decreased cardiac output and mechanical reduction of the blood flow due to increased intraabdominal pressure , and humoral vasoconstriction . \n however , as we will describe , progression to critical ischemia seems to rely on underlying vasculopathy or an inciting event . in this report \n , we describe the first case in gynecology of small bowel ischemic necrosis following laparoscopic surgery . \n our intention is to increase awareness about this catastrophic complication and to examine the literature for possible causes and preventive measures . \n a 34-year - old african - american woman , gravida 3 , para 1 , presented to the first author with a newly developed 4- to 5-cm left uterine mass associated with menorrhagia and pelvic pain of several months duration . \n a diagnostic laparoscopy with possible myomectomy was planned pending a second opinion consult and medical clearance . \n past surgical history included laparotomy with left oophorectomy for ovarian cysts and 2 elective terminations of pregnancy , the last of which may have been associated with an inflammatory process that contributed to development of the pelvic mass not appreciated during previous pelvic examinations . \n past medical history included asthma and use of nonsteroidal anti - inflammatory drugs for pain . \n a laboratory workup showed mild iron deficiency anemia ( hemoglobin 11.6 g / dl ; hematocrit 35.3% ) and inconclusive thyroid studies with the patient being clinically euthyroid . at laparoscopy , the open entry method was used . \n adherent small bowel loops were noted at the entry site requiring careful dissection . with the establishment of an adequate pneumoperitoneum at a pressure of 15 mm hg , \n careful port placement and adhesiolysis using disposable scissors were performed to expose the pelvic organs . \n because of safety considerations , myolysis was selected as the procedure of choice with serial bipolar needle punctures placed into the perceived junction between each lesion and uterus as well as in its dome . \n the abdomen was lavaged with copious amounts of saline , leaving behind a generous amount to minimize adhesion reformation . \n the operative time was 75 minutes , the anesthetic inhalation agents used were sevoflurane and nitrous oxide , and the estimated blood loss was < 30 ml . \n the first author kept in contact with the patient on a daily basis because of the extensive dissection . \n she was afebrile throughout , voiding well , and tolerating oral intake on the first postoperative day . \n she passed flatus on the second day and had a small bowel movement on the third day . \n she had a bloody drainage from the incision attributed to hydroflation on day 1 and a black and blue mark around the umbilicus on day 2 , probably representing a bruise from surgical dissection . \n however , on the morning of the fourth postoperative day , the patient reported that she had vomited and was told to go to the emergency room immediately . \n the patient presented to the emergency room several hours later and was met by the first author as she was being admitted . \n an acute abdomen was diagnosed , and the surgical team mobilized for an immediate laparotomy . shortly thereafter , \n the patient experienced a sudden onset of atrial fibrillation and hypotension . a triple lumen catheter was placed , and the patient was taken to the operating room for exploratory laparotomy . \n enteric fluid was noted to exit from the umbilical port site while the patient was being prepped and draped . a midline laparotomy was performed . \n gross spillage of enteric fluid was noted as well as extremely dense adhesions between the bowel and anterior abdominal wall mostly in the midline . \n mobilization of the bowel was difficult and required repeated releasing incisions due to foreshortening of the mesentery . \n the pathology report showed a 3-cm defect in the center of the resected bowel associated with changes consistent with ischemic necrosis . \n for instance , no coagulation necrosis , inflammatory reaction , or granulation tissue was present . \n following surgery , the patient was transferred to the intensive care unit ( icu ) and treated with broad - spectrum antibiotics and activated protein c ( xigris ) . \n it was complicated by candidemia , unsuccessful extubation trials , profound multiple neuropathy , nosocomial pneumonia , pressure ulcer , drug - induced thrombocytopenia , and gastrointestinal bleeding , precipitating shock and 2 cardiopulmonary arrests . \n gastrostomy and tracheostomy tubes were placed , as well as an inferior vena cava filter when she developed a deep - vein thrombosis . \n this case represents a rare catastrophic complication that is not attributable to operative laparoscopic manipulations . as with all previously reported 6 cases for whom detailed information was available , \n none of the procedures lasted longer than 85 minutes ; 4 took less than 1 hour to complete . \n the procedures were variably described as elective , routine , uneventful , or without apparent complications . \n extensive lysis of adhesions was carried out in our case ; multiple adhesions were also noted in another report . \n different anesthetic agents were utilized , sevoflurane in our case , isoflurane , deslfurane , and halothane in others . \n the intraabdominal pressure was set at 15 mm hg in our case as well as in all communications specifying this parameter . \n the case reports are from different countries : 3 ( including ours ) from the usa , 2 from germany , 2 from the uk , 1 from australia , and 1 personal communication from an unspecified source . in the usa , \n the length of stay following the laparoscopic procedure was consistent with an outpatient approach as opposed to in - patient hospitalization considered standard elsewhere . \n our 34-year - old patient was discharged the same day of surgery and returned to the hospital on day 4 after having vomited and reporting a few other symptoms . \n the 72-year - old reported by andrei et al was discharged on day 1 and returned to the hospital on day 8 . \n her symptoms consisted of diffuse abdominal pain associated with nausea , vomiting , and bloody diarrhea that started 1 or 2 days after the operation . \n she was reoperated upon on day 9 , following marked deterioration in her general condition . \n schorr 's 62-year - old patient was ready for discharge on day 1 , but could not be because of family matters . \n while awaiting her family on day 2 , she developed severe abdominal pain and tachycardia and was kept in the hospital . \n autopsy revealed infarction of the small bowel and colon to the hepatic flexure due to superior mesenteric artery thrombosis . \n the artery was not constricted by plaque , and no evidence was present of an embolic phenomenon anywhere . the first case in the literature \n their 68-year - old patient , who was kept in the hospital , developed uncharacteristic abdominal pain and intermittent diarrhea . \n he was reoperated on on day 4 following increasing abdominal pain and general deterioration including renal failure . \n the patient died on day 45 from a septic complication . the 76-year - old patient reported by jaffe and russell from the uk \n was discharged on day 2 and readmitted on day 3 complaining of vague periumbilical discomfort . \n she was observed and investigated for 4 days and reoperated on on day 7 following rapid hemodynamic deterioration . \n a diagnosis of superior mesenteric artery thrombosis was made , intestinal resection was not done , and the patient died 12 hours later . the 55-year - old patient from australia reported by mitchell and jamieson developed on day 1 deep epigastric pain with nausea , retching , and itching . on day 2 \n she improved on day 3 but suffered sudden left subcostal pain at midnight and was operated on on day 4 following increasing abdominal pain and rigidity . \n subsequently , the patient underwent 7 reoperations , resecting ischemic tissue and ultimately died from overwhelming sepsis and hepatic infarction . \n postmortem examination revealed congenital narrowing of the celiac artery , which contained an organized thrombosis . \n the 36-year - old english patient reported by dwerry - house et al was discharged on day 1 . \n he returned to the hospital on day 2 complaining of colicky pain , abdominal distention , and vomiting . \n the terminal ileum was stenosed and thickened with associated fat wrapping suggestive of crohn 's disease . \n histologic examination of the resected ileum showed features of a venous infarction with patchy mucosal necrosis . \n some abdominal pain was experienced by all patients , including ours , but was considered severe in only 2 . \n nausea and vomiting occurred in 4 of the 7 patients , including ours , diarrhea in 2 , and abdominal distention in 1 patient . \n reintervention laparotomy was carried out in our patient as soon as she was admitted and on the same day of readmission in dwerryhouse et al 's patient . \n it was delayed in the other cases from 1 day to 4 days because of vague presentations and inconclusive investigations . \n it is generally agreed that the co2 pneumoperitoneum is a predisposing factor for the development of intestinal ischemia because it causes the following physiologic alterations : a ) decreased cardiac output due to increased systemic vascular resistance , decreased venous return , and elevated intrathoracic pressure , b ) significant reduction in the splanchnic blood flow , resulting from mechanical compression of the mesenteric veins , humoral vasoconstriction of the mesenteric bed and increased portal venous pressure caused by hypercapnia , local absorption of the co2 , and increased release of vasopressin . \n however , reduction in cardiac output and mesenteric blood flow associated with the co2 pneumoperitoneum should be insufficient to cause significant intestinal ischemia in healthy patients . \n critical intestinal ischemia following co2 pneumoperitoneum may be triggered by thrombosis of an arterial trunk , acute compromise of the mesenteric circulation of undetermined nature , or a focal lesion caused by a local insult ( present report ) . \n arterial thrombosis or acute insufficiency causes gangrene or infarction , with or without full thickness necrosis , of the gastrointestinal segment supplied by the compromised arterial trunk . \n superior mesenteric artery thrombosis was diagnosed clinically in 2 instances and acute insufficiency in one . \n these were attributed to atherosclerosis on the basis of age and history of cardiovascular disease . \n arterial thrombosis noted at autopsy was deemed to be due to unknown causes in 1 patient , and to congenital stenosis of the artery in another . in our case and that of dwerryhouse et al \n , the triggering mechanism appears to have been related to a local insult : dense adhesions pulling against a foreshortened mesentery in our case and abnormal ileum suggestive of crohn 's disease in theirs . \n the severity of insult in our case caused transmural necrosis , consistent with the description of severe focal segmental ischemia provided by greenwald et al . \n the gravity of the clinical situation resulting from a 3-cm open defect in the small bowel may be compared to that associated with a grossly ruptured appendix . \n free flow of intestinal contents into the peritoneal cavity would soon lead to generalized peritonitis and major third space losses into the peritoneum promptly resulting in dehydration , hemodynamic instability , and toxic shock . \n the profound multiple neuropathy noticed in our patient represents a peculiar phenomenon that has been linked to bowel ischemia . \n woodward et al recently described a previously fit 23-year - old man who presented with a sudden onset of profound autonomic neuropathy of unknown origin and subsequently developed ischemic enterocolitis requiring colectomy and subtotal enterectomy . \n the approach to this rare catastrophic complication should be directed to prevention rather than treatment . \n analysis of the reported cases indicate that signs and symptoms existing prior to rapid patient deterioration were generally insufficient to warrant intervention by the reporting teams , and surgical management after the fact was frequently futile . \n however , a high index of suspicion and early evaluation by ultrasound followed by laparotomy may be helpful in improving survival rates . \n it is generally agreed that periodic decompression of the pneumoperitoneum gas while maintaining the intraabdominal pressure at 15 mm hg or less represents the best available option for preventing this complication . \n patients in whom the splanchnic circulation is suspected of being impaired , based on preexisting atherosclerosis or other risk factors , should be handled with great care or offered an open laparotomy rather than the laparoscopy approach .", "role": "user" }, { "content": "background and objective : small bowel ischemia following laparoscopy was described recently as a rare fatal complication of the co2 pneumoperitoneum . of the 8 cases reported in the surgical literature , \n 7 were fatal , 1 was not . in this report \n , we describe the first gynecological case.methods:a 34-year - old woman who underwent laparoscopy with extensive adhesiolysis and myolysis was re - admitted with an acute abdomen on postoperative day 4 . \n immediate laparotomy revealed acute peritonitis , extensive adhesions , and a 3-cm defect in the small bowel . \n tissue examination showed ischemic necrosis of edematous , but essentially normal , bowel mucosa . \n the postoperative course was extremely complicated . \n she was discharged after a 2-month hospital stay in the intensive care unit for rehabilitation.results:data are available on 7 patients ( including ours ) . \n all procedures were described as uneventful . \n the intraabdominal pressure was set at 15 mm hg when specified . \n some abdominal pain occurred in all , nausea and vomiting in 4 , diarrhea in 2 , abdominal distention in 1 , fever in none . \n quick reintervention laparotomy was performed in 2 and delayed in 5 ( up to 4 days).discussion : the co2 pneumoperitoneum is a predisposing factor for intestinal ischemia as it reduces cardiac output and splanchnic blood flow . \n however , critical ischemia relies on underlying vasculopathy or an inciting event.conclusion:patient selection , maintaining intraabdominal pressure at 15 mm hg or less , and intermittent decompression of the gas represent the best options for preventing this complication .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: their consequences may range from small tooth fractures to complete displacement from the socket , characterizing tooth avulsion1 . \n root surface and endodontic treatments attempt to increase the occurrence of repair and reduce the presence of ankylosis and resorption1 . \n sodium hypochlorite is employed for removal of necrotic tissues present on the root surface before replantation , followed by washing in saline10 . \n the use of fluoride strengths the tooth structure by formation of fluorapatite , yet it is toxic to the resorption cells in the hard tissues1,14 . \n the use of calcium hydroxide as a root canal dressing may avoid or limit inflammatory resorption17 due to its alkaline ph and bactericidal effect15 , and therefore it may eliminate any infection . despite such treatment possibilities , there are still many cases of root resorption , and affected teeth usually must be extracted in a mean period of 4 - 6 years13 . \n hence , the search for new substances that may inhibit or delay the effects of root resorption is demanding . due to the similar morphology , enzymatic properties and function of cells leading to resorption of dentin , cementum and bone , the processes of root and bone resorption \n thus , drugs or other substances that may inhibit bone resorption may also be effective for the treatment of root resorption . \n thus , the newly formed bone becomes more resistant to the action of clast cells and resorption is inhibited3,4,5,19 . \n gallium nitrate may act on the proton pump present on the clast membrane , on the genetic expression of protein synthesis , or even on the carbonic anhydrase present on those cells2 . \n bockman , et al.4 ( 1995 ) verified that low doses of gallium nitrate are enough for its activity . assuming that gallium nitrate participates in the formation of hydroxyapatite and that the processes of bone and root resorption are similar , ghazi , et al.6 , in2000 , proposed the use of a gallium nitrate solution as a root canal dressing in teeth susceptible to root resorption . \n the authors reported that the gallium nitrate solution was capable of diffusing through the dentinal tubules and reach the external root surface . \n the authors have also reported that the ideal concentration of this solution is 10 m . \n based on the aspects presented , the purpose of this study was to evaluate the effect of gallium nitrate solution as a root canal dressing in avulsed replanted teeth , compared to calcium hydroxide paste . \n thirty male wistar rats ( rattus norvegicus , albinus ) weighing approximately 250 - 300 grams were used in this investigation . the study design was revised and approved by the institutional review board on animal experimentation of bauru dental school , university of so paulo , brazil ( process # 032/2003 , october 31 , 2003 ) . \n the animals were anesthetized by intramuscular injection of a mixture of ketamine ( dopalen , sespo indstria e comrcio ltda , jacare , sp , brasil ) and xylazine ( anasedan , agribrands do brasil ltda , jacare , sp . brasil ) at a dose of 0.05ml/100 g of weight for each drug . asepsis of the anterior portion of the maxilla was performed with periogard ( pfizer ltda , so paulo , sp , brasil ) . \n the maxillary right central incisor was then extracted , simulating a case of tooth avulsion . \n the extracted teeth were kept dry , attached to a sheet of pink wax by their crowns for 30 minutes . \n the dental papillae of teeth were excised with a # 11 surgical blade ( embramac exportao e importao , ribeiro preto , sp , brasil ) for exposure of root canals . \n the pulp was removed through the apical foramen with a slightly curved # 15 flexofile ( dentsply- maillefer , ballaigues , switzerland ) . \n canals were thoroughly irrigated with 1% hypochlorite solution ( probem produtos farmacuticos e odontolgicos ltda , catanduva , sp , brasil ) using a luer look syringe and a 30x4 gauge needle . after preparation , the teeth were immersed in 50ml of 1% hypochlorite solution for 30 minutes , washed in sterile saline and then immersed in 20ml of sodium fluoride solution ( ph 5.5 ) ( pharmcia specfica , bauru , so paulo , brasil ) for 20 minutes . \n the sodium fluoride solution was prepared from a mixture of 0.1 m phosphoric acid ( ph 2 ) and 2% sodium fluoride ( ph 8) . \n root canals were further irrigated with sterile saline and filled with edta ( odahcam dentsply indstria e comrcio ltda , petrpolis , rio de janeiro , brasil ) for 3 minutes and once again washed with sterile saline . suction was performed with a 30x4 gauge needle attached to a luer look syringe and the root canals were dried with sterile absorbent paper points ( tanariman industrial ltda , manacapuru , amazonas , brasil ) . \n the teeth were divided into two groups according to the root canal dressing used , as follows : group i - 10 m gallium nitrate solution ( acros organics , pittsburg , usa ; cas n 13494 - 90 - 1 ) ; and group ii - calcium hydroxide paste ( s.s . \n the animals received a single dose of 20,000 u.i . of benzathine g penicillin via intramuscular injection . \n five animals in each group were killed at 15 , 30 and 60 days after replantation by anesthetic overdose . \n the right side of the maxilla was separated from the left through the median line with a # 15 surgical blade . \n the maxilla was further sectioned close to the third molar to separate the hemimaxilla containing the replanted tooth . \n the specimens were fixed in 10% buffered formalin for seven days and decalcified in 4.13% edta ( ph 7 ) . \n the specimens were then processed , embedded in paraffin , and transversally sectioned through the cervical , middle and apical tooth thirds ; 5-m sections were obtained at each 100m with a microtome , adding up to 20 sections for each specimen . \n the following parameters were considered for microscopic analysis : integrity of tooth structure , presence of cementum , characteristics of the connective tissue formed in the periodontal space , presence of tooth ankylosis and occurrence of inflammatory or replacement resorption . \n morphometric analysis was also performed , with aid of the imagelab 2000 software version 2.4 ( diracom 3 , vargem grande so sul , sp , brasil ) , specific for measuring areas and perimeters . \n fifteen histological sections of each study period were photographed with a digital camera ( jvc tk-1270 color video camera , tokyo , japo ) attached to a microscope ( carl zeiss , axiolab , jena , germany ) connected to a pc computer . \n the images were stored as figures ( tif 24 ) for analysis in the imagelab software . \n the perimeter of areas showing ankylosis , connective tissue formed in the periodontal space , the amount of remaining cementum and the areas of inflammatory or replacement resorption were measured . \n the results were organized in tables and statistically analyzed by the kruskal - wallis test at a significance level of 5% . \n whenever significant differences were found , the dunn 's test was used to assay individual comparisons . \n analysis of histological sections of specimens in group i at 15 days revealed formation of a dense connective tissue , parallel ( figure 1 ) or perpendicular to the root surface ( figure 2 ) . \n the presence of a periodontal ligament - like connective tissue ( perpendicular direction ) was decreased over time and was no longer observed at 60 days ( table 1 ) ( figure 3 ) . \n tooth ankylosis was observed in all study periods in both groups ( table 1 ) . \n is statistically significantly different from examination of the root surface revealed the presence of cementum covering great part of the root surface , which decreased over time in both groups ( table 2 ) . \n inflammatory resorption lacunae were observed in all study periods ( table 2 ) ( figure 1 and 3 ) . in group \n i , replacement resorption lacunae were present at 60 days , different from the 15 and 30-day periods , when no replacement resorption was observed ( table 2 ) . in group \n ii , replacement resorption was present at 30 and 60 days ( table 2 ) . \n is statistically significantly different from comparison of results obtained in groups i and ii revealed that the presence of tooth ankylosis and replacement resorption lacunae was more evident in group ii at 30 days ( p<0.05 ) . \n in general , no statistically significant differences were observed between groups i and ii , demonstrating a similar mechanism of action of both chemicals when used as root canal dressings ( tables 1 and 2 ) . \n the frequency of occurrence of root resorption in the present study groups was low , although resorption events were increasingly observed over time . \n this emphasizes the characteristic of gallium nitrate2,3,4,5,6,19 and calcium hydroxide7,8,16,18 in limiting the resorption process . \n table 2 shows the effectiveness of gallium nitrate in inhibiting replacement resorption at 15 and 30 days , since resorption was greatly increased at 60 days ( p<0.05 ) . \n this observation may indicate the need for placement of successive dressings , which might inhibit the occurrence of replacement resorption for a longer period . \n the presence of cementum could be detected on the root surface of most specimens in both groups . \n this suggests the efficacy of sodium hypochlorite for removal of necrotic periodontal ligament present on the root surface with no injuries to the cementum layer9 . \n a direct proportion between the occurrence of root resorption and the absence of cementum was also noticed ( table 2 ) . regarding the histological events occurring in the periodontal space , both groups exhibited tooth ankylosis . \n the occurrence of tooth ankylosis is related to the loss of epithelial rests of malassez present in the periodontal ligament as a consequence of tooth avulsion , since these cells are responsible for maintaining the periodontal space11 . \n the low frequency of tooth ankylosis affecting the present study groups is described in table 1 . \n this finding may have influenced the occurrence of replacement resorption , which is a consequence of tooth ankylosis1,12,17 . \n the results of this study also demonstrate discrete presence of periodontal ligament - like connective tissue at 30 and 60 days ( table 1 ) , which was considerably higher at 15 days . \n this may be explained by the presence of periodontal tissue remnants from the alveolar wall reaching the root surface of the replanted tooth , and not by formation of new tissue . \n such periodontal tissue remnants were replaced over time by a newly formed connective tissue arranged in parallel direction to the root surface . \n according to the present results , both gallium nitrate and calcium hydroxide were able to limit root resorption in teeth submitted to replantation , yet they did not prevent its occurrence . concerning the periodontal space , formation of a periodontal ligament - like connective tissue was not favored in any group . \n hence , it may be concluded that the gallium nitrate solution and calcium hydroxide paste demonstrated similar performance .", "role": "user" }, { "content": "the purpose of this study was to test a gallium nitrate solution , a resorption inhibitor , employed as a root canal dressing in teeth submitted to late replantation . \n thirty maxillary right central incisors of rats were avulsed and kept dry for thirty minutes . \n the teeth were instrumented and the root surfaces were treated with 1% hypochlorite solution followed by application of 2% sodium fluoride . \n thereafter , the teeth were divided into two groups according to the root canal dressing : group i , solution of gallium nitrate ; and group ii , calcium hydroxide paste . \n the teeth were then replanted in their respective sockets . \n the animals were killed at 15 , 30 and 60 days after replantation and the samples were processed for morphometric and microscopic analysis . \n the results demonstrated that the gallium nitrate solution and the calcium hydroxide paste limited the root resorption , yet they did not impair its occurrence . it may be concluded that gallium nitrate solution and calcium hydroxide paste demonstrated similar performance .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: mountainous ecosystems are highly sensitive to any disturbances in natural balance , what makes them especially interesting for observations of chemical and radiochemical contamination . \n the area of the babia gra national park ( bpn ) , being located along the main ridge of the flysch carpathian mountains , is just the proper place in this respect . \n the terrain belongs to the young folded mountains , built mainly of flysch and some other lithology . \n the highest summit of the babia gra mountain s massif ( west part of the beskid ywiecki chain of mountains ) is diablak reaching 1,725 m asl . to protect this unique area \n the area of the bpn , like other regions of poland , was exposed to substantial contamination with radionuclides due to the chernobyl accident in 1986 and nuclear weapon tests since the fifties of the twentieth century [ 16 ] . \n researches on radionuclides distribution in the environment are important not only because these are perfect markers of the environment pollution but also because radioactive isotopes are not immobilized in soil but are constantly interchanged between inorganic matter and living organisms . \n the aim of the work was to establish the spatial distribution of -radionuclides i.e. anthropogenic cs ( t1/2 = 30.07 years ) and natural k ( t1/2 10 years ) in the soil samples collected in the described above area . \n 10 cm thick soil core samples were collected with the use of a cylindrical corer ( 10 cm in diameter ) . \n 13 sampling points were selected in the studied area , most of them localized along the mountains main ridge . \n each of the sampling points was described by its geographical coordinates and height above the sea level . in case of few points \n it was impossible to determine their precise coordinates ( the lack of gps signal ) , so the closest available coordinates were interpolated . in the laboratory , the cores were divided into three sub - samples each to enable radioisotopes determination in various depths . \n the samples were then dried at 105 c and the total weight was determined . \n after removal of organic macro - particles and small stones , the samples were sieved mechanically ( 2 mm mesh ) . \n after preparation , the samples were analysed by means of gamma ray spectrometry with the use of hpge ( high purity germanium ) coaxial detectors of relative efficiency 7 and 21 % . prior to the measurements , \n each measurement lasted 72 h. the reference materials iaea-154 , iaea-375 , and iaea-447 of the international atomic energy agency ( vienna , austria ) were used in the measurements . in the present work cs radioactivity in the \n for the whole core samples , cs and k activities were expressed in bq m units . \n after gamma spectrometric measurements organic matter content in the soil samples was determined by means of incineration at 600 c . to extract relevant information out of the obtained data , \n chemometric tools i.e. cluster analysis ( ca ) and principal components analysis were used ( using statistica 10 software ) . \n the most important values are written in bold.table 1the babia gra national park sampling points descriptionpoint no.geographical coordinatesheight ( m asl)place description1n 49st 34,7971,515red hiking trail , after sokolica , right from this red routee 19st 33,4142n 49st 34,6161,577near zimny staw lake , at the top of babia grae 19st 32,7723n 49st 34,3411,707by green hiking trail from babia gra to lipnicae 19st 31,8244n 49st 34,4261,701the yellow hiking trail from babia gra , on the right sight of this routee 19st 31,7695n 49st 34,3861,695the route from diablak ( red hiking trail)e \n 19st 31,6896no data1,600the route from diablak below cross ( red hiking trail)7no data1,530krzywka szlakw na markowe szczawiny i ma babi.8no data950mokry stawek lake , from the site of krowiarki pass9no data951brona pass10n 49st 34,5281,463by yellow hiking trail , dwarf pine zonee 19st 31,45311n 49st 34,6671,350yellow hiking trail , at the end of higher montane zonee 19st 31,30112n 49st 34,9531,257at the intersection of szumica woda valley with the route to markowe szczawinye 19st 31,46713n 49st 34,293836hala mietanowa , above little bridge on syhlec streame 19st 35,609table \n 2 \n cs and k radioactivities , density and organic matter content in the soil samples collected in the babia gra national park areapoint no.height ( m asl)soil density ( g cm)organic matter content ( % ) activity cs ( bq kg ) ( layer a)activity cs ( bq m ) ( whole core)activity k ( bq m ) ( whole core)11,5150.7017416 118,837 2416,900 61921,5770.1790405 163,757 2114,127 37431,7070.5520304.1 9.26,591 2247,914 71541,7010.6515 \n 831 21 \n 28,551 72016,477 1,49751,6950.7014246.2 8.48,166 27510,848 97761,6000.2473697 217,231 2192,134 \n 19471,5300.2281214 103,276 1777,869 71389500.99936.8 2.11,783 119 \n 25,654 2,28899510.,703765.8 6.42,119 15212,907 1,164101,4630.5021137.3 6.11,916 838,368 775111,3500.5719309 105,472 \n 230 \n 25,200 2,255121,2570.1782326 152,030 951,642 \n 151138360.4023408 1316,152 48417,480 1,573 the babia gra national park sampling points description \n cs and k radioactivities , density and organic matter content in the soil samples collected in the babia gra national park area soil in the bpn originates from the carpathian flysch waste . as in the area vegetation levels \n the resulting soil type classification is given in table 3 .table 3prevailing soil types of the considered sampling pointsvegetation levelheight ( m asl)prevailing soil typesampling pointsalpine zone1,6501,725lithic leptosol,3 , 4 , 5regosol skeletic , podzolic ranker ( leptosol)dwarf pine zone1,3901,650haplic podzol,1 , 2 , 6 , 7 , 10tangel \n ranker(umbric leptosol)higher montane zone1,1501,390dystric cambisol12 , 11partially haplic podzollower montane zone7001,150eutric and dystrict cambisol13 , 8 , 9 prevailing soil types of the considered sampling points chemometric tools are widely used in the environmental data analysis especially when high variability and uncertainty of the data may be expected . for the obtained set of data the ca according to ward and principal components analysis ( pca ) after initial normalization / auto - scaling , \n the outcome of the analyse is presented in figs . 1 and 2.fig . \n activity of cs [ bqm ] represents activity in the whole profile ; activity of cs [ bqkg ] represents activity in a layerfig . \n activity of cs [ bqm ] represents activity in the whole profile ; activity of cs [ bqkg ] represents activity in a layer cluster analysis ( samples ) in fig . \n the first one represents soil density and k radionuclide activity ( k activity is in constant relation to the total potassium , about 31 bq k per 1 g of potassium ) . \n these two variables belong to the same cluster what suggests that potassium is incorporated mainly in mineral components of the soil . \n the higher is mineral content of soil , the higher is soil density ( table 2 ) . in the second cluster , radioactivities of artificial cs in the a soil layer and in the whole core as well as the sampling point height asl and organic matter content , \n similar course of changes of cs activity and soil organic matter content can be attributed to the sorptive properties of this soil fraction . as cs presence in \n the soil results only from the distant transport of the contaminant , its retention in soil is a direct result of its sorption . \n the other important factor is the amount of precipitation in the area ( rain , snow ) . \n the higher is the location of the sampling point in the mountains , the higher is the amount of precipitation . \n thus , the variable altitude belongs to the same cluster . in fig . \n but the chosen criteria i.e. analysed variables , enabled proper and logical division of the sampling points into groups ( clusters ) for which the soil type is a common feature ( table 3 ) . \n however , it should be noted that in mountains , soil types are not sharply defined . as a result \n the revealed clusters do not classify soil types directly , but the obtained classification is relevant for a given soil complex structure ( fig . 2 ; \n principal components are the linear combination of the previous variables . taking into consideration the kaiser criterion in further analysis only the first and the second principal component should be used ( table 4 ) . \n the scree diagram suggests however , that it would be reasonable to consider three components as they describe 92.36 % of global variation of the data ( fig . \n 3scree diagram in table 5 factor loadings ( after varimax rotation ) for the three principal components are given.table 4eigenvaluesprincipal components analysiseigenvalues% total variancecumul .- eigenvaluecumul.-%12.7946.482.7946.4821.9732.784.7679.2630.7913.105.5592.3640.254.235.8096.5950.132.125.9398.7160.071.296.00100.00table 5factor loadings ( varimax)vf-1vf-2vf-3altitude0.0900.232 \n 0.942 \n density \n 0.950 \n 0.1070.096radioactivity of cs ( a \n layer)0.235 \n 0.906 \n 0.271radioactivity k \n 0.721 \n 0.0940.556radioactivity cs ( whole core)0.247 \n 0.945 \n 0.025organic matter \n 0.956 \n 0.1080.050significant values are given in bold factor loadings ( varimax ) significant values are given in bold the first component ( explaining 46 % of variability ) covers mainly the following variables : density , organic matter content and potassium concentration . \n all these parameters are connected with the physical properties and chemical composition of the soil . the second principal component , explaining 33 % of variability , considers only anthropogenic variable i.e. radioactivity of cs . \n third principal component explaining 13 % of variability refers to the height asl of the sampling point . \n it should be stressed , that this component supplies additional information related to some aspects explained by the first two principal components . \n the height above sea level has the influence on such soil properties like density and organic matter content . \n with increasing altitude the pace of organic matter decomposition decreases ( the same tendency is noted for temperature ) . \n simultaneously , anthropogenic cs was introduced to the soil mainly with rainfall in the mountains , the higher located is the place the more rain is precipitated . in fig . 4 \n it may be noted that the variables ( soil density and potassium content ) are positively correlated witch each other and negatively correlated with organic matter content . on the other hand \n it is clear that cs radioactivity is orthogonal to those variables ( soil density and potassium content)no correlation was found.fig . \n 4projection of the variables onto the plane of the first two principal components projection of the variables onto the plane of the first two principal components \n it was established that : caesium radioactivity in the soils of the bpn is changeable . in the top ( a layer ) 3 cm thick soil layer it is in the range from 36 to 831 bq kg , while in the whole core ( 10 cm ) it spans from 1,916 to 28,551 bq m. ca enabled grouping of the sampling points according to the soil type . \n it may be assumed that the sampling points are better characterized by the cs radioactivity expressed in surface units rather than radioactivity of the first layer only.the pca reduced the number of analysed variables to three principal components , explaining 92 % of the total variance of the variables . \n first three principal components differentiate the variables into : natural ( connected with physical and chemical soil properties ) like soil density , organic matter and potassium contents , anthropogenic cs radioactivity , and sampling points characteristic altitude above sea level . \n ( a layer ) 3 cm thick soil layer it is in the range from 36 to 831 bq kg , while in the whole core ( 10 cm ) it spans from 1,916 to 28,551 bq m. ca enabled grouping of the sampling points according to the soil type . \n it may be assumed that the sampling points are better characterized by the cs radioactivity expressed in surface units rather than radioactivity of the first layer only . \n the pca reduced the number of analysed variables to three principal components , explaining 92 % of the total variance of the variables . \n first three principal components differentiate the variables into : natural ( connected with physical and chemical soil properties ) like soil density , organic matter and potassium contents , anthropogenic \n cs radioactivity , and sampling points characteristic altitude above sea level . presented above conclusions \n it is true especially for the neighbouring tatra mountains national park [ 3 , 5 , 12 ] . \n the similar positive correlation of cs activity with the sampling point altitude and organic matter content in the soil were found in the tatras . \n soils of the mountains neighbouring to the bpn constitute currently the subject of complementary research projects .", "role": "user" }, { "content": "the aim of this work was to determine 137cs and 40k radioactivities in soil samples taken from the babia gra national park ( bpn ) in south poland . \n the cluster analysis ( ca ) and principal component analysis ( pca ) were used to discuss the obtained data . \n 10 cm thick soil cores were collected from the bpn area . \n each sample was divided into three sub - samples . \n the samples were dried , homogenized and packed in polyethylene containers . \n the radioactivities of 137cs and 40k were measured by means of gamma spectrometry . \n it was found that 137cs radioactivity in the whole 10 cm soil cores was in the range from 1,916 to 28,551 bq m2 . \n the radioactivity of 40k varied from 1,642 to 25,654 bq m2 . using ca it was possible to diverse the soils taking into account soil types . by use of the pca method , \n it was chosen three factors which are appropriate to characterize researched parameters .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: the medial vestibulospinal tract ( mvst ) , which descends in the medial longitudinal fasciculus ( mlf ) , may mediate the vestibular evoked myogenic potentials ( vemps ) in the contracting sternocleidomastoid muscle . \n we report herein abnormal vemps in a patient with medial medullary infarction ( mmi ) that appeared to involve the mlf . \n a patient with infarction involving the right medial medulla showed decreased p13-n23 amplitude and increased p13/n23 latencies of the vemps on the right side . \n these abnormal vemps recorded in an mmi patient support the theory that vemps are mediated by the mvst contained within the mlf . \n vemps may represent a valuable tool for investigating vestibular dysfunction originating from the saccule , even in patients with central vestibulopathies , which is not readily defined by conventional vestibular function tests . \n the term \" vestibular - evoked myogenic potentials ( vemps ) \" refers to inhibitory potentials recorded in the contracting muscles , usually the sternocleidomastoid muscles ( scm ) , when sound stimuli are applied.1 - 4 the main pathway for saccule - induced inhibitory postsynaptic potentials to ipsilateral scm motoneurons appears to be the medial vestibulospinal tract ( mvst ) , which descends within the medial longitudinal fasciculus ( mlf).5 however , vemps have not been measured in lesions involving the medullary mlf . \n we report herein abnormal vemps in a patient with medial medullary infarction ( mmi ) that appeared to involve the mlf . \n a 67-year - old man developed sudden vertigo and vomiting followed by dysarthria and left - sided weakness 9 days before admission to hospital . \n he had a history of diabetes , hypertension , and chronic renal failure . on admission \n , he had a blood pressure of 153/79 mmhg , a pulse rate of 75 beats / min , a respiration rate of 20 breaths / min , and a body temperature of 37.0. neurological examination revealed gaze - evoked nystagmus ( gen ) during both horizontal and upward gazes . \n both eyes deviated to the left during eye closure , and saccades were hypermetric to the left and hypometric to the right . \n this patient also exhibited dysarthria , dysphagia , and medical research council grade i weakness on the left side . \n the deep tendon reflexes were hypoactive in the extremities , but the toe sign was extensor in the left foot . \n laboratory tests were notable for increased serum glucose ( 279 mg / dl , normal range=70 - 110 mg / dl ) , hemoglobin a1c ( 7.7% , normal range=4.0 - 6.4% ) , blood urea nitrogen ( 71 mg / dl , normal range=10 - 26 mg / dl ) , creatinine ( 2.9 mg / dl , normal range=0.7 - 1.4 mg / dl ) , and uric acid ( 8.8 mg / dl , normal range=3.0 - 7.0 mg / dl ) . \n the results of bithermal caloric tests and measurements of ocular torsion and tilt of the subjective visual vertical ( svv ) were all normal . \n vemps exhibited decreased p13-n23 amplitude and increased p13/n23 latencies on the right side ( fig . \n 1 ) . mri documented a right mmi extending from the pyramid to the medullary tegmentum ( fig . \n 2a and b ) . magnetic resonance ( mr ) angiography disclosed diffuse narrowing of the right distal vertebral artery and focal narrowings of the left vertebral artery ( fig . \n our patient , who had an mmi on the right side , showed decreased vemp p13-n23 amplitude and increased p13/n23 latencies on the right side in addition to gen , saccadic contrapulsion , dysarthria , hemiparesis , and hemihypesthesia , which are characteristic findings of mmi.6 since the mlf contains the mvst , which is known to mediate vemps recorded in the contracting scms , the abnormal vemps recorded in our patient with mmi indicate disruption of the mlf in the medullary tegmentum where the mlf is located . \n vemps can be used to assess the sacculocollic reflex pathway , which travels downward in the lower brainstem.7 the saccule , which is a linear accelerometer in mammals , is an acoustic receptor in lower species , and the vestibular neurons sensitive to click sounds are of otolithic origin.5,8 in peripheral vestibulopathy , abnormal vemps are usually associated with canal paresis and the ocular tilt reaction ( otr).4 however , recordings of vemps in our patient with mmi disclosed decreased p13-n23 amplitude and increased p13/n23 latency on the ipsilesional side in the absence of caloric paresis , otr , or svv tilt . \n the principal brainstem areas of saccular nerve termination are the inferior vestibular nucleus and the lateral portion of the superior vestibular nucleus , while the principal projections from the utricle are to the laterodorsal medial vestibular nucleus , the ventrolateral superior vestibular nucleus , and the rostral portion of the inferior vestibular nucleus.9 the dissociation of abnormalities between the vemps and otr / svv tilt suggests differential involvement of the otolithic pathways in central vestibulopathies . in primates , saccule projects to the superior and inferior vestibular nuclei,9 and \n however , sound - evoked vemps recorded from the neck are almost completely unilateral.2,4,10 since the mvst contains inhibitory fibers , the sacculocollic inhibitory signal may travel down from the inferior vestibular nucleus mostly along the mvst to the ipsilateral scm,2,10 - 12 while the lateral vestibulospinal tract is excitatory . \n however , since the mvst originates mostly from the medial vestibular nucleus , the inhibitory sacculocollic responses may also be mediated by the medial vestibular nucleus.13,14 because the mlf is supplied by the anteromedial medullary arteries and is usually involved in mmi,6 damage to the mlf is most likely to be responsible for the abnormal vemps recorded in our mmi patient . \n this interpretation is also consistent with the associated gen and the lesion observed on mri extending to the medullary tegmentum , where the mlf is located . \n gen in mmi is ascribed to damage to the nucleus prepositus hypoglossi ( nph ) or cell groups of the paramedian tracts , which are also located in the medullary tegmentum.6 in conclusion , the recording of abnormal vemps in our mmi patient supports the theory that these potentials are mediated by the mvst contained within the mlf . \n the association between abnormal vemps and gen also indicates that the lesion in our patient involved the medullary tegmentum where the mlf and the nph are located . \n vemps may represent a valuable tool for investigating vestibular dysfunction originating from the saccule , even in patients with central vestibulopathies , which is not readily defined by conventional vestibular function tests .", "role": "user" }, { "content": "backgroundthe medial vestibulospinal tract ( mvst ) , which descends in the medial longitudinal fasciculus ( mlf ) , may mediate the vestibular evoked myogenic potentials ( vemps ) in the contracting sternocleidomastoid muscle . \n we report herein abnormal vemps in a patient with medial medullary infarction ( mmi ) that appeared to involve the mlf.case reporta patient with infarction involving the right medial medulla showed decreased p13-n23 amplitude and increased p13/n23 latencies of the vemps on the right side . \n these abnormal vemps recorded in an mmi patient support the theory that vemps are mediated by the mvst contained within the mlf.conclusionsvemps may represent a valuable tool for investigating vestibular dysfunction originating from the saccule , even in patients with central vestibulopathies , which is not readily defined by conventional vestibular function tests .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: antineutrophil cytoplasmic autoantibody ( anca)-associated vasculitis ( aav ) is a systemic vasculitis that predominantly affects the elderly ( 1,2 ) . for the primary induction of remission , \n aav is generally treated with glucocorticoids , cyclophosphamide , or rituximab , a newly available therapy ( 3,4 ) . as initial therapy , \n the japanese society of nephrology also recommends daily treatment with oral cyclophosphamide ( 25 - 100 mg / day ) or intravenous pulses of cyclophosphamide ( 250 - 750 mg / m ) with corticosteroids ( 5 ) . \n initial therapy with corticosteroids alone is indicated for cases in which aggressive treatment is required ; however , the use of immunosuppressive agents is not desirable , particularly in patients older than 70 years . \n diffuse alveolar hemorrhage ( dah ) is well known as a serious aav symptom associated with high mortality ( 6,7 ) . in a study of 107 patients with biopsy - proven anca - associated microscopic polyangiitis and glomerulonephritis , \n the relative risk of death was 8.65 times higher in patients with pulmonary hemorrhage than in those without ( 7 ) . \n however , the adverse effects of this therapy can not be overlooked , and glucocorticoids and immunosuppressive agents may increase the susceptibility to infection , which is a strong indicator of mortality in patients with aav ( 8) . \n plasma exchange is a promising option for the treatment of aav and is recommended in the guidelines ( 3,5,9 ) . because anca causes necrotizing glomerulonephritis and small vessel vasculitis , the removal of anca may contribute to the control of disease activity and the prevention of organ damage . although the guidelines recommend the addition of plasma exchange to corticosteroid and cyclophosphamide therapy in patients with advanced kidney dysfunction , whether or not plasma exchange is effective in patients with alveolar hemorrhage is unclear ( 10 ) . \n we herein present the case of a 76-year - old woman with severe aav and dah who successfully achieved remission following prompt initiation of plasma exchange and low - dose prednisolone . \n one year before admission , she had presented with a three - month history of dry cough and was admitted to a local hospital for an examination . \n chest - x ray showed ground glass opacity , and respiratory function testing showed an obstructive pattern . \n she was diagnosed with idiopathic pulmonary fibrosis and discharged with home oxygen therapy only when she experienced shortness of breath . at the time , her serum creatinine level was 0.56 mg / dl , and there were no abnormalities on urinalysis . \n three months before admission , she was again admitted to the local hospital for shortness of breath , anorexia , and weight loss ( 15 kg over 3 months ) , and her serum creatinine level was elevated to 2.23 mg / dl . \n in addition , urinary protein , hematuria ( 20 - 29 per high - power field ) , and markedly elevated myeloperoxidase ( mpo)-anca were detected . \n she was transferred to our hospital for further examination and treatment of suspected mpo - anca - associated glomerulonephritis . on admission , \n a physical examination revealed fine crackles , pale conjunctivae , clubbed fingers , body weight of 62.0 kg , body temperature of 36.7 , blood pressure of 132/60 mmhg , pulse rate of 80 beats / min , percutaneous oxygen saturation of < 90% on atmospheric air with a respiratory rate of 12 - 16 breath / min , and her birmingham vasculitis activity score was 24 . as shown in table , \n the serum creatinine level was 2.35 mg / dl , and the mpo - anca titer was 336 u / ml ( normal range , 0 - 3.5 u / ml ) ; normocytic anemia was detected . \n mcv : mean corpuscular volume , mch : mean corpuscular hemoglobin , mchc : mean corpuscular hemoglobin concentration , egfr : estimated glomerular filtration rate , tibc : total iron binding capacity , aptt : activated partial thromboplastin time , pt : prothrombin time , pr3 : proteinase-3 , mpo : myeloperoxidase , anca : antineutrophil cytoplasmic autoantibody , gbm : glomerular basement membrane , rbc : red blood cell , upcr : urinary protein to creatinine ratio on the day following admission , percutaneous renal biopsy was performed ( fig . \n 1 ) . the histopathological findings revealed cellular crescents in 5 out of 10 glomeruli . \n the microscopic findings of the glomerulus showed cellular crescent formation and disruption of the bowman s capsule ( a : periodic acid - schiff staining ; original magnification , 400 ) . \n high disease activity was indicated by fibrinoid necrosis in the lobular artery ( b : masson s trichrome staining ; original magnification , 400 ) . \n the immunofluorescence studies revealed no glomerular deposition ( c : igg , d : iga , e : igm , f : c3 , and g : c1q ) . \n 2 , intravenous methylprednisolone pulse therapy was administered every 24 hours for 3 consecutive days ; however , on the day following the pulse therapy ( day 5 ) , the patient 's respiratory condition suddenly worsened , and she required ventilatory support . \n dah was diagnosed on bronchoscopy and chest computed tomography , which showed broad infiltrative shadows and ground glass opacity in her lungs ( fig . \n , plasma exchange was initiated for a total of 7 times for 2 weeks , with approximately 1 time the predicted plasma volume ( estimated by the following formula : [ 0.065body weight ( kg)][1-hematocrit ] ) ( 11 ) per session , using freshly frozen plasma as the replacement solution . during this period , water - soluble prednisolone \n 60 mg ( equivalent oral prednisolone dose : 30 mg / day ) was administered intravenously . \n methylprednisolone pulse therapy was initiated to treat rapidly progressive glomerulonephritis due to anti - nuclear cytoplasmic antibodies . \n however , diffuse alveolar hemorrhaging occurred , and the respiratory condition rapidly worsened on day 5 after admission . \n there was an improvement in the respiratory condition , and the patient was successfully extubated on day 14 . \n her serum creatinine level decreased from a maximum of 2.99 mg / dl to 1.48 mg / dl , and the mpo - anca titer also decreased to the normal range . \n chest computed tomography on the sudden onset of dyspnea showed broad infiltrative shadows and ground glass opacity in the lungs ( left panel ) . \n the therapy with plasma exchange and low - dose prednisolone was dramatically effective , and these shadows were considerably attenuated after one month ( right panel ) . \n after 4 sessions of plasma exchange were performed , the fio2 decreased from 100% to 30% , and the positive end - expiratory pressure decreased from 10 cmh2o to 5 cmh2o . \n her respiratory condition improved , and she was successfully extubated on day 14 . the intravenous water - soluble prednisolone was discontinued , and oral prednisolone at a dose of 30 mg per day was started on day 29 . without using additional immunosuppressive agents , the serum creatinine level returned to 1.48 mg / dl , and the mpo - anca titer returned to the normal range . \n after extubation , the percutaneous oxygen saturation was 100% at rest on 1 - 2 l of oxygen per minute ; however , on exertion , the oxygen saturation dropped to < 90% , and she was discharged with home oxygen therapy . \n we successfully treated an elderly patient with severe aav and dah via the immediate introduction of plasma exchange . \n therefore , the combination of plasma exchange and low - dose glucocorticoid therapy without additional immunosuppressive agents may be a safe and effective treatment for aav in elderly patients . \n mechanical ventilation and admission to the intensive - care unit ( icu ) have been reported to be strong predictors of increased mortality within 28 days after admission ( 12 ) . however , despite requiring mechanical ventilation and icu admission , the present patient survived . \n high disease activity and the need for high doses of immunosuppressive agents have also been associated with a higher risk of death , probably due to infection and active vasculitis ( 13 ) . \n it is difficult but critically important to achieve an optimal balance of immunosuppression and avoid infection during the treatment of severe aav . \n although the mpo - anca titer had decreased before the initiation of plasma exchange , our patient still required mechanical ventilation and icu management for dah . \n after 4 sessions of plasma exchange were performed , the mpo - anca titer decreased from 188 u / ml to 26.2 u / ml . in a previous study , \n patients with dah treated by ventilator support needed at least five sessions of plasma exchange for recovery ( 14 ) . \n we performed seven sessions of plasma exchange in total , in accordance with the protocol of the ongoing pexivas study ( 15 ) . \n the pexivas study is being conducted to clarify the effectiveness and safety of adjunct plasma exchange and oral glucocorticoid regimens in patients with severe aav . \n patients with aav accompanied by kidney dysfunction ( egfr < 50 ml / min ) , necrotizing glomerulonephritis , or lung hemorrhage are included in this study and have been randomized to receive one of the following treatment regimens : 1 ) plasma exchange and either standard or low - dose glucocorticoids or 2 ) no plasma exchange and either standard or low - dose glucocorticoids . \n although the study is still ongoing , klemmer et al . reported that dah accompanied by aav resolved during plasmapheresis in all 20 patients evaluated ( 14 ) . according to a previous study ( 9 ) \n , 137 newly diagnosed aav patients with elevated serum creatinine levels were randomly assigned to receive 7 sessions of plasma exchange or 3,000 mg of intravenous methylprednisolone ( both groups received oral cyclophosphamide and prednisolone ) . \n most deaths occurred during the first three months , when steroid dosages were the highest and vasculitis was the most active . at three months , however , plasma exchange was associated with a reduction in risk of death and progression to end - stage kidney disease compared with methylprednisolone . based on these results , we performed combined therapy of 7 plasma exchanges over 2 weeks and methylprednisolone pulse therapy , followed by low - dose prednisolone ( 0.5 mg / kg / day ) in the present study . \n although aav is commonly treated with oral prednisolone ( 1 mg / kg / day ) in combination with other immunosuppressants , low - dose prednisolone alone was prescribed because of our patient 's age and the preceding lung injury . as a result \n , the disease was controlled with plasma exchange and low - dose prednisolone only , and we succeeded in reducing the risk of treatment - related toxicity , such as infection . in conclusion , our present findings suggest that the treatment protocol of seven plasma exchanges for two weeks and low - dose prednisolone may be an effective and safe means of inducing the remission of severe aav in elderly patients . \n ", "role": "user" }, { "content": "antineutrophil cytoplasmic autoantibody ( anca)-associated vasculitis is a life - threatening disease characterized by rapidly progressive glomerulonephritis ( rpgn ) and diffuse alveolar hemorrhage ( dah ) . glucocorticoids and immunosuppressants are commonly used to treat this disease but may induce irreversible side effects , particularly in elderly patients . \n we herein report the case of a 76-year - old woman with rpgn . \n after methylprednisolone pulse therapy , dah occurred , and she required ventilatory support . \n after plasma exchange , her serum creatinine level improved , and she was discharged with home oxygen therapy . \n immunosuppressive agents other than glucocorticoids were not required . in conclusion , \n plasma exchange with glucocorticoid therapy may be effective in treating severe anca - associated vasculitis in elderly patients .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: obesity is associated with an increased risk of multiple health problems and many comorbid conditions , such as cardiovascular diseases , type 2 diabetes , hypertension , certain cancers , and sleep apnea / sleep - disordered breathing.14 in addition , obesity is becoming a global pandemic,5 and there has been an explosive increase in its prevalence worldwide in recent times.6,7 this has led to increased morbidity and mortality globally.6,7 cardiovascular diseases produce the greatest morbidity and mortality among these comorbidities , and the greater the obesity , the higher the morbidity and mortality rates.1 cardiovascular diseases that have been associated with obesity include hypertension , heart failure , arrhythmias , ischemic heart disease , obesity - related cardiomyopathy , stroke , and sudden death.5 the end result of this is loss of productivity and increased burden on health care systems . altered metabolic profile and changes in the structure and function of the cardiovascular system \n have been suggested to be responsible for the morbidities seen in obesity.5 the development of obesity has been attributed to the combination of environmental and genetic factors . \n presently , there over 35 independent genetic risk loci that have been identified for obesity.8 the genetic loci regulate eating habits as well as body metabolism and the body s response to exercise and eating.5 these loci lead to the development of obesity in environments favorable to it.5 increasing numbers of communities worldwide are becoming favorable to the development of obesity or overweight as a result of dietary changes and sedentary living . the prevalence of obesity is rising in urban adult nigerians911 and preschool children,12 and mirrors the ongoing global epidemic in children and adults.5 in many communities in nigeria , there are no inbuilt community facilities to encourage regular exercise , and the combination of poverty and poor nutritional education make individuals adopt poor dietary habits . \n some data exist on the prevalence of overweight and obesity among urban and semiurban adult nigerians.10,11,1319 however , limited information exists on the prevalence of cardiometabolic risk factors , including overweight and obesity , in the adult nigerian population , especially in the rural setting.2025 this study therefore assessed the prevalence and pattern of overweight and obesity in adult populations living in three rural communities of the ife north local government area ( lga ) of osun state , southwest nigeria . \n this will further help define the prevalence and pattern of overweight and obesity in this rural population , and also add to the national and global database . \n a total of 777 adults between 20 and 90 years of age from three rural towns of the ife north lga of osun state , southwest nigeria , were recruited into this cross - sectional study , which was carried out over a 6-month period using a multistage proportional stratified random sampling technique.26 the minimum sample size was determined using the formula for calculating sample size in the population,27 and the population of the lga was 153,274 , according to the 2006 population census.28 the study procedure followed the world health organization ( who ) guidelines for conducting community surveys.29,30 the dominant occupation and economic activities of the people of the lga center around farming , agro - allied productions , trading , artisanship , school administration , teaching , and cottage industries.31 there are civil servants in the services of local , state , and federal governments with other corporate and private businesses . \n the dominant language is yoruba , and the lga is the home of yoruba ethnic group.31 the permission of the traditional rulers of each community was sought and informed consent obtained from each participant . \n the three major towns ipetumodu , edunabon , and moro were randomly selected from the seven major towns of ife north lga . \n ten percent of the census enumeration areas ( eas ) were randomly selected from each town which translated to 80 eas for ipetumodu , 34 eas for edunabon , and 13 eas for moro . \n six households per ea were randomly selected in ipetumodu , ten households per ea in edunabon , and 23 households per ea in moro . in each household , at least one eligible and consented respondent ( aged 20 years and older ) was interviewed . \n ethical approval was given by the ethics and research committee of the obafemi awolowo university teaching hospitals complex , ile - ife . \n all sociodemographic data were obtained by using a standard questionnaire , and anthropometric variables were measured.32 the international classification of adult overweight and obesity according to body mass index ( bmi ) adapted from the who33 was used to define overweight and obesity . \n weight was measured in kilograms with a bathroom weighing scale , with subjects standing , arms hanging naturally by the sides , and footwear off . \n height was measured using a stadiometer in meters at the crown of the head without any footwear or headgear and the patient looking straight ahead.32 using this classification,33 bmi was categorized as follows : underweight = < 18.5 kg / m normal = 18.524.9 kg / m overweight = 2529.9 kg / m class i obesity = 3034.9 kg / m class ii obesity = 3539.9 kg / m class iii ( commonly called severe or morbid obesity ) = 40 kg / m waist circumferences ( wc ) and hip circumference ( hc ) were also measured according to the who protocol.30,34 wc values 102 cm for males and 88 cm for females were regarded as abdominal obesity.24 the data were summarized using descriptive statistics ( mean , standard deviations , and percentages ) . \n independent t - tests were used to compare the age and physical characteristics of males and females . \n significant differences of values of variables across the age - groups were determined by analysis of variance . \n multiple comparisons between the groups were performed by one - way analysis of variance with duncan s post hoc test . \n the data were analyzed using spss 16 ( spss , chicago , il , usa ) software . \n the data were summarized using descriptive statistics ( mean , standard deviations , and percentages ) . \n independent t - tests were used to compare the age and physical characteristics of males and females . \n significant differences of values of variables across the age - groups were determined by analysis of variance . \n multiple comparisons between the groups were performed by one - way analysis of variance with duncan s post hoc test . \n the data were analyzed using spss 16 ( spss , chicago , il , usa ) software . \n a total of 777 adults participated in the study ( 385 [ 49.5% ] men and 395 [ 50.5% ] women ) , as shown in table 1 . \n the meanage and bmi of the participants were 36.314.3 years and 23.534.6 kg / m , respectively . \n the mean bmi for males and females was 23.84.6 kg / m and 23.24.6 kg / m , respectively , as shown in table 2 . \n the overall crude prevalence of overweight and obesity in the total population was 20.8% and 8.4% , respectively . \n obesity using bmi increased across the age gradient , peaking in the 51- to 60-year age - group in men and women ( table 3 ) . \n mean wc and hc also increased across the age gradient , peaking in the 51 to 60-year age - group in the total population ( table 4 ) . among the overweight and obese subjects ( n=227 ) , \n 70.9% of them were overweight and the remaining 29.1% were obese ( table 5 ) . \n class i obesity was the most common obesity pattern , accounting for 20.7% among these overweight and obese subjects ( table 5 ) , 71.4% of the obese subjects , and 6% of the total study population ( table 3 ) . \n the prevalence of overweight and obesity in these three rural communities of 20.8% and 8.4% , respectively , is less than the prevalence rates of overweight and obesity of 32.7% and 22.2% in an urban community in southwest nigeria,35 35.1% and 9.8% in north - central nigeria,36 and 23.4% and 17.1% by amoah in ghana,37 respectively . \n this finding thus corroborates earlier studies that observed a higher prevalence of obesity and/or overweight in urban and semiurban populations compared with rural populations in africa.10,11,13,15,17,20,3841 therefore , the prevalence of obesity in these rural communities is less than the 12.5% we obtained in the neighboring semiurban community in ile - ife,11 the 21.3% obtained by sani et al in an urban community in katsina , northern nigeria,15 and the 8.82% by olatunbosun et al obtained in an urban community in ibadan , southwest nigeria.10 oyeyemi et al38 also obtained a overweight prevalence of 22.8% among urban adults in maiduguri , northern nigeria , higher than the prevalence of 20.8% in this study . \n in addition , adediran et al,13 in their recent study of anthropometric differences among natives of abuja , nigeria living in urban and rural communities , found that all anthropometric indices of overweight and obesity considered in their study were significantly higher in the urban than in the rural population . \n overweight and obesity were significantly prevalent among their urban population.13 ntandou et al in the republic of benin also showed a positive rural \n urban gradient in the prevalence of abdominal obesity,40 and mean bmi in this study was lower than that reported among adults in an urban area of abuja , nigeria39 and us blacks.41 it has been suggested that better nutrition and increased physical activity are responsible for this positive rural \n urban gradient.40 mean wc and hc increased with age , and peaked in the 51- to 60-year age - group . \n mean bmi increased with age in both sexes , and peaked in the 51- to 60-year age - group in males and females . \n this peak is similar to the findings of adediran et al39 in abuja , who found the peak in the 50- to 59-year age - group in both sexes . \n earlier reports indicated that the prevalence of obesity in the rural population in nigeria is relatively low . \n for example , okesina et al23 in 1999 reported obesity prevalence of 2% in three rural villages around maiduguri in northeastern nigeria . \n however , recent prevalence reports indicated that the prevalence is increasing , and they confirmed our finding of increased prevalence of overweight and obesity in this rural adult nigerian population.14,18,20,21,24 iloh et al25 also reported a high prevalence of obesity among rural adult nigerians in imo state , southeastern nigeria , of 6.0% . \n they also found that class i obesity ( 86.1% ) was the most common pattern among this rural population , which supported our finding that class i obesity ( 71.4% ) was the most common obesity pattern in our obese subjects . \n this study finding of increased prevalence of overweight and obesity in the rural adult nigerian population , supported by other recent reports,14,18,20,21,24,25 should alert the health authorities at all levels of government and community leaders on the need to include the prevention and control of overweight and obesity as well as the risk of obesity comorbidities in the planning of health services for these rural communities . \n there should be regular community education on lifestyle modification , and regular health screening to facilitate the early detection of overweight and obesity , as well as the institution of appropriate preventive and therapeutic measures.20,25 \n the prevalence of overweight and obesity in the three rural communities was 20.8% and 8.4% respectively , indicating a trend towards increased prevalence . \n class i obesity is the most common obesity pattern , and obesity increased across the age gradient , peaking in the 51 to 60-year age - group in both men and women . \n there is a need for regular community education on healthy lifestyles , and regular health screening to control the rising prevalence of overweight and obesity , as well as to prevent or reduce the risk of obesity comorbidities in these communities .", "role": "user" }, { "content": "backgroundlimited data exist on the prevalence of overweight and obesity in the nigerian adult rural population . \n this study therefore assessed the prevalence and pattern of overweight and obesity in adults in three rural communities of the ife north local government area , nigeria.materials and methodsa total of 777 adults between 20 and 90 years of age were recruited into this cross - sectional study , which was performed over a 6-month period using a multistage proportional stratified random sampling technique . \n sociodemographic data and anthropometric variables were obtained.resultsa total of 385 ( 49.5% ) men and 395 ( 50.5% ) women participated in the study . \n the mean age and body mass index of the participants were 36.314.3 years and 23.534.6 kg / m2 , respectively . \n the overall crude prevalence of overweight and obesity in the total population were 20.8% and 8.4% , respectively . \n obesity increased across the age gradient , peaking in the 51- to 60-year age - group in men and women . among the overweight and obese subjects ( n=227 ) , 70.9% of them were overweight and the remaining 29.1% were obese , with class i obesity accounting for 20.7% of these overweight and obese subjects.conclusionthe prevalence of overweight and obesity in these communities was 20.8% and 8.4% respectively , indicating a trend towards increased prevalence . \n class i obesity is the most common obesity pattern , and obesity increased across the age gradient , peaking in the 51- to 60-year age - group . \n there is a need for regular community education on healthy lifestyles , and regular health screening to control the rising prevalence of overweight and obesity , as well as to prevent or reduce the risk of obesity comorbidities in these communities .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: chronic fatigue syndrome ( cfs ) is characterized by severe and disabling fatigue in addition to other disturbances in concentration , short - term memory , sleep disruption , and musculoskeletal pain . community prevalence rates of cfs range between 0.2% and 2.6% . in the united states \n women are between 1.3 and 1.7 times more likely to be diagnosed with the syndrome than men . \n cfs patients are heavy users of health services and almost every family doctor has patients with cfs . in the united states , cfs results in an annual cost of $ 9.1 billion due to medical expenses and lost productivity . \n several studies suggest that serious childhood stressors such as physical abuse , parental addictions and lower socioeconomic status increase an individual 's vulnerability to developing cfs . \n one factor that , to our knowledge , has not been examined as a risk factor for cfs is parental unemployment . \n there is considerable evidence to suggest that parental unemployment is associated with a wide range of negative educational , social , health behaviors , and mental health outcomes for children , adolescents , and adults [ 5 , 6 ] . \n parental unemployment is associated with poorer school performance and negative health outcomes among children and adolescents . \n children of unemployed parents are more likely to report being bullied and socially isolated at school , and to experience multiple changes of schools . \n this may significantly impact learning , as children of unemployed parents have poorer grades and drop out of school earlier than their peers with employed parents . in cases where unemployment leads to extreme poverty , children are vulnerable to food insecurity and nutritional problems which may also influence school performance . young children with unemployed parents appear to be more susceptible to psychosomatic symptoms such as headaches , stomach - aches , and problems with eating and sleeping . in comparison to children of employed parents , those with parents who are unemployed are more likely to have accidents and to be hospitalized . \n in addition to unintentional injuries , unemployed fathers are three times more likely to abuse their children than employed fathers . \n parental unemployment is also associated with lower levels of self - esteem and higher levels of anxiety and mental illness in children and adolescents . \n the strong association between parental unemployment and child and adolescent depression , suicidal ideation , self - harming behaviors , and suicide has been well established . \n christoffersen suggests that the increased risk of suicide is due to the loss of family support caused by the upheaval of unemployment . \n other factors such as depression , anxiety and substance abuse and the physical and sexual abuse that sometimes accompanies parental unemployment may also influence children 's suicidal behaviour . \n these factors in turn may result in increased parental conflict , higher rates of divorce , and poorer parenting behaviors exacerbating mental health concerns in children . \n a few panel studies have followed the children of unemployed and employed parents into later adolescence and adulthood to document the long - term social and educational outcomes of parental unemployment [ 5 , 9 ] . \n these studies indicate that the children of unemployed parents are more likely to leave high school before graduation , to experience higher rates of unemployment themselves ( particularly true of men ) , and to have lower incomes than their peers who did not experience parental unemployment [ 5 , 9 ] . according to flouri and buchanan , \n these children are not exposed to working role models in the home and may not develop the necessary behavioral correlates of work . \n parental unemployment has been demonstrated to reduce social networks for youth and adults thereby decreasing social capital . \n the literature examining the long - term outcomes of childhood parental unemployment on mental health and health is limited . \n kaltiala - heino and colleagues report that children who have experienced parental unemployment are more likely to experience depression in adulthood . \n although negative health behaviors such as smoking are associated with parental unemployment in childhood , surprisingly little is known about links between parental unemployment and physical health . \n this gap in the literature extends to the relationship between parental unemployment and subsequent chronic fatigue syndrome . \n other adverse childhood experiences , such as physical abuse and parental addictions , have been identified as important risk factors for chronic fatigue syndrome . in this study \n we examine the association between parental unemployment and subsequent chronic fatigue syndrome , while controlling for a wide range of possible mediating factors including demographics , other childhood adversities , adult stressors and socioeconomic status , adult health behaviors and hypertension , and mental health . \n childhood parental unemployment often occurs with or exacerbates the presence of poverty , physical abuse , sexual abuse , parental mental illness , parental addictions , parental conflict , divorce , step - family situations , and single parent families . \n a number of adverse childhood experiences have been implicated in the development of chronic fatigue syndrome in adulthood . \n known risk factors for cfs include poverty , childhood physical and sexual abuse , and low socioeconomic status . \n parental unemployment has been associated with adult health behaviors such as smoking . in women , \n chronic fatigue syndrome has also been associated with adverse health behaviors such as tobacco use . \n kennedy and colleagues report some emerging studies suggesting that obesity and hypertension may be additional factors that impact the development of cfs . as documented earlier , childhood experiences of parental unemployment influence adult education , lower household income , job status , daily stress , and divorce [ 5 , 18 ] . \n marital status is used as a proxy for social support and has also been identified as an important factor in determining severity and functional impairment in cfs patients . \n jason and colleagues in a community - based study of cfs found those with lower levels of education and occupational status had the highest prevalence of cfs . \n the long - term impacts of parental unemployment on mental health include but are not limited to depression , anxiety , and lower self - esteem [ 1416 ] . in turn , the prevalence of chronic fatigue syndrome is markedly higher among those with anxiety disorders and depression . \n reports suggest that individuals who suffer from cfs also have a high prevalence of trauma and stressful life events . \n it is important to understand the relationship between parental unemployment and cfs in the light of the present economic climate . \n there are few studies that have examined the long - term health outcomes of childhood parental unemployment . to our knowledge \n , no studies have examined the link between childhood parental unemployment and cfs . using a large regionally representative community sample , this study seeks to examine the association between parental unemployment and cfs in adult women , while controlling for risk factors including demographics , other childhood adversities , adult health behaviors and hypertension , adult stressors and ses and mental health . \n we conducted a secondary data analysis of female respondents from the manitoba and saskatchewan sample of the 2005 canadian community health survey ( n = 7 , 244 ) . \n respondents were asked whether , as a child or a teenager , their father or mother ( did ) not have a job for a long time when they wanted to be working ? \n individuals were also asked to indicate whether they had long - term conditions lasting six months or more that were diagnosed by a health professional . specifically they were asked do you have chronic fatigue syndrome ? \n the decision about which variables to include was hypothesis driven , based on our review of the literature . in our first and all subsequent models , we controlled for age and race . in the second model we controlled for the number of other childhood adversities including childhood physical abuse , parental divorce , and parental addictions . \n the third model included adult health behaviours such as obesity , smoking , and alcohol use . in the fourth model we controlled for adult stress and socioeconomic status , which included factors such as self - reported stress , education , and income and marital status ( a proxy for social support ) . \n the fifth model controlled for mental health conditions that were indicated through self - report of a medical diagnosis of ( 1 ) mood disorders and ( 2 ) anxiety disorders . \n finally we adjusted for all of the above - mentioned risk factors in our last model . \n figure 1 illustrates the odds ratios associated with our key variable of interest : parental unemployment . \n this figure presents the association between parental unemployment and cfs varied when we controlled for each cluster of risk factors . due to \n missing data for the independent variables , the sample sizes varied from n = 7 , 193 in the age and race adjusted model to n = 7 , 088 in the fully adjusted model . \n we used a weighting variable created by statistics canada to adjust for the probability of selection and nonresponse in all our models . \n respondents were asked if they had experienced the following stressors during childhood and adolescence : ( 1 ) were you ever physically abused by someone close to you ? ( 2 ) \n did your parents get a divorce ? ( 3 ) did either of your parents drink or use drugs so often that it caused problems for the family ? respondents answers were summed to get the number of other childhood adversities . \n body mass index ( bmi ) was divided into four categories ( bmi < 25 , overweight , obese , and missing data category ) . \n hypertension was based on the respondent 's self - report that they had been diagnosed with high blood pressure by a health professional . \n educational attainment , self - reported stress , marital status , and household income were examined under adult stressors . \n these stressors were categorized as following : educational attainment ( < high school graduation , high school graduation , postsecondary graduation ) ; daily stress ( no stress to a bit of stress , quite a bit to extreme stress ) ; marital status ( not married , married ) ; income ( < $ 15,000 , $ 15,000$29,999 , $ 30,000$49,999 ; $ 50,000 or more , missing data category ) . \n the two mental health conditions of interest in our study were depression and anxiety disorders . \n women were asked ( 1 ) do you have a mood disorder such as depression , bipolar disorder , mania or dysthymia ? \n ( 2 ) do you have an anxiety disorder such as a phobia , obsessive compulsive disorder , or a panic disorder ? . \n we found a strong and significant association between long - term parental unemployment in childhood and chronic fatigue syndrome in women ( see figure 1 ) . \n when we adjusted for age and race , we found the odds ratio for cfs among those women who reported parental unemployment to be 4.12 ( 95% ci : 2.60 , 6.52 ) when compared to those who did not report parental unemployment . \n when we adjusted for number of other childhood adversities , the odds ratio for cfs among those reporting parental unemployment dropped substantially to 3.13 ( 95% ci : 1.91 , 5.13 ) . \n adjustments for adult health behaviors and hypertension did not change the odds ratio when compared to the age and race adjusted model ( or = 4.15 , 95% ci : 2.60 , 6.62 ) . \n the odds ratios for cfs among women who had experienced parental unemployment in the models controlling for adult stressors and socioeconomic status and adult mental health were relatively similar ( or = 3.79 , 95% ci : 2.35 , 6.1 and or = 3.61 , 95% ci : 2.26 , 5.77 ) . \n finally when controlling for all the above risk factors , the odds ratio for cfs among women reporting parental unemployment dropped to 3.05 ( 95% ci : 1.81 , 5.14 ) , but remained statistically significant . \n table 1 shows the age - race adjusted model and the fully adjusted logistic analysis of chronic fatigue syndrome . in the fully adjusted model , in addition to parental unemployment five other characteristics \n were associated with cfs : white women had five times higher odds of cfs than did non - whites . in comparison to those who had not finished high school , those who had finished high school had twice the odds of cfs . in comparison to respondents who had a household income of $ 50,000 or more , \n women who had an income of less than $ 15,000 had almost four times the odds of cfs and women with an income of $ 30,000$49,999 had twice the odds . \n those reporting more stress had three times the odds of cfs compared to women with lower levels of stress . \n women with anxiety disorders had three times the odds of cfs in comparison to those who had not been diagnosed with anxiety disorders . \n this study examined the association between long - term parental unemployment and chronic fatigue syndrome when controlling for other childhood adversities , adult health behaviors , adult stressors , adult socioeconomic status , and adult mental health . \n we found a strong and significant association between parental unemployment and cfs after adjusting for these factors . \n our analysis suggests that adjusting for number of other childhood adversities substantially reduced the parental unemployment - cfs relationship although parental unemployment was still significantly associated with cfs . in the literature \n , one childhood stressor in particular , physical abuse , has been implicated as an important risk factor for cfs [ 16 , 24 ] . \n albeit substantially reduced , the relationship between parental unemployment and cfs remains strong and significant after controlling for the number of other childhood adversities including physical abuse , parental divorce , and parental addictions . \n adult health behaviors and hypertension do not appear to affect the relationship between parental unemployment and cfs . \n felitti and colleagues suggested that health behaviors were the major pathways that mediated the relationship between adverse childhood experiences such as parental unemployment and adult health conditions . \n adult health behaviors such as smoking , alcohol use , and drug abuse may be used as coping mechanisms in adulthood . \n although adult health behaviors have been associated with both cfs and parental unemployment , it appears that these variables do not influence the relationship directly . \n including adult stressors resulted in a slight decrease in the relationship between parental unemployment and chronic fatigue syndrome . \n we anticipated that parental unemployment would lead to lower socioeconomic status and greater distress in adulthood , which in turn would influence cfs . \n christoffersen hypothesized that the relationship between parental unemployment and a child 's eventual career trajectory may be mediated through self - esteem and self - image , thus impacting social and economic well - being in adulthood . \n mental health conditions such as mood and anxiety disorders have been strongly linked to both cfs and parental unemployment [ 9 , 14 , 26 , 27 ] . \n however the association between parental unemployment and cfs remained robust after controlling for both these mental health conditions . to our knowledge \n this was the first study to examine the association between parental unemployment and cfs in a large population - based study of women . \n even after controlling for the number of other childhood adversities , adult health behaviors and hypertension , adult stressors , socioeconomic status , and adult mental health , there remains a strong relationship between parental unemployment and cfs . \n future research would benefit from examination of the potential role that the hypothalamic - pituitary - adrenal ( hpa ) axis plays in the relationship between parental unemployment and cfs . \n although our research could not explore this issue , hpa axis dysfunction has been previously implicated in functional somatic syndromes such as cfs . \n there appears to be a strong and significant correlation between childhood adversity and the development of cfs . \n heim and her colleagues have examined the role of the hpa axis in influencing the relationship between another childhood stressor ( i.e. , abuse ) and cfs . \n they found early childhood adversity and , in particular , abuse early in life to be associated with cfs . \n they hypothesized that the relationship may be mediated through physiological pathways that cause hypocortisolism , which , in turn , may cause fatigue . \n accordingly , they found that individuals who were abused in childhood had between 3 and 8 times the risk of developing cfs in comparison to those who were not abused . \n the cchs survey is a cross - sectional study and therefore does not permit causal analysis of the data . \n retrospective data was used to assess childhood parental unemployment ; prospective studies would minimize the likelihood of differential recall among participants . \n chronic fatigue syndrome status was based on the respondents ' report of a diagnosis by a health professional . \n the chances of underreporting are high as research suggests that fewer than 20% of individuals with cfs have been diagnosed . \n childhood parental unemployment remained an independent risk factor for chronic fatigue syndrome even after adjustment for most of the known risk factors for cfs . \n it may therefore be useful for clinicians and researchers to assess patients exhibiting symptoms of cfs for childhood experiences of parental unemployment and other adverse childhood experiences . \n future research should also examine the role of other potential risk factors in mediating the relationship between parental unemployment and cfs . \n the risk factors we examined in this study vary in their importance ; however , childhood physical abuse appears to play an important role in development of cfs and should be examined further . \n this study provides additional support to the emerging literature on the long - term mental and physical health consequences of parental unemployment . in conjunction with the substantial literature suggesting short - term negative outcomes for the well - being and mental and physical health of the children and adolescents of unemployed parents , the study underlines the urgency of adopting policies and programs to decrease unemployment levels and to provide an expanded social safety net for those who are out of work . \n the intergenerational transfer of the negative consequences of unemployment makes it imperative that as a nation we move quickly to improve employment opportunities .", "role": "user" }, { "content": "objective . \n the association between long - term parental unemployment in childhood and chronic fatigue syndrome was examined in a population - based sample of women . methods . a secondary analysis of data from a regionally representative sample of women ( n = 7 , 288 ) from the canadian community health survey ( 2005 ) was conducted using logistic regression . age and race as well as the following clusters of factors were controlled for : ( 1 ) other childhood adversities , ( 2 ) adult health behaviors and hypertension , ( 3 ) adult stressors and socioeconomic status , and ( 4 ) adult mental health . results . when adjusting for age and race only , the odds ratio of chronic fatigue syndrome among those reporting parental unemployment was 4.12 ( 95% ci : 2.60 , 6.52 ) compared to those not reporting parental unemployment . when controlling for age and race plus all four clusters of factors the odds ratio for chronic fatigue syndrome dropped slightly to 3.05 ( 95% ci : 1.81 , 5.14 ) , but remained statistically significant . \n conclusions . \n this study provides evidence for a significant association between long - term parental unemployment in childhood and chronic fatigue syndrome even after controlling for a wide range of potential risk factors .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: burning mouth syndrome ( bms ) is a chronic pain syndrome primarily affecting elderly women with hormonal changes or psychological disorders . \n it is multifactorial in origin , often idiopathic and its etiopathogenesis , majorly being neuropathic , largely remains enigmatic . \n the etiopathogenesis seems to be complex and in a majority of patients involves interaction among local , systemic or psychogenic factors . \n pathological lesions of the oral mucosa are usually not evident while the primary complaints are burning and dryness . \n stated that patients with burning mouth complain of intense to unbearable burning or pain that may interfere with eating . \n other common lesions associated with burning sensation include lichen planus , oral submucous fibrosis , candidiasis , allergy , nutritional factors , diabetes , menopause and psychogenic factors . \n when no clinical lesions are found in accordance with the burning sensation in the oral cavity the condition is termed as bms and is said to have a strong psychological relationship with the patients being under anxiety and stress . this can be either attributed to age related or hormonal related factors . \n the diagnostic label of bms should be given after a thorough clinical and laboratory workup , which does not reveal any possible etiological explanation towards the symptom of burning sensation . \n . therefore they comment that a pure psychogenic origin can not be inferred in all cases of burning mouth , and organic cause must be sought . \n pajukoski et al , stated that burning sensation can be a very distressful condition especially when its etiology can not be elucidated . if the burning is due to the visible lesions , treating it will resolve the problem . \n reiss and reiss mention that for appropriate management of a patient with burning , a multidisciplinary approach is needed which includes dentistry , neurology and internal medicine . \n lowental and pisanti say that local measures with reassurance and minor tranquilizers can treat this condition . \n the dentist should rule out any pathological process seen in the oral cavity , which is liable to cause burning . \n common causes of burning sensation in the oral cavity are discussed in the upcoming text . \n as the age advances , there are invariable changes occurring in the body . particularly in women \n moreover , as one ages the mucosa becomes thinner and dry , so complains such as burning , dryness and distorted taste becomes far more common . \n xerostomia is also an important factor in postmenopausal women as salivary secretion plays a vital role in the integrity of the oral tissues , in the selection and preparation of food for digestion . in patients with sjogren 's syndrome or treated with radiotherapy for oral cancer , there is virtually no salivary secretion . \n they also complain of burning sensation , dysguesia , difficulty in speech and there may be lobulation and fissuring of the tongue.[79 ] a number of medicines like antidepressants , antipsychotics , anti - histaminics and anti - diarroheal cause burning sensation in the oral cavity . \n brown uses the term scaled mouth syndrome ( sms ) to describe the burning caused by angiotensin converting enzymes inhibitors ( ace ) . \n lotti et al , states that lithium , grisiofulvin , antibiotics and metronidazole cause burning sensation in the oral cavity . to determine the prevalence of burning mouth symptom , in elderly women and evaluate local and systemic causes responsible for burning sensation \n to determine the prevalence of burning mouth symptom , in elderly women and evaluate local and systemic causes responsible for burning sensation . \n 105 elderly postmenopausal women were included in the study , out of which 05 dropped out and 56 ( out of remaining 100 ) had a chief complaint of burning sensation . \n patients were assessed by a strictly coordinated management protocol based on conventional guidelines namely history and clinical examination . in this study , burning mouth as a symptom was evaluated and the prevalence of burning mouth in middle and elderly aged postmenopausal women was calculated . \n the questionnaire was classified as burning sensation associated with visible lesions and burning sensation not associated with visible lesions . \n only those patients who did not have any visible lesions for burning sensations were diagnosed as bms patients . \n these patients were also evaluated for the levels of anxiety and depression by means of standardized questionnaires i.e. had scale and ghq-28 . \n the severity of burning and the response to the treatment was assessed with a five point visual analog scale ( vas ) . \n chi - square test was used to evaluate psychological parameters and the results were expressed in percentage , mean , standard deviation ( sd ) , quartiles and graphs . \n in the present study the total sample size was 100 out of which 56 patients had a chief complaint of burning sensation [ table 1 ] . \n all the patients ranged from age 43 to 85 years . out of this sample size , 43 patients ( 40.95% ) were found to have burning sensation as there primary chief compliant for which they sort expert opinion and another 13 patients ( 12.38% ) complained about burning as there secondary problem [ table 2 ] . out of these 43 patients with primary complaint of burning sensation 7 patients ( 16.27% ) had oral lichen planus and 3 patients ( 6.97% ) had oral ulcers ( rau ) . \n 11 patients ( 25.58% ) were taking some medications which are proved to cause burning sensation as there common side effects [ table 3 ] . \n remaining 22 patients ( 51.16% ) did not have any visible oral mucosal lesions but had burning sensation of 3 and above on a vas ( scale from 1 - 5 ) and thus were diagnosed as bms cases based on the principle of exclusion . \n prevalence of burning sensation patients with primary and secondary bms conditions associated with primary bms when tested for psychological variables in 43 patients with burning sensation as their primary compliant it was noticed that 9 patients ( 20.93% ) had no anxiety , 20 patients ( 46.51% ) had mild levels of anxiety and another 14 patients ( 32.55% ) had moderate anxiety [ table 4 ] . \n when tested for depression 8 patients ( 18.60% ) of the postmenopausal group showed no depression , 12 patients ( 27.90% ) showed mild depression , 13 patients ( 30.23% ) of the showed moderate depression and 10 patients ( 23.25% ) showed severe depression [ table 5 ] . \n the present study indicates that most of the patients had moderate to severe levels of depression , which suggests that anxiety and depression are constant features in postmenopausal female patients . \n zageralli stated that bms was a frequent feature seen in postmenopausal women which was consistent with our results . in the present study , it was found that burning sensation of the mucosa was present in 43% patients . according to gallagher et al . \n burning sensation in the oral cavity is due to atrophic mucous membrane of the oral cavity and the depletion of the iron stores in the body . \n lamey and lamb call bms as a multifactorial condition that generally affects women more commonly than men . in the present study \n two inventories namely ghq-28 and had were used to check the loading of anxiety and depression in the patients . \n ghq-28 was given by goldberg and hiller in 1978 and is a screening questionnaire , which measures the anxiety and depression scores . \n hospital anxiety and depression inventory ( had ) was given by snaith and taylor in 1985 . \n this inventory helps the clinician to know the levels of anxiety and depression in an individual . \n eli et al . analyzed 65 postmenopausal women and found visible oral mucosal lesions along with burning sensation as primary complaint , in our study out of 43 patients with primary complaint of burning sensation , 7 patients had oral lichen planus and 3 patients had oral ulcers ( rau ) . in the present study 32.5% \n patients showed moderate anxiety and 23.2% patients showed severe depression , the results are consistent with the study done by friedlander et al . \n where he stated anxiety and depression are features of post menopausal women ; however , forbabosco et al . mentioned that the increase of oral discomfort in postmenopausal women is due to hormonal modification and psychological factors play a secondary role . \n the study revealed most of the patients had moderate to severe level of depression , similar finding was observed by pisanty et al . \n the oral mucosal changes reported to be associated with menopause are many and varied with stress in association with oral symptoms . \n mention that there are specific estrogen receptors proteins in human gingiva which can effect cellular proliferation and keratinization in oral epithelium . \n bms is a fascinating , though poorly understood , condition in the field of oral medicine . \n possible causal factors include hormonal disturbances associated with the menopause , psychogenic factors ( including anxiety , depression , stress , life events , personality disorders , and phobia of cancer ) , and neuropathy . to conclude it can be said that psychological aspect plays a major role in the etiological niche amongst postmenopausal women .", "role": "user" }, { "content": "introduction : burning mouth syndrome is a chronic pain syndrome primarily affecting elderly women with hormonal changes or psychological disorders . it is multifactorial in origin , often idiopathic and its etiopathogenesis , majorly being neuropathic , largely remains enigmatic.aim:to determine the prevalence of burning mouth symptom , in elderly women and evaluate local and systemic causes responsible for burning sensation.materials and methods:100 elderly postmenopausal women were included in the study out of which 56 had a chief complaint of burning sensation . \n these patients were evaluated for the levels of anxiety and depression by means of questionnaire . \n the severity of burning and the response to the treatment was assessed with a five point visual analog scale . \n the results were analyzed using chi - square test.results:there was statistically significant increase in the levels of anxiety and depression in the study group.conclusion:the present study clearly indicates that most of the patients had moderate to severe levels of depression , which suggests that anxiety and depression are constant features in postmenopausal female patients .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: treatment of the edentulous mandible has always been one of the most challenging issues in dentistry . \n however , due to reduced stability when compared with natural teeth or fixed partial dentures , patients often complain of difficulty adapting to a removable prosthesis . \n there are different approaches , to improve denture stability such as , rebasing and surgical techniques ; yet , they do not create a high level of comfort or function for many . \n the advent of osseointegrated implants has vastly improved treatment outcomes in patients with complete edentulism . increased stability and retention of prostheses \n can be accomplished by one of two means , either an implant - retained removable overdenture or an implant - supported fixed prosthesis with distal cantilevers . \n the most outstanding merit of fixed implant - supported prosthesis over the other implant option is the psychological advantage of being fixed versus having a removable overdenture prosthesis . \n in addition , fixed prosthesis with posterior cantilevers gains bone in posterior regions 80% of the time ; whilst , implant overdenture with posterior soft tissue support loses bone in posterior regions almost 75% of the time . \n the biomechanical needs of a patient may necessitate the elimination of cantilevering and the placement of several implants in the posterior mandible ; but most of the time , it is impossible without advanced surgical procedures . on the other hand , \n fixed implant prosthesis has been one of the most controversial and expensive reconstructions for mandibular edentulism and it may be contraindicated in cases , where the replacement of lost soft and hard tissue and facial support by a prosthesis flange is critical . \n fewer implants , lower cost and esthetic and speech improvements are the advantages of implant - retained overdenture over fixed implant prosthesis . but like any other treatment option , it has its own drawbacks namely having to remove the prosthesis during the night , continued posterior bone loss , food impaction and long term maintenance [ 79 ] . \n an alternative treatment option can be a bilateral removable partial denture anchored to a fixed prosthesis , supported by implants placed in the interforaminal region , with stress directors near the connection with the fixed prosthesis . \n this prosthetic option , regardless of its clinical application , has not been fully validated in the dental literature and only one case report with 3 years of follow up has been published . \n the authors claim that this treatment method does not require grafting or multiple surgical procedures . \n furthermore , it has the esthetic and functional advantages of an overdenture , decreases compression of the edentulous ridge in function and the patient can benefit from the fixed anterior segment while removing the posterior part at night . the finite element method ( fem ) , which was first introduced in aerospace modeling , \n three - dimensional fem is an acceptable , powerful , replicable , and reliable tool in research protocols in dentistry and has proven its efficiency in solving numerous problems [ 1116 ] . \n the main goal of this study was to evaluate stress distribution differences when the number of implants under an anterior bridge supporting a removable partial denture in the posterior region varies . \n four three - dimensional finite element models ( 3d fem ) were designed of mandibles containing an implant - supported bridge extending between the canines , and a removable partial denture for premolars and first molars . \n nonrigid connection ( dalbo mini system , swiss ) was selected as the attachment method between the partial denture and canines of the anterior bridge . \n the lingual plate was designed as the major connector . the number of implants to support the bridge varied between the models from 2 to 5 with a length of 10 mm and a diameter of 3.8 mm . in the 4 and 5-implant models , \n the distances were 7.74 mm and 7.76 mm ( center - to - center ) , respectively for the implants moving from the canine position towards the midline . \n inter - implant distances for 2- and 3-implant models were 31 and 15.50 mm ( center - to - center ) , respectively from the canine position towards the midline . \n the distances were the same for both sides due to the symmetry of the 3d models ( figure 1a d ) . \n the next phase was to transfer the models for calculation to the ansys workbench ver . \n the corresponding elastic properties such as young s modulus and poisson s ratio were determined according to recent research [ 1115 ] ( table 1 ) . \n models were meshed , between 88900 and 114124 node- and between 49828 and 64740 10-node - quadratic tetrahedron body elements were used in the models with the highest and the lowest number of elements , respectively ( figure 2a d ) . \n all nodes at the distal and also the inferior surface of the models were restrained so that all rigid - body motions were prevented . \n von mises stresses on spongy bone , crestal bone and on implants for each model were evaluated . \n stress amounts showed a decrease from 2 implants to 4 implants but showed an increase in the 5-implant model . \n stresses on distal and mesial of terminal implants were in similar range when 2 , 3 and 4 implants were used . while , in the 5-implant model the amount of stress on terminal implants increased dramatically . \n the stresses on implants were similar in all models , with the greatest force on terminal ones . \n stress amounts showed a decrease from 2 implants to 4 implants but showed an increase in the 5-implant model . \n stresses on distal and mesial of terminal implants were in similar range when 2 , 3 and 4 implants were used . while , in the 5-implant model the amount of stress on terminal implants increased dramatically . \n the stresses on implants were similar in all models , with the greatest force on terminal ones . \n the stress levels that actually cause biological complications , such as resorption or bone remodeling , have yet to be comprehensively verified . \n there is no physiological limit for bone in regard to von mises stress in the literature . \n the physiological limit of bone strain , reported by frost ( 3000 ) , can not be compared with our results . \n application of removable partial dentures in combination with implants , which serve as abutments , is rarely seen in dental articles . \n jang et al ( 1998 ) used a single implant crown as an abutment for a removable cast partial denture . \n there is only one article in which , a distal - extension removable partial denture is designed on anterior fixed implant - supported prosthesis . \n this was done by pellacchia and emtiaz in 2000 . in this clinical case report with 3 years of follow up , the possibility of achieving positive results with a rpd connected to an implant - supported fixed prosthesis was demonstrated . \n they reported an increase in stability of rpd during function . regarding the optimum number of implants in distal - extension fixed implant - supported prosthesis , reports are limited to cantilever designs . \n number of implants , in ranges of 4 to 6 , has been reported with variable results in terms of the magnitude and distribution of forces and survival rates of implants [ 20 , 21 ] . \n misch reported 5 implants to be the best number . in his point of view , increasing the number up to 7 implants , may be more beneficial . while , ogawa showed no significant biomechanical difference between 4 and 5 implants in hybrid models . in overdenture models , \n klemetti in his literature review reported that patient satisfaction and function are not dependent on the number of implants or attachment type . \n thus , the number of implants necessary to provide adequate implant support for mandibular overdenture remains open to debate . in our study , \n stress transferred to spongy bone decreased from 2 to 4 implants ; but it dramatically increased in 5-implant models . in crestal bone of terminal implant models with 2 , 3 and 4 implants similar stresses were shown ; but in the 5-implant model the stresses on terminal implants increased significantly . \n the probable reason for this stress increase in models with 5 implants could be due to inter - implant distance . \n considering the advantages of an implant - supported anterior bridge supporting a removable partial denture posteriorly ( namely the function of an overdenture , decreased compression of the edentulous ridge in function and esthetics of having a fixed anterior segment when removing the posterior part during the night ) , it seems that this option can be practical with 2 , 3 or 4 implants ; because , our study showed that there was no significant difference regarding stress transferred to cortical bone and implants in models with 2 , 3 or 4 implants . \n thus , in financially compromised patients it can be achieved with fewer implants , without fear of placing excess stress on the implants . \n nevertheless , all patients must be assessed for other factors , including opposing dentition , parafunction , occlusion , etc . \n additionally , implant diameter and length may also affect the pattern of stress distribution [ 26 , 27 ] . \n a nonrigid connection was designed between anterior implant - supported fixed prosthesis and posterior removable partial denture . since in implant/ tooth connections stress values \n were decreased in bone when using non - rigid connections , application of rigid or non - rigid attachments can be based on the posterior bone support . \n within the limitations of this study , 2- , 3- and 4-implant models showed less stress on cortical and spongy bone in comparison with 5-implant model .", "role": "user" }, { "content": "objective : the main goal of this study was to evaluate differences in stress distribution relevant to the number of implants under an anterior bridge when combined with a removable partial denture in the posterior region.materials and methods : four three - dimensional finite element models ( 3d fem ) were designed from a mandible containing an implant - supported bridge extending between canines , and a bilateral distal extension removable partial denture . \n a nonrigid connection was selected as the attachment method between the partial denture and the anterior implant - supported fixed prosthesis ; 2 , 3 , 4 and 5 implants supporting the bridge all with 10 mm length and 3.8 mm diameter were assessed . with the aid of the finite element program ansys 8.0 , \n the models were loaded and von mises stresses were evaluated.results:in spongy bone , stress forces showed a decrease from 2 implants to 4 implants but showed an increase in the 5-implant model . stresses on cortical bone of terminal implants were in similar range in the 2- , 3- and 4-implant models . while , in the 5-implant model the amount of stresses on terminal implants increased dramatically . \n the stresses on implants were nearly similar in all models , with the greatest amount on terminal implants.conclusion:within the limitations of this study , 2- , 3- and 4-implant models showed less stress on cortical and spongy bone in comparison with the 5-implant model . \n the stresses transferred to implants were nearly similar .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: uveal melanoma ( um ) is an ocular cancer involving the uveal tract ( iris , ciliary body , and the choroid ) . \n the incidence is approximately 4.3 per million / year in the united states occurring mostly amongst white males . with treatment um \n prognosis depends on a number of factors including primary tumor size , time of diagnosis , and whether metastasis is present [ 5 , 6 ] . \n larger tumors have a mortality rate of approximately 50% shortly after diagnosis , and medium sized tumors also have a similar mortality rate when measured 15 years after primary tumor treatment [ 4 , 7 , 8 ] . \n metastatic disease usually involves the liver and almost always leads to death within 15 months . \n it is believed that that most choroidal , perhaps most uveal , melanomas origin from transformed benign naevus . \n clinically the primary diagnosis of um of the choroid often involves decreased visual acuity and scotoma secondary to retinal detachment , with slit lamp biomicroscopy showing melanotic or amelanotic tumor with or without orange dusting . \n the diagnosis is often supported by an ultrasound investigation showing acoustic hollowness [ 911 ] . \n primary tumor is treated with either brachytherapy using radioactive plaques in an effort to preserve the tissues of the eye or enucleation . \n there is no difference in survival rate , between these two treatments , for medium and large sized tumors . \n metastasis is searched for but seldom found at the primary diagnosis , which often develops in the following months to years . \n so far the best methods for detecting metastasis are nonspecific liver enzymes serum concentration ( lactate dehydrogenase ( ld ) , alkaline phosphatase ( ap ) , aspartate aminotransferase ( asat ) , alanine aminotransferase ( alat ) and -glutamyl transpeptidase ( -gt ) ) , ultrasound , computed tomography ( ct ) , and magnetic resonance imaging ( mri ) [ 12 , 13 ] . \n these methods have a sensitivity of 2767% and a specificity of 9096% with ct being significantly more sensitive than ultrasound and lactate dehydrogenase ( ld ) being the most sensitive serologic marker [ 1315 ] . \n treatments with liver resection , if there is only a small number of metastasis , have proven useful in increasing survival time 3.7-fold . thus far , to our knowledge , chemotherapy has not been proven as an effective treatment in um metastasis [ 12 , 16 ] . \n there is an urgent need for early diagnosis of the primary tumor , as well as its metastatic spread , and for the development of improved methods of treatment . \n indeed it is likely that micrometastasis may be present in the majority of patients at the time of the primary diagnosis , but due to their small size and reduced cell turnover they are undetectable by conventional methods . \n therefore , there is understandably a great deal of research aimed at identifying primary and metastatic um tumor biomarkers , to which discovery based proteomics has contributed significantly which we will discuss in this present review . \n the search for tumor biomarkers and therapeutic targets has largely been centered on genomics and transcriptomic studies , though in the last few years proteomics has emerged as a strong player in this field . \n proteomics was originally defined as the study of the total protein complement of the genome , but the definition has now broadened to include the study of single proteins and their variants and modifications . \n secretomics is a subgroup of proteomics , which studies the protein secretion from the cells that is measurable in the serum and represents an especially promising method to study the development of metastasis in patients with um ( figure 1 ) . \n normal cells protect their genome from damage by a complicated caretaker system , which involves enzymes that monitor and repair dna and act at certain checkpoints during mitosis . \n cells can become cancerous following changes in the normal cellular genome , which enables the cell to avoid the caretaker system by a number of ways such as changes in growth factor response , apoptosis regulation , and ability to spread throughout the tissues and body ( table 1 ) . a malignant tumor comprises of a mixture of cancerous and normal cells . this enables the cancerous cells to be oxygenated , nourished and near capillaries to ease migration . \n the growth of the normal cells is probably stimulated by growth signals from the cancerous cells . \n the change in gene expression is converted to a change in the tumor cells proteomic profile enabling the tumor to grow and spread . in the following segments \n , we systematically review the current proteomics studies , seen in the light of the cancer process described above many of the proteins depicted below falls into these categories . \n the changes in protein profile when a nevus changes into a choroidal melanoma were investigated by bande et al . who examined serum concentrations of an oncoprotein dj-1 in 53 nevi patients and 32 healthy controls with the enzyme - linked immunosorbent assay ( elisa ) method . \n 7 ( park 7 ) also known as dj-1 is an oncogene which is transcribed into an oxidative stress response protein ( dj-1 ) ; this protein defends the cell against oxidative species . \n bande et al . showed that dj-1 concentrations were not significantly elevated in nervi patients when compared to the controls ( 37,39 ng / ml versus 32,98 ng / ml ) . \n but when they examined concentrations of nervi patients with ocular symptoms at time of um diagnosis ( 69,79 ng / ml ) , ultrasound diagnosed acoustic hollowness ( 86,95 ng / ml ) , nevus thickness > 1,5 mm ( 73,22 ng / ml ) , and large basal diameter > 8 mm ( 73,03 ng / ml ) and compared them to nervi patients without symptoms and objective findings , they found a statistically significant elevated dj-1 concentration . \n three of the 53 patients had a nevus transformed into a um during the studied period . \n the baseline average serum level of these patients was 89,56 ng / ml compared to the nevus group ( 37,39 ng / ml ) and the control group ( 32,98 ng / ml ) . \n though the number of the studied groups was small with very few nevus transformations , dj-1 could be a useful biomarker in predicting nevus transformations and indeed has also been shown to be elevated in breast cancer . \n the protein profile of a single um cell culture was investigated by pardo et al . in 2005 . \n the investigators used two - dimensional gel electrophoresis ( 2-de ) and mass spectrometry ( ms ) to analyse the protein in human um cell culture . \n about 69 of these proteins were related to the carcinogenesis as shown in figure 2 . \n this is of course a momentary picture of a protein profile which is in constant change ; however , their findings appear to greatly correlate with previous studies . \n the proteins found include ezrin which is involved in connecting the cytoskeletal to the plasma membrane and has a role in cell division and proliferation . \n a number of heat shock proteins ( hsp ) , including hsp 90 , help with protein folding thereby functioning as a chaperone and defending the protein from damage such as that originating from oxidative stress . \n . also found proteases and peptidases involved in degeneration of extra- and intracellular proteins and matrix , which include the family of cathepsins , thereby helping the invasion and metastasis process ; this also includes the s100 proteins ( discussed later ) . \n furthermore , oncoproteins such as dj-1 , which defends the cell against oxidative species , and proteins that conferred resistance to cancer drug treatment were also found . \n what is quite striking is that 33% of the 69 cancer related proteins were involved in tumor invasion and metastasis . though um tumors are in constant change reflecting their genetic and subsequent protein expression , this momentary picture gives an indication of how much efforts are necessary for the cancer cell to change its surroundings and the intracellular environment . as mentioned previously the diagnosis of uveal melanoma \n is mostly based on clinical observations , and there is often a need to distinguish atypical melanoma such as amelanotic / hypomelanotic melanomas and other types of ocular tumors including ocular metastasis and retinal detachments with subretinal hemorrhages or exudation . \n therefore , one of the main aims of proteomics is to establish a biomarker that can be used along with the clinical and histological presentation of the tumor . \n one of the earliest studied biomarkers is the acidic protein s-100 , which is a dimer comprised of two almost identical subunits named and . this marker has been shown immunohistochemically to be present in 70% to 91% of primary tumor biopsies and um cell cultures with the strongest correlation with s-100 subunit [ 2629 ] . \n s-100 protein has been highly elevated in all ocular melanomas , inversely proportionally to the amount of melanin . \n it is not consistently accumulated in ocular fluids or in serum , where it to our knowledge has not proven useful as a diagnostic or prognostic marker . \n s-100 is not a unique marker for um and is also changed in many other tissues and tumors including cutaneous melanomas . \n this makes it unsuitable for fluid screenings and may also question its usefulness as a histopathologic marker . \n other protein markers investigated include hmb-45 which has been found in approximately 93 to 100% of all um , correlated to melanocytic tumors [ 27 , 30 ] and melanoma inhibitory activity ( mia ) found in 4 out of 5 primary tumors . \n p53 is a well known modulator of cellular proliferation , and it has also been shown to inhibit angiogenesis in the normal cell . \n the relationship between p53 expression in um was investigated by chowers et al . in 98 um tissue blocks . \n it was found that increased p53 expression was significantly associated to tumors with high proliferative activity and epithelioid cell type , though not related to other microcirculating patterns ( normal blood vessels , tumor silenced area , loops , cross - linking , and much more ) previously shown to be associated with death from um metastasis . \n it is now believed that p53 mutation is associated with a high proliferative activity . as an involvement of previous work pardo et al . \n studied the proteomics of two in vitro cell lines with different tumor progressive status before ( um < 7 ) and after 7 passages ( um > 7 ) using 2-de and ms . \n they found that um < 7 contained more actin related and actin - binding proteins , such as vimentin and adhesive proteins such as ezrin , when compared to um > 7 . \n in contrast , um > 7 had the presence of proteins related to disassembly of actin filaments , in addition to an increase in glycolytic enzymes , such as alpha enolase . \n some proteins decreased or disappeared , while new proteins , namely , high mobility group protein 1 ( hgm-1 ) , were identified in the um > 7 cell line . \n they also found proteins expressed in um > 7 that promoted cell survival such as hs1-binding protein and new adhesive proteins such as melanoma associated antigen 18 ( muc18 ) . \n they identified hmg-1 , which is involved in transcription of genes in the metastatic cascade , in different human cancers , and muc18 , a cell adhesive molecule ( cam ) involved in endothelium adhesion , that may be involved in allowing hematogenous spread . both hmg-1 and muc18 \n are suggested as potential invasive markers as they are expressed only in um > 7 and not in um < 7 . \n finally , dj-1 oncogene was identified as a potential biomarker for um as it is present in all um cell lines ( including um < 7 and um > 7 ) though not in non - um cell lines . \n dj-1 was confirmed in the um cell culture medium by western blot ( wb ) , thereby identifying it as a secretory molecule . \n pardo et al . also used wb and ms to identify potential biomarkers in um . by studying um cell line secretomes and autoantibodies against secreted um protein in 11 patients , they identified three potential biomarkers . \n cathepsin is a lysosomal acid proteinase that was overexpressed in the um cell line compared to non - um cell line , even more in um < 7 passage compared to um > 7 passages . \n the same was found with melanoma specific antigen gp100 and adapter protein syntenin 1 , though syntenin 1 was not detected in patient sera . \n the proteomic profiles of metastasis compared to primary tumor in um were investigated by zuidervaart et al . . \n they compared the proteomic profiles of 2 liver metastasis to those of the primary um cells with the use of 2-de and ms . \n out of 1184 spots on the gels they identified 24 proteins that differed in metastasis when compared to primary tumor . \n they found no difference between the two metastatic cell lines but an increase in cellular defense , apoptosis , proliferation , and migration . \n as depicted in table 1 , the proteins shown to increase include heat shock protein 27 ( hsp-27 ) , involved in protein conformation stabilization , and cathepsin z , involved in lysosomal proteolysis , while the production of proteases is generally decreased compared to primary tumor . \n this study gives some important insight into the differences between primary tumor and the metastasis ( table 2 ) . \n the proteomic differences between metastasizing um and nonmetastasizing um have also been investigated by linge et al . . instead of comparing the primary tumor to the metastatic \n , they looked at 25 primary um patients with a minimum followup of 7 years and subsequently identified 9 patients who developed metastatic disease . \n they used 2-de and ms to identify 14 proteins which significantly differed in amount between the two groups ( table 3 ) . amongst those identified they made an in vitro function study by silencing the fatty acid - binding protein , heart - type ( fabp3 ) , and triosephosphate isomerase ( tpi1 ) gene . \n their results showed that significantly less cells invaded through the underlying membrane but also a significant decrease in cell motility was found . \n as mentioned earlier the leading cause of death in um patients is metastasis that is primarily to the liver . \n although micrometastasis at the time of the primary diagnosis is thought to be present , most metastasis is usually only found later at more advanced stages of um disease . in recent years \n proteomic investigations have been used to try and identify biomarkers that could help to determine which um patients have micrometastasis at the time of primary diagnosis and have increased risk of developing metastasis , and therefore those patients that are in need of adjuvant treatments including liver resection . \n firstly , those studies upon the primary tumor to try to determine if there is metastasis at the time of enucleation ( all these studies performed on enucleated tumors ) ; and secondly , those studies on the proteomic expression of the serum , vitreous or aqueous humour associated with the primary tumor and/or its metastasis . later we discuss the potential biomarkers that have been investigated using these two methods . as shown in table 4 a number of potential biomarkers associated with um metastasis have been identified , most of them investigated by immunohistochemistry ( ich ) . \n some of the most promising candidates include melanoma cell adhesion molecule ( mcam ) , stem cell factor ( scf ) , hsp-27 , and vimentin [ 3941 ] since these proteins have a crucial role in normal , as well as pathological cell functions and can be found within the primary tumor . \n use prior knowledge from the genomics field and search for monosomy 3 which is significantly associated with hepatic metastasis in um [ 42 , 43 ] . during the last couple of years \n there has been an extensive search for serum biomarkers , since early diagnosis is the key to successful treatment of metastatic um . \n as mentioned earlier , current blood biomarkers and imaging techniques have a sensitivity of 2767% , with computed tomography ( ct ) and lactate dehydrogenase ( ld ) being the most sensitive [ 13 , 15 , 44 ] . \n often the metastatic burden is too great at the time of the diagnosis , and early diagnosis allows more treatment options that give better chance of curing the patient [ 16 , 18 ] . \n studies show that melanoma inhibitory activity ( mia ) , osteopontin ( opn ) and tissue polypeptide - specific antigen cytokeratin 18 ( tsp ) are significantly elevated in serum of metastatic um patients when compared with nonmetastatic patients ( table 5 ) . \n they more than double in measurable concentration in serum , and a threshold has even been suggested for mia of 8,3 ng / ml giving it a sensitivity of 82% and a negative predictive value of 97% as shown by klingenstein et al . \n the size of the tested groups ( 503 um patients in klingenstein et al . ) also makes it ready for clinical trials , and it is likely that the same goes for opn and maybe even for tsp . \n vascular endothelial growth factor ( vegf ) has not been shown to have a large increase with metastasis as one might expect , due to the wide interpersonal variability in serum vegf levels . \n barak et al . suggest that this may be due to the fact that vegf has two very different splicing products that occur naturally . \n these two vegf products have very different abilities regarding pro- and antiangiogenic signals , and a change in ratios vegf types can not be distinguished by current tests such as elisa . \n prognostic factors include clinical and histomorphological appearance ( tumour size , the relation to surrounding tissue , cellular type , etc . ) , in addition to monosomy of chromosome 3 that is associated with a decreased survival due to hepatic metastasis [ 42 , 43 ] . in recent years \n have investigated the prognostic value of hsp-27 , a well known chaperone involved in cell proteolysis inhibition . \n perform ihc upon enucleated eyes of 99 um patients , 44 of which were predicted dead within a follow - up period of 8 years by conventional methods . \n they only identified 5 deaths due to metastatic disease and 5 patients with developed metastasis all of which were monosomy 3 positive of which 6 were found to have a low to negative hsp-27 score ( score 6 ) . \n they found that hsp-27 6 score significantly predicted a decreased < 8 years survival rate and when combined with clinicopathological factors predicted monosomy 3 with a sensitivity of 78% and a specificity of 72% [ 40 , 42 , 43 , 47 ] . \n healthy cells become cancerous through genetic mutations , which are implemented through splicing in effector proteins found intra- as well as extracellularly including vitreous body and serum . through proteomic analysis \n these proteins may be identified in primary tumor , ocular fluids , and in the patient 's serum , thereby assisting in diagnosis , primary tumor classification , and metastatic identification , together with conventional methods . \n the proteomic profiles of two um cell lines and tissue samples are never identical , but as proteomic studies have shown us there are many similarities , which may make it possible to one day identify clinically useful protein biomarkers and therapeutic targets . many of the current biomarkers have only been found in small pilot studies and are in need of validation in larger prospective patient cohort studies . \n this has proven to be difficult given that the incidence of um is quite low compared to that of other tumors and that the followup of metastatic patients has proven difficult because of large mortality rates as described by reiniger et al . . \n however , this task is essential if potential proteomic markers will have a chance to be implemented in everyday clinical assessments . \n there is an especially important need for further diagnostic aids in nevi - transformation , atypical melanoma , micro- and macrometastasis . \n serum biomarkers have an obvious advantage over histopathological biomarkers in that it is not necessary to obtain tissue samples from the primary tumor or metastasis . \n when it comes to diagnosing metastasis at the time of primary um treatment , serum biomarkers can be used as a prospective tool to monitor the um patients , while histopathologic biomarkers of primary um biopsies can be used for prognostic staining and to identify those um patients of high risk or high probability of micro- and macrometastasis and therefore in need of additional treatments . \n studies performed with the potential um biomarkers of mia and opn show especially promising results in terms of increased sensitivity of detecting um metastasis compared with conventional methods [ 45 , 49 ] . \n however , there remains a need for an in - depth understanding of the um metastasizing process and the identification of further potential biomarkers with proteomics centered uponcharacterization of primary um at differing stages including metastatic um prospective studies of biomarkers in um patients using quick and hopefully cost effective tests implementation into effective clinical practices . \n characterization of primary um at differing stages including metastatic um prospective studies of biomarkers in um patients using quick and hopefully cost effective tests implementation into effective clinical practices . \n optimal um management may be to combine proteomics , genomics , and transcriptomics in order to identify high risk um patients and establish their surveillance together with early and individualized treatment . \n furthermore , as proteomic analysis becomes more cost effective and rapid in the future , it may even be possible to ascertain an individual set of biomarkers for each um patient . \n however , this will require better knowledge of um development to which proteomics studies will also surely continue to contribute .", "role": "user" }, { "content": "uveal melanoma ( um ) continues to be associated with a high mortality rate of up to 50% due to metastatic spread primarily to the liver . \n currently there are relatively effective treatments for the primary tumor , though the management of the metastatic disease remains inadequate . \n conventional diagnostic tools have a low sensitivity for detecting metastasis , and early detection of metastatic spread would allow more treatment options that could ultimately increase survival of um patients . \n advanced proteomic methods have already helped to find potential biomarkers associated with um pathogenesis and metastasis . in the present review \n we discuss the field of proteomics in relation to studies elucidating biomarkers of um , where proteins such as s-100 , osteopontin ( opn ) , and melanoma inhibitory activity ( mia ) have been shown to be associated with metastasis .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: stroke is the second highest cause of death in the world and the main cause of disability in people in productive ages . in indonesia , \n the average age of stroke patients is 58.8 years old and stroke is the leading cause of high mortality rate in people above 50 years old . \n statistics of heart disease and stroke2013 update reveals that number of stroke patients aged between 2045 years has increased significantly in recent years . according to world health organization , stroke is rapidly developing clinical signs of focal ( or global ) disturbance of cerebral function . \n the symptoms last in 24 hours or longer and may lead to death , with no apparent cause other than of vascular origin . \n more than 87% of stroke cases are caused by ischemia due to thrombosis or cardio embolisms . \n ischemic stroke is atherosclerosis - associated complication [ 35 ] which is progressive in process . \n it requires 3 to 4 decades or more since the period of endothelial dysfunction and carotid intimamedia thickness ( cimt ) to develop until clinical manifestations appear . \n a study on american children aged 1014 years old who died from motorcycle accidents showed that 50% of them have early atherosclerosis evidence . \n similarly , bogalusa heart study revealed that 50% of its 204 subjects aged 215 years old had fatty streak in their coronary arteries . \n thus , detecting atherosclerotic vascular changes in childhood helps identifying high - risk groups which is very useful for the early stroke management . \n cimt is a surrogate marker that reveals atherosclerosis in general and constitutes a predictor of blood vessels condition in the future . \n the increase of cimt has been proven to be related with the raise of ischemia stroke risk [ 1113 ] . in indonesia \n , cimt measurement has not yet become a routine screening procedure for atherosclerosis prevention . to date , there is no normal value data of cimt for population in indonesia . \n , this sign becomes very useful to be intermediate phenotype in genetic studies [ 14 , 15 ] . \n epidemiological data show that the increase of cimt is influenced by genetic factors , with the prediction that 30% to 40% is heritable . in 2006 , \n genetique de l'infarctus cerebral [ genic ] study conducted a research on genetic factors related with cimt of cerebral infraction patients . \n the study examined polymorphism association with vascular pathologies which incorporate inflammation , hypertension , coagulation , and lipid metabolism . \n the findings revealed that there was a significant relationship between cimt with monocyte chemoattractant protein-1 [ mcp-1 ] and osteopontin ( opn ) polymorphisms . \n mcp-1 is synthesized by various cells associated with atherosclerosis , including endothelial cells , muscle cells , fibroblasts , and macrophages . in early stages of atherogenesis , mcp-1 is primary chemokine which recruits monocytes into arterial subendothelium . \n mcp-1 protein and rna are highly evidenced in atherosclerotic vessels , but not in normal vessels . \n the mcp-1 polymorphism in the forms of snp-957 , snp-2518 , and snp-2578 causes earlier occurrence of cimt faster progression [ 12 , 1619 ] in chinese , japanese , french , and slovakian people . \n however , the mechanism of earlier and faster cimt development could not be identified yet . \n some researchers suggest that ethnic and geographic factors give different effects on genetic polymorphisms taking in mcp-1 polymorphism . thus far \n , there has never been a study on mcp-1 polymorphism and its contribution toward cimt conducted for indonesian population . \n it functions in atherosclerosis , in cell - mediated immunity , and in macrophage recruitment and activation . in early stages of atherosclerosis , \n opn attracts inflammatory cells , promotes the releases of proteinolytic enzymes , and stimulates smooth muscle cell proliferation . \n the opn promoter polymorphism is studied in several areas , such as in snp-443 and snp-66 [ 12 , 21 ] \n . this research attempts at explaining the effect of mcp-1 and opn promoter polymorphism on cimt changes in javanese indonesian children . \n subjects were 54 javanese indonesian children who were classified into two groups : case and controlled . \n the case group consisted of 27 children aged 1021 years whose parents had ischemic stroke historical background . \n the controlled group consisted of 27 javanese children aged 1021 years whose parents had no ischemic stroke history . \n all participants were required to complete a standardized questionnaire designed to obtain information about family history . \n plasma concentrations of total cholesterol , high - density lipoprotein [ hdl ] , low - density lipoprotein [ ldl ] cholesterol concentrations , triglycerides , and fasting blood glucose were measured in accordance with standardized protocols in the physiology laboratory of medical faculty , university of brawijaya malang . \n the research had been approved to be conducted by ethical committee , medical faculty , university of brawijaya , malang , indonesia . to measure cimt , all subjects went through carotid ultrasound process in the radiology department of saiful anwar general hospital in malang , indonesia . \n high resolution carotid ultrasound measurements were performed using logiq s6 ultrasound ( ge healthcare ) , with 10 mhz linier transducer . \n the measurement was done by scanning the far wall of common carotid arteries in 1.0 cm distal . \n the crest at the origin of the bifurcation was used as an anatomical mark to identify segment to be visualized . in each examination , \n the sonographer used three different scanning angles , namely , transversal , anterolateral , and posterolateral angles , to record the greatest cimt ( figure 1 ) [ 2224 ] . \n genomic dna was isolated from 1 ml blood using extraction kit ( wizard genomic dna purification kit / promega ) . \n mcp and opn promoters were amplified by using specific primer mcp forward 5-ccgagatgttcccagcacag-3 and mcp reverse : 5-ctgctttgcttgtgcctctt-3 . \n amplified dna was sequenced by employing automated sequencer ( macrogen , seoul , korea ) . \n data characteristics , laboratory test results and ultrasound results were tested with data normality test ( kolmogorov - smirnov ) and data homogeneity test ( levence statistics ) prior to comparison test by using independent t - test . \n the calculation of mutation influence on carotid artery toward cimt - increasing risks was done using the odd ratio [ or ] in a 2 2 table . \n each sample received the same treatment in which right and left common carotid arteries were checked , and cimt and diameter of the carotid artery were measured and calculated . \n result of correlation analysis on the average cimt with clinical characteristic variables is presented as in table 2 . \n result of correlation test carried on using pearson product moment test showed that there was no correlation between average cimt with age , height , weight , body mass index , fasting glucose , total cholesterol , hdl , and ldl , as well as triglycerides . \n mutations took place in base number-2138 in which adenine base had substituted to thymine base . \n the mutations occurred in two controlled samples ( 14.3% ) and one case sample ( 5% ) ( figure 2 ) . \n further mutations took place in base number-2464 in which guanine base mutated into adenine base . \n the calculation of mutation influence on cimt - increasing risks measured through odd ratio [ or ] using a 2 2 table appears as in table 3 . based on previous research \n [ 11 , 24 , 26 ] , 0.5 mm was set as the normal value of cimt for odd ratio analysis . \n this means that children with mutation have 1.471 times higher tendency of having cimt thickening than children without mutation . \n all samples were analyzed for identifying t-66 g opn polymorphism promoter , but there was no mutation found in all samples . \n from the research , two types of point mutation in mcp-1 gene promoters were found , that is , a-2138 t and g-2464a . either base-2138 or base-2464 was located in promoter site . \n the promoter determines the efficiency of rna polymerase binding and , thus , it also determines the transcription efficiency . \n the transcription factor is attached to the binding site in the promoter region and stimulates rna polymerase to bind with the promoter site . \n in japan , mcp-1a-2138 t polymorphism has been reported to be identified in myocardial infarction patients above 65 years old . \n it was found out that the mcp-1 a-2138 t polymorphism was significantly associated with mcp-1 serum level and incidence of myocardial infarction . in this research , mcp-1a-2138 t polymorphism \n was found in children from parents with ischemic stroke history and children whose parents do not have ischemic stroke history as well . \n the polymorphism has never been reported to be found in people with no ischemic stroke history previously . \n children with a-2138 t polymorphism , either from the case or the controlled group , had a thicker cimt than the normal value . \n the two children with a-2138 t polymorphisms had a family history of myocardial infarction disease with body mass index over 25 . \n ( 2006 ) informed that in japan a-2138 t polymorphism was found in patients with body mass index of more than 25 kg / m with myocardial infarction . \n , the polymorphism was found in the case sample of a 19-year - old girl who inherited it from the mother suffering from ischemic stroke at the age of 32 . \n her grandfather also suffered from ischemic stroke with hypertension , while her grandmother suffered from hypertension . \n her total cholesterol was 208 mg / dl and ldl cholesterol was 136 mg / dl , which indicated dyslipidemia . \n her average cimt was 0.60 mm , or thicker than normal average cimt [ 11 , 26 ] . \n results of this research reveal that children with polymorphism had thicker cimt than children without mutation . \n risk measured with odd ratio shows that children with mcp-1 polymorphism promoter had 1.471 times possibility to have thicker cimt than children without polymorphism . \n these results confirm the importance of genetic examination , especially the mcp-1 polymorphism promoter analysis in children with atherosclerosis risk factors . \n result of this interaction is mcp-1/ccr2 messenger system increasing activity which leads to the local recruitment of monocytes at the site of injury in the arterial wall . \n this , in turn , will lead to increasing atherosclerosis incidence [ 29 , 30 ] . \n the relation of mcp-1 promoter polymorphism with increasing cimt has been reported in several studies , in which the mcp-1 g-927c and a-2578 g polymorphisms are significantly associated with the increasing cimt in patients with ischemic stroke . \n the mcp-1 g-362c polymorphism in black population is associated with the increasing cimt and atherosclerosis risk . \n the mcp-1 g-928c polymorphism is associated with the increased cimt and the atherosclerosis risk [ 29 , 30 ] . \n in contrast to some other studies conducted on atherosclerosis - related conditions , this research did not result in any finding on mcp-1 a-2518 g polymorphism [ 16 , 31 ] . \n the difference among the above genetic polymorphism findings is likely to be due to the ethnic and geographical differences . \n high mcp-1 transcription rate contributes to the severity of stroke [ 32 , 33 ] . \n the mcp-1 transcription rate is influenced by several risk factors such as hypertension , hypercholesterolemia , smoking habit , and diabetes . \n in summary , two mcp-1 promoter polymorphisms were found in the research , namely , a-2138 t and g-2464a . \n children with polymorphism demonstrate increasing cimt and have 1.471 times higher possibility to have thicker cimt . \n cimt of javanese indonesian children from parents with ischemic stroke is thicker than cimt of children from healthy parents .", "role": "user" }, { "content": "carotid intima media thickness ( cimt ) is clearly associated with atherosclerosis . \n studies in ischemic stroke patients reveal that there is a significant association between cimt with monocyte chemoattractant protein-1 ( mcp-1 ) and osteopontin ( opn ) promoter polymorphism . \n this research aims to explain the effect of mcp-1 and opn promoter polymorphism toward cimt changes identified in javanese indonesian children . \n subjects were 54 children : 27 were from parents with ischemic stroke ( cases ) , and 27 were from healthy parents ( controlled ) . \n the cimt was examined by utilizing high resolution b - mode ultrasound . physical examination and genotyping analysis of mcp-1 promoter \n were conducted by employing pcr method . \n research results indicate that two polymorphisms were obtained , that is , a-2138 t and g-2464a , respectively . \n a-2138 \n t polymorphism was found in 5% of case children and in 14.3% of controlled children . \n g-2464a polymorphism was found in 5% of case children . \n cimt of case children was significantly different from that of controlled children ( 0.61 0.012 mm versus , 0.52 0.015 mm , p = 0.021 ) . \n subjects with mcp-1 promoter polymorphism have 1.471 times higher tendency to have thicker cimt than subjects with no polymorphism in mcp1 promoter . \n opn promoter t-66 g was also studied but it did not indicate occurrence of polymorphism in samples .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: in the era of minimally invasive technologies , laparoscopic transperitoneal left adrenalectomy ( ltla ) has become the standard treatment for adrenal masses < 6 cm . \n however , masses > 6 cm can be successfully treated using the laparoscopic approach as well , replicating principles of open surgical oncology . \n splenorenal and splenoparietal ligaments are dissected during ltla with the lateral approach , and if necessary , the left colic flexure is mobilized . \n the middle adrenal vein is identified behind the pancreas so that , after its ligature and dissection , the adrenal gland can be removed . \n the dissection of splenic ligaments replicates their congenital absence or weakening , present in cases of wandering spleen ( ws ) . \n other possible approaches are laparoscopic transperitoneal anterior adrenalectomy ( during which the patient is supine ) and posterior retroperitoneal laparoscopic adrenalectomy ; neither involves the dissection of splenic ligaments . \n ws , also known as splenoptosis or ectopic , free - floating , or aberrant spleen , is the migration of the spleen from the left upper quadrant to a more caudal location in the abdomen . a unique case of ws after ltla was described by corcione et al , who reported the case of a 57-year - old woman who underwent ltla because of a 4-cm nonfunctional mass . \n three months later , she was admitted to the surgical emergency department with acute abdominal pain , nausea , and vomiting . \n she underwent emergency laparotomy , which revealed an organoaxial gastric volvulus caused by the hypermobile spleen . \n another case report , by le et al , showed ws as a rare complication of splenic torsion after laparoscopic nissen fundoplication . \n ws , with an incidence of < 0.5% among splenectomies , is rare and is often due to the congenital absence or weakening of splenic ligaments . \n the spleen develops from a mesenchymal condensation in the dorsal mesogastrium of the lesser sac in the fourth week of gestation . during fetal development , rotation of the intestinal tract and growth of the dorsal mesentery carry the spleen to the left side of the abdominal cavity , where it is anchored to the left kidney and stomach by gastrosplenic and splenorenal ligaments , respectively . \n ws , as well as intestinal malrotation , congenital short bowel , and other abnormalities , has been associated with mutations of filamin a in male patients affected by x - linked periventricular nodular heterotopia . \n failure of fusion of the mesogastrium to the posterior body wall leads to incomplete formation of splenocolic , splenorenal , and splenophrenic ligaments , and this is considered the mechanism by which ws , defined as the spleen 's lying outside the left upper quadrant suspended only by hilar vessels , occurs . \n consequently , ws has a long mobile mesenteric pedicle that is prone to torsion and ischemia . \n this mechanism can also be responsible for gastric rotation ( volvulus ) , rarely associated with ws . \n when present in adults , ws may be the delayed onset of congenital conditions or an acquired disease from abdominal wall laxity due to the hormonal effects of pregnancy . \n some authors have associated it with splenomegaly , although there is no higher incidence of ws in areas of endemic splenomegaly . in adults \n , it usually occurs as a mobile abdominal mass , whereas in children , it presents as acute or chronic abdominal pain and can be associated with a history of congenital diaphragmatic hernia or prune belly syndrome . \n the clinical presentation is vague , with symptoms including nausea , vomiting , early satiety , and acute , cramplike abdominal pain due to splenic ischemia and infarction . \n bowel obstruction may occur because of the compression of abdominal organs ; enuresis and hydronephrosis may be other consequences , as well as gastric fundal varices , mesenteric varices , and acute pancreatitis . \n the progressive splenic enlargement can lead to rare splenic ruptures and to possible spontaneous massive hemoperitoneum . \n ultrasonography is considered efficacious in diagnosing ws , as it is a noninvasive , inexpensive test that can confirm both the ectopic location of the spleen and blood flow pattern . \n both grayscale and color doppler sonography can easily show the migratory nature and perfusion status of a ws in real time , in the right decubitus position . \n sometimes it is necessary to examine patients with computed tomography ( ct ) , which provides information on the location , vascular integrity , and size of the spleen as well as the surrounding organs . with ws , \n ct may show elongated or tortuous splenic vessels and may also reveal multiple collaterals of the hilum of the spleen . \n with torsion of the splenic pedicle , a whirled appearance of the splenic vessels may be noticed on ct . despite the advantages offered by ultrasonography and ct , \n the aim of this prospective study was to determine whether the mobilization of the spleen during ltla , replicating the congenital condition of absence or weakening of the splenic ligaments , could be recognized as a cause of postoperative ws . \n between january 2006 and may 2011 , 59 patients underwent laparoscopic lateral adrenalectomy ; 30 right adrenalectomies and 29 left adrenalectomies were performed . \n we took into consideration the 29 patients who underwent ltla : 9 men ( 31% ) and 20 women ( 69% ) . \n twenty - four patients ( 83% ) were included in our ultrasonographic follow - up . \n five patients were not included , 3 with terminal cancer and the remaining 2 who had undergone multiple upper gastrointestinal surgical procedures . \n therefore , we analyzed 24 patients , 8 men ( 33% ) and 16 women ( 67% ) , with a mean age of 59 13 years ( range , 3784 years ) . among these 24 patients , \n 13 were affected by nonfunctional adenoma ( 54% ) , 4 by pheochromocytoma ( 17% ) , 3 by metastasis ( 12% ) , 3 by adrenocorticotropic hormone dependent hyperplasia with cushing 's disease ( 12% ) , and 1 by conn 's adenoma ( 4% ) ( table 1 ) . \n demographic and clinical data of 24 patients undergoing ltla acth , adrenocorticotropic hormone ; ltla , laparoscopic transperitoneal left adrenalectomy . \n each patient underwent preoperative abdominal ultrasonography , ct , or nuclear magnetic resonance , as well as hormonal laboratory exams . \n all ltlas were performed using the same laparoscopic technique : right lateral decubitus pneumoperitoneum induced by a veress needle ; 3 trocars , placed on the midclavicular line ( t3 , 5 mm for the forceps and divaricators ) , midaxillary line ( t2 , 1012 mm for the radiofrequency dissector , scissors , irrigation - aspiration cannula , clips , etc ) , and anterior axillary line in the middle of the line joining the umbilicus to the subcostal margin ( t1 , 1012 mm for the optic ) ( figure 1 ) ; dissection of the splenoparietal and splenorenal ligaments ; en bloc mobilization and medialization of the spleen and pancreas ( figure 2 ) ; sectioning using clips or a radiofrequency dissector of the middle adrenal vein ; and removal of the adrenal gland into an endobag ( covidien , dublin , ireland ) . \n mobilization of the left colic flexure was performed only when absolutely necessary , and exposure of the adrenal lodge was limited to the field of view . \n furthermore , care was taken in repositioning the spleen and pancreas to their correct seats without using fibrin glue or other devices . \n the 24 selected patients were studied postoperatively using abdominal ultrasonography after 2 and 4 months ( 4 patients [ 17% ] ) and after 1 year ( 20 patients [ 83% ] ) , with a 3.5-mhz convex probe , always by the same radiologist , experienced in ultrasonography , who evaluated the following parameters : morphologic and dynamic aspects of the spleen in the supine position , in the right lateral position , and in orthostatism ; potential mobilization of the spleen related to the diaphragmatic dome ; and flow direction and resistive index at the origin of the splenic artery and at the splenic hilum . \n the average operation length was 89 35 minutes ( range , 45180 minutes ) ; the average diameter of the lesions was 4 0 cm ( range , 38 cm ) ; and the average postoperative stay was 2 1 days ( range , 15 days ) . \n postoperative courses were free of complications in 22 patients . in the remaining 2 patients ( overall morbidity , 8% ) , 1 patient \n had left pneumothorax treated with thoracic drainage , and 1 patient had right pleural effusion with hematoma in the left renal region , conservatively treated . \n clinical and ultrasonographic follow - up showed an absence of symptoms attributable to ws in each patient , confirming normal position of the spleen with normal blood flow in the supine position , in the right lateral position , and in orthostatism ; an absence of spleen mobilization related to the diaphragmatic dome ; and an absence of free or loculated collections . \n the average longitudinal spleen diameter was 9.9 1.3 cm ( range , 7.812.2 cm ) , and the average anteroposterior spleen diameter was 4.8 0.6 cm ( range , 3.85.8 cm ) . \n the average resistive index at the origin of the splenic artery was 0.6 0.1 ( range , 0.40.7 ) , and the average resistive index at the splenic hilum was 0.6 0.1 ( range , 0.50.7 ) . \n hepatic steatosis was seen in 3 patients ( 12% ) , and ptosis of the right kidney was seen in 1 patient ( 4% ) . \n to evaluate whether ws can be considered a frequent postoperative complication of ltla , we considered 29 patients who underwent this surgical procedure , but 5 patients were not included in the study . for the remaining 24 patients , postoperative ultrasonographic follow - up was performed to evaluate the morphologic and dynamic aspects of the spleen , its potential mobilization , and the flow direction and resistive index at the origin of the splenic artery and at the splenic hilum . ws was completely absent , despite the dissection of ligaments and the female majority . \n although there are surgical techniques that do not involve dissection of the splenic ligaments , ltla seems to be free of complications related to the supposed hypermobile spleen . given the rarity of ws , we are aware that the number of patients in our study is too small to reach any definitive conclusions , and our results should be regarded as purely exploratory . \n ws should not be considered a common complication of ltla , especially in patients with small adrenal masses , which generally do not require extensive dissection ; in our patients , the spleen was repositioned in its correct seat without the necessity of glue or splenopexy , mobilization of the left colic flexure was performed only when absolutely necessary , and exposure of the adrenal lodge was limited to the field of view . \n in addition , we have no data regarding postoperative adhesion formation , but we suppose that the healing process of dissected structures may be sufficient to fix the spleen in its natural place . \n ws may occur if the dissection is extensive ; for this reason , in these cases we recommend the use of glue or other methods to reposition the spleen in its correct seat . \n clinical and ultrasonographic follow - up of 24 patients who underwent ltla showed no symptoms or signs of migration of the spleen . \n ltla remains widely recognized as a safe approach , at least as effective as other techniques , such as laparoscopic transperitoneal anterior adrenalectomy and posterior retroperitoneal laparoscopic adrenalectomy . in any case \n , dissection should be limited and should not exceed too much in the mobilization of the left colic flexure . \n furthermore , it is necessary to pay attention when repositioning the spleen and pancreas in their correct seats . \n ws should be therefore considered a very rare complication of ltla that , however , can not be ignored . \n it would be useful for other authors to signal this complication , should it occur , so that different approaches and consequent results may be compared .", "role": "user" }, { "content": "background and objectives : laparoscopic transperitoneal left adrenalectomy ( ltla ) has become the standard treatment for adrenal masses < 6 cm . \n ltla involves the dissection of splenic suspensory ligaments , which replicates their congenital absence or weakening , present in cases of wandering spleen ( ws ) . \n ws is a rare condition in which the spleen migrates from the left upper quadrant to a more caudal location in the abdomen . a unique case of ws after ltla was described by corcione et al . in this prospective study , we investigated the possibility of ws as a consequence of ltla.methods:twenty-four patients , 8 men and 16 women , who underwent ltla with the dissection of splenoparietal and splenorenal ligaments were selected.results:clinical and ultrasonographic follow - up showed no evidence of postoperative ws.conclusions:in the literature , ws is not commonly reported as a postoperative complication of ltla . in effect , especially in the case of small adrenal masses , \n the spleen 's repositioning in its seat is autonomous . \n however , the alarming possibility of ws should not be ignored , especially in the case of extensive dissection of the left colic flexure . it would be useful for other authors to signal this complication , so that different approaches and consequent results may be compared .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: before the arrival of new direct - acting antiviral agents ( daas ) , triple therapy combining pegylated interferon ( peg - ifn ) , ribavirin ( rbv ) , and protease inhibitors ( boceprevir or telaprevir ) was the standard of care for the treatment of hepatitis c virus ( hcv ) infection ( 1 - 4 ) . \n anemia is commonly observed during rbv - based therapy and often results in dose reduction and decreased antiviral efficacy . \n because of a large inter - individual variability in exposure ( 5 - 7 ) , the weight - based rbv daily dose is poorly correlated with the rbv plasma concentration ( 8) . \n previous studies have indicated that anemia is about twice more frequent in patients receiving telaprevir with peg - ifn / rbv than in those receiving peg - ifn / rbv alone ( 9 ) . \n the objective of this study was to determine whether telaprevir - based therapy was associated with an increased rbv plasma exposure , which could in part explain the increased incidence of anemia . in parallel \n hcv patients with a previous non - response to peg - ifn / rbv therapy and re - treated with peg - ifn / rbv and telaprevir were prospectively recruited from the department of hepatology , croix - rousse hospital , lyon , france . \n non - response was defined as a failure to achieve a sustained virological response and could thus include null response , virological breakthrough , and virological relapse . \n patients were selected if they had consecutive assessment of plasma rbv trough concentrations ( prescribed at the clinician s discretion ) . \n patients with chronic kidney disease ( ckd ) stage 4 ( estimated glomerular filtration rate ( egfr ) < 30 ml / minutes/1.73 m ) were excluded from the study . \n plasma rbv trough concentrations were measured using a validated high - performance liquid chromatography - diode array detector method ( 10 ) . \n mg / l ) , and precise ( total imprecision , calculated by measuring the coefficient of variation of the internal quality control values , was < 10% for concentrations from 0.20 - 5.00 mg \n plasma rbv trough concentrations were measured before telaprevir initiation during the previous course of peg - ifn / rbv combination therapy ( t-1 ) , during the early phase ( week ( w ) 4 2 weeks ) , the later phase ( w 8 2 weeks ) , and after telaprevir cessation ( at least 4 weeks after telaprevir withdrawal ( w16 ) ) . \n as rbv daily doses may vary among patients and during the study course , rbv exposure was calculated using the ratio concentration / dose / body weight and used as a mean of estimated inter - individual bioavailability . \n the commonly used modification of the diet in renal disease ( mdrd ) study equation was used for the egfr as follows : in males : 186 ( creatinine 0.0113 ) age . if the patient was of african origin or female , multiplicative factors of 1.21 and 0.742 were applied , respectively . \n the occurrence of renal insufficiency during treatment was defined as an egfr < 60 ml / minutes/1.73 m. the nominal and categorical parameters were expressed as absolute numbers and percentages . \n means and standard deviations ( sd ) were calculated for the normally distributed variables , and medians and interquartile ranges ( iqr ) for the skewed variables . the non - parametric test for paired data ( wilcoxon test ) \n was used to analyze the variations in rbv bioavailability and the egfr before , during , and after triple therapy . \n all the analyses were performed using statistical package for the social sciences ( spss ) , version 19.0 for windows ( spss inc . , \n all the patients included in this study gave their written inform consent to allow the use of their personal clinical data in accordance with the ethics regulation defined in france by the cnil ( commission nationale de linformatique et des liberts ) . \n hcv patients with a previous non - response to peg - ifn / rbv therapy and re - treated with peg - ifn / rbv and telaprevir were prospectively recruited from the department of hepatology , croix - rousse hospital , lyon , france . \n non - response was defined as a failure to achieve a sustained virological response and could thus include null response , virological breakthrough , and virological relapse . \n patients were selected if they had consecutive assessment of plasma rbv trough concentrations ( prescribed at the clinician s discretion ) . \n patients with chronic kidney disease ( ckd ) stage 4 ( estimated glomerular filtration rate ( egfr ) < 30 ml / minutes/1.73 m ) were excluded from the study . \n plasma rbv trough concentrations were measured using a validated high - performance liquid chromatography - diode array detector method ( 10 ) . \n this method is highly specific , sensible ( limit of quantification = 0.05 mg / l ) , and precise ( total imprecision , calculated by measuring the coefficient of variation of the internal quality control values , was < 10% for concentrations from 0.20 - 5.00 \n plasma rbv trough concentrations were measured before telaprevir initiation during the previous course of peg - ifn / rbv combination therapy ( t-1 ) , during the early phase ( week ( w ) 4 2 weeks ) , the later phase ( w 8 2 weeks ) , and after telaprevir cessation ( at least 4 weeks after telaprevir withdrawal ( w16 ) ) . \n as rbv daily doses may vary among patients and during the study course , rbv exposure was calculated using the ratio concentration / dose / body weight and used as a mean of estimated inter - individual bioavailability . \n the commonly used modification of the diet in renal disease ( mdrd ) study equation was used for the egfr as follows : in males : 186 ( creatinine 0.0113 ) age . if the patient was of african origin or female , multiplicative factors of 1.21 and 0.742 were applied , respectively . \n the occurrence of renal insufficiency during treatment was defined as an egfr < 60 ml / minutes/1.73 m. \n means and standard deviations ( sd ) were calculated for the normally distributed variables , and medians and interquartile ranges ( iqr ) for the skewed variables . the non - parametric test for paired data ( wilcoxon test ) was used to analyze the variations in rbv bioavailability and the egfr before , during , and after triple therapy . \n all the analyses were performed using statistical package for the social sciences ( spss ) , version 19.0 for windows ( spss inc . , chicago , illinois , usa ) . \n all the patients included in this study gave their written inform consent to allow the use of their personal clinical data in accordance with the ethics regulation defined in france by the cnil ( commission nationale de linformatique et des liberts ) . \n male gender was predominant ( 73.0% ) , and mean age was 56.5 years ( 10.7 ) . \n thirty - one per cent of the patients were cirrhotics , among whom 18% had some sign of decompensation . \n three ( 8.1% ) patients had renal insufficiency defined as 30 < egfr < 60 ml / minutes/1.73 m ( ckd stage 3 ) . \n ( % ) except body mass index and age that are presented as mean sd . \n defined by 30 < estimated glomerular filtration rate ( egfr ) < 60 ml / minutes/1.73 m . \n a significant increase in rbv bioavailability was observed between t-1 and w4 ( n = 37 , 0.06 \n this increase was even stronger between t-1 and w8 ( n = 20 , 0.18 0.21 mg / l / dose / kg ; p < 0.001 ) . \n a significant increase was also observed between w4 and w8 ( n = 20 , 0.11 0.17 mg / l / dose / kg ; p < 0.001 ) . among these patients , 13 had measures of rbv bioavailability before , during , and after triple therapy . in this subgroup of patients , similar results were observed with an increase after telaprevir initiation ( 0.08 0.10 mg / l / dose / kg between t-1 and w4 ; p = 0.017 and 0.07 0.12 mg / l / dose / kg between w4 and w8 ; p = 0.012 ) . a decrease in bioavailability , \n albeit non - significant , was observed after telaprevir cessation ( -0.03 0.14 mg / l / dose / kg between w8 and w16 ; p = 0.839 ) ( figure 1 ) . \n eleven patients had a rbv dose reduction between w4 and w8 , and all of them still increased their rbv bioavailability despite a reduced plasma concentration in five of them . \n none of the following factors was predictive of the variations in rbv bioavailability : gender ( p = 0.837 ) ; il-28b genotype(p = 0.630 ) ; cirrhosis versus non cirrhosis ( p = 0.414 ) ; fibrosis stage ( p = 0.858 ) ; hcv genotype 1a versus 1b ( p = 0.937 ) ; and body mass index ( bmi ) ( p = 0.436 ) , and nor was the egfr at baseline associated with rbv bioavailability ( p = 0.527 ) . only age was significantly associated with a higher increase in rbv bioavailability in the older patients ( p = 0.011 ) . in parallel to these variations , \n a decrease in the egfr was observed between t-1 and w4 ( n = 37 , -6.8 25.21 ml / minutes/1.73 m ; p = 0.109 ) and an increase after telaprevir cessation in the subgroup of 13 patients ( 6.34 23.7 ml / minutes/1.73 m2 ; p = 0.243 ) ( figure 1 ) . \n when comparing between three different groups of patients characterized by their egfr at t-1 ( i.e. egfr < 60 , 60 - 90 , and > 90 ml / minutes/1.73 m , respectively ) , the group of patients with an initial egfr > 90 was associated with the most important decrease in the egfr between t-1 and w4 compared with the other two groups ( p < 0.001 ) . \n no significant difference in the mean rbv bioavailability increase was observed between these three groups ( p = 0.555 ) . \n anemia was also assessed during the different phases of therapy . a significant decrease in the hemoglobin level \n g / l ; p = 0.004 ) and also between w4 and w8 ( n = 20 , 8.5 18.8 \n however , no significant variations were observed between w8 and w16 for the subgroup of 13 patients . \n the assessment of rbv bioavailability measures the actual exposure to rbv , whereas the trough plasma concentration varies with posology . \n rbv bioavailability is known to range between 45% and 65% with an important variability within and between individuals , which can reach 30% after an oral single dose ( 11 ) . \n equilibrium state is reached after 4 weeks of multiple dosing and the half - life is 300 hours . \n our results showed that during triple therapy , rbv bioavailability was significantly increased after the introduction of telaprevir irrespective of the variations in the rbv doses . \n this increase was progressive throughout exposure to telaprevir . after the cessation of the hcv protease inhibitor , a 4-week period was necessary to observe a decrease in rbv bioavailability . \n in parallel to these variations , the egfr also seemed to have been impaired by telaprevir . \n recently , some authors have shown in hiv - hcv coinfected patients that telaprevir enhances rbv - induced anemia through renal function impairment ( 12 ) . \n telaprevir was already known to play a role in the inhibition of renal drug transporter in the in vitro system ( 13 ) . in hek 293 cell lines , telaprevir exposure seems to result in the inhibition of the organic cation transporter 2 ( oct2 ) and of the multidrug and toxin extrusion ( mate)-type transporter 1 ( mate1 ) ( 14 ) . \n this mechanism could explain the egfr decrease in patients treated with telaprevir , which in turn is a risk factor for anemia ( 15 ) . \n rbv - induced anemia is thought to result from molecular mechanisms such as the inhibition of intracellular energy metabolism and oxidative membrane damage ( 16 ) . \n it is thus possible that renal dysfunction due to hcv protease inhibitor increases rbv exposure , thereby inducing anemia . \n our study , including 37 patients , was possibly underpowered to detect potential associations between rbv bioavailability and co - factors such as gender , bmi , and other clinical factors . in a previous study , jen et al . \n ( 6 ) reported associations between the rbv apparent clearance and body weight , gender , and age and showed that the rbv apparent clearance was increased among the patients with a higher bmi and among the males , but decreased among the patients above 40 years of age . \n this latter observation is in accordance with our results , showing a higher increase in rbv bioavailability in the older patients . \n a more recent study also showed that among the hcv genotype 2/3 patients treated with peg - ifn / rbv , obesity ( bmi > 30 ) was associated with lower rbv concentrations ( 17 ) . \n 18 ) , who observed that lean body weight was the only covariate with a clinically significant influence on rbv pharmacokinetics and that rbv exposure decreased as weight increased . in the present study \n , we did not include data on albumin , bilirubin , or prothrombin time , involved in the hepatocellular function . \n however , various studies have demonstrated that the liver function has no effect on the rbv concentration ( 19 - 21 ) . \n although conducted on a limited number of patients , our study clearly shows a reversible increase in rbv exposure during telaprevir treatment , which might be linked to the impairment of the egfr . \n this also suggests a rbv - telaprevir pharmacological interaction , a possible source of severe anemia observed under triple therapy and already reported in hcv - monoinfected ( 22 ) or hiv - hcv coinfected cirrhotic patients ( 23 ) . \n the persistence of a high rbv exposure despite rbv dose reductions recommended for severe anemia could explain why these dose reductions do not impact the virological response . \n as recently suggested , rbv pharmacological monitoring may still be clinically relevant even in the context of direct - acting antivirals - based therapy ( 24 , 25 ) . \n this monitoring , potentially followed by rbv dose adaptations , could ensure an optimal rbv exposure , thereby improving safety and reducing the relapse risk . \n cautious clinical monitoring of patients focusing not only on hematological parameters but also on the renal function is warranted .", "role": "user" }, { "content": "background : anemia is more frequent in patients receiving telaprevir with pegylated interferon / ribavirin ( peg - ifn / rbv ) than in those receiving peg - ifn / rbv alone.objectives:the objective was to measure the impact of telaprevir on rbv bioavailability and to assess the concomitant renal function.materials and methods : thirty - seven hepatitis c virus ( hcv ) patients non - responders to a previous course of peg - ifn / rbv therapy and re - treated with triple therapy combining peg - ifn / rbv and telaprevir were analyzed . \n rbv bioavailability was measured before the triple therapy initiation , during telaprevir treatment at week ( w ) 4 and w8 , and after telaprevir cessation ( post w16 ) . \n the renal function was assessed by estimating the glomerular filtration rate ( egfr).results : at w4 , rbv bioavailability , expressed as mg / l / daily dose / kg body weight , was significantly increased ( median increase = 0.06 mg / l / dose / kg ; p < 0.001 ) . in parallel , \n the renal function was impaired with a mean egfr decrease of -6.8 ml / minutes/1.73 m ( p = 0.109 ) . between w4 and w8 , \n rbv bioavailability continued to increase ( p < 0.001 ) but subsequently decreased slightly after telaprevir discontinuation with a concomitant restoration of the renal function ( egfr increase of 6.34 ml / minutes/1.73 m).conclusions : our results indicated a reversible increase in rbv bioavailability after telaprevir exposure , which might be linked to the impairment of the gfr . \n this also suggests a rbv - telaprevir pharmacological interaction , a possible source of severe anemia observed under triple therapy . \n these results suggest that rbv pharmacological monitoring may be clinically relevant , especially in the context of first - generation hcv protease inhibitor - based therapy .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: a 29-year - old male reported to the department of dental surgery , cmch , vellore , tamil nadu , india , with an inability to sleep for the since the past 1-year with a reduced mouth opening from childhood . \n a detailed history revealed that he was diagnosed with ja and was managed with methotrexate for a year , before 5 years . \n he had also been diagnosed with severe sleep apnea syndrome and had an epworth sleepiness scale of 24/24 . on examination , he was found to have a retrognathic mandible , proclined anteriors , decreased chin - neck angle ( andy gump deformity ) , hypotonic / hypoplastic lower facial musculature , increased overjet and overbite , deranged occlusion with retruded chin , and a mouth opening of 16 mm [ figures 1 and 2 ] . \n polysomnography was done to assess the degree of sleep apnea and a bone isotope scan was needed to evaluate residual tmj arthritis . \n a computed tomography ( ct ) scan showed the extent of mandibular deformity which was resulting in the mechanical obstruction to breathing [ figure 3 ] . \n bone isotope scan revealed the absence of arthritis in the tmj and the rest of the skeleton , based on tracer uptake . \n preoperative view of straight and lateral profile preoperative mouth opening preoperative sagittal view of computed tomography scan a three - dimensional ct revealed the absence of both the condyles . \n condylar atrophy secondary to ja had resulted in a decrease in the mandibular ramal height as well as the length of the body [ figure 4 ] . on the basis of the presentation \n , a dual stage surgery was planned wherein a pair of titanium condylar analogues would be used to replace the missing condyles followed by a mandibular advancement at a later period . \n the stage i ( condylar analogue ) surgery was expected to improve the mouth opening as well as provide a posterior stop to the ramus ( proximal segment ) while advancing the distal segment of the mandible anteriorly in the second stage . before surgical intervention , \n a pair of titanium condylar analogue was used to reconstruct bilateral tmj [ figure 5 ] . following the surgery , \n after 4 weeks , epworth sleepiness scale showed marginal improvement from 24/24 to 16/24 , but was not satisfactory as the polysomnography revealed an apnea - hypopnea index ( ahi ) of 47.8/h , with a desaturation index ( dsi ) of 50.7/h indicating persistence of severe sleep apnea status . \n the body osteotomy was performed anterior to the mental foramen and fixed using titanium reconstruction plate with inter - positioning iliac graft bilaterally [ figure 6 ] . \n postoperatively the patient was very comfortable with his sleep pattern and very much satisfied with his facial appearance because he was no longer a \n postoperative lateral cephalometric view of stage 2 surgery postoperative view of straight and lateral profile the postoperative cephalogram confirmed the skeletal changes which were evident clinically . \n there was also an increase in the retrolingual - pharyngeal space ( between the back of the tongue and posterior pharyngeal wall ) from 2.7 mm to 10.2 mm , which is an index of the level of respiratory obstruction . \n a polysomnography revealed marked improvement with an ahi of 7/h and dsi of 6/h , indicating only mild osas . \n obstructive sleep apnea is a growing public health concern because it can cause hypertension , cardiac arrhythmias , stroke , and in rare circumstances , sudden death if untreated . \n although nasal continuous positive airway pressure therapy is considered the first - line treatment of osa , surgery has been shown to be a valid option for patients who are intolerant to positive pressure therapy . \n orthognathic surgery for repositioning the mandible and the genial tubercles for forward positioning of genioglossus has been recommended as a successful method of increasing the post glottic space and reducing the respiratory obstruction during sleep . \n the use of orthognathic surgery to treat osas began toward the end of the 1970s when mandibular surgery was reported to have reversed the symptoms of sleep apnea . \n advancement of the mandible and maxilla increase the available tongue space and have a positive influence on the pharyngeal airway . \n maxillomandibular advancement decreases the collapsibility of the velopharyngeal and suprahyoid musculature and improves lateral pharyngeal wall collapse , all of which have been shown to be significant components contributing to the upper airway obstruction in osa . \n the outcome of maxillomandibular advancement has been extensively reported , with success rates of 57% to 100% . \n the cephalometric radiograph , although rarely used by other specialties , is very valuable because it demonstrates both bony and soft tissue abnormalities related to the soft palate , pharyngeal airway , and maxillomandibular complex . \n if the radiographic and clinical examinations of the soft tissues of the pharynx reveal a narrow airway in conjunction with retrognathia of the maxilla and mandible , the patient should be considered a candidate for mma surgery . because an underlying skeletal abnormality is sometimes hard to detect clinically , it is reasonable for an oral and maxillofacial surgeon to assess these patients . \n the surgeon 's goal is to optimize the advancement of the deficient structures while maintaining a normal facial balance for each patient . \n it is , therefore , beneficial to use a cephalometric analysis , which will clearly demonstrate all the maxillofacial abnormalities and provide a visual treatment objective . \n cephalometric analysis provides a reliable method of evaluating the changes in the tongue base before and after surgery . \n it is estimated that for each millimeter of maxillary and mandibular bone advancement there was a 0.76 mm increase in the retropalatal region and a 1.2 mm increase in the pharynx in the retrolingual region , respectively . \n bilateral tm joint reconstruction has been recommended for the replacement of the joint for the restoration of function and also for elongation of the shortened mandible in joint destructive diseases like juvenile rheumatoid arthritis . \n the use of custom made two part total joint reconstruction of the cranial articular surface as well as the condylar head has been advocated . in this case , \n consequently , the length of the vertical ramus could be increased by 6 mm , which could be maintained by a splint and intermaxillary traction . \n the mandible with the new condyles was made to function after a period of 1-month of traction . \n the patient was able to open his mouth more than before and the full range of mandibular movements was possible , proving that the condylar prostheses were functioning well . \n further increase in length of the mandible could be achieved ( 10 mm ) through the second procedure for a single stage mandibular advancement with bone grafting and rigid plating . \n the forward repositioning of the chin enabled the posterior pharyngeal space to be enlarged in the anteroposterior direction by 7.5 mm , as measured cephalometrically . \n this resulted not only in a cosmetically pleasing facial appearance but a significant relief from the osa . \n review of the patient over a period of 10 months has shown that the mandibular advancement procedure had a stable outcome . through the management of the described case it 's our observation that osa secondary to ja could be effectively managed with condylar replacement and mandibular advancement to produce a predictable outcome . however , early identification , diagnosis , and treatment planning could alleviate the agony manifested by these conditions . \n \n ", "role": "user" }, { "content": "variations affecting the growth centers can severely affect the normal formation and subsequent function of vital musculoskeletal structures . \n we report a case of bilateral condylar atrophy with a history of juvenile arthritis ( ja ) resulting in progressive obstructive sleep apnea ( osa ) in adulthood . \n in addition to this , the case report emphasizes the role of temporomandibular joint replacement and advancement of the mandible to correct progressive osa secondary to idiopathic ja . \n computed tomography revealed micrognathia , condylar hypoplasia , and decreased pharyngeal airway space . \n the resultant increase in the retrolingual - pharyngeal airway space following the surgery , helped to completely resolve the presenting symptoms . \n it is hoped that the described technique could be used in similar cases with a predictable outcome .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: human cytomegalovirus ( hcmv ) prevalence in the general population is about 70% ( 1 ) . the infection is an important cause of morbidity and mortality in infants and immunocompromised individuals such as solid - organ and hematopoietic stem cell transplant recipients , aids patients and patients taking chemotherapy or medicines with suppressed immune system ( 2 - 7 ) . \n the hcmv accounts for major viral complications in the post - transplant period ( 6 ) and is also the most common intrauterine infection that may lead to sensorineural hearing loss and mental retardation ( 8 , 9 ) . \n these medicines require close monitoring for side effects and although they can help stop the virus from copying itself within the body yet do not eliminate it from the body ( 10 - 12 ) . \n immunotherapy is a promising new treatment for infectious diseases involving both therapeutic vaccines and monoclonal antibodies ( 13 ) . \n glycoprotein 55 ( gp55 ) is part of glycoprotein b ( gb ) , which is the most abundant constituent of hcmv envelope and the major target for neutralizing antibodies ( 14 , 15 ) . \n several monoclonal antibodies have been generated against gb epitopes and showed some degree of neutralization ( 16 ) . \n recombinant antibodies ( rab ) are becoming the preferred therapeutic substitutes to full - length monoclonal antibodies due to their characteristics . \n single - chain fragment variable antibodies ( scfv ) , which are composed of variable heavy ( vh ) and variable light ( vl ) domains linked together by a flexible polypeptide linker , are one of the most desired rab formats . \n the smaller size , human origin , high affinity and specificity and also better penetration to their targets , are beneficial characteristics of human scfvs in medical applications ( 17 - 20 ) . \n this study aimed to select specific single chain antibodies against the conserved sequence of gp55 and to evaluate the virus neutralization effects of the antibodies in cell culture using the phage display technology . \n a phage antibody display library of scfv was produced , as described previously ( 16 , 17 ) . \n escherichia coli bacteria containing phagemid were cultured overnight on 2tyg agar /ampicillin ( tryptone , yeast extract , glucose , agar and ampicillin ) ( merck , germany ) plates at 30c . \n the bacterial pellet was transferred to 2ty broth containing ampicillin ( 100 g / ml ) and kanamycin ( 50 l / ml ) , and incubated with constant shaking , at 30c overnight . \n the supernatant was passed through 0.3 m filters and stored at 4c ( 22 ) . \n peptides ( lvsadgttvtsgstkdtslq ) at a concentration of 10 g / ml were coated on polystyrene immunotubes ( nunc , denmark ) at 4c overnight . \n the tubes were washed four times with phosphate buffered saline ( pbs ) , and 4 ml of blocking solution ( 2% skimmed milk ) was added and incubated at 37c for two hours . \n the tubes were washed four times with pbs/ tween and four times with pbs . in the next step \n , the diluted phage supernatant in the blocking solution ( 1/1 ) was added to the tubes and incubated at room temperature for one hour with occasional inversions . \n the tubes were washed and logarithmic phase e. coli was added , followed by incubation at 37c for one hour and centrifugation at 3500 rpm for five minutes . the supernatant was discarded and the bacterial pellet was re - suspended in 2ty broth ( tryptone , yeast extract ) ( merck , germany ) , plated on 2ty agarose / ampicillin plate and incubated at 30c , overnight . \n after the panning process , the inserts of selected clones were amplified by pcr ( 94c for one minute , 55c for one minute and 72c for two minutes , performed in 30 cycles ) . \n furthermore , 17 l of the pcr product was mixed with 1 l restriction enzyme ( mva - i ) ( roche applied science , germany ) and 2 l of restriction enzyme buffer . \n the mixtures were placed in a dry block heater at 37c for two hours and run on a 2% agarose gel . \n peptides ( 100 g / ml ) as an epitope were coated in 96 wells of a polystyrene plate and incubated at 4c overnight . after washing with pbs , 150 l of blocking solution ( \n 2% skimmed milk ) was added to each well and incubated at 37c for two hours . \n phage rescue supernatant containing the appropriate scfv , diluted with blocking solution ( 1:1 ) , was added to each well and incubated at room temperature for two hours . \n nonbinding phages were removed by washing the wells three times with pbs / tween , and three times with pbs . \n anti - fd bacteriophage ( 1:100 ) ( sigma , germany ) was added to each well and incubated at room temperature for 1.5 hours . \n the wells were washed three times with pbs / tween and three times with pbs . \n horseradish peroxidase ( hrp ) conjugated anti - rabbit antibody ( 1:1000 ) ( sigma , germany ) was added to each well and incubated at room temperature for one hour . \n the wells were then washed and 150 l of azino - bis-3-ethylbenzothiazoline-6-sulfonic ( abts ) acid ( sigma - aldrich , germany ) solution ( 10 mg abts , 20 ml citrate buffer ( ph4 ) and 6 ml h2o2 ) was added to each well . \n the absorbances were read at 405 nm after 30 minutes using an elisa reader . the wells without peptide , with unrelated peptide ( prostate stem cell antigen peptide ) , unrelated scfv ( scfv to her2 ) and with m13ko7 , \n the hcmv ( ad169 ) was diluted in dulbecco s modified eagle s medium ( dmem ) ( sigma - aldrich , germany ) to 50 pfu / ml . \n equal amounts of virus and each scfv phage rescue supernatant were mixed and incubated at 37c for one hour . \n the mixtures were added to the wells of 24-well plates containing confluent human fibroblast , which were then rocked gently for 45 minutes at 37c . \n the mixtures were removed and 1 ml of 1% w / v carboxy methyl cellulose ( cmc ) in 1x dmem , supplemented with 2% fetal calf serum , 0.14% v / v sodium bicarbonate , 100 u penicillin ml , 100 g streptomycin sulfate ml , 0.25 g amphotericin b ml and 0.1 m sodium hydroxide , was added . \n the number of micro plaques were counted after ten days and compared with the number of plaques seen in the virus control wells containing virus in dmem . \n a phage antibody display library of scfv was produced , as described previously ( 16 , 17 ) . \n escherichia coli bacteria containing phagemid were cultured overnight on 2tyg agar /ampicillin ( tryptone , yeast extract , glucose , agar and ampicillin ) ( merck , germany ) plates at 30c . \n the bacterial pellet was transferred to 2ty broth containing ampicillin ( 100 g / ml ) and kanamycin ( 50 l / ml ) , and incubated with constant shaking , at 30c overnight . \n the supernatant was passed through 0.3 m filters and stored at 4c ( 22 ) . \n peptides ( lvsadgttvtsgstkdtslq ) at a concentration of 10 g / ml were coated on polystyrene immunotubes ( nunc , denmark ) at 4c overnight . \n the tubes were washed four times with phosphate buffered saline ( pbs ) , and 4 ml of blocking solution ( 2% skimmed milk ) was added and incubated at 37c for two hours . \n the tubes were washed four times with pbs/ tween and four times with pbs . in the next step \n , the diluted phage supernatant in the blocking solution ( 1/1 ) was added to the tubes and incubated at room temperature for one hour with occasional inversions . \n the tubes were washed and logarithmic phase e. coli was added , followed by incubation at 37c for one hour and centrifugation at 3500 rpm for five minutes . the supernatant was discarded and the bacterial pellet was re - suspended in 2ty broth ( tryptone , yeast extract ) ( merck , germany ) , plated on 2ty agarose / ampicillin plate and incubated at 30c , overnight . \n after the panning process , the inserts of selected clones were amplified by pcr ( 94c for one minute , 55c for one minute and 72c for two minutes , performed in 30 cycles ) . \n furthermore , 17 l of the pcr product was mixed with 1 l restriction enzyme ( mva - i ) ( roche applied science , germany ) and 2 l of restriction enzyme buffer . \n the mixtures were placed in a dry block heater at 37c for two hours and run on a 2% agarose gel . \n peptides ( 100 g / ml ) as an epitope were coated in 96 wells of a polystyrene plate and incubated at 4c overnight . after washing with pbs , \n 150 l of blocking solution ( 2% skimmed milk ) was added to each well and incubated at 37c for two hours . \n phage rescue supernatant containing the appropriate scfv , diluted with blocking solution ( 1:1 ) , was added to each well and incubated at room temperature for two hours . \n nonbinding phages were removed by washing the wells three times with pbs / tween , and three times with pbs . \n anti - fd bacteriophage ( 1:100 ) ( sigma , germany ) was added to each well and incubated at room temperature for 1.5 hours . \n the wells were washed three times with pbs / tween and three times with pbs . \n horseradish peroxidase ( hrp ) conjugated anti - rabbit antibody ( 1:1000 ) ( sigma , germany ) was added to each well and incubated at room temperature for one hour . \n the wells were then washed and 150 l of azino - bis-3-ethylbenzothiazoline-6-sulfonic ( abts ) acid ( sigma - aldrich , germany ) solution ( 10 mg abts , 20 ml citrate buffer ( ph4 ) and 6 ml h2o2 ) was added to each well . the absorbances were read at 405 nm after 30 minutes using an elisa reader . the wells without peptide , with unrelated peptide ( prostate stem cell antigen peptide ) , unrelated scfv ( scfv to her2 ) and with m13ko7 , were also considered as negative controls ( 22 ) . \n the hcmv ( ad169 ) was diluted in dulbecco s modified eagle s medium ( dmem ) ( sigma - aldrich , germany ) to 50 pfu / ml . \n equal amounts of virus and each scfv phage rescue supernatant were mixed and incubated at 37c for one hour . \n the mixtures were added to the wells of 24-well plates containing confluent human fibroblast , which were then rocked gently for 45 minutes at 37c . \n the mixtures were removed and 1 ml of 1% w / v carboxy methyl cellulose ( cmc ) in 1x dmem , supplemented with 2% fetal calf serum , 0.14% v / v sodium bicarbonate , 100 u penicillin ml , 100 g streptomycin sulfate ml , 0.25 g amphotericin b ml and 0.1 m sodium hydroxide , was added . \n the number of micro plaques were counted after ten days and compared with the number of plaques seen in the virus control wells containing virus in dmem . \n dna fingerprinting of selected clones was performed to determine the common patterns after the panning process . as shown in figure 1 , dna fingerprinting of the selected clones demonstrated pattern 1 ( scfv1 ) , ( lanes : 1 , 2 , 5 , 8 , 13 ) with frequency of 25% , pattern 2 ( scfv 2 ) , ( lanes : 4 , 7 , 11 , 14 ) with frequency of 20% and pattern 3 ( scfv3 ) , ( lanes : 3 , 15 , 17 , 20 ) with frequency of 20% . \n dominant patterns ( patterns 1 , 2 and 3 ) were selected for further evaluation . \n three predominant patterns were differentiated including pattern 1 ( lanes 1 , 2 , 5 , 8 and 13 ) , pattern 2 ( lanes 4 , 7 , 11 and 14 ) and pattern 3 ( lanes 3 , 15 , 17 and 20 ) . \n specific binding of the selected scfv antibodies to the corresponding peptide was determined by phage elisa . \n the absorbance of wells coated with the corresponding peptide for the three selected scfvs were significantly higher than that of the wells containing no peptide . \n table 1 , shows the percentages of plaque reduction when the virus was incubated with scfv . \n as the results indicate only scfv2 efficiently neutralized the virus and a neutralizing effect of 68.8% was obtained . \n number of plaques are shown in each experiment . following incubation of virus with scfv2 , \n dna fingerprinting of selected clones was performed to determine the common patterns after the panning process . as shown in figure 1 , dna fingerprinting of the selected clones demonstrated pattern 1 ( scfv1 ) , ( lanes : 1 , 2 , 5 , 8 , 13 ) with frequency of 25% , pattern 2 ( scfv 2 ) , ( lanes : 4 , 7 , 11 , 14 ) with frequency of 20% and pattern 3 ( scfv3 ) , ( lanes : 3 , 15 , 17 , 20 ) with frequency of 20% . \n dominant patterns ( patterns 1 , 2 and 3 ) were selected for further evaluation . \n three predominant patterns were differentiated including pattern 1 ( lanes 1 , 2 , 5 , 8 and 13 ) , pattern 2 ( lanes 4 , 7 , 11 and 14 ) and pattern 3 ( lanes 3 , 15 , 17 and 20 ) . \n specific binding of the selected scfv antibodies to the corresponding peptide was determined by phage elisa . \n the absorbance of wells coated with the corresponding peptide for the three selected scfvs were significantly higher than that of the wells containing no peptide . \n table 1 , shows the percentages of plaque reduction when the virus was incubated with scfv . \n as the results indicate only scfv2 efficiently neutralized the virus and a neutralizing effect of 68.8% was obtained . \n number of plaques are shown in each experiment . following incubation of virus with scfv2 , \n glycoprotein b is known as the most abundant constituent of the hcmv envelope and the major target for neutralizing antibodies . \n it contains three major neutralizing determinants including ad1 , ad2 and ad3 domains ( 23 - 25 ) . \n the hcmv peptides derived from the n - terminal part of gb ( netiynttlkygdv ) and c - terminal part of gb ( vtsgstkd ) in the ad2 region and n - terminal part of gh ( paasealdphafhlllntygr ) have been used to develop recombinant anti - hcmv gb and gh . \n this resulted in the isolation of eight neutralizing clones with neutralizing effect of 51% - 63% ( 16 ) . \n due to recent reports showing the association of cmv diseases with the lack of antibodies against the ad2 region in renal transplant individuals ( 23 ) . in the present study , a well - known peptide in the ad2 region \n was considered as the target epitope and a phage display library of scfvs was applied to select specific antibodies against this epitope . the peptide ( lvsadgttvtsgstkdtslq ) , amino acids 789 - 805 derived from the c - terminal end of the part of gb ( gp55 ) was used as an epitope . \n it has been shown that the sequence of ad2 domain is conserved among cmv isolates ( 26 ) , which makes it more attractive for cmv targeted therapy . \n the vtsgstkd epitope of ad2 has been reported as a major antigenic region on gp55 with the vtsg part , which is essential for recognition . \n it was found that 55% of the anti - hcmv seropositive blood samples recognize the tsgstkd epitope , which is part of the applied peptide . in order to select specific scfv antibodies against the epitope , \n after four rounds of panning against the derived peptide , 20 clones were selected and mva- fingerprinting demonstrated the common patterns . \n three specific scfvs were selected with frequencies of 25% ( scfv1 ) , 20% ( scfv2 ) and 20% ( scfv3 ) . \n lillo et al . , ( 27 ) selected scfv antibodies against a capsular protein ( f1 antigen ) of yersinia pestis by enrichment of a phage library using the panning process . \n krishnaswamy et al . , ( 28 ) identified a specific scfv antibody against a. fumigatus membrane fraction ( amf ) as an antigen by panning a phage antibody . \n the results of the phage elisa showed that the panning procedure was successfully performed and the selected clones reacted specifically with the peptide . \n the three selected scfvs ( scfv1 , scfv2 , scfv3 ) produced positive elisa with the cmv peptide , while no reactivity was detected for the control peptide , helper phage and no peptide wells as negative controls . \n furthermore , the unrelated scfv ( scfv to prostate stem cell antigen peptide ) did nt react with the gp55 peptide . \n the absorbances of wells coated with peptide of interest for the three selected scfvs were three folds higher than that of the wells containing no peptide . \n the specificity of selected scfvs to the corresponding peptides has been shown in different phage elisa assessments . \n thathaisong et al . , ( 29 ) showed that the optical density ( od ) of specific scfvs against influenza - a virus h5n1 subtype at 405 nm was two folds higher than negative controls in a positive phage elisa . \n specific single chain antibodies to outer membrane protein d ( ompd ) of salmonella typhimurium ( 30 ) , staphylococcal enterotoxin b ( seb ) ( 31 ) and hepatitis b virus ( hbv ) surface antigen ( 32 ) have also been confirmed by phage elisa . \n the scfv2 showed a mean plaque reduction of 68.8% while the mean of plaque reduction for scfv1 and scfv3 were 23.7 and 11.6% , respectively . \n although monoclonal antibodies have been produced against human cytomegalovirus gb and gh ( 33 , 34 ) , the application of these antibodies in clinical settings is limited due to human anti - mouse antibody ( hama ) response and most of these antibodies have been used for diagnosis purposes . \n new treatment options for cmv are urgently needed because currently available therapies have major limitations . \n it has also been reported that the management of solid organ transplant recipients for prevention and treatment of cmv disease is a major strategy since a significant correlation between steroid administration for acute cellular rejection ( acr ) and cmv reactivation and disease has been shown in liver transplant recipients ( 35 ) . \n the results suggest that the specific scfv2 with neutralizing effect of 68.8% has the potential to be used for prophylaxis and treatment of cmv infections . \n as the epitope used for selection of the specific antibodies is a conserved epitope among different cmv strains , the antibody is able to neutralize different cmv strains and would be helpful for clinical applications . \n the specific scfvs 1 and 3 with lower neutralizing effects can be used for diagnosis of cmv - related diseases . a combination of the selected neutralizing scfv in this study with the previously selected neutralizing scfvs and scfv against the n - terminal part of gb with 57% and scfv against gh of cmv with 63% neutralizing effects ( 16 ) , could be an effective alternative cocktail for therapy of at risk patients such as transplant recipients and aids patients .", "role": "user" }, { "content": "background : immunotherapy is a promising prospective new treatment for cytomegalovirus ( cmv ) infections . neutralizing effects have been reported using monoclonal antibodies . \n recombinant single chain antibodies ( scfvs ) due to their advantages over monoclonal antibodies are potential alternatives and provide valuable clinical agents.objectives:the aim of this study was to select specific single chain antibodies against gp55 of cmv and to evaluate their neutralizing effects . in the present study , we selected specific single chain antibodies against glycoprotein 55 ( gp55 ) of cmv for their use in treatment and diagnosis.materials and methods : single chain antibodies specific against an epitope located in the c - terminal part of gp55 were selected from a phage antibody display library . after four rounds of panning , \n twenty clones were amplified by the polymerase chain reaction ( pcr ) and fingerprinted by mvai restriction enzyme . \n the reactivities of the specific clones were tested by the enzyme - linked immunosorbent assay ( elisa ) and the neutralizing effects were evaluated by the plaque reduction assay.results:fingerprinting of selected clones revealed three specific single chain antibodies ( scfv1 , scfv2 and scfv3 ) with frequencies 25% , 20 and 20% . \n the clones produced positive elisa with the corresponding peptide . \n the percentages of plaque reduction for scfv1 , scfv2 and scfv3 were 23.7 , 68.8 and 11.6 , respectively.conclusions:gp55 of human cmv is considered as an important candidate for immunotherapy . in this study , we selected three specific clones against gp55 . \n the scfvs reacted only with the corresponding peptide in a positive elisa . \n the scfv2 with 68.8% neutralizing effect showed the potential to be considered for prophylaxis and treatment of cmv infections , especially in solid organ transplant recipients , for whom treatment of cmv is urgently needed . \n the scfv2 with neutralizing effect of 68.8% , has the potential to be considered for treatment of these patients . \n the specific scfv1 and scfv3 with lower neutralizing effects can be used for diagnostic purposes .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: chronic hepatitis c ( chc ) is a major cause of chronic liver disease and affects > 185 million people worldwide.1 the most serious complications of chc are high mortality from decompensated liver cirrhosis and hepatocellular carcinoma ( hcc).2,3 age and age at infection are two of the most important risk factors for the progression to liver cirrhosis during chronic hepatitis c virus ( hcv ) infection.4,5 patient age seems to have a greater effect than age at infection , and is especially significant for patients older than 65 years.6 many countries have aging cohorts of people with hcv infections because of previous peaks in the incidence of hcv infections.2,4 a sustained virologic response ( svr ) , achieved by interferon - based therapies , can slow the progression of liver fibrosis710 and reduce the development of hcc and liver - related mortality.1113 thus , antiviral treatments may provide the greatest benefit for elderly patients . \n however , the high prevalence of cirrhosis and comorbidities in elderly patients leads to higher rates of treatment discontinuation and dose reduction.14,15 in particular , the svr rate is lower in elderly patients than young patients due to their poor tolerance to treatment.15 thus , the decision of treatment for an elderly patient must consider the long - term benefits and the patient s life expectancy . \n few studies have examined the occurrence rate of liver cirrhosis after achieving svr in elderly patients with chc . the aspartate aminotransferase - to - platelet ratio index ( apri ) \n is a noninvasive marker that has been validated for the diagnosis of both significant fibrosis and cirrhosis.16,17 apri is a useful marker for the prognosis in chc patients . \n however , the prognostic value of apri in elderly patients for predicting the occurrence of liver cirrhosis is uncertain . \n the aim of this study was to evaluate the risk of the progression to liver cirrhosis after combined therapy consisting of pegylated interferon ( peg - ifn ) plus ribavirin ( rbv ) in elderly patients ( 65 years old ) with chc . \n patients with hcv infection who were aged 65 and underwent treatment with either peg - ifn--2a or peg - ifn--2b plus rbv from january 2005 to december 2011 were enrolled from the dalin tzu chi general hospital in this retrospective study . \n all the patients were positive for anti - hepatitis c antibody for > 6 months , had alanine aminotransferase ( alt ) levels higher than the upper limit of normal ( uln ) , and had detectable serum hcv rna . \n patients with decompensated liver cirrhosis , malignant neoplasms , incomplete medical records , autoimmune diseases , hiv infection , neutropenia ( < 1,500 neutrophils / ml ) , thrombocytopenia ( < 75,000 platelets / ml ) , anemia ( < 12 g hemoglobin / dl in females and < 13 g / dl in males ) , or poorly controlled psychiatric diseases were excluded . \n fifty - three patients with compensated liver cirrhosis , five patients with hcc , 16 patients with posttreatment follow - up times < 1 year , and six patients without complete medical records were excluded . \n there were 801 person - years of follow - ups , and the average follow - up time was 5.5 years ( standard deviation [ sd ] : 2.5 years , range : 1.112.3 years ) . \n this study was approved by the ethics committee of dalin tzu chi general hospital ( b10303008 ) , all patients provided written consent , and the local ethics committee approved the consent process . \n oral rbv plus subcutaneous peg - ifn--2a ( 180 g ; pegasys ; hoffmann - la roche , basel , switzerland ) or subcutaneous peg - ifn--2b ( 1.5 g / kg ; pegintron ; schering - plough corp . , \n the fixed duration ( 6 months ) without consideration of hcv genotypes is due to restrictions imposed by the reimbursement policy of the bureau of national health insurance in taiwan . \n the dose of oral rbv was 800 mg / day for patients who weighed < 55 kg , 1,000 mg / day for patients who weighed 5575 kg , and 1,200 mg / day for patients who weighed > 75 kg . \n dose adjustments of all drugs and administration of supplemental erythropoietin or blood transfusion were determined according to published practice guidelines.1820 the primary outcome was time to liver cirrhosis . \n all patients were given liver function tests ( serum aspartate aminotransferase [ ast ] , alt , total bilirubin , creatinine , hemoglobin , white blood cell count , and platelets ) , and abdominal sonography was performed at the gastrointestinal outpatient clinic at baseline , end of treatment ( 6 months ) , 24 weeks after the end of treatment , and every 36 months thereafter . \n liver cirrhosis and associated liver complications were evaluated every 36 months after the end of treatment . \n radiologic cirrhosis was defined as two documented ultrasonograms of liver cirrhosis ( included coarse liver echotexture with nodularity and small liver size ) combined with features of portal hypertension ( splenomegaly , ascites , hepatic encephalopathy , or varices).21 liver biopsy was an optional procedure and was performed in 108 patients ( 74.5% ) at baseline with consent . \n hcv rna was quantified at baseline and at 24 weeks after the end of treatment . \n a diagnosis of fatty liver was based on results from a biopsy and/or abdominal ultrasound . \n other clinical factors , including diabetes mellitus , chronic hepatitis b , and alcoholism , were recorded by chart review . \n chronic hepatitis b was diagnosed if a patient had seropositivity for hepatitis b surface antigen for at least 6 months . \n the apri score was calculated as : [ ( ast / uln)/platelet count ( 10/l ) ] 100.22 the apri was used as a noninvasive maker for fibrosis . \n serum hcv rna was quantified at baseline and at 24 weeks after the end of treatment by use of real - time polymerase chain reaction , which had a detection limitation of 15 iu / ml.23 the threshold for discriminating low and high baseline hcv rna was 400,000 iu / ml.16 hcv genotyping was performed by melting curve analysis ( roche lightcycler ; biotronics tech corp . , \n lowell , ma , usa).24 svr was defined as undetectable hcv rna for at least 24 weeks after completion of the combined treatment . \n patients who were positive for hcv rna at week 24 after the end of treatment were considered non - svr . \n spss 19.0 for windows ( spss inc , chicago , il , usa ) was used for all statistical analyses . the chi - square test or fisher \n age , sex , diabetes , svr , other risks of hepatitis ( alcoholism , hepatitis b virus [ hbv ] coinfection , and fatty liver ) , hcv rna load , hcv genotype , and baseline apri score were considered as potential confounders for cirrhosis progression . in order to identify risk factors for cirrhosis , \n the proportional hazards cox regression model was used to control for those possible confounding factors , regardless of whether they were significant predictors in the univariate analyses.25 results were shown as hazard ratios ( hrs ) with 95% confidence intervals ( cis ) . \n patients with hcv infection who were aged 65 and underwent treatment with either peg - ifn--2a or peg - ifn--2b plus rbv from january 2005 to december 2011 were enrolled from the dalin tzu chi general hospital in this retrospective study . \n all the patients were positive for anti - hepatitis c antibody for > 6 months , had alanine aminotransferase ( alt ) levels higher than the upper limit of normal ( uln ) , and had detectable serum hcv rna . \n patients with decompensated liver cirrhosis , malignant neoplasms , incomplete medical records , autoimmune diseases , hiv infection , neutropenia ( < 1,500 neutrophils / ml ) , thrombocytopenia ( < 75,000 platelets / ml ) , anemia ( < 12 g hemoglobin / dl in females and < 13 g / dl in males ) , or poorly controlled psychiatric diseases were excluded . \n fifty - three patients with compensated liver cirrhosis , five patients with hcc , 16 patients with posttreatment follow - up times < 1 year , and six patients without complete medical records were excluded . \n there were 801 person - years of follow - ups , and the average follow - up time was 5.5 years ( standard deviation [ sd ] : 2.5 years , range : 1.112.3 years ) . \n this study was approved by the ethics committee of dalin tzu chi general hospital ( b10303008 ) , all patients provided written consent , and the local ethics committee approved the consent process . \n oral rbv plus subcutaneous peg - ifn--2a ( 180 g ; pegasys ; hoffmann - la roche , basel , switzerland ) or subcutaneous peg - ifn--2b ( 1.5 g / kg ; pegintron ; schering - plough corp . , \n the fixed duration ( 6 months ) without consideration of hcv genotypes is due to restrictions imposed by the reimbursement policy of the bureau of national health insurance in taiwan . \n the dose of oral rbv was 800 mg / day for patients who weighed < 55 kg , 1,000 mg / day for patients who weighed 5575 kg , and 1,200 mg / day for patients who weighed > 75 kg . \n dose adjustments of all drugs and administration of supplemental erythropoietin or blood transfusion were determined according to published practice guidelines.1820 \n the primary outcome was time to liver cirrhosis . all patients were given liver function tests ( serum aspartate aminotransferase [ ast ] , alt \n , total bilirubin , creatinine , hemoglobin , white blood cell count , and platelets ) , and abdominal sonography was performed at the gastrointestinal outpatient clinic at baseline , end of treatment ( 6 months ) , 24 weeks after the end of treatment , and every 36 months thereafter . \n liver cirrhosis and associated liver complications were evaluated every 36 months after the end of treatment . \n radiologic cirrhosis was defined as two documented ultrasonograms of liver cirrhosis ( included coarse liver echotexture with nodularity and small liver size ) combined with features of portal hypertension ( splenomegaly , ascites , hepatic encephalopathy , or varices).21 liver biopsy was an optional procedure and was performed in 108 patients ( 74.5% ) at baseline with consent . \n hcv rna was quantified at baseline and at 24 weeks after the end of treatment . \n a diagnosis of fatty liver was based on results from a biopsy and/or abdominal ultrasound . \n other clinical factors , including diabetes mellitus , chronic hepatitis b , and alcoholism , were recorded by chart review . \n chronic hepatitis b was diagnosed if a patient had seropositivity for hepatitis b surface antigen for at least 6 months . \n the apri score was calculated as : [ ( ast / uln)/platelet count ( 10/l ) ] 100.22 the apri was used as a noninvasive maker for fibrosis . \n serum hcv rna was quantified at baseline and at 24 weeks after the end of treatment by use of real - time polymerase chain reaction , which had a detection limitation of 15 iu / ml.23 the threshold for discriminating low and high baseline hcv rna was 400,000 iu / ml.16 hcv genotyping was performed by melting curve analysis ( roche lightcycler ; biotronics tech corp . , lowell , ma , usa).24 \n svr was defined as undetectable hcv rna for at least 24 weeks after completion of the combined treatment . \n patients who were positive for hcv rna at week 24 after the end of treatment were considered non - svr . \n spss 19.0 for windows ( spss inc , chicago , il , usa ) was used for all statistical analyses . the chi - square test or fisher s exact test was used for nominal variables . \n age , sex , diabetes , svr , other risks of hepatitis ( alcoholism , hepatitis b virus [ hbv ] coinfection , and fatty liver ) , hcv rna load , hcv genotype , and baseline apri score were considered as potential confounders for cirrhosis progression . in order to identify risk factors for cirrhosis , \n the proportional hazards cox regression model was used to control for those possible confounding factors , regardless of whether they were significant predictors in the univariate analyses.25 results were shown as hazard ratios ( hrs ) with 95% confidence intervals ( cis ) . \n there were 64 males and 81 females , and the mean patient age was 69.13.3 years ( table 1 ) . \n eighty - nine of 145 patients ( 61.4% ) were infected with hcv genotype-1 , and 56 patients ( 38.6% ) were infected with other hcv genotypes . \n ninety - five patients ( 65.5% ) , including 45 with hcv genotype-1 and 50 with other hcv genotypes , achieved svr . \n twenty - six patients ( 17.9% ) did not complete the full 6-month treatment , and the patients had mean treatment duration of 84 days . \n twenty - four patents did not accept complete treatment due to treatment side effect . among those patients , \n table 1 shows the baseline characteristics and treatment outcomes of patients who did and did not progress to liver cirrhosis . \n there were 801 person - years of follow - up data , and the average follow - up time was 5.5 years ( sd : 2.5 years , range : 1.112.3 years ) . during the follow - up , \n 27 patients ( 18.6% ) developed liver cirrhosis , 12 patients developed hcc , three patients developed varices , two patients developed ascites , and one patient developed spontaneous bacterial peritonitis . \n one death that was unrelated to liver disease occurred during the follow - up period . \n the average age of patients with liver cirrhosis was 70.23.3 years , and 17 of these patients ( 62.9% ) were females . among patients who progressed to cirrhosis ( n=27 ) , 14 patients ( 51.9% ) \n univariate analysis of patients who did and did not progress to cirrhosis indicated no significant differences in age , sex , diabetes , smoking , other risks of hepatitis ( alcoholism , hbv coinfection , and fatty liver ) , hcv rna load , hcv genotype , baseline apri , and svr . \n there was a trend of a higher baseline apri score in patients who progressed to liver cirrhosis compared with those who did not progress ( 2.11.2 vs 1.61.3 , p=0.055 ) . \n table 2 shows the result of cox regression analysis with adjustment for age , sex , diabetes , other risks of hepatitis ( alcoholism , hbv coinfection , and fatty liver ) , hcv rna load , hcv genotype , and baseline apri . \n the results show a significant association between progression to liver cirrhosis and no svr ( hr : 3.39 , 95% ci : 1.3128.761 , p=0.012 ) . \n there was also a trend of an association between liver cirrhosis and low hcv rna load ( hr : 0.392 , 95% ci : 0.1521.009 , p=0.052 ) . \n there was no significant association between progression to liver cirrhosis and high baseline apri by cox regression ( adjusted hr : 1.285 , 95% ci : 0.9211.791 , p=0.14 ) . \n the risk of liver cirrhosis was significantly greater in patients without svr ( 3-year cumulative incidence : 35.2% , 95% ci : 13.057.5 , p=0.012 ) than in those with svr ( 3-year cumulative incidence : 10.4% , 95% ci : 3.117.7 ) . \n decrease of progression to liver cirrhosis occurred in one patient for every four chc patients who achieved svr within 3 years . \n there were 64 males and 81 females , and the mean patient age was 69.13.3 years ( table 1 ) . \n eighty - nine of 145 patients ( 61.4% ) were infected with hcv genotype-1 , and 56 patients ( 38.6% ) were infected with other hcv genotypes . \n ninety - five patients ( 65.5% ) , including 45 with hcv genotype-1 and 50 with other hcv genotypes , achieved svr . \n twenty - six patients ( 17.9% ) did not complete the full 6-month treatment , and the patients had mean treatment duration of 84 days . \n twenty - four patents did not accept complete treatment due to treatment side effect . among those patients , \n table 1 shows the baseline characteristics and treatment outcomes of patients who did and did not progress to liver cirrhosis . \n there were 801 person - years of follow - up data , and the average follow - up time was 5.5 years ( sd : 2.5 years , range : 1.112.3 years ) . during the follow - up , 27 patients ( 18.6% ) developed liver cirrhosis , 12 patients developed hcc , three patients developed varices , two patients developed ascites , and one patient developed spontaneous bacterial peritonitis . \n one death that was unrelated to liver disease occurred during the follow - up period . \n the average age of patients with liver cirrhosis was 70.23.3 years , and 17 of these patients ( 62.9% ) were females . among patients who progressed to cirrhosis ( n=27 ) , 14 patients ( 51.9% ) \n univariate analysis of patients who did and did not progress to cirrhosis indicated no significant differences in age , sex , diabetes , smoking , other risks of hepatitis ( alcoholism , hbv coinfection , and fatty liver ) , hcv rna load , hcv genotype , baseline apri , and svr . \n there was a trend of a higher baseline apri score in patients who progressed to liver cirrhosis compared with those who did not progress ( 2.11.2 vs 1.61.3 , p=0.055 ) . \n table 2 shows the result of cox regression analysis with adjustment for age , sex , diabetes , other risks of hepatitis ( alcoholism , hbv coinfection , and fatty liver ) , hcv rna load , hcv genotype , and baseline apri . \n the results show a significant association between progression to liver cirrhosis and no svr ( hr : 3.39 , 95% ci : 1.3128.761 , p=0.012 ) . \n there was also a trend of an association between liver cirrhosis and low hcv rna load ( hr : 0.392 , 95% ci : 0.1521.009 , p=0.052 ) . \n there was no significant association between progression to liver cirrhosis and high baseline apri by cox regression ( adjusted hr : 1.285 , 95% ci : 0.9211.791 , p=0.14 ) . \n the risk of liver cirrhosis was significantly greater in patients without svr ( 3-year cumulative incidence : 35.2% , 95% ci : 13.057.5 , p=0.012 ) than in those with svr ( 3-year cumulative incidence : 10.4% , 95% ci : 3.117.7 ) . \n decrease of progression to liver cirrhosis occurred in one patient for every four chc patients who achieved svr within 3 years . \n patients older than 65 years are often excluded in clinical trials of hcv therapies because they often have significant comorbidities and experience adverse effects during or after treatment . \n therefore , few studies have examined the progression of chc in elderly patients.6,14,15,2628 previous research indicated that the progression of chc to liver cirrhosis was associated with certain baseline host factors and time - dependent risk factors.2,3,29,30 the cumulative nature of fibrosis progression and the potential for more rapid progression in patients older than 40 years mean that cirrhosis is increasingly significant in elderly patients.2,5 in fact , previous research has documented that a longer duration of chc infection and advanced patient age are associated with more rapid progression of chc to liver cirrhosis.2,31 our findings support the rapid progression of liver cirrhosis in patients aged 65 years and older . in particular , the 3-year cumulative risk of liver cirrhosis was 35.2% in our elderly patients who had persistent viremia . \n previous studies examined the efficacy of peg - ifn plus rbv for treatment of chc in elderly patients.4,15,26,28,3235 although such patients have higher rates of dose reduction and adverse effects , some of them achieve svr.4,15,32,35 a recent meta - analysis indicated that the overall rate of svr in elderly patients was significantly lower than in young patients based on intention - to - treat analysis ( 42.0% vs 60.1% , p<0.00001 ) and per - protocol analysis ( 54.4% vs 67.4% , p=0.002).15 the rates of drug discontinuation and rbv dose reduction in elderly patients were significantly higher than in younger patients.15 the poor svr rate in elderly patients may be related to the high prevalence of liver cirrhosis and treatment intolerance.4,14,33 our study demonstrated the efficacy of peg - ifn plus rbv treatment in elderly patients . \n in particular , 65.5% of patients overall achieved svr , and this included 50.6% of those with hcv genotype-1 and 89.3% of those with other hcv genotypes . \n these data support the use of our treatment regimen in elderly patients with chc who do not have liver cirrhosis or hcc . \n furthermore , there were no deaths or severe treatment - related complications , and most patients who could not tolerate treatment stopped treatment near the beginning of the regimen ( mean treatment duration : 84 days ) . \n this suggests that the standard peg - ifn plus rbv treatment is safe in elderly patients if they are closely monitored . from our previous study , the patients who had absence of liver cirrhosis , low baseline hcv rna levels , high baseline alt level , no hcv genotype-1 , or were treated with peg - ifn--2a were good responders to peg - ifn plus rbv treatments.36 thus \n , it seems that treatment will be effective in selected elderly patients with chc , such as those with good response predictors ( genotype 2/3 , low viral load , il28b - cc ) , good performance status , and without major concomitant diseases . \n previous long - term follow - up studies showed the benefits of svr in patients with chc.711 the achievement of an svr through interferon - based therapy slows the progression from chronic hepatitis to liver fibrosis710 and reduces the subsequent development of hcc and liver - related mortality.1113 these effects are universal and were stronger in patients with advanced fibrosis.11 a large - scale study that followed 1,386 patients with chc but not liver cirrhosis over 5 years reported that the annual incidence of liver cirrhosis in untreated patients and patients treated with interferon - based therapy ( n=892 ) was 2.26% and 1.11% ( nonresponders : 1.99% , sustained responders : 0.74% ) , respectively.9 in addition , the 14.5-year cumulative incidence of cirrhosis was significantly lower in sustained responders ( 4.8% ) than in nonresponders ( 21.6% , p=0.0007 ) and in untreated patients ( 36.6% , p<0.0001 ) . \n the present study showed that cirrhosis progressed rapidly and that an svr slowed the progression to fibrosis in elderly patients with chc . \n in particular , patients without svr had a significantly higher risk of liver cirrhosis than those with svr , and a significantly higher 3-year cumulative incidence of liver cirrhosis than those with svr . \n these results suggest that suppression of hcv rna can dramatically reduce the risk of liver cirrhosis in elderly patients . \n a previous study reported that high apri score was associated with significant fibrosis or cirrhosis.22 our patients also had elevated apri scores ( mean sd : 1.71.3 ) , confirming the presence of advanced fibrosis at baseline and consistent with the rapid progression of fibrosis in elderly patients . \n in addition , there was a trend of a higher baseline apri score in patients who progressed to liver cirrhosis than those who did not progress ( 2.11.2 vs 1.61.3 , p=0.055 ) . \n nonetheless , after adjusting for apri score by cox regression , svr remained the most important factor for progression to liver cirrhosis . \n antiviral treatments for chc have evolved since 2011 , and several new drugs , including direct - acting antivirals and host - targeted agents , are marketed or under clinical development . \n interferon - free regimens are expected to cure > 90% of patients with chc,37 and the newly available drugs have fewer side effects than interferon - based regimens . \n however , the new hcv therapies are very expensive and have not been thoroughly evaluated in elderly patients . \n thus , additional research is needed to evaluate the cost - effectiveness of these new therapies in elderly patients . before adoption of the new therapies in elderly patients \n , the peg - ifn plus rbv regimen should continue to be used to prevent the rapid progression to cirrhosis . \n first , this was a retrospective observational study . some information such as age at infection \n , the number of patients who underwent therapy evaluation , the excluded patient number , and the reasons for excluding can not be obtained from this study . \n however , it is difficult to perform a prospective controlled trial in elderly patients , so our findings nonetheless provide important information regarding treatment decisions in this population . \n second , only 108 patients ( 74.5% ) underwent liver biopsy at baseline , and we can not provide paired biopsy data . thus , most patients with cirrhosis were diagnosed using clinical criteria . \n however , our diagnostic criteria for cirrhosis included two documented ultrasonograms of liver cirrhosis with solid clinical end points ( splenomegaly , ascites , hepatic encephalopathy , or varices).9 although ultrasound is inaccurate in detecting the early stages of liver cirrhosis , combination of the morphologic finding can improve the performance of ultrasound to detect cirrhosis , reaching an accuracy of over 80%.38 in conclusion , liver fibrosis progressed rapidly in elderly patients . \n an svr following peg - ifn combination treatment can reduce the risk of liver cirrhosis in elderly chc patients .", "role": "user" }, { "content": "objectivewe studied the effect of sustained virologic response ( svr ) after treatment with pegylated - interferon ( peg - ifn ) plus ribavirin on the development of liver cirrhosis in elderly patients with chronic hepatitis c ( chc).patients and methodsthis retrospective study enrolled 145 elderly chc patients ( aged 65 years ) who were treatment - nave and were treated with peg - ifn plus ribavirin for 6 months between january 2005 and december 2011 . \n abdominal sonography was performed and liver biochemistry was studied at baseline , at the end of treatment , and every 36 months thereafter . \n the development of liver cirrhosis and related complications was evaluated at the follow - ups . \n the aspartate aminotransferase - to - platelet ratio index was used as a noninvasive maker for fibrosis.resultsthe mean patient age was 69.13.3 years , and the average follow - up time was 5.5 years ( standard deviation : 2.5 years , range : 1.112.3 years ) . \n ninety - five patients ( 65.5% ) achieved svr , and 26 ( 17.9% ) discontinued treatment . \n twenty - seven patients ( 18.6% ) developed liver cirrhosis after treatment . \n patients without svr had significantly greater risk of liver cirrhosis than those with svr ( hazard ratio [ hr ] : 3.39 , 95% confidence interval [ ci ] : 1.3128.761 , p=0.012 ) . \n the difference in 3-year cumulative incidence of liver cirrhosis was 24.8% greater for patients without svr ( 35.2% , 95% ci : 13.057.5 , p=0.012 ) compared with those with svr ( 10.4% , 95% ci : 3.117.7 ) . \n there was a trend of a higher baseline aspartate aminotransferase - to - platelet ratio index score in patients who progressed to liver cirrhosis compared with those who did not progress ( 2.11.2 vs 1.61.3 , p=0.055 ) , but the difference failed to reach significance by cox regression ( adjusted hr : 1.285 , 95% ci : 0.9211.791 , p=0.14).conclusionan svr following peg - ifn combination treatment can reduce the risk of liver cirrhosis in elderly chc patients .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: leukotrienes ( lts ) are an important family of eicosanoid lipid \n mediators derived from the metabolism of arachidonic acid ( aa ) and \n associated with asthma and allergic reactions . \n in contrast to prostaglandins ( pgs ) , which are produced from aa by the action of cyclooxygenase \n ( cox ) enzymes , lts are made predominately by inflammatory cells \n like polymorphonuclear leukocytes , activated macrophages \n , and mast cells \n . \n dendritic cells ( dc ) are the \n major players in both innate and adaptive immunity . \n despite their important role as the most professional \n antigen - presenting cells ( apc ) of the immune system , \n dc possess \n the enzymatic machinery to convert aa to proinflammatory lts \n [ 6 , 7 , \n 8 , 9 ] . \n since , all human dc phenotypes examined until now \n constitutively express the 5-lo pathway , the role of dc \n 5-lo - derived lipid mediators in the regulation of the proximal \n steps of the immune responses was recently suggested . \n primary immune response involves phenomena as diverse as antigen \n uptake by dc , their subsequent migration through multiple tissue \n barriers , homing in lymphoid organs , and antigen presentation to \n t cells [ 8 , 9 ] . \n it is known that lts exert their effects through extracellular \n g - protein - coupled receptors . among human tissues studied by \n northern blot analysis , ltb4 receptor messenger rna ( mrna ) \n is strongly expressed in both thymus and lymph nodes \n ( spanbroek r. unpublished data ) , while cyslt1 receptor mrna is \n expressed in spleen . \n in addition , 5-lo - deficient mice \n show altered ovalbumin - dependent cellular and humoral \n immune responses [ 12 , \n 13 ] . these observations provided \n substantial support for an indispensable role of 5-lo pathway in \n the regulation of adaptive immunity . \n since lts display a great \n variety of biological effects , it is not surprising that cellular \n lt biosynthesis must be tightly regulated . \n based in the current \n knowledge of 5-lo pathway regulation , it becomes obvious that \n cellular lt biosynthesis is modulated by multiple mechanisms , \n including gene expression , cytokine effects , enzyme movement , and \n compartmentalization of the 5-lo pathway . \n thus the understanding \n of the precise regulatory mechanism of the 5-lo activity might \n provide new concepts for the development of anti - inflammatory \n drugs . \n lts are lipid messengers that play central role in immune \n responses and tissue homeostasis . \n biosynthesis of lts \n from aa was initially described in polymorphonuclear leukocytes \n and monocytes . \n first described in 1937 as the slow reacting \n substances of anaphylaxis ( srs - a ) , these lipid mediators are now \n known as the cysteinyl lts ( cyslts ) , ltc4 , ltd4 and \n lte4 . \n synthesis of lts can be divided in two \n pathways : one to create cyslts and another to create ltb4 \n ( figure 1 ) . \n 5-lo is \n the key enzyme in lt biosynthesis and is located in the nucleus \n in some cell types and in the cytosol of others . \n 5-lo is \n 72- to 80-kd monomeric soluble protein containing one nonheme \n iron believed necessary for catalysis . \n this \n enzyme possesses an nh2-terminal domain that \n binds to calcium iron and is essential for nuclear membrane \n translocation . \n active mainly in myeloid cells , such as \n monocytes , macrophages , b lymphocytes , granulocytes , mast cells , \n and dc , 5-lo requires ca and is stimulated by atp , \n phosphatidylcholine , lipids , and hydroperoxides . \n following cellular activation , 5-lo translocates to the nuclear \n membrane where it is able to interact with an 18-kd \n membrane - associated protein referred to as \n five - lipoxygenase - activating protein ( flap ) . \n flap is an \n aa - binding protein whose function is to optimally present \n substrate to 5-lo . \n the first evidence that the 18-kd \n protein is absolutely required for cellular 5-lo activity \n comes from transfection studies in human osteosarcoma cell lines \n . in this system , \n expression of 5-lo alone resulted in no \n detectable cellular 5-lo activity following challenge with \n calcium ionophore a-23187 . \n lt synthesis only occurred when 5-lo \n and flap were co - expressed in these cells . \n flap is expressed on t \n cells , macrophages , and dc \n , but not on \n erythrocytes or endothelial cells . \n arachidonic acid is \n liberated from membrane phospholipids by the action of cytosolic \n phospholipase a2 ( cpla2 ) and presented to 5-lo enzyme \n by flap . \n 5-lo enzyme converts arachidonic acid to an unstable \n intermediate termed 5-hydroperoxyeicosatetraenoic acid ( 5-hpete ) . \n subsequently , 5-hpete is dehydrated to yield the epoxide \n lta4 , a pivotal intermediate in the biosynthesis of \n inflammatory and anaphylactic mediators . \n enzymatic hydrolysis of \n lta4 by lta4 hydrolase results in the formation of \n ltb4 . alternatively , ltc4 synthase catalyzes the \n conjunction of lta4 with glutathione to form ltc4 , \n which may be converted to ltd4 and lte4 by the actions \n of the transferase and peptidase enzymes , respectively . \n cellular activation by immune complexes , bacterial peptides , and \n other stimuli elicit a sequence of events that include cytosolic \n phospholipase a2 ( cpla2 ) and 5-lo translocation to the \n nuclear envelope to produce 5-hydroperoxy eicosatetraenoic acid \n subsequently , 5-hpete is dehydrated to yield \n the epoxide lta4 , a pivotal intermediate in the biosynthesis \n of inflammatory and anaphylactic mediators . \n lta4 undergoes \n transformation by one or more of three possible fates depending \n on the cellular context : hydrolysis , conjugation with \n glutathione , or transcellular metabolism to generate bioactive \n lipid mediators . in neutrophils and monocytes , \n lta4 is converted predominately to the chemoattractant \n ltb4 by lta4 hydrolase , but in human \n eosinophils , mast cells , and basophils , lta4 is conjugated \n with reduced glutathione by ltc4 synthase to form the first \n of the cyslts , ltc4 . after carrier - mediated \n cellular \n export , sequential cleavage of the glutathionyl side \n chain of ltc4 generates the extracellular metabolites \n ltd4 and lte4 . \n ltb4 may be degraded by microsomal \n -oxidation and peroxisomal -oxidation in myeloid \n cells and hepatocytes . \n it is important to note that degradative \n enzymes are increased by the transcription factor peroxisome \n proliferator - activated receptor ( ppar ) and this \n nuclear hormone receptor is in turn activated by binding with \n ltb4 . \n the actions of lts require the expression of extracellular \n g - protein - coupled receptors ( gpcr ) which are members of the \n rhodopsin - like receptor superfamily . \n ltb4 is a potent \n chemoattractant that is primarily involved in inflammation , \n immune responses , and host defense against infection . \n ltb4 \n activates inflammatory cells by binding to its cell - surface \n receptor ( designated blt ) via two known receptors , blt1 and blt2 , \n according to the affinity of ltb4 for the receptor . \n blt1 has \n been characterized as a 43-kd gpcr expressed only in inflammatory \n cells , including neutrophils , alveolar macrophages , and \n eosinophils and with a high affinity for only ltb4 \n . \n the second g - protein - coupled receptor for ltb4 , \n blt2 receptor , has only recently been described and is a \n low - affinity receptor expressed more ubiquitously [ 28 , \n 29 ] . \n both receptors are found in lymph node on the same \n chromosome , but intriguingly , the promoter region of the blt1 \n gene is localized within the open reading frame ( orf ) of the blt2 \n gene , which encodes a low - affinity receptor for ltb4 \n . the high - affinity receptor \n blt1 mediates chemotaxis , chemokinesis , aggregation , and \n adherence to endothelium . \n overexpression of ltb4 receptors in leukocytes leads to \n increased neutrophil influx into the lungs after \n ischemia - reperfusion in mice . \n it is also important to \n indicate that ltb4 can bind and activate the intranuclear \n transcription factor ppar , resulting in the activation of \n genes that terminate inflammatory processes . \n the \n receptor for cysteinyl lts , ltc4 , ltd4 , and lte4 \n are named cyslt receptors and are distinct from the ltb4 \n receptors . \n two classes of cyslt receptors have been identified : \n cyslt1 and cyslt2 according to their sensitivity to selective lt \n receptor antagonists . \n cyslt1 has been identified in human smooth \n muscle cells , vascular endothelial cells \n , \n lung macrophages , and peripheral blood leukocytes . \n cyslt2 \n receptor , originally found in pulmonary vein preparations \n by pharmacological assays , is detected in spleen , \n purkinje fibers of the heart , and discreet region of the adrenal \n gland by molecular methods . \n abnormal production of lt contributes to a variety of \n inflammatory diseases , including asthma , glomerulonephritis , \n psoriasis , inflammatory bowel diseases , and acute lung injury \n . \n considering their important role in host defense \n against microbial infections , biosynthesis of lts must \n be tightly regulated . \n two steps seem to be involved in the \n control of lt biosynthesis , that is , liberation of aa and \n regulation of 5-lo activity . \n moreover , the expression \n of 5-lo and flap , which are subject to regulation in monocytic \n cells , have been considered as the major determinant of \n cellular lt biosynthesis . \n 5-lo activity and product \n generation at sites of inflammation might be modulated in \n multiple ways . \n another possibility \n is modulating the expression of enzymatic components \n through transcriptional or posttranscriptional mechanisms . \n recently , it has been reported that cyclic amp ( camp ) elevating \n agents , such as pge2 , inhibited lt biosynthesis and 5-lo \n translocation to the nucleus in cytokine - primed human neutrophils \n . \n the intracellular compartmentalization of the 5-lo \n metabolism affects the integrated output of this biosynthetic \n pathway . \n nuclear import of 5-lo can modulate lt \n production , since nuclear import can strongly enhance \n or suppress 5-lo activity . in considering the question \n of 5-lo translocation and its cellular localization , it is \n necessary to consider that 5-lo can move in or out of the nucleus \n in response to in vivo or in vitro experimental conditions \n . \n cytokines and other components may also affect 5-lo \n pathway , and the expression of 5-lo and flap can be controlled by \n specific molecules . \n for example , it has been reported that the \n th2-derived cytokines , il-4 , and il-13 inhibited the release of \n lt from monocytes by decreasing flap mrna and protein levels , \n whereas , il-1 and ifn ( th1 cytokines ) stimulate 5-lo \n expression . \n 5-lo activity may be modulated by nitric \n oxide ( no ) since prolonged exposure to lipopolysaccharide inhibits \n macrophage 5-lo metabolism via induction of no synthesis \n . in bone marrow - derived dc \n , we have recently \n demonstrated that exogenous or pge2-induced il-10 inhibited \n flap expression without any effect on 5-lo enzyme . \n the \n regulatory mechanisms of the 5-lo pathway appear to be complex \n and depends on the composition of the investigated cell \n populations . \n specific control of 5-lo pathway has long been \n suspected because expression of 5-lo activity is tissue specific \n and narrowly distributed to inflammatory cells . \n it is also \n important to note that there is wide variation in 5-lo activity \n and in 5-lo and flap levels even among different inflammatory \n cell types . \n ltb4 is produced by many cells of the immune \n system , including activated neutrophils stimulated with aa or \n calcium ionophore , macrophages , dc , and t cells \n [ 46 , 47 ] . \n this lipid mediators has diverse effects on the \n function and activities of cells of the immune system \n ( figure 2 ) . \n several observations have suggested that \n ltb4 , in cooperation with monocytes , stimulates \n t - suppressor - cell functions [ 48 , \n 49 ] . \n these effects appeared \n to be exerted through inhibition of t cd4 ( helper / inducer ) \n subsets and enhancement of t cd8 ( suppressor / cytotoxic ) \n subsets . \n the suppression required the presence of \n monocytes and was induced by the cyclooxygenase inhibitors \n . \n it has been suggested that the mechanism of action \n involved induction of monocytes to produce both prostaglandins \n ( metabolites of cyclooxygenase pathways ) and cytokines , such as \n ifn- , il-2 , which in turn lead to the production of il-1 \n [ 52 , 53 ] . \n thus , the effects of ltb4 on t lymphocytes are \n not the result of a direct action on lymphocytes themselves but \n an indirect action via monocytes . on natural killer ( nk ) cells , \n ltb4 significantly augmented nk cell cytotoxicity and \n stimulated the expression of il-2 receptors when added to \n effector target cocultures [ 54 , \n 55 ] . \n ltb4 is also known \n to enhance b cell differentiation and secretion of ige via an \n il-4-dependent mechanism . \n ltb4 causes adhesion and \n chemotactic movement of leukocytes and stimulates aggregation , \n enzyme release , and generation of superoxide in neutrophils . \n leukotrienes c4 , d4 , and e4 , which are released \n from the lung tissue of asthmatic subjects exposed to specific \n allergens , seem to play a pathophysiological role in immediate \n hypersensitivity reactions . \n the expression of 5-lo and flap genes \n have been reported in human b cells by reversed transcription \n chain analysis . \n several aa - derived lipid mediators \n possess immunoregulatory properties in vivo and in vitro . \n the most \n interesting example is pge2 , which clearly affects the \n functions of dc and other cell types of the immune system \n . \n in contrast to pge2 , ltb4 is considered as a \n mainly immunopotentiating mediator , which stimulates the activity \n of nk cells and b lymphocytes , although it also induces \n suppressor function among human mononuclear leukocytes . \n it also stimulates the release of il-1 and \n tnf- in human monocytes , and il-8 from human \n polymorphonuclear leukocytes . \n cells that produce lts or become targets to their actions can \n also produce other proinflammatory molecules , such as pgs , and \n various cytokines . \n for example , the exogenously added or the \n endogenously produced ltb4 can enhance the release of il-6 \n from human monocytes and mice dc \n . \n this may \n constitute an important mean by which monocytes and dc modulate \n inflammatory responses . \n several lines of evidence support an \n expanded role of the 5-lo pathway in the regulation of \n antigen - specific adaptive immunity . \n epidermal langerhans cells \n ( lc ) , a member of the dc family capable of initiating \n antigen - specific immune responses in naive lymphocytes , markedly \n express the 5-lo pathway . \n we have recently demonstrated \n that mice bone marrow - derived dc ( bm - dc ) produce ltb4 and \n other aa metabolites , which have an autocrine actions in \n apc - mediated immunity [ 6 , \n 7 ] . \n other investigators have \n reported the expression of 5-lo in human dc generated from \n cd34 hematopoietic progenitors and in lymphoid organs \n . \n it has been recently reported that bm - dc express \n cyslt1 receptors , 5-lo , flap , and ltc4 synthase , \n and produce cyslts in response to aeroallergen . \n the produced cyslts play a crucial role in regulating dc function \n in a murine model for asthma . \n the ability of dc to migrate to \n lymphoid nodes may be affected by 5-lo products since the \n ltc4 transporter mrp1 may regulate ccl19-dependent \n mobilization of dc from peripheral tissues to lymph nodes \n . \n taken together , these data suggest that by producing \n proinflammatory lts and other components , dc and other apc may \n play important role in inflammatory processes and allergic \n diseases . \n the main function of the immune system is the defence of the host \n against pathogens and tumors . \n two major defence systems are \n involved in host defence against infectious agents and tumors : \n the innate and adaptive immunity , which differ in the means by \n which they recognize antigens . \n however , it is important to \n indicate that innate immune system activates , orientates , and \n regulates the adaptive immunity , which through clonal expansion \n of lymphocytes will respond to pathogen in antigen specific \n manner . \n both innate and adaptive immunity are closely linked , and \n the interactions between the two systems of the host defence are \n regulated by a variety of cells and their mediators in particular \n cytokines and aa - derived lipid mediators , such as lts . \n the development of specific agonists and antagonists for each lt \n receptors will provide important reagent for further defining the \n biological and pathological actions of proinflammatory lts . \n other \n lipoxygenase products will also certainly receive attention in \n the year to come . to delineate molecular mechanisms within immune \n responses that are affected by 5-lo pathway products , \n future \n studies should be directed toward dc / t lymphocytes interactions , \n the identification of lt receptor - expressing cells in lymphoid \n organs , and immune response studies of 5-lo knockout mice . \n finally , further studies of lt receptors expression in different \n types of dc and other apc are needed to truly assess the \n potential roles of 5-lo pathway in dc biology and apc - mediated \n immunity .", "role": "user" }, { "content": "5-lipoxygenase ( 5-lo ) pathway is the major source of potent \n proinflammatory leukotrienes ( lts ) issued from the metabolism of \n arachidonic acid ( aa ) , and best known for their roles in the \n pathogenesis of asthma . \n these lipid mediators are mainly released \n from myeloid cells and may act as physiological autocrine and \n paracrine signalling molecules , and play a central role in \n regulating the interaction between innate and adaptive immunity . \n the biological actions of lts including their immunoregulatory \n and proinflammatory effects are mediated through extracellular \n specific g - protein - coupled receptors . despite their role in \n inflammatory cells , such as neutrophils and macrophages , lts may \n have important effects on dendritic cells ( dc)-mediated adaptive \n immunity . \n several lines of evidence show that dc not only are \n important source of lts , but also become targets of their actions \n by producing other lipid mediators and proinflammatory molecules . \n this review focuses on advances in 5-lo pathway biology , the \n production of lts from dc and their role on various cells of \n immune system and in adaptive immunity .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: endoscopic variceal sclerotherapy has become widely used to treat patients with portal hypertension and esophageal varices , either during acute variceal bleeding or as elective chronic therapy . but \n this report deals with an esophageal intramulal hematoma , an unusual complication of endoscopic variceal sclerotherapy that is not previously described in korea . \n a 35-year - old man was admitted to the hospital because of hematemesis one month prior to admission . \n he had been treated with nph insulin , 30 units daily , due to known diabetes mellitus for the past 2 years . on admission , \n hemoglobin was 11.1 g / dl and alkaline phosphatase was 100 iu / l , but other liver function tests were within normal ranges . \n serum hbs ag was negative and alpha - fetoprotein was below 5 ng / ml . \n bleeding time , prothrombin time , activated partial thromboplastin time and platlet counts were all normal . \n upper gastrointestinal fiberscopy revealed slightly bluish , beaded esophageal varices , located in the lower esophagus , with no sign of redness ( fig . \n after discharge , 2 more sessions of sclerotherapy were done with intravariceal injections of 5% ethanolamine oleate in one week intervals . \n on endoscopy , a huge submucosal bulge in the lumen of the esophagus 25 cm distal to the upper incisors was found with dark bluish mucosal discoloration ( fig . \n barium esophagogram revealed a large smooth filling defect in the lower two thirds of the esophagus with near total occlusion ( fig . \n because of the submucosal nature of the mass , an intramural hematoma was presumed as the cause . \n supportive care with parenteral nutrition was given and liquids could be swallowed with mild discomfort on the seventh day after sclerotherapy . \n two weeks after the third sclerotherapy , repeat endoscopy revealed that the hematoma had completely disappeared and was replaced with a longitudinal mucosal defect ( fig . \n minor complications such as retrosternal discomfort , fever , mild tachycardia , and ulceration of the esophageal wall as well as major complications such as bleeding , esophageal wall necrosis , perforation , empyema , mediastinitis , and late onset stenosis have been described . \n many causes of intramural hematoma have been described as resulting from emetics , after ingestion of a foreign body and endoscopic instrumentation . following remote trauma , after medication , or as spontaneous events in a patient with impaired hemostasis such as in thrombocytopenia , in hemophiliacs and in patients receiving anticoagulation therapy , as well as in patients with normal hemostasis . \n only 4 cases of intramural hematoma of the esophagus after sclerotherapy have been described in the english literature harris et al . reported a case of intramural hematoma after sclerotherapy with sodium tetradecyl sulphate and the patient died of bacteremia . \n van steenbergen et al . described a case after the fourth sclerotherpy session with 1 % polidocanol paravariceally and intravariceally in a 43-year - old woman with hematemesis and she recovered with conservative management . \n korula described a case after sclerotherapy with 1.5% sodium tetradecyl sulphate intravariceally in a 45-year - old man . \n conservative treatment was done and no stricture or mucosal lesion was found 5 months later . \n jones et al . experienced a case after the second injection with 5% ethanolamine oleate intravariceally in a 50-year - old woman with alcoholic cirrhosis . \n coagulation profiles in van steenbergen s case and ours were normal . in two cases and \n ours intravariceal technique was used and in one case both intravariceal and paravariceal techniques were used . \n the intravariceal technique was found to cause less retrosternal pain than the paravariceal technique . even if intravariceal injection is attempted , extravasation of the sclerosant into the surrounding tissue has been noted . \n it seems probale , therfore , that injection in the present case and in the others resulted in intramural extravasation of blood and sclerosant leading to mucosal dissection . \n diagnosis of esophageal hematoma can readily be made radiologically and endoscopically . on an esophagogram the unusual \n finding is an elongated radiolucent filling defect with smooth borders . since posterior hematomas are relatively common , this appearance is often best shown on a lateral view . from a review of 26 patients with esophageal hematoma , shay et al \n . found that in patients with normal hemostasis , the hematoma always presented radiologically as a single lesion involving the distal esophagus and in patients with impaired hemostasis , the hematoma either spared the distal esophagus or occured at multiple sites in 60% of the cases . \n intraluminal bulging of the mucosa with a characteristic dark bluish discoloration indicates the presence of a submucosal hematoma . \n the hematoma usually resolves spontaneously within 1 to 3 weeks , so management of such lesion is preferably conservative , as is evident in our case .", "role": "user" }, { "content": "the authors report a case of esophageal hematoma in a 35-year - old man after esophageal variceal sclerotherapy with intravariceal injection of 5% ethanolamine oleate . a huge submucosal hematoma of the esophageal wall was demonstrated endoscopically and radiologically . \n resolution occurred spontaneously after conservative treatment .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: bacterial meningitis , tetanus and cerebral malaria have been reported as the commonest causes of neuroinfection in developing countries.1,2 mainly resulting from african countries , these data may not only reflect the poorest communities of developing countries , but also the inadequacy of hospital infrastructure and laboratory resources associated with civil war in many of these countries.3 however , differences in economical , political and climatic characteristics , and quality of health assistance , could contribute to distinct epidemiological data . \n the current study was undertaken to determine the frequency and characterize clinical aspects of neuroinfection admitted to our tropical neurological ward . \n this retrospective study was approved by the ethics committee of hospital das clnicas , so paulo university school of medicine , brazil . \n our tertiary teaching hospital has 970 beds , of which 26 belong to the neurology division which also has an infectious diseases ward that receives the majority of infectious cases . from 12 december 1999 to 12 december 2007 , all patients admitted because of neuroinfection had their medical records reviewed . \n the main inclusion criterion was positive infectious pathogen identification , including our previously reported cases.4,5 although we had patients with undetermined infectious agents , we also included patients who had neurological manifestations and cerebrospinal fluid ( csf ) abnormalities that suggested infectious disease . \n age , gender , immunological status , neurological syndrome , infectious agent and clinical outcome were recorded when the inclusion criteria had been fulfilled . \n neurological syndromes were classified as : meningitis ; meningoencephalitis ; stroke ; and , myelitis.6 confirmed by electroneuromyographic studies , peripheral nervous system ( pns ) encompassed spinal roots , plexuses or peripheral nerve affection.7 clinical involvement of cerebellar and/or brain stem structures was recorded as posterior fossa and brain space occupying lesion was applied when abscess or granuloma were seen during imaging studies despite neurological signs . simultaneous or serial inflammation of the meninges , brain , medulla and spinal roots were classified as overlap involvement . \n computerized tomography ( ct ) , magnetic resonance imaging ( mri ) and extensive blood and csf analyses were carried out in all patients . \n the infectious agent was identified from csf ( gram staining , culture , antibody detection or protein chain reaction ) or nervous tissue samples ( histology and/or culture ) . \n the outcome was classified as death , and unaltered or improved based on neurological examination at hospital discharge . \n cerebrovascular diseases and pulse therapy for multiple sclerosis and non - infectious inflammatory neuropathies were the main reasons for admission \n . initial diagnosis of neuroinfection occurred in 416 patients , but 42 were excluded after laboratorial investigation : systemic lupus erythematosus ( 3 ) , chronic idiopathic meningitis with low glucose level ( 1 ) , vogt - koyanagi - harada disease ( 2 ) , behet disease ( 17 ) , sarcoidosis ( 5 ) , meningeal carcinomatosis ( 2 ) , undetermined eosynophilic meningitis ( 2 ) and rasmussen 's encephalitis ( 10 ) . \n the inclusion criteria were fulfilled by 374 patients ( 4.2% , 374/8760 ) , mean age was 40.513.4 years ( range : 1388 years ) , and 63.8% were male . \n sixty - nine ( 18.4% , 69/374 ) patients were immunocompromised as a result of solid organ transplantation ( 7 ) , acquired immunodeficiency syndrome ( aids ; 42 ) , diabetes mellitus ( 7 ) , chronic hepatopathy ( 3 ) , systemic lupus erythematosus ( 4 ) , aplastic anemia ( 2 ) , neoplasia ( 2 ) and t - cell dysfunction ( 2 ) . \n the most common clinical presentation was meningoencephalitis ( 211/374 ) in immunocompromised and non - mmunocompromised patients ( table 1 ) . \n the infectious pathogen was identified in 81% ( 303/374 ) of cases , and presumed viral and bacterial neuroinfections were assumed in 8% ( 30/374 ) and 10.9% ( 41/374 ) of patients , respectively . \n human immunodeficiency virus ( hiv ) was the most commonly identified virus with self - limited aseptic meningoencephalitis during the seroconversion period , followed closely by herpes simplex virus 1 ( hsv1 ) . \n three non - immunocompromised - patients with hsv1 infection died as a result of hemorrhagic form despite serial treatment with acyclovir , intravenous methylprednisolone , pulse therapy and plasmapheresis . \n jc virus ( jcv ) infection occurred in association with immunodeficiency ( one patient had ovarian cancer and two had aids ) . \n also , 29.4% ( 110/374 ) of admissions were as a result of bacterial infection ( table 3 ) . \n mycobacterium tuberculosis was the main pathogen identified by positive polymerase chain reaction ( pcr ) in 82.6% ( 19/23 ) of patients and histological study in the remainder . \n although intravenous penicillin g ( 24 million / day ) had been administered for 21 days , no response was observed in two patients with progressive general paresis and only slight improvement occurred in patients with meningovascular syphilis . \n polyradiculitis was diagnosed in two patients with tuberculosis and peripheral neuropathy appeared with mycobacterium leprae ( three ) , and bartonella henselae ( one ) infection . \n parasites comprised 28% ( 105/374 ) of neurology ward ( nw ) admissions , of which taenia solium and schistosoma mansoni were the most common . \n accounting for 64.8% of parasitic infection , all cases of t. solium presented with meningoencephalitis in 66 non - immunocompromised and two immunocompromised patients . \n status epilepticus owing to the degenerating intraparechymatous larval form of t. solium was the main clinical presentation ( n = 55 ) . \n moreover , intraventricular vesicles caused ventriculitis in three non - immunocompromised patients , two of whom underwent endoscopic resection and one died . \n in addition , two other non - immunocompromised patients developed hydrocephalus owing to racemous cysts in the spinal cord with clinical improvement after ventricular drainage . s. mansoni was identified in 21 ( 20% ) non - immunocompromised patients representing the most important cause of myelitis ( n = 20 ) in our series ; however , one presented with stroke . \n antiparasitic drugs were only used in neurocysticercosis and schistosomiasis when parasitic eggs had been found in stools or a rectal biopsy . \n schistosoma japonicum caused brain space occupying lesion in one non - immunocompromised patient who had just returned from japan . triggering meningoencephalitis in immunocompromised - patients , \n toxoplasma gondii , trypanosoma cruzi and strongyloides stercoralis were isolated in 11 , 2 , and 2 patients , respectively . accounting for 9.6% ( 36/374 ) of cases \n , fungi caused more severe disease , longer hospital stay ( data not shown ) and higher mortality rate than other infectious agents . \n brain space occupying lesion owing to penicillium marffenei , cryptococcus neoformans and paracoccidioides braziliensis occurred in one immunocompromised , two non - immunocompromised and three non - immunocompromised patients , respectively . presenting with meningoencephalitis , \n the identified agents were c. neoformans ( four non - immunocompromised , five immunocompromised patients ) , histoplasmosis capsulatum ( five non - immunocompromised patient , one immunocompromised patient ) , zygomicetes ( five immunocompromised - patients ) , pseudallescheria boydii ( two non - immunocompromised patients , one immunocompromised patient ) , and aspergillus fumigatus ( two immunocompromised patients ) . \n in addition , cladophialophora batiana , phialophora fonsecae and chromomycetes accounted with one non - immunocompromised patient each and candida sp . and rhodotorula sp . with one immunocompromised - patient each . particularly , zygomicosis occurred in five diabetic patients with disseminated cerebral presentation . \n progressive dementia , myoclonus and periodic sharp wave complexes on electroencephalogram ( eeg ) were present in all patients with prion disease ( n = 13 ) . moreover , clinical and electrophysiological features of inferior motor neuron disorder were seen in three patients , and 14.3.3 protein was present in four patients in whom pcr analysis was carried out . considering the clinical outcome , 14.2% ( 53/374 ) of patients died , of whom 62.3% ( 33/53 ) were non - immunocompromised patients ( table 4 ) . despite having neurological disability at hospital discharge , 82% ( 307/374 ) of patients \n all patients with presumed viral infection ( n = 30 ) and 34% ( 14/41 ) of patients with undetermined bacterial meningitis were asymptomatic at hospital discharge . \n in our study , neuroinfection predominated in young adults and males . considering similar results published by chapp - jumbo ( mean age 34.1 years , 62.9% males ) in africa and tan et al . \n ( mean age 42.5 years , 56% males ) in north - america,8,9 these data did not seem to be influenced by geographical area or quality of health assistance and could reflect high environmental exposition ( recreational and working activities ) of young persons , particularly males . in comparison to developed countries ' data , \n our admission and agent identification rates were closer to the data series published by tan et al ( 3.8% and 68.9% , respectively ) than african teaching infectious disease ward , where admission rates ranged from 11% to 23% and the identification rate was 45%.8,9,10 in addition , the frequency of undetermined neuroinfection was less than 31% in tan et al.9 as reported by these authors , undetermined neuroinfection was frequently diagnosed in non - immunocompromised patients in our series . \n indeed , our neuroinfection mortality was closer to rates in north america that ranged from 1.7% to 12% than the 47% reported by chapp - jumbo.8,9,11 analogously to north - american nw results,9 prion and fungi were the more important lethal agents in our patients . \n however , a high mortality of fungal infection was a common finding , not only in north - american , but also in african studies.8,10,12 in our study , pathogen identification occurred in 81% of patients , being higher than results reported in developed countries.9,11 it could possibly be an overestimation as a result of our inclusion criterion requiring positive agent identification , which could also explain 100% identification in parasitic and fungal infections . \n another contributing factor was virus recognition in more than 72% of meningoencephalitis cases . according to the literature , \n virus identification ranged from 15.3% to 81.5% with the highest values being observed in industrialized countries.13 particularly in suspected viral neuroinfection , we follow the international consensus that recommends repetition of virological tests in serum and cfs.14 ordinary agents causing meningococcal , pneumococcal , streptococcal , staphylococcal and hemophilus meningitis were not observed in our series possibly as a result of referral bias . \n for example , common causes of meningitis could be treated by the internal medicine or infectious diseases services of our hospital whose patients were not included in our study or could be referred to hospital emlio ribas , which is an infectious diseases center located in same geographical area as our institution . \n some infectious diseases are commonly associated with under - development . considering that more than 90% of tuberculosis cases in the world are found in developing countries,15 it is not surprising that neurotuberculosis was the most frequent bacterial infection in our series . \n moreover , neglected tropical diseases have high morbidity with neuropaludism and schistosomiasis being the first and second identified causes in africa , respectively.10,16 similar to these reports , our series showed a great number of parasitic diseases in which cysticercosis and schistosomiasis were the main causes with low mortality and high morbidity . \n cysticercosis has been considered as a biological marker of social and economic development , an identified cause of epilepsy in 26.3% to 53.8% of patients , and is endemic in most of developing countries.17,18 in addition , more than 200 million people have schistosomiasis in tropical countries and 600 million people live in its transmission zones , where the snail habitat has increased as a result of the implementation of agriculture and fresh water exposition combined with lack of sanitary systems.19 brazil is considered the country most affected by schistosomiasis in the americas and takes part in one of the american developing vaccine programs.20,21 schistosomiasis has been associated with liver fibrosis , portal hypertension and hepatosplenomegaly as the more severe clinical impairments , but our study showed myelitis as another infection - associated disability . according to wadhwa et al,22 c. neoformans was the most common cause of fungal meningitis in our series . \n however , some reports showed other agents , such as candida albicans in an african study or aspergillosis and zygomicosis in an indian pathological report , in which cryptococcosis accounted for 2% of histological samples.12,23 histoplasma capsulatum was the second more frequent fungus in our cases , possibly because of its high prevalence in brazil.24 also , regional variability in the fungal aetiological spectrum would possibly be expected in countries that have continental dimensions with distinct ecological settings . \n this retrospective study had methodological limitations related to scarce epidemiological reports from tropical countries . taking into account the characteristics of our institution \n , our results did not represent a tropical neuroinfection profile but could be considered as a panel commonly found in other tropical tertiary nw and certainly different from infectious disease ward admissions at same institution . although comparison with literature data could be a matter of debate owing to differences in health facilities , period and design of studies , and type of population or ward , it could be used as a tool to control the quality of health assistance .", "role": "user" }, { "content": "objectives : this study was undertaken to characterize the neuroinfection profile in a tertiary neurological ward.introduction:neuroinfection is a worldwide concern and bacterial meningitis , tetanus and cerebral malaria have been reported as the commonest causes in developing countries.methods:from 1999 to 2007 , all patients admitted to the neurology ward of hospital das clnicas , so paulo university school of medicine because of neuroinfection had their medical records reviewed . \n age , gender , immunological status , neurological syndrome at presentation , infectious agent and clinical outcome were recorded.results:three hundred and seventy four cases of neuroinfectious diseases accounted for 4.2% of ward admissions and the identification of infectious agent was successful in 81% of cases . \n mean age was 40.513.4 years , 63.8% were male , 19.7% were immunocompromised patients and meningoencephalitis was the most common clinical presentation despite infectious agent . \n viruses and bacteria were equally responsible for 29.4% of neuroinfectious diseases ; parasitic , fungal and prion infections accounted for 28% , 9.6% and 3.5% respectively . \n human immunodeficiency virus ( hiv ) , herpes simplex virus 1 ( hsv1 ) , mycobacterium tuberculosis , treponema pallidum , taenia solium , schistosoma mansoni , cryptococcus neoformans and histoplasma capsulatum were the more common infectious pathogens in the patients . \n infection mortality rate was 14.2% , of which 62.3% occurred in immunocompetent patients.conclusion:our institution appeared to share some results with developed and developing countries . \n comparison with literature may be considered as quality control to health assistance .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: disbursement in antiretroviral treatment ( art ) drugs within the context of a public health shortage of resources has increased the interest in pharmacoeconomical analyses of the available regimens for both treatment - nave and -experienced individuals.1,2 in western europe , the classic concepts of efficacy , effectiveness , and efficiency supported by the european medicines agency s actions at the time of commercialization must be completed with equity of care , the efficiency of drug packages , and their potential impact on public health budgets.3 in spain , there is universal access to art , and drugs are delivered by law only through hospital - based pharmacies . not infrequently , \n patients return opened packages to the pharmacy due to unexpected toxicity , treatment changes , pharmacokinetic interactions , pregnancy , or incomplete adherence.4 the efficiency of these packages , which vary significantly in their quality standards , has an influence on their potential for reuse , and the impact on the cost of treatment for the health system may eventually be considerable.5 socially efficient packaging in ambulatory care hospital - dispensed drugs should be the one that is best adapted to unit - dose dispensation . \n in addition , it should facilitate patient compliance , allowing for an easy and hygienic review of the ingested and remaining doses with no need to manipulate either the packaging or the pharmaceutical forms , particularly those that are unused . \n moreover , in directly observed art , drugs that are properly packed permit better control , allowing the delivery of just the needed doses in every dispensation ( ie , adjusted to weekly methadone delivery in drug users ) . \n we established antiretroviral package categories based on the excellence of drug packaging and analyzed the number of pills and the cost of drugs returned during a period of 1 year in a hospital - based hiv unit . \n we defined a classification system for the so - called social efficiency of antiretroviral drug packages that could also be applicable to any other family of high - cost drugs used for outpatients or inpatients ( ie , hepatitis c direct antivirals ) . the classes were as follows : \n class a : drug packed in unit doses with complete information ( name of the drug , dosage in mg , lot number , and expiry date ) in each unit , thus maintaining complete information if returned when the external package is opened . \n no unit doses are opened in the returned packages ( figure 1).class b : drug packed in blisters with complete information only in the blister , but not in every unit dose , without special conservation conditions . \n should be initially repackaged in unit doses in the pharmacy before dispensation to ensure class a excellence , with complete tracking information in every unit.class c : drug packed in plastic containers with complete information written only on a label over the container , with no special conservation conditions . \n it would allow to supply a repackaged drug to the patient.class d : drug packed in plastic containers with any manufacturer s warning that the product can not be placed outside of the original package due to special conditions of conservation ( fridge , humidity ) that will not allow either initial unit dose repackaging or reusing an opened container . \n class a : drug packed in unit doses with complete information ( name of the drug , dosage in mg , lot number , and expiry date ) in each unit , thus maintaining complete information if returned when the external package is opened . \n class b : drug packed in blisters with complete information only in the blister , but not in every unit dose , without special conservation conditions . should be initially repackaged in unit doses in the pharmacy before dispensation to ensure class a excellence , with complete tracking information in every unit . \n class c : drug packed in plastic containers with complete information written only on a label over the container , with no special conservation conditions . \n class d : drug packed in plastic containers with any manufacturer s warning that the product can not be placed outside of the original package due to special conditions of conservation ( fridge , humidity ) that will not allow either initial unit dose repackaging or reusing an opened container . \n class b and class c packing increase pharmacy department budgets in terms of human resources , time , and repacking materials . in class d packing , \n all of the contents will be lost once the container is opened , regardless of being returned . in order to deliver only the necessary amount of pills until the next visit , or to deliver a predefined schedule of medication via each pharmacy department , class \n a packages would exhibit optimal social efficiency , whereas class d would demonstrate the worst . \n depending on the social efficiency of the drug that patients withdraw , the returned old one would immediately become a wasted drug . \n the change of a class b or class c drug into a class a drug requires internal pharmacy technician work . \n the current cost of this change is approximately 0.034 per pill , according to conditioning material suppliers and human resources obtained through our hospital price list . \n an experienced technician repackages approximately 214 pills per hour ( 1,500 pills / day ) . \n the hospital - based pharmacy in our hiv unit served 2,413 treated individuals during the year of study . \n patients generated 23,574 visits to the pharmacy during this period , and they experienced a total of 1,051 treatment changes for any reason . \n the pharmacy department delivered 48,325 antiretroviral drug packages ( 2,529,137 pills ) . a total amount of 122,945 was returned in opened antiretroviral packages . \n of these packages , 6,090 pills ( 34% of all returned antiretrovirals ) with a cost of 47,139.91 would be totally lost , mainly due to being packed up in class c and class d boxes . this would be the equivalent to treating 78 individuals with a standard coformulation of rilpivirine / tenofovir / emtricitabine during 1 month . \n we have classified all the current art specialties depending on their social efficiency as supplied in spain , the uk , and the usa ( table 1 ) . \n us packaging shows a greater deal of class c and class d bottles when compared to europe . class \n a and class b packages returned in bad condition represented only 1.1% of the cost and 75,805 came from returned packages in good condition that could potentially be reused . \n during a period of 1 year , the percentage of antiretroviral drugs returned to the hospital pharmacy was low . \n however , the cost of these drugs is high , and therefore there is a significant economic budget loss through unused antiretroviral medicines returned to the hospital pharmacy that can not be reused due to being supplied in low - excellence packages ( approximately one - third of all returned packages ) . for hospital inpatients , \n the excellence in use of art requires class a packed drugs that allow for administration in unit doses , thus minimizing time hours consumed by nurses in drug administration , reducing medication errors , and improving drug stocks in hospital wards . \n the european commission has singled out specific countries to make particular recommendations to increase the cost effectiveness of the health care sector , and in the case of spain , there is particular interest in further rationalizing pharmaceutical spending , including ambulatory drugs dispensed in hospitals.6 further to this point , the european association of hospital pharmacists has encouraged european authorities to identify the drug with all the items we define as class a efficiency , and also to include a barcode to allow unit dose administration in hospitals . \n barcode technology substantially reduces the rate of errors in order transcription and medication administration , and it reduces potential adverse drug events and interactions.7 establishing strategies to reduce the waste of unused medicines is an unmet need.4,5,8 unopened unit dose ( class a ) drugs that are returned inside the hospitals can be reused . \n however , in ambulatory care , this reusing is usually not allowed due to the impossibility of granting the quality of drugs once delivered to outpatients . \n unfortunately , newer antiretrovirals mainly , the new three drug combos , eviplera / complera , stribild , and triumeq , are commonly packaged in class c and class d categories , thus promoting the maintenance of these hidden costs in the near future . \n any improvement in the excellence of packaging by the manufacturer , and favoring the choice of drugs supplied through more efficient packages ( whenever efficacy , toxicity , and convenience are similar ) , should minimize treatment expenditures paid by national health budgets . \n in summary , while the overall percentage of unused anti - retroviral medicines returned to the hospital pharmacy is low , their cost is quite high . \n therefore , the hidden costs of this low - efficiency drug packaging in wasted drugs in these kinds of medications are substantial and should be properly addressed . drug regulatory authorities should consider including the concept of social efficiency of the packaging in the authorization process of high - cost specialty medicines delivered through hospital dispensing to ambulatory patients , as many of these drugs are actually manufactured in europe .", "role": "user" }, { "content": "while the overall percentage of unused antiretroviral medicines returned to the hospital pharmacy is low , their cost is quite high . \n adverse events , treatment failure , pharmacokinetic interactions , pregnancy , or treatment simplification are common reasons for unplanned treatment changes . \n socially inefficient antiretroviral packages prevent the reuse of drugs returned to the hospital pharmacy . \n we defined antiretroviral package categories based on the excellence of drug packaging and analyzed the number of pills and costs of drugs returned during a period of 1 year in a hospital - based hiv unit attending to 2,413 treated individuals . \n a total of 6,090 pills ( 34% of all returned antiretrovirals ) with a cost of 47,139.91 would be totally lost , mainly due to being packed up in the lowest efficiency packages . \n newer treatments are packaged in low - excellence categories of packages , thus favoring the maintenance of these hidden costs in the near future . \n therefore , costs of this low - efficiency drug packaging , where medication packages are started but not completed , in high - cost medications are substantial and should be properly addressed . \n any improvement in the packaging by the manufacturer , and favoring the choice of drugs supplied through efficient packages ( when efficacy , toxicity , and convenience are similar ) , should minimize the treatment expenditures paid by national health budgets .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: colonoscopy has been used for colorectal cancer screening worldwide . however , the miss rate for advanced colorectal adenoma and cancer is known to be 2% to 6%.1 the locations of lesions , the technique of the endoscopist , and normal colorectal lesion image intensity are associated with the miss rate of colorectal adenoma and cancer . \n image - enhanced endoscopy ( iee ) highlights lesions , which can improve the detection rate and diagnostic accuracy for colon polyps . \n iee methods can be classified into dye - based ( chromoendoscopy ) , and equipment - based using optical techniques.2 using a visual filter and magnification , equipment - based iee is used to observe the mucosal epithelium of the microstructure and the capillaries of the lesion . \n in addition , lesions can be evaluated by iee though the measurement of fluorescence intensity emitted in tissues . in this section , the role of equipment - based iee in the detection and estimation of histological findings for colorectal polyps will be discussed . \n narrow band imaging ( nbi ) separates white light into red , green , and blue using a special optical filter , and reconstructs the image while enhancing blue light , which has a relatively short wavelength.3,4 blue light penetrates the mucosal surface , and is reflected at the mucosal surface , highlighting the superficial vasculature . \n since mucosal hemoglobin selectively absorbs blue light , and the mucosa surrounding blood vessels reflects it , the contrast of the image is increased and the mucosal micro - vessel architecture can be estimated in fine detail . in the colon and rectum , microvessels form a ring , and each ring surrounds its respective gland . \n neoplastic changes result in a change in the form , density , and size of microvessels , and colorectal lesions can therefore be diagnosed.5 since hirata et al.6 reported that chromoendoscopy is similar to nbi in colorectal adenoma detection , six large - scale studies have been reported . \n three of them demonstrated that nbi in fact has a higher adenoma detection rate.7,8,9 the adenoma detection rate of nbi is 27% to 40% . \n however , in a meta - analysis , the accuracy , sensitivity , and specificity of nbi were not significantly different from those of chromoendoscopy.10 according to the surface color , capillary shape , and surface structure of the colorectal mucosa , several nbi classifications have been proposed.11,12 based on the classification type , nbi could be helpful in the differentiation between colorectal adenomas and hyperplastic polyps . \n recently , to create an easy - to - use nbi classification system , professionals around the world have developed the nbi international colorectal endoscopic ( nice ) classification , which is based on surface color , the structure of blood vessels , and surface structure ( table 1).13 it has been reported that nice is useful in the pathologic diagnosis of lesions.13,14 \n fig . \n a meta - analysis of several studies concerning the differentiation between colorectal adenomas and hyperplastic polyps showed that the accuracy of predictions of lesion pathology were 62% to 93% for nbi , 65% to 82% for while light endoscopy ( wle ) , and 69% to 96% for chromoendoscopy.10,15 the accuracy of pathological prediction by nbi was higher than that of wle , and similar to that of chromoendoscopy . in terms of the prediction of the invasion of malignant tumors with nbi \n , it has been reported that the density and regularity of the vascular structure are useful.11,16,17,18 wada et al.18 have reported that the vascular structures of submucosal invasive carcinoma are irregular , and that they can not be classified by the type of blood vessel . depending on vascular structural findings , \n the sensitivity and specificity of the diagnosis of submucosal invasive cancer were 100% and 95.8% , respectively . \n since hirata et al.6 reported that chromoendoscopy is similar to nbi in colorectal adenoma detection , six large - scale studies have been reported . \n three of them demonstrated that nbi in fact has a higher adenoma detection rate.7,8,9 the adenoma detection rate of nbi is 27% to 40% . \n however , in a meta - analysis , the accuracy , sensitivity , and specificity of nbi were not significantly different from those of chromoendoscopy.10 \n according to the surface color , capillary shape , and surface structure of the colorectal mucosa , several nbi classifications have been proposed.11,12 based on the classification type , nbi could be helpful in the differentiation between colorectal adenomas and hyperplastic polyps . \n recently , to create an easy - to - use nbi classification system , professionals around the world have developed the nbi international colorectal endoscopic ( nice ) classification , which is based on surface color , the structure of blood vessels , and surface structure ( table 1).13 it has been reported that nice is useful in the pathologic diagnosis of lesions.13,14 \n fig . 1 shows two cases classified by nice , representing a benign neoplasm ( fig . \n a meta - analysis of several studies concerning the differentiation between colorectal adenomas and hyperplastic polyps showed that the accuracy of predictions of lesion pathology were 62% to 93% for nbi , 65% to 82% for while light endoscopy ( wle ) , and 69% to 96% for chromoendoscopy.10,15 the accuracy of pathological prediction by nbi was higher than that of wle , and similar to that of chromoendoscopy . \n in terms of the prediction of the invasion of malignant tumors with nbi , it has been reported that the density and regularity of the vascular structure are useful.11,16,17,18 wada et al.18 have reported that the vascular structures of submucosal invasive carcinoma are irregular , and that they can not be classified by the type of blood vessel . \n depending on vascular structural findings , the sensitivity and specificity of the diagnosis of submucosal invasive cancer were 100% and 95.8% , respectively . \n fujinon intelligent color enhancement ( fice ) emphasizes subtle changes of the mucosal surface through the reconstruction of a certain wavelength ( computed spectral estimation technology ) , without using an optical filter such in nbi.19 fice utilizes a simple switching operation to create the various wavelengths , and can choose the most appropriate wavelength to produce the image of the digestive tract . \n for the differentiation of colorectal adenomas from hyperplasic polyps , pohl et al.20 reported that the sensitivity , specificity , and accuracy of fice were 90% to 97% , 74% to 80% , and 83% to 90% , respectively , and were better than those of wle . \n however , fice was not significantly different from chromoendoscopy in the differentiation of adenomas and nonneoplastic polyps.21 \n i - scan utilizes the same spectral estimation technology as fice technology , and applies a digital color filter to images to emphasize lesions . in comparisons of i - \n scan with nbi and high - resolution colonoscopy , i - scan was similar in the differential diagnosis of colon polyps and the prediction of pathological results . \n this revealed that i - scan is also helpful in the differential diagnosis and prediction of the pathology of colorectal lesions.22,23 \n equipment - based iee methods such as nbi , fice , and i - scan , are useful for the detection and differential diagnosis of colorectal tumors . despite the adenoma detection rate of nbi being similar to that of chromoendoscopy \n , nbi can be used to differentiate colorectal polyps and to predict the submucosal invasion of malignant tumors . despite a lack of studies \n , it is known that fice and i - scan are both similar to nbi in the detection rate of colorectal lesions . to successfully use equipment - based iee \n , endoscopists must first be aware of the features of the devices involved , and with the device 's uses , applications , and limitations . in the future , diagnostic accuracy \n could be improved , and new treatment paradigms developed through more effective and advanced endoscopic equipment .", "role": "user" }, { "content": "the use of colonoscopy for the screening and surveillance of colorectal cancer has increased . however , the miss rate of advanced colorectal neoplasm is known to be 2% to 6% , which could be affected by the image intensity of colorectal lesions . \n image - enhanced endoscopy ( iee ) is capable of highlighting lesions , which can improve the colorectal adenoma detection rate and diagnostic accuracy . \n equipment - based iee methods , such as narrow band imaging ( nbi ) , fujinon intelligent color enhancement ( fice ) , and i - scan , are used to observe the mucosal epithelium of the microstructure and capillaries of the lesion , and are helpful in the detection and differential diagnosis of colorectal tumors . \n although nbi is similar to chromoendoscopy in terms of adenoma detection rates , nbi can be used to differentiate colorectal polyps and to predict the submucosal invasion of malignant tumors . \n it is also known that fice and i - scan are similar to nbi in their detection rates of colorectal lesions . through \n more effective and advanced endoscopic equipment , diagnostic accuracy could be improved and new treatment paradigms developed .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: to report a case of recurrent nontuberculous mycobacterial endophthalmitis in the context of neurotrophic keratopathy secondary to herpes zoster ophthalmicus that had an atypical presentation and complex course , and highlights the challenges of causative organism identification and therapeutic interventions in this condition . \n \n a 68-year - old pseudophakic male with long - standing neurotrophic keratopathy and perforated descemetocele managed with cyanoacrylate glue and a contact bandage lens in the left eye , began experiencing recurrent episodes of endophthalmitis after undergoing a penetrating keratoplasty . \n several therapeutic procedures including an anterior chamber washout , two pars plana vitrectomies , explantation of the posterior chamber intraocular lens and capsular bag , and multiple intravitreal antimicrobial injections , were performed to which he has ultimately responded favorably , with no signs of infection to date and stable visual acuity . \n the causative organism of his recurrent infections was initially identified as mycobacterium abscessus through biochemical testing and 16s ribosomal ribonucleic acid gene sequencing ; however , repeat polymerase chain reaction ( pcr ) and sequencing of the 65 kda heat shock protein ( hsp65 ) gene for experimental purposes confirmed the accurate identification of the organism to be mycobacterium chelonae . given the greater reliability of pcr and sequencing of the hsp65 gene over traditional biochemical tests and culture techniques , m. chelonae was likely the infectious agent all along , and the organism was originally misidentified on the basis of less accurate tests . \n standard biochemical testing can identify the causative organism but is limited by the inability to distinguish between nontuberculous species reliably . \n we recommend the use of pcr in conjunction with sequencing of the hsp65 gene for reliable differentiation of m. chelonae and m. abscessus in atypical mycobacterial ocular infections . \n minimum inhibitory concentration antibiotic susceptibility tests on cultured strains are the best guide to antibiotic selection , given the rapidly rising resistance to antimicrobials in atypical mycobacterial species . \n rapidly - growing nontuberculous mycobacteria ( rgntm ) include bacteria such as mycobacterium chelonae , mycobacterium abscessus , and mycobacterium fortuitum.1,2 these atypical mycobacteria can be acquired from the natural environment and nosocomially . \n the mycobacteria can infect the eye , resulting in keratitis , conjunctivitis , and endophthalmitis among other ocular infections.1 risk factors for ocular rgntm infections include trauma , previous corneal infection or surgery , corticosteroid use , and systemic immunosuppression.3 we report a case of recurrent atypical mycobacterial endophthalmitis in the context of neurotrophic keratopathy secondary to herpes zoster ophthalmicus ( hzo ) that had an atypical presentation and complex course , and highlight the challenges of causative organism identification and therapeutic interventions in this condition . \n a 68-year - old pseudophakic male with long - standing neurotrophic keratopathy secondary to hzo characterized by a non - healing corneal ulcer ( figure 1a ) , was referred with a recurrent seidel - positive cornea of the left eye . \n medical history was notable for chronic lymphocytic leukemia that was in remission after chemotherapy treatment ( oral chlorambucil and combination fludarabine and cyclophosphamide ) . \n prior management of the neurotrophic cornea included topical lubricating ; antibiotic and steroid drops ; oral antivirals ; injections of botulinum toxin for induction of ptosis ; and amniotic membrane grafts ( amg ) . \n a corneal descemetocele had perforated 6 months prior and was managed with cyanoacrylate glue and a bandage contact lens . \n examination revealed cyanoacrylate glue on the central cornea with slow aqueous leakage , diffuse deep vascularization , an inferior stromal infiltrate , and a 1.5 mm layered hypopyon ( figure 1b ) . \n the glue was removed and re - applied after obtaining corneal scrapings , and topical vancomycin and tobramycin were administered empirically . \n initial aerobic cultures were positive for micrococcus species , and acid - fast bacilli ( afb ) stain was 3 + positive . \n a therapeutic penetrating keratoplasty ( pk ) with anterior chamber ( ac ) washout , amg , subconjunctival bevacizumab injection , and permanent tarsorrhaphy were performed . \n amg was used for its anti - inflammatory , anti - scarring , and wound healing properties on the ocular surface,4 and subconjunctival bevacizumab was used to help decrease corneal neovascularization.5 initially , the patient recovered well but returned 20 days post - pk with painless 3 + white blood cells in the ac . \n b - scan ultrasonography showed dense retrolental opacity , concerning for endophthalmitis ( figure 2a ) . \n vitreous tap and intravitreal injection of ceftazidime ( 2 mg/0.1 ml ) and vancomycin ( 1 mg/0.1 ml ) were performed with rapid improvement . \n histopathology from the host corneal button subsequently demonstrated afb , and cultures were reported as m. abscessus ( figure 2b ) . \n there was no corneal infiltrate , but 4 + white blood cells in the ac , recurrent layered hypopyon ( figure 1c ) , mild anterior vitritis , and unaffected posterior vitreous were observed . a second vitreous tap and intravitreal injections of ceftazidime ( 2 mg/0.1 ml ) and vancomycin ( 1 mg/0.1 ml ) \n were administered based on prior responsiveness ; however , this time , the patient did not improve . \n the tap revealed no organisms or cells , and cultures ( aerobic , anaerobic , afb , and fungal ) and viral polymerase chain reaction ( pcr ) ( to test for herpes simplex virus 1 and 2 , and varicella zoster virus ) were negative . \n anterior and intermediate uveitis work - ups were performed ( to test for tuberculosis , sarcoidosis , human leukocyte antigen b27 , lyme disease , and syphilis ) , and were negative . based on the recent negative \n work - up and failure to respond to therapy , as well as prior culture results , vitreous and ac biopsies , pars plana vitrectomy ( ppv ) , ac washout , and injection of intravitreal antimicrobial and fungal agents ( amikacin , 400 g/0.1 ml ; vancomycin , 1 mg/0.1 ml ; and amphotericin b , 5 g/0.1 ml ) , were performed . \n antibiotic susceptibility testing was performed prior to this procedure on m. abscessus isolates grown from the prior host corneal button cultures , and showed high sensitivity to amikacin . \n internal stains and cultures of the vitreous sample showed 2 + afb positive staining , and culture results were reported as m. abscessus complex . \n vitreous samples were also sent out to a validated laboratory ( the university of washington s , clinical microbiology laboratory ; seattle , wa , usa ) for pcr and gene sequencing . \n pcr using primers targeting the 65 kda heat shock protein ( hsp65 ) gene and the beta subunit of ribonucleic acid polymerase ( rpob ) gene was negative . however , sequencing of the 16s ribosomal ribonucleic acid ( rrna ) gene was positive for m. abscessus / chelonae complex . based upon the biochemical testing and gene sequencing results , the presumed cause of infection was m. abscessus . after surgery , the eye again returned to a quiet state without evidence of infection or inflammation ( figure 1d ) , until the patient returned 3 months post - vitrectomy with painless clouding of vision in the left eye . \n examination revealed presence of a layered hypopyon in the ac and 2 + inflammatory cells in the vitreous . \n concern for recurrence of m. abscessus endophthalmitis prompted intravitreal injection of amikacin ( 400 g/0.1 ml ) , but activity persisted despite an additional injection of amikacin ( 400 g/0.1 ml ) the following week . \n ppv and explantation of the posterior chamber intraocular lens and capsular bag with intravitreal injection of amikacin ( 400 g/0.1ml ) , were then performed to eradicate mycobacteria that could be sequestered within the capsular bag . \n pathologic examination of the capsular bag showed rare neutrophils , and brown - brenn and afb stains were negative for bacteria and acid - fast organisms . \n however , afb cultures of the vitreous cassette from the second vitrectomy were again reported as positive for m. abscessus , further substantiating the presumptive diagnosis of recurrent m. abscessus endophthalmitis . since then \n , the eye has remained quiet without signs of infection for 11 months , and visual acuity is 20/70 with contact lens correction . \n given the challenges encountered in identifying this organism , repeat pcr and gene sequencing were performed at the university of rochester s clinical microbiology laboratory on culture growths from previously obtained vitreous samples from the two ppv procedures , for experimental purposes . \n 16s rrna gene sequencing was first performed , which identified m. abscessus / chelonae complex . \n pcr as well as sequencing of the hsp65 gene was subsequently performed to differentiate between the two organisms . \n since all prior biochemical testing had identified the organism as m. abscessus , and the initial send - out sequencing of the 16s rrna gene had identified m. abscessus / chelonae complex , m. abscessus had been considered the most likely causative organism throughout the patient s clinical course . \n however , this discrepancy in identification upon repeat pcr and gene sequencing of the hsp65 gene was surprising . it is unlikely that both organisms co - existed in the patient s eye during his episodes of recurrent endophthalmitis . given the greater reliability of pcr and sequencing of the hsp65 gene over traditional biochemical tests and culture techniques,68 we now believe that m. chelonae was likely the infectious agent all along and that the organism was initially misidentified on the basis of less accurate tests . \n m. chelonae endophthalmitis is a grave condition associated with poor visual outcomes.915 causes of m. chelonae endophthalmitis include prior cataract surgery,912 glaucoma surgery,13 iridocyclitis,14 intravitreal steroid injections,9,15 neurotrophic cornea,9 and systemic immunosuppression.16,17 the patient had several risk factors for an ocular rgntm infection : 1 ) an immunosuppressed state after completion of chemotherapy during which herpes zoster infections are more common ; 2 ) a neurotrophic cornea secondary to hzo with a non - healing ulcer and recurrent perforations ; and 3 ) multiple diagnostic and therapeutic ocular surgeries , long - term antibiotic therapy , and steroid immunosuppression during periods of active infection . \n the long - standing cyanoacrylate glue and bandage contact lens used to manage the open globe may have created the milieu for the mycobacteria to thrive . given the neurotrophic cornea and the difficulty in visualizing corneal changes , a smoldering mycobacterial keratitis may have been long - standing but difficult to detect and \n absence of the crystalline lens may have allowed passage of the organism into the vitreous during the therapeutic pk . \n the cause of the early endophthalmitis is unclear , as the vitreous tap was negative for bacterial , viral , and fungal agents . \n although mycobacteria may have been implicated , the eye cleared in response to vancomycin and ceftazidime , antibiotics to which m. chelonae has a low response due to its relatively impermeable lipid cell envelope , drug export systems , biofilm development , and intrinsic b - lactamase activity which would inactivate ceftazidime.18 it is possible that the patient had a polymicrobial infection . nonetheless , despite complete resolution of initial activity , endophthalmitis recurred 9 months later , this time clearly caused by rgntm . m. chelonae may have been sequestered in the eye for the duration , initially kept indolent by low susceptibility to the vancomycin , ceftazidime , and poorly penetrant topical amikacin , and then reactivated as these medications were discontinued . \n additionally , as the topical steroids were tapered , the local inflammatory reaction may have increased , resulting in the return of clinical signs of the latent infection . \n although the ppv , ac washout , and intravitreal injection of antimicrobials were initially effective , endophthalmitis recurred and was unresponsive to additional intravitreal amikacin injections , perhaps due to development of microbial resistance . \n explantation of the posterior chamber intraocular lens and capsular bag was performed to clear a potential nidus of residual mycobacterial infection . \n yet , the pathology results did not identify any mycobacteria within the intraocular lens / capsular bag complex . \n alternative foci of sequestration of the mycobacteria could have been in areas posterior to the iris or near the ciliary body . \n patients with rgntm endophthalmitis typically present with pain , decreased vision , and irritation.3,9,19 nerve injury from the underlying hzo and the previous pk may have altered this classic presentation.20 post - operative recovery of m. chelonae endophthalmitis often results in markedly diminished vision911,13,15 and enucleation9,12,14 due to delayed diagnosis and the devastating nature of the infection . \n fortunately , the patient was able to achieve a highly favorable visual outcome despite his protracted clinical course . \n correct identification of the causative organism throughout the patient s clinical course proved to be an unforeseen challenge . \n m. abscessus , while initially categorized as a subspecies of m. chelonae , was determined to be a separate species in 1992 after dna hybridization studies.2 given the many biochemical similarities between m. abscessus and m. chelonae , clinical laboratories often fail to distinguish between these organisms at the species level with standard biochemical testing.68 two biochemical tests that are often useful for identifying these organisms at the species level are 5% sodium chloride tolerance and utilization of sodium citrate.6,7 \n m. abscessus has been shown to grow on lowenstein - jensen ( lj ) medium with 5% sodium chloride but fails to grow on lj medium with sodium citrate , while m. chelonae tends to grow in lj medium with sodium citrate but is unable to grow on lj medium with 5% sodium chloride.6 however , these tests require up to 4 weeks to complete,6 which can delay proper identification and introduce opportunity for error . in this case , the results from these phenotypic tests had guided the initial identification of m. abscessus from cultures , but the discrepancy in the species identification was likely due to inherent inaccuracies in these tests . \n molecular methods are the most reliable means for the identification of rgntm.68 pcr is a rapid , specific , and cost - effective means of identification of rgntm to the species level that can be performed in parallel with standard biochemical tests;2,6,7 however , it often requires outsourcing to validated laboratories , requiring additional time and resources . \n gene sequencing also serves as an effective option for identification of rgntm.6 genetic loci typically used for mycobacteria species identification with pcr or gene sequencing include the 16s rrna gene , the 16s23s spacer region , the rpob gene , and the hsp65 gene.7 sequencing of the 16s rrna gene is insufficient to differentiate m. abscessus and m. chelonae due to high sequence conservation throughout this gene between the two species , with the sequence being identical in the proximal 500 bases of the gene that are sequenced for species identification.7 pcr with primers targeted toward the hsp65 gene followed by direct sequencing of this gene is the most reliable method of differentiation , given the high sequence variation in the hsp65 gene between m. abscessus and m. chelonae as well as other rgntm.6,8 in this case , pcr plus an important second step , gene sequencing of the hsp65 gene , helped clinch the identification of the causative organism . \n the send - out pcr of the rpob and the hsp65 genes from the vitreous sample from the initial ppv was negative , perhaps due to the limited concentration of the organism in the diluted vitreous sample . \n sequencing of the 16s rrna gene identified the organism as m. abscessus / chelonae complex but did not differentiate between the two . \n repeat testing performed on culture growths from the same samples likely had increased concentrations of the organisms that allowed for positive hsp65 gene pcr , as well as sequencing results for m. chelonae . while culture growths have been widely used to identify organisms with biochemical testing , and to guide antibiotic selection , their use for pcr and gene sequencing may facilitate accurate identification of bacterial species and warrants further exploration . \n while initial identification of the organism was incorrect , all therapy was primarily guided by clinical observation and antibiotic susceptibilities of the cultured organisms . \n a study by brown - elliot et al found that the most active antimicrobials for m. abscessus were amikacin and clarithromycin / azithromycin , and for m. chelonae were clarithromycin / azithromycin , amikacin , tobramycin , and quinolones.1 our patient was treated with topical and intravitreal amikacin based on antibiotic susceptibility testing , and this antimicrobial is an appropriate choice for either m. abscessus or m. chelonae . \n although now equipped with the knowledge of the correct identification of the causative organism , clinical management in retrospect would not have been changed . \n rgntm endophthalmitis should be highly considered in the differential diagnosis of refractory ac and vitreous inflammation , as expedient identification and management is necessary to prevent poor visual outcomes . \n standard biochemical testing can identify the causative organism but is limited by the inability to distinguish between rgntm species reliably , and should not be used as the sole means of organism identification . \n we recommend the use of pcr in conjunction with sequencing of the hsp65 gene for reliable differentiation of m. chelonae and m. abscessus in rgntm ocular infections.68 while pcr and gene sequencing have conventionally been performed directly on patient samples , performing such testing on culture growths can be considered to increase the bacterial load and facilitate the identification of the organism . \n cultures remain critical to therapy , as minimum inhibitory concentration antibiotic susceptibility tests on cultured strains are the best guide to antibiotic selection , given the rapidly rising resistance to antimicrobials within rgntm strains.18", "role": "user" }, { "content": "objectiveto report a case of recurrent nontuberculous mycobacterial endophthalmitis in the context of neurotrophic keratopathy secondary to herpes zoster ophthalmicus that had an atypical presentation and complex course , and highlights the challenges of causative organism identification and therapeutic interventions in this condition.methodsa retrospective chart review was conducted to determine the visual outcomes of the patient.resultsa 68-year - old pseudophakic male with long - standing neurotrophic keratopathy and perforated descemetocele managed with cyanoacrylate glue and a contact bandage lens in the left eye , began experiencing recurrent episodes of endophthalmitis after undergoing a penetrating keratoplasty . several therapeutic procedures including an anterior chamber washout , two pars plana vitrectomies , explantation of the posterior chamber intraocular lens and capsular bag , and multiple intravitreal antimicrobial injections , were performed to which he has ultimately responded favorably , with no signs of infection to date and stable visual acuity . \n the causative organism of his recurrent infections was initially identified as mycobacterium abscessus through biochemical testing and 16s ribosomal ribonucleic acid gene sequencing ; however , repeat polymerase chain reaction ( pcr ) and sequencing of the 65 kda heat shock protein ( hsp65 ) gene for experimental purposes confirmed the accurate identification of the organism to be mycobacterium chelonae . given the greater reliability of pcr and sequencing of the hsp65 gene over traditional biochemical tests and culture techniques , m. chelonae was likely the infectious agent all along , and the organism was originally misidentified on the basis of less accurate tests.conclusionrecurrent atypical mycobacterial endophthalmitis requires expedient identification and management to prevent poor visual outcomes . \n standard biochemical testing can identify the causative organism but is limited by the inability to distinguish between nontuberculous species reliably . \n we recommend the use of pcr in conjunction with sequencing of the hsp65 gene for reliable differentiation of m. chelonae and m. abscessus in atypical mycobacterial ocular infections . \n minimum inhibitory concentration antibiotic susceptibility tests on cultured strains are the best guide to antibiotic selection , given the rapidly rising resistance to antimicrobials in atypical mycobacterial species .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: spontaneous intracerebral hemorrhage ( sich ) is a severe type of stroke with high morbidity and mortality throughout the world . \n some indicators could expectantly identify sich patients with high risk of hematoma expansion and to some extent improve patient prognosis . in 2007 , wada et al . \n suggested that the spot sign on computed tomography angiography ( cta ) was associated with hematoma expansion . \n the meta - analysis by du et al . showed that the spot sign seemed to be a reliable neuroimaging predictor for hematoma expansion . \n the cta spot sign was also found to be associated with higher risk of intraoperative bleeding , postoperative rebleeding , and large residual sich volumes in sich patients undergoing hematoma evacuation . \n moreover , the spot sign score was found to be an independent predictor for mortality in hospital and poor outcomes in patients with sich . although the spot sign may be a practical indicator \n , many factors could affect its accuracy , such as hematoma volume , history of anticoagulants , and onset - to - cta time . \n however , when cta was unavailable , potential neuroimaging predictors on non - contrast ct ( ncct ) were necessary to predict hematoma expansion . \n the blend sign on ncct , which was the blending of the hypoattenuating area and the hyperattenuating region with a clear margin , was introduced as a predictor for hematoma expansion by li et al . . \n however , no studies have compared the predictive value of the cta spot sign and the ncct blend sign in the same cohort of patients . \n thus , we performed this retrospective cohort study to compare the accuracy of cta spot sign and the ncct blend sign in predicting hematoma expansion . \n this was a retrospective study based on the prospective database of the ich patients at the department of neurosurgery of west china hospital , sichuan university . \n all procedures performed in this study were in accordance with the 1964 helsinki declaration and its later amendments or comparable ethical standards . \n the inclusion criteria were the following : ( 1 ) adult patients had sich confirmed by ct or mri scans ; ( 2 ) cta was performed within 6 hours after the onset of sich ; and ( 3 ) follow - up ncct scan was performed within 24 hours after the cta . \n the patients were excluded if ( 1 ) secondary intracerebral hemorrhage was caused by tumor , aneurysm , or arteriovenous malformation ( avm ) ; or ( 2 ) there were no available imaging studies , including initial cta or follow - up ncct . \n patients who received emergency hematoma evacuation before follow - up ncct was performed were also excluded . \n the management of blood pressure followed the recommendations of the latest edition of american heart association / american stroke association ( aha / asa ) and european stroke organisation ( eso ) guidelines . \n baseline information including sex , age , admission blood pressure , and medical history were collected . \n any of the following histories was recorded : hypertension , diabetes mellitus , previous stroke , previous acute coronary event , smoking , and alcohol abuse . \n in addition , results of admission coagulation tests including platelet count , prothrombin time ( pt ) , activated partial thromboplastin time ( aptt ) , and international normalized ratio ( inr ) were also collected . \n cta was performed when the patients were admitted to the emergency department as a part of standard clinical care on a dual - source 64-slice ct scanner ( somatom definition flash ; siemens healthcare sector , forchheim , germany ) , including ncct scan ( 120 kv , 340 ma , contiguous 5-mm axial slices ) and cta scan . for the cta , 100 ml of ioversol ( iopamidol , bracco pharma co , shanghai , china ; \n 370 mg i / ml ) was intravenously injected at a rate of 4.8 ml / s via a power injector through an intravenous line using the following parameters : 80 kvp ; 110 ma ; slice thickness , 1 mm ; and pitch , 1 : 1 . \n ncct was performed within 24 hours after the first ct to evaluate the hematoma size . \n all the ct scans were reviewed independently by two neuroradiologists who were blinded to the clinical condition of the patients . \n disagreements about the occurrence of the spot sign or the blend sign were settled by joint discussion of the two readers . \n radiological criteria for the spot sign were in accordance with those reported previously : at least 1 focus of contrast pooling within the ich : high hounsfield unit ( hu ) value ( > 120 ) ; discontinuous from normal or abnormal vasculature adjacent to the ich ; and any size and morphology . \n the hematoma blend sign was defined as follows : ( 1 ) there is blending of the relatively hypoattenuating area with the adjacent hyperattenuating region within a hematoma ; ( 2 ) there is a well - defined margin between the hypoattenuating area and the adjacent hyperattenuating region that is easily recognized by the naked eye ; ( 3 ) the hematoma should have at least an 18 hu difference between the 2 density regions ; and ( 4 ) the relatively hypoattenuating area is not encapsulated by the hyperattenuating region . \n the hematoma has to meet the 4 criteria mentioned above to be defined as blend sign . \n illustrative spot sign positive cta images and blend sign - positive ncct images are showed in figure 1 . \n volume of hematoma was calculated from ct scans using the formula a*b*c/2 , where a was the largest diameter on the largest hemorrhage slice , b was the maximal diameter perpendicular to a , and c was the vertical hematoma depth . \n the hematoma expansion was defined as a > 33% relative increase or > 12.5 ml absolute increase in hematoma size on follow - up ct scan . \n baseline characteristics including demographics , history , admission blood pressure , admission hematoma characteristics , and coagulation tests were compared between patients with hematoma expansion and those without hematoma expansion . \n continuous values were expressed as mean and standard deviation ( sd ) and analyzed by the t - test . \n discontinuous variable data were expressed as median and interquartile range ( iqr ) and analyzed by the wilcoxon rank sum test . \n multivariable logistic regression was performed to adjust the odds ratio ( or ) and 95% confidence interval ( ci ) of the spot sign and the blend sign on hematoma expansion . \n the value of the spot sign and the blend sign for predicting hematoma expansion was analyzed by receiver - operator analysis . \n the area under the receiver - operating characteristic ( roc ) curves of the spot sign and the blend sign were compared by using the z test . \n the interobserver reliability for the identification of the spot sign and the blend sign was determined by calculation of values . \n this was a retrospective study based on the prospective database of the ich patients at the department of neurosurgery of west china hospital , sichuan university . \n all procedures performed in this study were in accordance with the 1964 helsinki declaration and its later amendments or comparable ethical standards . \n the inclusion criteria were the following : ( 1 ) adult patients had sich confirmed by ct or mri scans ; ( 2 ) cta was performed within 6 hours after the onset of sich ; and ( 3 ) follow - up ncct scan was performed within 24 hours after the cta . \n the patients were excluded if ( 1 ) secondary intracerebral hemorrhage was caused by tumor , aneurysm , or arteriovenous malformation ( avm ) ; or ( 2 ) there were no available imaging studies , including initial cta or follow - up ncct . \n patients who received emergency hematoma evacuation before follow - up ncct was performed were also excluded . \n the management of blood pressure followed the recommendations of the latest edition of american heart association / american stroke association ( aha / asa ) and european stroke organisation ( eso ) guidelines . \n baseline information including sex , age , admission blood pressure , and medical history were collected . \n any of the following histories was recorded : hypertension , diabetes mellitus , previous stroke , previous acute coronary event , smoking , and alcohol abuse . \n in addition , results of admission coagulation tests including platelet count , prothrombin time ( pt ) , activated partial thromboplastin time ( aptt ) , and international normalized ratio ( inr ) were also collected . \n cta was performed when the patients were admitted to the emergency department as a part of standard clinical care on a dual - source 64-slice ct scanner ( somatom definition flash ; siemens healthcare sector , forchheim , germany ) , including ncct scan ( 120 kv , 340 ma , contiguous 5-mm axial slices ) and cta scan . for the cta , 100 ml of ioversol ( iopamidol , bracco pharma co , shanghai , china ; \n 370 mg i / ml ) was intravenously injected at a rate of 4.8 ml / s via a power injector through an intravenous line using the following parameters : 80 kvp ; 110 ma ; slice thickness , 1 mm ; and pitch , 1 : 1 . \n ncct was performed within 24 hours after the first ct to evaluate the hematoma size . \n all the ct scans were reviewed independently by two neuroradiologists who were blinded to the clinical condition of the patients . \n disagreements about the occurrence of the spot sign or the blend sign were settled by joint discussion of the two readers . \n radiological criteria for the spot sign were in accordance with those reported previously : at least 1 focus of contrast pooling within the ich : high hounsfield unit ( hu ) value ( > 120 ) ; discontinuous from normal or abnormal vasculature adjacent to the ich ; and any size and morphology . \n the hematoma blend sign was defined as follows : ( 1 ) there is blending of the relatively hypoattenuating area with the adjacent hyperattenuating region within a hematoma ; ( 2 ) there is a well - defined margin between the hypoattenuating area and the adjacent hyperattenuating region that is easily recognized by the naked eye ; ( 3 ) the hematoma should have at least an 18 hu difference between the 2 density regions ; and ( 4 ) the relatively hypoattenuating area is not encapsulated by the hyperattenuating region . \n the hematoma has to meet the 4 criteria mentioned above to be defined as blend sign . \n illustrative spot sign positive cta images and blend sign - positive ncct images are showed in figure 1 . \n volume of hematoma was calculated from ct scans using the formula a*b*c/2 , where a was the largest diameter on the largest hemorrhage slice , b was the maximal diameter perpendicular to a , and c was the vertical hematoma depth . \n the hematoma expansion was defined as a > 33% relative increase or > 12.5 ml absolute increase in hematoma size on follow - up ct scan . \n baseline characteristics including demographics , history , admission blood pressure , admission hematoma characteristics , and coagulation tests were compared between patients with hematoma expansion and those without hematoma expansion . \n continuous values were expressed as mean and standard deviation ( sd ) and analyzed by the t - test . \n discontinuous variable data were expressed as median and interquartile range ( iqr ) and analyzed by the wilcoxon rank sum test . \n multivariable logistic regression was performed to adjust the odds ratio ( or ) and 95% confidence interval ( ci ) of the spot sign and the blend sign on hematoma expansion . \n the value of the spot sign and the blend sign for predicting hematoma expansion was analyzed by receiver - operator analysis . \n the area under the receiver - operating characteristic ( roc ) curves of the spot sign and the blend sign were compared by using the z test . the interobserver reliability for the identification of the spot sign and the blend sign \n from february 2015 to march 2016 , a total of 115 consecutive patients met the inclusion criteria and were enrolled in this study . in this study , the median time interval from sich onset to initial cta was 4 hours ( 3 , 5 hours ) . \n the range of age was between 36 and 83 years ( mean 58.811.6 years ) . \n positions of hematoma included lobar ( 17 , 14.78% ) , basal ganglia ( 64 , 55.65% ) , thalamus ( 14 , 12.17% ) , cerebellum ( 6 , 5.22% ) , and brain stem ( 14 , 12.17% ) . \n the baseline characteristics of the expander group and the non - expander group are shown in table 1 . in our study , the spot sign was observed in 25 patients , whereas the blend sign was observed in 22 patients . \n the prevalence of the spot sign and the blend sign was significantly higher in patients with hematoma expansion compared with those without hematoma expansion . \n interobserver reliability for the identification of the spot sign and the blend sign was excellent between the two neuroradiologists ; the was 0.924 and 0.946 , respectively . for the multivariable analysis , prevalence of the spot sign and the blend sign \n was independently associated with hematoma expansion ( table 2 ) . in this study , the sensitivity , specificity , positive predictive value , and negative predictive value of the spot sign for predicting hematoma expansion were 57.14% , 89.66% , 64.00% , and 86.67% , respectively . by contrast , the sensitivity , specificity , positive predictive value , and negative predictive value of the blend sign were 42.86% , 88.51% , 54.55% , and 82.80% , respectively . \n the roc curves of the two procedures for predicting hematoma expansion in sich patients are shown in figure 2 . \n no significant difference was found between the area under roc curves of the spot sign and the blend sign ( p=0.383 ) \n this is the first study comparing the predictive value of the cta spot sign with that of the ncct blend sign in the same cohort of patients with sich . \n both the spot sign and the blend sign had good predictive value for hematoma expansion . \n although there was no significant difference in the predictive accuracy of the spot sign and the blend sign , the spot sign seemed to be a better neuroimaging marker for predicting hematoma expansion . \n the cta spot sign was shown to be a potential predictor for hematoma expansion in recent studies . in accordance with previous studies \n , our study also showed that the cta spot sign is a good predictor for hematoma expansion . \n the cta spot sign was considered to be associated with bleeding from ruptured vessels , but the underlying mechanism is still unclear . \n although the results of these studies supported the predictive role of the cta spot sign in hematoma expansion , the values were different . \n found the sensitivity , specificity , positive predictive value , and negative predictive value of the spot sign to be 91% , 89% , 77% , and 96% . \n however , in demchuk et al.s study , the sensitivity , specificity , positive predictive value , and negative predictive value of the spot sign were 51% , 85% , 61% , and 78% . \n suggested the sensitivity , specificity , positive predictive value , and negative predictive value of the spot sign to be 57.38% , 90.48% , 74.47% , and 81.43% . \n the meta - analysis by du et al . showed the pooled sensitivity of the spot sign was 53% and the specificity was 88% . \n first , prevalence of the spot sign might be interfered with by different hematoma volumes or history of anticoagulant drugs . \n it has been found that the cta spot sign can predict expansion of hematoma reliably in patients with hematomas > 30 ml . however , small hematomas have a lower prevalence of the spot sign and a lower risk of expansion . ich patients with a history of warfarin use and the apolipoprotein \n second , it was found that spot signs identified in earlier phases might be associated with greater absolute expansion . with the increase of sich onset - to - cta time , the frequency of the cta spot sign increases and the positive predictive value of the spot sign for hematoma expansion decreases . in spot sign - positive patients , a shorter onset - to - cta time and higher hu of spot signs are factors for prediction of hematoma expansion . \n third , the different scan timing could lead to different prevalence and predictive accuracy of the cta spot sign . \n a 90-second delay in time of cta could improve the sensitivity of the spot sign for predicting hematoma expansion . \n in addition , the spot sign identified on cta obtained at high levels of tube was found to have better predictive accuracy for hematoma expansion . \n thus , due to these various factors a standardized protocol is necessary to improve the practicability and accuracy of the cta spot sign . unlike the spot sign \n compared to the spot sign on cta , the blend sign is easier to identify . \n were the first to report this novel neuroimaging predictor . in their study , the sensitivity , specificity , positive predictive value , and negative predictive value of the blend sign for predicting hematoma expansion were 39.3% , 95.5% , 82.7% , and 74.1% , respectively , which suggested that the blend sign seemed to be a good predictor for hematoma expansion . in the present study , \n the sensitivity and the negative predictive value were higher and the specificity and the positive predictive value were lower . \n just like the spot sign , many underlying factors might influence the predictive accuracy of the blend sign . \n the time to baseline ct of our study ( 3.32 hours in expanders and 3.94 hours in non - expanders ) was longer than that in li et al.s study ( 1.67 hours in expanders and 2.76 hours in non - expanders ) . \n moreover , the initial volume of hematoma was larger in our study ( 31.99 ml in expanders and 23.06 ml in non - expanders ) than that in li et al.s study ( 24.31 ml in expanders and 13.12 ml in non - expanders ) . \n the difference in baseline ct time and initial hematoma volume might be associated with the difference in predictive accuracy . \n further research is needed to investigate the exact factors affecting the predictive value of the blend sign . \n it was suggested that the ctp dynamic spot sign had a higher predictive value for hematoma expansion than cta . \n fluid levels , density heterogeneity , and margin irregularity on ncct were found to be associated with hematoma expansion at 24 hours . \n similarly , the black hole sign on ncct , which was defined as the hypoattenuating area within the hyperattenuating hematoma with a clear border , was found to be a predictor for hematoma expansion . \n although some scoring systems seemed to be effective in hematoma expansion prediction , they did not include these new imaging markers . a comprehensive predictive scoring system including more neuroimaging markers is expected to be built in the future . \n first , this was a single - center retrospective cohort study , and the sample size was relatively small . moreover , only the cta spot sign and the blend sign were investigated . \n furthermore , the interval between onset and baseline cta was relatively long , which might influence the predictive accuracy of both signs . \n further multi - center prospective studies with larger cohorts and shorter onset - to - cta time are needed . \n our study compared the predictive value of the spot sign and the blend sign in the same cohort of sich patients . \n both the spot sign and the blend sign seemed to be good predictors for hematoma expansion , and the spot sign appeared to have a better predictive accuracy . \n since more and more neuroimaging predictors for hematoma expansion have been developed , it is hoped that a comprehensive predictive scoring system including more neuroimaging markers will be built in the future .", "role": "user" }, { "content": "backgroundhematoma expansion is associated with poor outcome in intracerebral hemorrhage ( ich ) patients . \n the spot sign and the blend sign are reliable tools for predicting hematoma expansion in ich patients . \n the aim of this study was to compare the accuracy of the two signs in the prediction of hematoma expansion.material/methodspatients with spontaneous ich were screened for the presence of the computed tomography angiography ( cta ) spot sign and the non - contrast ct ( ncct ) blend sign within 6 hours after onset of symptoms . \n the sensitivity , specificity , and positive and negative predictive values of the spot sign and the blend sign in predicting hematoma expansion were calculated . \n the accuracy of the spot sign and the blend sign in predicting hematoma expansion was analyzed by receiver - operator analysis.resultsa total of 115 patients were enrolled in this study . \n the spot sign was observed in 25 ( 21.74% ) patients , whereas the blend sign was observed in 22 ( 19.13% ) patients . \n of the 28 patients with hematoma expansion , the cta spot sign was found on admission ct scans in 16 ( 57.14% ) and the ncct blend sign in 12 ( 42.86% ) , respectively . the sensitivity , specificity , positive predictive value , and negative predictive value of the spot sign for predicting hematoma expansion were 57.14% , 89.66% , 64.00% , and 86.67% , respectively . \n in contrast , the sensitivity , specificity , positive predictive value , and negative predictive value of the blend sign were 42.86% , 88.51% , 54.55% , and 82.80% , respectively . \n the area under the curve ( auc ) of the spot sign was 0.734 , which was higher than that of the blend sign ( 0.657).conclusionsboth the spot sign and the blend sign seemed to be good predictors for hematoma expansion , and the spot sign appeared to have better predictive accuracy .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: a retrospective study was conducted of all the patients who were treated in the department of surgery , sheri kashmir institute of medical sciences ( medical college ) , bemina , srinagar , kashmir , india , from june 2003 to may 2009 for missed foreign bodies in the hand . \n data regarding all cases managed for missed foreign bodies in hands were retrieved from the databank and minor procedures registry of the hospital , and processed and analyzed with respect to patient demographics and management . for our study \n , we defined the missed foreign body as the one which had been overlooked at the time of injury , and all such cases which reported with various complications resulting from the foreign bodies were studied irrespective of the time - interval between injury and complication . \n over a period of six years from june 2003 to may 2009 , 61 cases with missed foreign bodies in hands were managed ( 52 males and 9 females ) . \n foreign bodies of different types were retrieved in patients of all age groups and belonging to all walks of life as shown in figs . 1 , 2 and tables 1 , 2 . \n the clinical presentation and the time elapsed since possible penetration of foreign bodies is depicted in table 3 . as shown in fig . \n 1 , wooden splinters ( n=37 ; 60.66% ) were the most common foreign bodies retrieved followed by metallic fragments ( n=13 ; 21.31% ) , glass fragments ( n=7 ; 11.47% ) , and stone fragments ( n=4 ; 6.56% ) . \n the metallic fragments included components of explosive devices like land mines and grenades ( n=4 ) , needles ( n=3 ) , iron / copper wire fragments ( n=3 ) , and aluminum wire fragment ( n=1 ) . \n . occupational status of the patients presenting features of missed foreign bodies time elapsed since possible penetration of foreign body as is shown in fig . \n 3 , among the 61 cases treated by us , 34 ( 55.74% ) patients had not taken any medical treatment before reporting to our department . in the rest of the 27 cases , \n basic wound care had been provided after imaging with plain radiograms in 18 ( 29.51% ) cases and in 9 ( 17.75% ) cases , only wound care had been provided without any imaging . among these 18 cases that had been imaged with only radiograms , \n five cases had been missed in our facility and 13 patients had been managed in other health centers . \n preoperative evaluation with ultrasonography and exploration led to recovery of wooden splinters in 17 cases and aluminum wire fragment in one case ( see fig . \n ( c ) plain radiograph showing a radio - opaque pellet along proximal phalanx of index finger ( arrow ) . \n besides clinical assessment , plain radiographs and ultrasound ( high resolution and high frequency ) were the tools utilized in the diagnosis and localization of the missed foreign bodies as shown in table 4 . \n diagnostic modalities used for localizing missed foreign bodies management of all the missed foreign bodies comprised of localization by imaging and removal . in 47 cases , removal of the foreign body was accomplished under local anesthesia on outpatient basis . in the rest of the 14 cases , \n the indications for resorting to general anesthesia included young age ( n=8 ) , failed exploration or incomplete removal under local anesthesia ( n=4 ) . \n there were no postoperative complications in 27 patients who remained under follow - up up to three months . \n sixteen patients were discharged from services within a month as their wounds had healed normally and they had no symptoms / signs to warrant further follow - up . \n penetrating injuries to hands are the most common cause of hospital emergency room visits ( 11 ) and an important health hazard associated with many occupations . \n the obvious foreign bodies might be removed by the patients themselves or the primary health providers . \n however , it is reported that 1538% of the foreign bodies get overlooked at the time of initial examination ( 1 , 2 ) , and wound management depending upon the clinical acumen of the health care provider and the availability and use of imaging facilities . \n the missed foreign body may remain asymptomatic for prolonged periods or else lead to a wide range of complications including pain , abscess , chronic discharging wound , necrotizing fasciitis ( 12 ) , bone and joint destructive lesions ( 13 ) , granulomas ( 14 ) with impairment of tendon mobility or triggering of digits ( 15 ) , migration ( 16 ) , delayed tendon ruptures ( 16 , 17 ) , neurodeficits ( 15 , 18 ) , pyogenic granulomas ( 19 ) , and vascular events ( 20 , 21 ) . in our series \n as is evident from table 1 , impairment of wound healing and chronic pain were the major presenting features . \n one of the cases reported with paresthesias in median nerve territory and had a small retained glass piece in the area of carpus impinging on the median nerve . \n the vascular complications secondary to retained foreign bodies include major arterial erosion , spontaneous hematoma , thromboemboli , digital ischemia , and skin necrosis . \n one of the cases of retained shrapnel reported with spontaneous hematoma and on exploration , erosion in a small branch of superficial palmar arch was documented . \n we neither encountered any spontaneous ruptures of tendons nor did we retrieve any foreign body of plastic origin which is commonly reported in the literature . depending upon the occupational status , \n a wide range of retained foreign bodies has been reported in the literature . in our experience , \n wooden splinters formed the most common foreign bodies retained ( n=37 , 60.65% ) , followed by needles / metal pieces , glass pieces , stone fragments , and glass pieces . \n the increased incidence of wooden splinters can be explained by location of our hospital in the proximity of the biggest fruit market in kashmir where wooden boxes are ferried . \n we also managed four cases of retained metallic fragments of explosives ( pellets / shrapnel ) in whom these foreign bodies had been ignored at the time of injury as they had reported to hospitals along with many other seriously injured victims of armed violence and had got less attention due to diversion of attention toward serious patients ( scenario at the time of mass casualties like land mine blast ) . \n it is the failure of the primary care physician to advise appropriate imaging due to lack of suspicion or the non - radio opaque nature of the foreign bodies , which lead to retention of foreign bodies and subsequent complications and litigations . \n we also managed 18 such cases in which non - radio opaque foreign bodies had got initially missed due to non - utilization of ultrasonography on part of physicians . recently , \n various studies have been published which have strongly proven the sensitivity and cost effectiveness of ultrasonography in detection of foreign bodies in hands ( 6 , 7 , 11 ) . \n it is against this background that many workers have suggested that ultrasound facilities and basic training be provided to emergency physicians so that foreign bodies can be ruled out in all cases of penetrating hand injuries ( 22 ) . \n this suggestion is supported by the successful application of focused assessment with sonography for trauma ( fast ) by emergency physicians ( 23 ) . in literature , depending upon the nature of foreign body and the available facilities , various modalities of imaging are recommended for diagnosis and localization , which include plain radiographs , ultrasonograms , ct scans , mri , fistulograms , and xerographs ( 6 , 7 , 22 , 24 , 25 ) . \n radiopaque material is usually easy to detect but it is the radiolucent bodies like wooden splinters which are difficult to detect and are usually missed ( 4 ) . \n high resolution ultrasonography and mri are useful in detection of these non - radio opaque foreign bodies ( 2630 ) . in our setting , \n free - of - cost mri and ct - scan facilities are not available and all of the missed foreign bodies were detected by plain images or ultrasound as shown in table 4 . \n the only metallic body which was missed on plain image was picked up by ultrasound and on exploration turned out to be a piece of aluminum wire . \n ct and mri if available can , however , provide additional information regarding the location and relationship of retained foreign body with adjacent tendons , neurovascular bundles , and muscle groups and thereby guide in selection of surgical approach and extent of exploration ( 29 , 30 ) . the missed foreign bodies may produce immediate symptoms like wound infections or may remain dormant for even decades ( 2 ) . in our series , 37 patients ( 61.6% ) \n were not aware of the traumatic event leading to entry of the foreign bodies as they belonged to poor labor class who frequently get injured and hence tend to ignore the injuries to avoid disturbance in their routine life . \n one of our patients who belonged to non - labor class reported after two years of getting injured with retained glass piece . once diagnosed , proper surgical exploration and removal of the missed foreign body is the treatment . \n but the key to success is accurate localization of the foreign body ( 6 , 10 , 31 , 32 ) using available imaging facilities . in most of the cases , the exploration can be done on outpatient basis under local anesthesia , but improper localization can end up in long and taxing explorations with no recovery of foreign bodies and extensive damage to soft tissues ( 25 ) . in our experience , 47 cases ( 77% ) \n were managed as outpatients and only 14 cases ( 23% ) required exploration under general anesthesia . among these 14 cases , \n four patients were such in whom exploration under local anesthesia had failed and intraoperative use of ultrasonogram was resorted to for successful recovery of foreign bodies . \n intraoperative use of ultrasonography has in fact been recommended in literature also as an aid in real - time localization of foreign bodies ( 31 ) . \n this accurate localization enables surgeons to make smaller and cosmetically appealing incisions and remove foreign bodies with a minimum amount of dissection and operation time ( 33 ) . \n a total of 61 cases of missed foreign bodies were managed over a period of six years in our hospital . \n foreign bodies need to be suspected and ruled out in every case of penetrating injury to hands by use of appropriate imaging modality . in missed foreign bodies , management \n the authors have not received any funding or benefits from industry to conduct this study .", "role": "user" }, { "content": "introductionpenetrating hand injuries are common and improper assessment can result in missed foreign bodies . \n these bodies can result in a wide range of complications.aimthe aim of our study was to study the profile of patients reporting with missed foreign bodies in the hand.materials and methodsall the cases treated in the department of surgery , sheri kashmir institute of medical sciences ( medical college ) , kashmir , for missed foreign bodies in hands from june 2003 to may 2009 were studied retrospectively.resultsa total of 61 cases with missed foreign bodies of different nature were treated over the period of six years . \n wooden splinters were the most common foreign bodies missed . \n preoperative localization was accomplished with plain radiographs and ultrasonograms . \n most of the cases were treated on outpatient basis.conclusionsforeign bodies should be suspected and ruled out in all cases of penetrating injuries of hands . \n missed foreign bodies need to be removed after proper localization by imaging .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: metal \n complexes have been studied for decades as potential cytotoxic \n agents , because of the unprecedented and continued success of cisplatin \n as a chemotherapeutic . \n investigations into ru(ii ) polypyridyl \n complexes have been particularly extensive , because of their ease \n of synthesis , appealing chemical , physical , and photophysical characteristics , \n and their high affinities for nucleic acids . \n most early studies focused \n on characterizing the in vitro interactions of these \n complexes with dna , and quantifying the potencies of the compounds both as traditional \n cytotoxic agents and as light - activated agents for photodynamic therapy \n ( pdt ) or phototherapy . in recent years \n , attention has shifted to understanding \n the cellular localization properties of ru(ii ) complexes , along with their \n mechanisms of cellular uptake and cytotoxicity , providing a deeper understanding of how these \n compounds elicit their biological activities . \n an attractive \n feature of ru(ii ) polypyridyl complexes that makes \n them particularly useful for applications as biological probes and \n effectors is the diversity of the chemical structures that are readily \n available through modifications of the coordinated ligands . \n however , most biological studies have focused \n on complexes that carry an overall charge of + 2 or greater . \n this significantly \n limits the chemical structures and physical properties of the molecules \n under investigation . \n a question that occurred to us was this : to what \n degree could the biological properties of a chemically inert ru(ii ) \n complex be tuned by chemical modification of the ligands surrounding \n the metal center ? to address this question , we have investigated \n the biological activities , \n cellular uptake , localization , and mechanism of cell killing of two \n simple ru(ii ) complexes that are commonly used dyes for solar cell \n research or biological staining , but have not been previously explored \n as pdt agents . \n ru(bathophenanthroline)3 ( 1 ; see chart 1 ) is a hydrophobic molecule \n with a high dna binding affinity , while ru(bathophenanthroline \n disulfonate)3 ( 2 ) is hydrophilic and possesses \n a high affinity for proteins . \n both complexes are efficient \n singlet oxygen ( o2 ) generators with the same \n quantum yields for o2 production ( ) and similar molar extinction coefficients ( ) \n . both are luminescent , allowing for analysis \n by fluorescence microscopy . however , although the photophysical properties \n of the compounds are almost identical , the physical properties of \n the two ru(ii ) compounds are quite dissimilar ( see table 1 ) . \n compound 1 carries an overall charge of + 2 , while 2 has \n an overall charge of 4 . \n they also have very different hydrophilicities , \n as indicated by their partition coefficient or log p values . \n given the radically different physical properties of the \n complexes , we anticipated differences in their biological effects \n that could provide information for future rational design of light - activated \n cytotoxic agents . considering the established dogma \n of the field , \n it would be expected \n that the negatively charged compound 2 would not enter \n cells and would suffer low efficacy , \n while the positively charged , dna - binding compound 1 would \n prove the more effective pdt agent . \n instead , our findings run counter \n to this prediction . here , we show large differences in potency , cellular \n uptake , localization , and mechanism of cytotoxicity of these agents , \n illustrating that radical modulation of biological properties is possible \n with ligand modifications of simple homoleptic ru(ii ) polypyridyl \n complexes that are substitutionally inert . \n most importantly , our results \n also prove that a significantly greater range of charge states and \n physical properties of ru(ii ) complexes are compatible with potential \n application as pdt agents . \n as both 1 and 2 are efficient catalysts \n for the light - activated generation \n of o2 ( = 0.42 , 0.43 ) , it was expected that the two compounds would \n act as sensitizers for pdt . \n accordingly , their dna damaging properties \n were assessed with puc19 plasmid dna and analyzed by gel electrophoresis \n ( figure 1 ) . \n each compound was incubated with \n plasmid and irradiated with 40 j / cm of visible light ( > 400 \n nm ) or kept in the dark . \n compound 1 is known to bind \n strongly with dna , and precipitation of the dna with the complex was \n observed at concentrations above 31.3 m both in the light and \n in the dark ( figure 1a ) . \n in contrast , 2 is a much more hydrophilic molecule , and the negatively \n charged sulfonate functional groups were anticipated to cause electrostatic \n repulsions between the complex and the negatively charged backbone \n of the dna . \n consistent with low dna affinity , no dna precipitation or smearing was observed with up to 500 m \n of 2 ( figure 1b ) . when exposed \n to light , both 1 and 2 induced single strand \n dna breaks , creating relaxed circular plasmid . \n this is likely due \n to the photogeneration of o2 that mediates \n the dna damage . \n however , for 2 , the amount of relaxed \n circle plasmid did not exhibit any concentration dependence above \n 125 m , suggesting either a reduction in as the concentration of the complex is increased , or alternative \n quenching mechanisms that impede dna damage . \n agarose \n gel electrophoresis of puc19 with increasing concentrations \n of ( a ) 1 and ( b ) 2 in the dark or irradiated \n ( > 400 nm ) . \n lanes 1 and 12 , dna molecular weight standard ; \n lane 2 , linear ( reaction with ecor1 ) ; lane 3 , relaxed circle ( reaction \n with cu(phen)2 ) ; lanes 411 , 0 , 8.25 , \n 16.5 , 31.3 , 62.5 , 125 , 250 , and 500 m compound . \n cytotoxicity \n dose response of ( c ) 1 and ( d ) 2 in the \n dark ( open squares ) or irradiated ( closed circles ) . \n hl60 cells were \n incubated for 72 h with compound prior to quantification of viability . \n as both compounds are capable \n of light - induced dna damage , the \n cytotoxicity of 1 and 2 were evaluated in \n the a549 human non - small cell lung cancer , the hl60 human promyelocytic \n leukemia , and the jurkat human t lymphoblastoid cell lines in the \n presence and absence of 7 j / cm of > 400 nm light . \n the \n ic50 values across the cell lines for 1 ranged \n from \n 0.62 to 3.75 m in the dark . upon irradiation \n , potency was increased \n to a range of 0.075 to 0.35 m , resulting in an average phototoxicity \n index ( pi = ic50 ( dark)/ic50 ( light ) of 10- \n to 20-fold ( see figure 1 , table 2 ) ) . \n the phototoxicity index ( pi ) is \n the ratio of the dark and light ic50 values . in marked contrast to the high toxicity \n of 1 in the \n absence of light \n , no toxicity was observed in the dark with 2 across all cell lines at concentrations up to 300 m . \n however , compound 2 was effective in killing cells when \n irradiated , with ic50 values ranging from 3.3 m \n to 17.3 m , consistent with the concentrations required for in vitro dna damage . \n this provides for a large therapeutic \n window , as no cell death is observed for samples in the absence of \n irradiation . surprisingly , not only was compound 1 toxic to cells \n upon irradiation , it also induced cell death far more rapidly in hl60 \n cells than traditional dna damaging agents such as cisplatin . \n complete \n cell death was observed within 2 h of irradiation with 1 ( see figure s1 in the supporting information ) . \n the compound also induced cell death in the dark , but more slowly , \n with 30% viable cells remaining at 24 h. in marked contrast \n to 1 , compound 2 induced cell death after \n irradiation only after a long delay , with 70% viable cells remaining \n at 24 h and 55% remaining at 48 h. the significant disparity \n in the potency , phototoxicity index ( pi ) , \n and rates of cell killing for the two compounds despite their equivalent \n abilities to sensitize o2 strongly suggested \n that they were acting through different cellular mechanisms , possibly \n by interacting with different biological targets . \n as the compounds \n are substitutionally inert , they are unlikely to covalently modify \n biomolecules . \n this complicates target identification by isolation \n of protein or nucleic acid components of the cells , so the subcellular \n localization was investigated instead . \n both 1 and 2 are emissive , \n allowing for direct visualization \n in cells . \n flow cytometry and fluorescence microscopy were used to \n provide relative uptake values , the time dependence of compound uptake , \n and information on the subcellular localization of the compounds . \n to optimize signal intensity , \n flow cytometry \n with a549 cells revealed greater uptake of 1 compared \n with 2 at time points of 2 and 24 h , with an 11.6-fold \n and 8.2-fold difference in signal , respectively ( see figure s2 in the supporting information ) . between the time \n points of 2 and 24 h , the average emission of cells incubated with 1 increased by 2.8-fold while the amount of 2 increased by 4-fold . \n this data was supported by direct quantification \n of the number of ruthenium atoms per cell using inductively coupled \n plasma optical emission spectroscopy ( icp - oes ; see tables s1 and s2 in the supporting information ) . \n the relative amount of compound 1 in cells versus \n the cell media increased 4-fold from 2 h to 24 h , from 5.4% \n to 22% . \n much less of compound 2 entered the cells , with a maximal uptake of 0.7% at 24 h. while \n low , this degree of uptake of the 4-charged 2 is comparable to cisplatin , a neutral compound , dosed at the same \n concentration ( 20 m , 0.8% at 24 h ) . \n the emission of 1 and 2 was also measured in a549 cells using \n an apotome microscope ( the adherent cell line was chosen to facilitate \n the required wash steps to allow for the use of fluorescent reporters ) . \n the relative rates of uptake of each of the complexes were analyzed \n as a function of time , and both 1 and 2 were \n visible inside cells as early as 2 h after compound addition ( see figure s3 in the supporting information ) , consistent \n with the flow cytometry and icp - oes results . \n images were taken at \n 2 , 8 , 18 , and 24 h , and intracellular levels of both complexes appeared \n to plateau at 8 h. the uptake data showed good agreement between the \n three techniques and two cell lines , suggesting similar behavior in \n adherent and suspension cell lines . \n differences in the intracellular \n localization of 1 and 2 were evaluated by \n determining colocalization \n of luminescence of the compounds and fluorescent markers of organelles \n in a549 cells . \n overlap in signals between the compounds , cellular \n nucleus , mitochondria , and lysosome was measured over a 24 h period . \n compounds 1 and 2 have very different localization \n profiles , as exemplified by the imaging 8 h after dosing ( figure 2 ) . \n compound 1 substantially localized \n to lysosomes and the mitochondria in the absence of light , while 2 remained primarily in the cytosol . \n exposure to light did \n have an impact on compound localization , where 1 induced \n nuclear localization of both the mitochondrial and lysosome markers \n ( see figures 2a and 2b , top ) , indicating that photoinduced damage mediated by 1 reduced the integrity of the nuclear membrane . \n in contrast , 2 was primarily observed in lysosomes after irradiation , and \n did not co - localize with mitochondria ( figures 2a and 2b , bottom ) . \n in addition , irradiation \n in the presence of 2 did not result in the appearance \n of organelle markers in the nucleus , suggesting the nuclear membrane \n remained intact . \n neither 1 nor 2 was found \n to associate with the plasma or nuclear membranes . \n apotome microscopy showing \n subcellular localization of 1 and 2 at 8 \n h. co - localization of 1 and 2 in mitochondria \n or lysosomes is indicated by the apparent \n yellow emission . \n red color \n denotes intrinsic emission of 1 and 2 , whereas \n blue color denotes hoechst staining of the nucleus . \n the yellow color \n occurs due to overlap of the red emission from the ruthenium complexes \n and green emission of the organelle - specific dyes , indicating colocalization . \n compound 1 localizes in both the mitochondria and the \n lysosomes while 2 was not predominantly found in either \n organelle . \n as compound 1 appeared to localize within mitochondria , \n it was hypothesized that this may account for its high toxicity in \n the absence of irradiation . to determine \n if either 1 or 2 caused a reduction in mitochondrial function , mitochondrial \n membrane potential was measured using tetramethylrhodamine ethyl ester \n ( tmre ) ( see figure 3 ) . \n tmre is a cationic dye \n that accumulates in active mitochondria as a result of the negative \n membrane potential ( m ) . \n inactive or depolarized mitochondria \n exhibit a decreased membrane potential , and tmre does not localize \n in these organelles . \n compound 1 induced rapid and complete \n depolarization of mitochondria both in the dark and upon irradiation \n ( figure 3b , 3c ) . however , \n surprisingly , cell viability did not parallel mitochondrial potential . \n while mitochondrial function was completely impaired at 2 h post treatment \n with 1 in the dark , viability decreased slowly , with \n 44% 6% viable cells remaining after 24 h. in contrast \n , both \n mitochondrial function and cell viability fell to 0% within 2 h of \n irradiation . \n thus , while 1 completely impedes mitochondrial \n function within 2 h even in the absence of light , irradiation induced \n additional damage that results in rapid cell death , along with the \n loss of mitochondrial function . ( a ) example images and quantification \n of the emission of tetramethylrhodamine \n ethyl ester ( tmre ) in the presence and absence of 1 and 2 . \n compound 1 does not show increased tmre emission \n over background emission of 1 , while compound 2 does not add to the tmre emission . \n mitochondrial potential and cell \n viability of a549 cells as a function of time for ( b ) 1 , in the dark ; ( c ) 1 , irradiated ; ( d ) 2 , in the dark ; and ( e ) 2 , irradiated . \n tmre was used \n to quantify membrane potential ; values are relative to a no - compound \n control value of 100 . \n in contrast to compound 1 , 2 did \n not \n significantly reduce the mitochondrial potential over a 24 h period \n either when irradiated or kept in the dark . \n irradiation did reduce \n cell viability to 70% 12% at 2 h and 59% 4% after 24 \n h , but this occurred without a significant decrease in the relative \n mitochondrial potential ( figure 3e ) , indicating \n that 2 does not act through inhibition of mitochondrial \n function . \n these results strongly suggest that mitochondrial failure \n plays a role in the dark toxicity of 1 , and the lack \n of mitochondrial localization and inhibition may explain the comparatively \n low dark toxicity of 2 . \n most compounds used for pdt \n that generate singlet oxygen induce apoptosis . given the different \n cellular localization properties and time profiles for cell death \n induced by 1 and 2 , the mechanism of cell \n death was investigated . \n indicators of apoptotic cell death ( activation \n of parp and caspase 3 through proteolysis ) were determined in hl60 \n cells treated with either 1 or 2 ( figure 4a ) . \n the known apoptotic - inducing compounds , cisplatin \n and doxorubicin , were run in parallel . \n compound 1 induced \n the proteolytic activation of both parp and caspase 3 within 2 h of \n irradiation ( figure 4a ) . in the absence of \n light , \n parp and caspase 3 were observed with 1 , but only \n after 18 and 24 h , and to a lesser degree . in the absence of irradiation , \n exposure \n of 2 to light also induced parp cleavage as early as \n 2 h , but unlike 1 , increasing amounts of cleaved parp \n were observed over the course of 24 h ( figure 4a ) . \n the increase in the level of activated caspase 3 also occurred \n on a slower time scale than parp cleavage , with the protein initially \n observed at 18 and 24 h. this suggests that the irradiated samples \n undergo apoptosis that is not primarily signaled through caspase 3 . \n conversely , cells treated with 2 and protected from light \n did not display parp or caspase 3 cleavage , which is consistent with \n viability measurements indicating no cytotoxicity in the absence of \n light . \n the level of procaspase 3 also did not change over 24 h , further \n confirming a lack of cytotoxicity under these conditions . \n since 2 showed strong parp induction without significant \n caspase 3 activation , as compared to 1 and cisplatin \n or doxorubicin , a mechanism of cell death through necrosis was explored . \n the level of an alternate 55 kda parp fragment was determined by immunoblot \n as a marker for necrosis ( see figure s6 in the \n supporting information ) , with 10% ( v / v ) ethanol used as a positive \n control . \n exposure of hl60 cells to 1 produced this fragment at significant levels both when protected \n from light and when irradiated , which is consistent with necrosis . \n cisplatin and doxorubicin also produced this cleavage product , indicating \n that some cells had progressed into necrosis ( figure s6 in the supporting information ) . \n in contrast , cells \n exposed to 2 , both in the presence and absence of irradiation , \n produced a lower level of the 55 kda parp fragment , similar to the \n untreated cells , suggesting necrosis is not a significant cell death \n pathway for this compound . \n ( a ) western blot analysis of cleaved parp , caspase \n 3 , and pro - caspase \n 3 . \n ( b ) agarose gel electrophoresis \n of genomic dna harvested from hl60 cells after treatment with various \n agents . \n lanes 1 and 9 , ladder ; lane 2 , no compound control ; lane 3 , \n 10% etoh , 24 h ( necrosis control ) ; lane 4 , cisplatin , 24 h ( apoptosis \n control ) ; lane 5 , 1 , irradiated , 8 h ; lane 6 , 1 , in the dark , 8 h ; lane 7 , 2 , irradiated , 24 h ; and \n lane 8 , 2 , in the dark , 24 h. to support the assessment of the disparate mechanisms of \n cell death \n induced by 1 and 2 , the degradation pattern \n of genomic dna was investigated . \n dna laddering is observed as a result \n of dna fragmentation stemming from the executionary phase of apoptosis . \n in contrast , necrotic cell death lacks this characteristic laddering \n effect , allowing differentiation between these two mechanisms . \n hl60 \n cells were exposed to the ru(ii ) complexes , cisplatin , and 10% ethanol , \n followed by genomic dna isolation and resolution by gel electrophoresis . \n as expected , the apoptosis inducer , cisplatin , initiated dna fragmentation , \n resulting in a laddering pattern on the gel ( figure 4b ) . \n this laddering was absent in the cells treated with ethanol \n and compounds 1 and 2 in the dark . \n however , \n both compounds 1 and 2 displayed similar \n laddering patterns as cisplatin when irradiated , suggesting apoptosis \n is a contributing cell death pathway for both compounds when irradiated . \n in contrast , given the cytotoxicity of 1 in the dark \n and the presence of the 55 kda parp fragment , it appears that necrotic \n cell death is a significant pathway for 1 . for compound 2 , the absence of dna laddering is a result of the lack of \n cytotoxicity of the negatively charged compound . \n flow cytometry \n was also employed to further corroborate the assessment \n of the mechanism of cell death using annexin v / propidium iodide ( pi ) \n or annexin v / hoechst staining to differentiate apoptotic versus necrotic \n cell death . \n fluorescent annexin v conjugates recognize the translocation \n of phosphatidylserine to the outer leaflet of the cell membrane during \n apoptosis . \n pi and hoechst were used as nuclear stains to distinguish \n between live and dead cells ; while pi is most commonly used , hoechst \n was also applied , because of spectral interference of 1 and 2 with pi . \n the presence of significant populations \n of pi positive cells in the absence of annexin v staining for 1 both in light and in the dark demonstrate that necrosis \n is a significant cell death pathway for this compound ( see figures s7 and s8 in the supporting information ; 5% and 14% , respectively , at 2 h ) . in contrast , compound 2 produced large populations of annexin v positive cells ( 55% \n at 24 h ) , showing an apoptotic pathway ( see figures \n s9 and s10 in the supporting information ) . \n similarly , hoechst \n staining was consistent with necrosis as a contributing pathway for 1 in the dark and in light , while 2 induced apoptosis . \n the goal of phototherapy is to achieve cell death \n in cancerous \n or abnormal tissues that are irradiated , while protecting healthy \n tissues that are not exposed to light . as a result , \n the compounds \n developed for this application should possess large therapeutic \n windows where the toxicity in the dark is minimized . \n this \n has proven to be challenging for many inorganic agents developed for \n pdt applications . \n the rational design of new and more efficacious \n compounds would be facilitated by ( 1 ) a better understanding of the \n mechanisms of action that induce dark toxicity for promising pdt agents , \n and ( 2 ) the identification of chemical features that eliminate dark \n toxicity . \n the investigations of the biological activities of these \n two simple ru(ii ) complexes provide guidance for both approaches toward \n improved pdt agents . while compound 1 demonstrated \n notable potency when \n irradiated ( 0.0750.35 m , depending on cell line ) , the \n toxicity of the compound in the dark ( 0.62 to 3.75 m ) reduces \n its potential as a pdt agent . \n imaging studies showed both mitochondrial \n and lysosomal localization , and assessment of mitochondrial potential \n indicated that 1 immediately inhibits mitochondrial function . \n however , the disconnect between the time dependence of the inhibition \n of mitochondrial function and the reduction in cell viability in the \n dark shows that disruption of mitochondrial membrane potential does \n not lead to rapid cell death . \n it is possible that the cells treated \n with 1 and kept in the dark survive for several hours , \n despite the complete abrogation of mitochondrial function , due to \n the warburg effect , where cancer cells exhibit a reduced reliance \n on oxidative phosphorylation and increased dependence on glycolysis \n for energy production . \n the obliteration of mitochondrial function would explain the observation \n of necrotic cell death in this case . in \n contrast , \n when 1 is irradiated , it appears that a combination \n of necrotic and apoptotic pathways are activated . \n cell death is so \n rapid ( with most cells undergoing death at 2 h , as indicated by trypan \n blue staining ) that necrosis is likely to be a primary pathway for \n a majority of cells . \n the breakdown of membrane integrity is apparent \n not only for the plasma membrane , but also for the membranes of organelles , \n as cells treated with 1 and irradiated showed nonspecific \n nuclear localization of both mitotracker and lysotracker . \n the high \n dark toxicity , mitochondrial localization , and induction of necrotic \n death pathways may reduce the potential of compounds structurally \n similar to 1 for pdt . \n in marked contrast , compound 2 was found to possess \n several features that encourage further exploration of derivatives \n or similar compounds . \n while uptake was low , it was comparable to that \n of cisplatin , despite the overall charge of 4 . \n most importantly , 2 exhibited ic50 values on the order of 3.317.3 \n m when exposed to light , with no observed toxicity in the dark . \n a slight increase in intracellular accumulation was observed upon \n irradiation , possibly due to induction of plasma membrane damage that \n facilitated compound uptake . \n once inside the cell , the compound remained \n in the cytosol , with no observable localization to the mitochondria \n or inhibition of mitochondrial function . \n furthermore , the light - induced \n cell death mediated by 2 occurred by apoptosis , in contrast \n to the mitochondrial targeting of 1 , which resulted in \n necrosis . \n one possibility to explore is that pdt compounds that avoid \n mitochondrial localization will exhibit lower toxicity in the absence \n of light than those that associate with the mitochondria . \n it is anticipated \n that structural modifications that result in a modest increase in \n cellular uptake could sufficiently drive down potency to make improved \n derivatives of 2 that maintain large therapeutic windows . \n given the high binding affinities of most ru(ii ) complexes for \n dna ( kb > 10 ) , it was previously \n believed that the compounds would preferentially localize in the nucleus , \n and indeed several do . \n however , recent fluorescence and electron microscopy \n studies have shown localization of several ru(ii ) compounds in the \n mitochondria , suggesting it is a common target . \n other reports indicate membrane accumulation and disruption , along with apoptosis pathways that are mediated by the mitochondria . gasser and co \n - workers have shown in a recent report that this mitochondrial \n localization was , in fact , required for cytotoxicity for a lead compound \n in a structure activity relationship study of a family of ru(ii ) \n polypyridyl complexes . while mitochondrial \n accumulation results in cytotoxicity , this \n mechanism is not likely to be compatible with a pdt - type approach \n where such redox - active compounds are required to be essentially nontoxic \n in the absence of photons . alternatively , \n targeting moieties such \n as nuclear localization signals can be conjugated to the coordinated \n ligands to affect the affinity of the complex for biological molecules \n and regulate cellular uptake and subcellulation localization . \n this approach requires significant \n chemical modifications , and the targeting often fails to increase \n cytotoxicity . while microscopy is a powerful tool to assess \n compound localization , \n imaging experiments can cause relocalization of compounds that induce \n production of o2 or perform other photochemical \n reactions when exposed to light . \n previous \n studies on porphyrins used for pdt applications also have demonstrated \n this phenomenon , including uptake and \n relocalization of an anionic tetrasulfonated porphyrin . \n for this reason , it is important to perform \n imaging using a minimum of light exposure , and to probe for compound \n relocalization by comparing to conditions where the treated cells \n have been exposed to significant light doses . \n this current study \n shows that simple ligand modifications produce \n complexes with divergent physical properties and , correspondingly , \n different biological activities . \n compound 1 , despite \n its high dna affinity , localizes to the mitochondria and induces rapid \n membrane depolarization and necrotic cell death . \n it is possible that \n this will be a common problem for compounds containing the bathophenanthroline \n ligand . \n compound 2 , despite its overall charge of 4 , \n is taken up into cancer cells to a sufficient degree to mediate light - induced \n cell death through an apoptotic pathway . \n the absence of mitochondrial \n localization may be the factor that eliminates the dark toxicity of \n this ru(ii ) complex . \n the incorporation of the sulfonic acids into \n the ligands is likely responsible for the alteration in subcellular \n localization , suggesting a possible general approach to reducing dark \n toxicity for other ru(ii ) complexes developed for applications in \n phototherapy . \n ru(bathophenanthroline)3 ( 1 ) and ru(bathophenanthroline disulfonate)3 ( 2 ) were synthesized using previously established \n procedures . \n all cell lines were purchased \n from atcc . cell culture media , heat - inactivated \n fetal bovine serum ( fbs ) , 420% tris - glycine precast gels , \n dulbecco s phosphate buffered saline ( dpbs ) , penicillin - streptomycin \n solution ( pen - strep ) , and 0.4% trypan blue solution were from invitrogen . \n 35 mm wide , 4-compartment cellview cell culture dishes were obtained \n from usa scientific . \n hoechst 33342 , \n lysotracker green dnd-26 and mitotracker green fm were purchased from \n invitrogen . \n propidium iodide ( pi ) and fitc - annexin v were obtained \n from bd science . \n antibodies for parp-1 , procaspase 3 , and \n gapdh were from santa cruz biotechnology , inc . \n compounds \n were mixed with 40 \n g / ml puc19 plasmid dna in 10 mm potassium phosphate buffer , \n ph 7.4 . to determine the effect of light \n , samples were irradiated \n with light ( > 400 nm ) from a 200 w light source for total light \n doses \n of 40 j / cm . \n single- and double - strand dna break controls \n were prepared , and the dna samples were resolved on agarose gels , \n as described previously . in brief , \n samples were resolved on a 1% agarose gels prepared in tris - acetate \n buffer with 0.3 g of plasmid / lane . \n the gels were stained with \n 0.5 g / ml ethidium bromide in tris - acetate buffer at room temperature \n for 40 min , destained with tris - acetate buffer , and imaged on a chemidoc \n mp system ( bio - rad ) . \n human alveolar adenocarcinoma \n cell line a549 , human promyelocytic leukemia cell line hl60 , and human \n t lymphocyte cell line jurkat cells were maintained in media supplemented \n with 10% fbs and 50 u / ml pen - strep at 37 c with 5% co2 , with dmem used for a549 cells and imdm and rpmi 1640 used for hl60 \n and jurkat cells , respectively . \n cells were assayed in opti - mem supplemented \n with 1% serum supreme and 50 u / ml pen - strep and seeded into 96 well \n plates at a density of 1.5 10 cells / well for a549 , \n 2 10 cells / well for hl60 , and 1 10 cells / well for jurkat followed by a 6 h incubation at 37 c , \n 5% co2 . \n cells were then dosed with serial dilutions of \n compound and incubated for 18 h. they were then irradiated with 7 \n j / cm light ( > 400 nm ) in 30 s pulses or kept in the \n dark . \n cell viability was determined 72 h later by measuring the conversion \n of resazurin to resorufin , using a spectrafluor \n plus plate reader ( tecan ) . \n a549 cells were seeded in opti - mem with \n 1% serum supreme at a concentration \n of 2 10 cells / ml in 25 cm cell culture \n flasks and incubated overnight . \n a concentration of 5 m of 1 and 20 m of 2 were added to the cells \n and incubated for 2 or 24 h protected from light ; the concentration \n of 20 m was selected for 2 to correspond with \n the ic50 of the complex when irradiated with light . \n a concentration \n of 5 m was used for 1 for compatibility with fluorescent \n imaging . \n after 2 and 24 h the media was removed , cells were washed \n twice with dpbs , trypsinized , and collected by centrifugation at 125 \n g for 4 min . \n the cells were resuspended in \n opti - mem , filtered through 40-m cell strainers , and analyzed \n on a facscalibur ( becton dickenson ) with an excitation wavelength \n of 488 nm and emission measured at 650 nm . \n this instrument averages the fluorescence of three separate \n images to greatly reduce out - of - plane fluorescence . \n a549 cells were \n seeded in 35 mm , four - compartment cellview culture dishes at a density \n of 5 10 cells per compartment in a 500 l \n volume and incubated for 24 h in opti - mem containing 1% fbs , followed \n by the addition of 5 m 1 or 20 m 2 and time points measured at 2 , 8 , 18 , or 24 h. media was \n removed at each time point , cells rinsed with dpbs , and incubated \n in opti - mem with 16 m hoechst 33342 , 16 m hoechst and \n 0.15 m lysotracker green dnd-26 , or 16 m hoechst and \n 0.2 m mitotracker green fm . \n cells were incubated for 30 min \n then washed three times with dpbs and imaged at 50 magnification \n using an apotome structured illumination fluorescent microscope ( carl \n zeiss ag ) . \n a549 cells \n were seeded at 2 10 cells / well in 24-well plates \n and incubated for 18 h , followed by the addition of 5 m 1 or 20 m 2 . \n they were incubated for an \n additional 8 h , and then irradiated with light as described for cell \n cytotoxicity measurements or kept in the dark . \n the cells were incubated \n for an additional 2 , 8 , 18 , or 24 h , washed with dpbs , followed by \n the addition of 0.5 m tmre in opti - mem , and incubated for 30 \n min . \n the change \n in tmre signal was determined by the difference in fluorescence between \n compound - treated cells in the presence and absence of tmre . \n fluorescence \n was measured at 10 magnification using an apotome microscope \n in normal fluorescence mode . \n samples dosed \n with 1 and 2 were compared to untreated \n a549 cells ( n = 3 ) to give the relative mitochondrial \n potential . \n hl60 cells were \n seeded in opti - mem at a density of 1 10 cells / ml \n and incubated for 2 h. the cells were then dosed 1 or 2 , incubated for 8 h , irritated as above or protected from \n light , and incubated for 2 , 8 , 18 , 24 , or 48 h. as hl60 cells grow \n in suspension , trypan blue staining was employed in place of resazurin \n to simplify and accelerate cell viability analysis . at each time point , \n a 10 l cell suspension was mixed with an equal amount of trypan \n blue solution and cell viability determined by manual counting with \n a hemocytometer . \n hl60 cells were cultured , \n dosed with 1 and 2 , then irradiated as above \n or protected from light . \n cells were harvested 0 , 2 , 8 , 18 , or 24 h \n after treatment , pelleted by centrifugation at 124 g for 5 min , washed twice with dpbs , and lysed in ripa buffer \n supplemented with 5 mm sodium pyrophosphate for 15 min on ice . \n the \n insoluble fraction was removed by centrifugation at 20,800 g for 10 min at 4 c . \n ten micrograms \n ( 10 g ) of protein was loaded onto 420% tris - glycine \n gels , followed by transfer to nitrocellulose membranes . \n after blocking \n with 5% nonfat milk in dpbs with 0.1% tween20 ( pbst ) for 1 h at room \n temperature , the membrane was immunoblotted with parp-1 at a 1:500 \n dilution , procaspase 3 , cleaved caspase 3 , and cleaved parp at 1:1,000 \n dilutions , or gapdh at a 1:2,000 dilution in 5% nonfat milk overnight \n at 4 c . \n immunoblots were washed with pbst for 10 min four times \n and incubated for 1 h with secondary antibodies at a 1:10,000 dilution \n for gapdh and at a 1:5,000 dilution for all other antibodies . \n detection \n was performed using clarity western ecl substrate and imaged with \n the chemidoc mp system . \n hl60 cells were cultured , \n dosed with 1 and 2 , then photoactivated \n or protected from light as detailed above . \n cells were harvested 2 \n h after photoactivation for 1 or after 24 h for 2 , pelleted by centrifugation at 124 g for 5 min , washed twice with dpbs , and prepared with an apoptotic \n dna - ladder kit as per manufacturer instructions . \n gel electrophoresis \n was carried out using a 1% agarose gel containing 0.5 g / ml \n ethidium bromide with 1 g dna loaded per lane and run for 90 \n min at 75 v. gel imaging was performed with the chemidoc mp system . \n cells were seeded in opti - mem at a density of 1 10 cells / ml in 25 cm cell culture flasks , cultured overnight , \n then dosed with 5 m 1 , 20 m 2 , or 20 m cisplatin . \n cells treated with ru complexes were \n incubated for 8 h , protected from light before irradiating or kept \n in the dark . \n cells were collected after 2 and 24 h by centrifugation \n at 124 g for 5 min . \n the cell media was transferred \n to separate 15-ml centrifuge tubes , and cells were washed twice with \n dpbs . \n two milliliters ( 2 ml ) of concentrated hno3 was added \n to media samples while cell pellets were resuspended in 5 ml of 20% \n ( v / v ) hno3 . \n all samples were heated at 110 c for \n 3 h. after digestion , the volume of all cell samples was adjusted \n to 5 ml and media samples were adjusted to 10 ml with deionized ( di ) \n water . \n the metal content was analyzed using a vista - pro ccd simultaneous \n inductively coupled plasma optical emission spectrometer ( varian , \n inc . ) with detection at 240.272 , 245.657 , and 267.876 nm for ruthenium \n and 214.424 , 217.468 , and 265.945 nm for platinum , with a replicate \n reading time of 60 s. yttrium ( 1 ppm ) in 1% nitric acid was employed \n as an internal standard . \n the percentage intracellular metal ratio \n in 10 cells was calculated by normalizing the metal amount \n to 10 cells , and then divided by the total metal amount \n in both media and cells . \n cells were cultured , \n dosed with 1 and 2 , then irradiated or protected \n from light . \n cells were harvested 2 and 24 h after treatment , pelleted \n by centrifugation at 124 g for 5 min , washed \n twice with dpbs , stained for 15 min with fitc - annexin v and pi or \n fitc - annexin v and hoechst 33342 , because of the interference between \n the emission of 1 and pi .", "role": "user" }, { "content": "compounds \n capable of light - triggered cytotoxicity are appealing \n potential therapeutics , because they can provide spatial and temporal \n control over cell killing to reduce side effects in cancer therapy . \n two simple homoleptic ru(ii ) polypyridyl complexes with almost - identical \n photophysical properties but radically different physiochemical properties \n were investigated as agents for photodynamic therapy ( pdt ) . \n the two \n complexes were identical , except for the incorporation of six sulfonic \n acids into the ligands of one complex , resulting in a compound carrying \n an overall 4 charge . \n the negatively charged compound exhibited \n significant light - mediated cytotoxicity , and , importantly , the negative \n charges resulted in radical alterations of the biological activity , \n compared to the positively charged analogue , including complete abrogation \n of toxicity in the dark . \n the charges also altered the subcellular \n localization properties , mechanism of action , and even the mechanism \n of cell death . \n the incorporation of negative charged ligands provides \n a simple chemical approach to modify the biological properties of \n light - activated ru(ii ) cytotoxic agents .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: parasites and animals - oncomelania hupensis hupensis \n snails infected with \n s. japonicum \n were purchased from the jiangsu institute of parasitic diseases , china . \n m. fortis \n in closed colonies , balb / c mice weighing 18 - 20 g each and new zealand white rabbits weighing 1 - 2 kg each were provided by the lab animal centre of central south university . \n the animal experiments were performed according to the protocols approved by the national animal care and use committee of china . \n serum preparation \n - blood samples from \n m. fortis \n and mice were collected using an eye - detachment technique , while the new zealand white rabbit blood samples were collected via heart puncture . \n the sera were separated by centrifugation and the sera of the rabbits and mice were then filtered ( millex \n\n gp filter unit , 0.22 m , millipore ) . \n schistosomula culture in vitro - \n cercariae were collected from infected \n o. hupensis \n snails as described previously ( \n salafsky et al . \n cercariae were washed four times with dulbecco s modified eagle s medium ( dmem ) ( gibco , usa ) containing 300 u / ml of penicillin g sodium and 300 g / ml of streptomycin . \n after centrifugation at 322 \n g \n for 10 min , schistosomula were transformed from cercariae by culturing in dmem containing 100 u / ml of penicillin g sodium and 100 g / ml of streptomycin at 37c , 5% co \n 2 \n with 25% new zealand white rabbit sera for 24 - 48 h. schistosomula were next cultured in 48-well cell plates with 100 20 schistosomula per well at 37c and 5% co \n 2 \n . \n the viability of the incubated schistosomula was assessed directly with light microscopy ( olympus co , tokyo , japan ) . \n dead schistosomula showed a loss of motility , an increase in opacity and occasionally gross morphological changes in shape or obvious lesions on the tegument . \n the viability assessment was ensured by methylene blue ( mb ) uptake testing after observation with the unstained schistosomula considered dead and the stained ones alive ( \n gold & flescher 2000 \n ) . \n screening of anti - schistosomula - associated protein \n - to obtain an effective schistosomula protein from \n m. fortis \n serum , molecular sieve chromatography was performed with a sephadex g50 column . \n chromatographic separations were performed by high - performance liquid chromatograph ( hplc ) ( waters company , usa ) . to confirm the anti - schistosome activity of hplc separations further , \n eluted peaks were pooled , desalted and lyophilised after the separations were gradient - eluted with 50 - 80% acetonitrile containing 0.1% trifluoroacetic acid . \n affinity chromatography \n - pre - swollen blue sepharose ff was mixed thoroughly and was poured into a chromatography column ( 3 cm i.d . \n 10 cm ) , which was then equilibrated thoroughly with solution a ( 0.05 m kh \n 2 \n po \n 4 \n , ph 7.0 ) . \n m. fortis \n serum was then diluted 1:1 ( v / v ) with solution a and was dialysed overnight against phosphate buffered saline ( pbs ) using dalton membranes ( 8,000 - 14,000 ) . \n after centrifugation at 8,050 \n g \n for 10 min , the supernatant was applied to the blue sepharose ff affinity chromatography column ( 4.8 cm 40 cm ) ( pharmacia ) at a flow rate of 2 ml / min . \n the column was eluted with solution a , followed by solution b ( 0.05 m kh \n 2 \n po \n 4 \n , 1.5 m kcl , ph 7.0 ) . \n the elution peak was pooled and dialysed overnight at 4c against pbs using dalton membranes ( 8,000 - 14,000 ) . \n finally , sodium dodecyl sulfate polyacrylamide gel electrophoresis ( sds - page ) analysis was conducted to ensure the purity of the \n m. fortis- \n albumin ( \n mf \n -albumin ) . \n anti - schistosomula experiment of mf - albumin in vitro - \n the inhibitory effects of \n mf- \n albumin on schistosomula were detected in vitro . \n after the live schistosomula were counted , samples corresponding to 1.25 m of \n mf- \n albumin were added and then incubated at 37c . \n m. fortis \n serum ( 40% , final concentration ) was used as the positive control and mice serum albumin was used as the negative control . \n schistosomula were cultivated for up to 96 h and the residual live schistosomula were counted and the mortality rate of schistosomula was calculated . \n the death rate of schistosomula was evaluated by the following formula : death rate % = ( total number of schistosomula - live number of schistosomula)/total number of schistosomula 100 . \n preparation of the conditioned medium - \n full - length \n mf \n -albumin genes ( genbank accession ay885264 ) were amplified from the total rna of \n m. fortis \n livers by reverse transcription polymerase chain reaction ( pcr ) using the following pcr primers : 5-tattggatccaccatgaggggcttgtttcgccgaga-3 and 5-tattctcgagagcaacaagttttggaccctctag-3 for \n mf \n -albumin and 5-tattggatccaccatgaggggtgtgtttcgccgag-3 and 5-tattctcgagagcaacaagttttggaccctctag-3 for mouse serum albumin ( \n ms \n -albumin ) . \n the generated dna fragment was cloned into plasmid pcdna3.1 ( + ) and was transferred into the \n escherichia coli \n strain dh5a . the recombinant plasmid ( pcdna3.1-albumin ) was sequenced and transiently transfected into 293 t cells using lipofectamine \n\n 2000 ( invitrogen , usa ) . \n twelve hours after dna transfection , the cells were cultured with 2 ml of serum - free dmem . \n supernatant ( conditioned medium ) from each pool was collected 36 h later and was tested at a concentration of 50% for its anti - schistosomula potential . \n protective experiment in mice with mf - albumin - \n thirty - six - week - old male balb / c mice were infected with 40 \n s. japonicum \n cercariae percutaneously and were randomly distributed into three groups containing 10 mice each : one blank pbs group , one \n ms \n -albumin control group and one \n mf \n -albumin test group . on the first day \n , the blank group was administered 0.3 ml of pbs , the control group was administered 0.3 ml of \n ms \n -albumin and the test group with 0.3 ml \n mf \n -albumin , all via intravenous injection . on the third and sixth days after the first injection \n s. japonicum \n adult worms in the hepatic and portomesenteric veins were recovered and counted . \n the number of eggs in the liver was estimated after the tissue was digested with 4% koh . \n the percentage changes in worm and egg burden were evaluated by the following formula : % change = ( mean number in infected controls - mean number in infected , treated mice)/mean number in infected control 100 . \n isolation of digestive tract excretions from schistosomula and adult worms \n - after schistosomula were harvested and washed three times with distilled water , they were placed in distilled water at room temperature for 20 min until the water became turbid with dark granular material from the parasites digestive tracts . \n these excretions were collected , adjusted to a ph of 4.9 with 0.1 m citrate and then centrifuged at 8,050 \n g \n for 10 min , after which the supernatant was collected , lyophilised , sealed and kept at -70c until further use ( \n chappell & dresden 1986 \n ) . \n immediately before use , the powder was dissolved with pbs and the protein content was measured using bovine serum albumin as a standard ( \n lowry et al . \n 1951 \n ) . \n on - site detection of mf - albumin in schistosomula \n - labelling of protein was performed with fluorescein isothiocyanate ( fitc ) according to a conventional protocol . \n fitc- \n mf- \n albumin was added to the schistosomula cultures and was incubated for 24 h and 48 h in vitro . \n fitc - labelled \n ms \n -albumin , fluorescein ( 30 g / ml ) and schistosomula without treatment were used as controls . to ascertain the site of fluorescence , \n schistosomula were collected after 24 h and 48 h of incubation , washed with 0.01 m pbs ( ph 7.0 ) three times and then were observed under an olympus fluorescence microscope . \n albumin digestion with digestive tract excretions \n in vitro - samples of 5 g/l \n mf- \n albumin and digestive tract excretions were incubated for 0 h , 1 h , 2 h , 4 h , 8 h , 16 h and 24 h , respectively , at 37c in citrate saline buffer solution , ph 4.0 , in the presence of 20 mm cysteine . after digestion , \n the reaction was terminated by 100 l of 0.5 m tris buffer ( ph 6.8 , 10% sds ) . \n twenty microlitres of digestion products and 10 l of loading buffer were run on 10% sds - page . \n statistical analysis - \n the differences between groups are presented as means standard deviation by one - way anova , using spss statistical software version 13.0 . \n schistosomula culture in vitro - \n the tails of cercariae were detached in dmem medium containing 25% fresh new zealand white rabbit serum for 24 - 48 h and the transformation rate remained greater than 90% with a less than 10% blank mortality rate . under light microscopy , the live schistosomula were very vivacious with semitransparent bodies , crawling unhindered on the bottom of culture plate , whereas the dead schistosomula showed ankylosis and immobility , increased opacity and obvious lesions on the tegument and they did not display mb staining ( \n fig . 1 \n ) . \n fig . 1 : micrographic images of schistosomula : dead schistosomula showed an increased opacity and obvious lesions in the tegumental outer membrane . \n the live schistosomula took up the dye and stained into blue - black with orbicular tegumental outer membrane , whereas the dead ones have not taken up the dye and unstained . \n a : schistosomula cultured in dulbecco s modified eagle s medium ( dmem ) without methylene blue ; b : live ( stained ) and dead ( unstained ) schistosomula after introduction of methylene blue into the dmem . \n identification of the protein associated with the inhibitory effects on schistosomula \n - the protein components of \n m. fortis \n serum were isolated by means of molecular sieve chromatography using a sephadex g50 column and hplc and the protein that induced resistance to schistosomula was acquired . \n the amino - terminal end was identified as dahkseiahr by protein sequencing and blast showed that it had 85% homology with mouse albumin and was named \n mf \n -albumin . at a protein concentration of 1 mg \n / ml , it induced a schistosomula mortality rate of 36.9% ( \n\n = 3.8594 , p \n < 0.05 ) in vitro , significantly greater than that of the negative control . \n albumin purification \n - blue sepharose ff was used to purify \n m. fortis \n and \n ms \n -albumin . \n there were two main peaks in the blue sepharose ff affinity chromatography ( data not shown ) . \n thus , it was accumulated and dialysed thoroughly with distilled water and then was placed in a -80c ultra - low temperature freezer for at least 4 h. the purity of the albumin was detected by sds - page and the result showed a single monomeric protein migration at approximately 66 kda ( \n fig \n the purified \n mf \n serum albumin was subjected to n - terminal amino acid sequencing for further verification . \n 2 : sodium dodecyl sulfate polyacrylamide gel electrophoresis of \n microtus fortis \n -albumin purified with blue sepharose ff affinity chromatography . \n lane m : broad range protein molecular weight marker ; 1 : \n m. fortis \n serum albumin purified . \n inhibitory effects of mf - albumin on schistosomula in vitro - \n to confirm the anti - schistosomula effects of \n mf \n -albumin , \n mf \n -albumin was added to cultured schistosomula at final concentrations of 0.31 , 0.62 , 1.25 , 2.5 , 5.0 , 10.0 and 20.0 mg / ml , with ms - albumin as a negative control and \n m. fortis \n serum as a positive control . \n in addition to the morphological changes of the dead schistosomula previously described , we also observed that there were one or two large blebs on the bodies of schistosomula during the period of \n mf \n -albumin treatment . \n the same change was observed in the positive control group , but no change was seen in the negative control group . \n most of the schistosomula in the negative control group were vivacious with semitransparent bodies and we observed them to be crawling on the bottom of the cell plate ( \n fig . \n the results of the morphological observations and schistosomula death rate data showed that \n mf \n -albumin had obvious inhibitory activity on schistosomula in vitro . \n our results showed that when the concentration of \n mf \n -albumin was 1.25 mg / ml , the death rate of schistosomula was 46.2% ( \n\n = 8.4703 , p \n this diagram showed that when the target protein concentration was equal or greater than 1.25 mg / ml , significant difference of schistosomula mortality rate was observed when compared with negative control and the dead rate of schistosomula raised markedly along with the rise of albumin concentration ( asterisk means p < 0.05 , compare with negative control ) . \n 3 : micrographic images of schistosomula cultured in 96 h : dead schistosomula showed an increased opacity and obvious lesions in the tegumental outer membrane . sometimes the blebs appeared in their tegumental outer membrane ( arrowed ) . \n a : schistosomula treated with 5 mg / ml \n microtus fortis \n ( \n mf \n ) -albumin ; b : schistosomula treated with 10 mg / ml \n mf \n -albumin ; c : schistosomula treated with represented 40% \n m. fortis \n serum ; d : schistosomula treated with 10 mg / ml mouse serum albumin . \n effects of conditioned media on schistosomula - \n to verify further the anti - schistosome effects of \n mf \n -albumin , conditioned medium containing \n mf \n -albumin or \n ms \n -albumin was added to schistosomula culture at a concentration of 50% . \n we observed and calculated the death rate of schistosomula at 96 h. our results showed the average schistosomula - killing rate of pcdna3.1/ \n mf \n -albumin conditioned medium was 38.7% , which was significantly higher than that of the negative control ( \n fig . \n fig . 5 : inhibitory effect of pcdna3.1/ \n microtus fortis \n conditioned medium on schistosomula after 96 h of incubation . a : schistosomula cultured with control pcdna3.1/mouse serum albumin ( \n ms \n -albumin ) conditioned medium ; b : schistosomula cultured with pcdna3.1/ \n m. fortis \n -albumin ( \n mf \n -albumin ) conditioned medium ; c : schistosomula cultured with pcdna3.1/amp conditioned medium ( negative control ) ; \n d : schistosomula cultured with 20% \n m. fortis \n sera ( positive control ) ( more dead schistosomula were observed in mf - albumin group than the negative control group in 96 h ) ; e : histogram of schistosomula mortality rate after incubation with conditioned medium ( p < 0.05 compared with negative control ) . results represented mean standard deviation for triplicates . \n anti - schistosome effects of mf - albumin in vivo \n - balb / c mice were randomly divided into three groups and were infected with \n s. japonicum \n cercariae . \n the resulting worm burden and number of lepg in balb / c mice treated with \n mf \n -albumin and \n ms \n -albumin are presented in table i. balb / c mice administered \n mf \n -albumin ( 15 mg / animal ) showed a significant worm burden reduction of 43.5% ( p < 0.05 ) compared with \n ms \n -albumin administration . \n moreover , we also observed a highly significant reduction in lepg in the \n mf \n -albumin treated group , which had a 48.1% ( p < 0.05 ) egg reduction . \n additionally , there were no differences in the numbers of egg granuloma observed between the controls ( data not shown ) . \n all of these results showed that \n mf \n -albumin possessed significant inhibitory activity on \n s. japonicum \n in vivo . \n tablethe anti - schistosome effect of \n microtus fortis ( mf \n ) -albumin on \n schistosoma japonicum \n cercariae infection in balb / c micegroupsworm burden ( mean se)worms reduction rate ( % ) lepg ( mean se)egg reduction rate ( % ) ppbs24.9 4.8 - 59.543 6.887- \n 6.256- > 0.05 \n mf \n -albumin13.9 2.643.530.900 3.29848.1 < 0.05worm burden rate and egg reduction rate were calculated relative to phosphate buffered saline ( pbs ) control . worm burden and liver eggs per gram ( lepg ) between pbs and mouse serum albumin ( \n worm burden and lepg of \n mf \n -albumin group were statistically different from those of pbs and \n ms \n -albumin groups ( p < 0.05 ) . \n worm burden rate and egg reduction rate were calculated relative to phosphate buffered saline ( pbs ) control . worm burden and liver eggs per gram ( lepg ) between pbs and mouse serum albumin ( \n worm burden and lepg of \n mf \n -albumin group were statistically different from those of pbs and \n ms \n -albumin groups ( p < 0.05 ) . \n detection of the deposit site of mf - albumin in schistosomula \n - schistosomula culture was incubated with fitc - labelled proteins or other control proteins . \n 6 \n , clearly showed that a fluorescent enrichment effect could be found in the gut lumen of schistosomula after incubation with fitc- \n mf \n -albumin for 24 h. after 48 h of incubation , the fluorescent enrichment effect became even stronger in the gut of schistosomula by fluorescence microscopy . \n however , there was no distinct fluorescence enrichment in schistosomula that were treated with the fitc - labelled \n ms \n -albumin , fluorescence and the blank control . \n fig . 6 : on - site of \n microtus fortis \n -albumin ( \n mf \n -albumin ) in schistosomula ( 24 h and 48 h post - treated ) . \n a1 : treated with fluorescein isothiocyanate ( fitc)-labelled \n mf \n -albumin after 24 h incubation ; a2 : treated with fitc - labelled \n mf \n -albumin after 48 h incubation ; b1 : treated with fitc - labelled mouse serum albumin ( \n ms \n -albumin ) after 24 h incubation ; b2 : treated with fitc - labelled \n ms \n -albumin after 48 h incubation ; c1 : fluorescein control after 24 h incubation ; c2 : fluorescein control after 48 h incubation ; d1 : blank control after 24 h incubation ; d2 : blank control after 48 h incubation . \n digestion of mf - albumin by digestive tract excretions from schistosomula \n - digestive tract excretions obtained from schistosomula were used to evaluate the ability of the digestion processes of \n m. fortis \n and mice albumin . \n the results showed that evident digestion could be observed when \n ms \n -albumin was incubated with digestive tract excretions of schistosomula for only 2 h , but no evident digestion of \n mf \n -albumin was detected until co - incubation for 24 h ( \n fig . \n fig . 7 : digestion of albumins by digestive tract excretions from schistosomula and adult worms . \n albumins \n microtus fortis \n and mice were incubated with digestive tract excretions from schistosomula and adult worms at 37c for 0 h , 1 h , 2 h , 4 h , 8 h , 16 h and 24 h , respectively . \n then the products were collected and tested by sodium dodecyl sulfate polyacrylamide gel electrophoresis . a : mouse serum albumin ( \n ms \n -albumin ) treated with distilled water ; b : \n ms \n -albumin treated with digestive tract excretions of schistosomula ; \n c : \n m. fortis \n -albumin ( \n mf \n -albumin ) treated with distilled water ; d : \n mf \n -albumin treated with digestive tract excretions of schistosomula . \n it is well known that \n m. fortis \n possesses natural resistance to \n s. japonicum and \n this characteristic could be steadily inherited . \n in addition , there can be variations in host resistance and parasite infectivity within a species or across different species ( \n schwarzenbach et al . \n however , the degree of resistance , which can be a plastic response to environmental variation in the natural population , is not well understood ( \n nagel et al . \n some studies have shown that environmental variation might increase ischaemia - modified albumin concentrations , which are a form of human serum albumin ( hsa ) ( \n bar - or et al . \n in addition , there has also been evidence of possible genetic variants of hsa , particularly structural changes at the n - terminus , which abolish transition metal binding ( \n sheat et al . \n therefore , we hypothesised that a genetic difference in albumin between \n m. fortis \n and mice would result in functional disparity between them , giving \n mf \n -albumin stronger anti - schistosome potential . \n laboratory studies have shown that \n m. fortis \n sera possessed anti - schistosome activity in vitro and in vivo . in the present investigation \n , we showed that extract of albumin from \n m. forti \n s sera exhibited anti - schistosomal activity both in vitro and in vivo . \n it is well known that serum albumin is the most abundant protein in the circulatory system and it possesses many important physiological functions that contribute to colloid osmotic blood pressure and aid in the transport , distribution and metabolism of many endogenous and exogenous substances . \n in addition , it was reported that albumin might play an important role in innate immunity ( \n fassi et al . \n giles and czuprynski ( 2003 ) \n showed that the albumin from several mammalian species had obvious inhibitory activity against \n blastomyces dermatitidis \n , while analbuminaemic rat serum failed to inhibit \n b. dermatitidis \n growth in vivo . \n ( 1992 ) \n indicated that maleylated hsa inhibited human immunodeficiency virus-1 infection and syncytia formation via binding to target cells with high affinity in vitro . \n recently , a series of studies suggested that fluid resuscitation with albumin was associated with higher mortality in patients ( \n schierhout & roberts 1998 \n , \n vincent et al \n all of these data indicated that , apart from the basic functions , albumin possesses an immunological effect related to the innate immunity of living things . \n m. fortis \n is a non - permissive host , the serum of which has been found to possess innate resistance to \n s. japonicum \n in vitro and in vivo . in this study , we isolated and purified \n m. fortis \n albumin . \n the inhibitory activity of the albumin to schistosomula was examined in vitro and in vivo . \n schistosomula culture treated with 1.25 mg / ml of \n mf \n -albumin caused a significantly higher schistosomula death rate when compared to the negative controls . \n the mortality rate of schistosomula approached 100% when the \n mf \n -albumin concentration was increased to 10 mg / ml . \n mf \n -albumin was administered to mice infected with cercariae by intravenous injection through the tail vein . \n after perfusion of the mice 42 d after infection , we found that mice injected with \n mf- \n albumin had 43.5% worm burden reduction and 48.1% lepg reduction compared with the control group . \n all of the results showed that \n mf \n -albumin possessed significant inhibitory activity against schistosomula . \n until now , the possible mechanism of \n mf \n -albumin anti - schistosomula had been unclear . \n it is well known that the definitive hosts of \n s. japonicum \n are widespread , infecting more than 40 mammalian species . \n however , \n m. fortis \n was shown to be the only non - permissive host of schistosome among mammalians in china ( \n li et al . \n when infected , the growth and development of \n s. japonicum \n are hindered and no adult worms and eggs could be found in both wild and laboratory - bred \n m. fortis \n ( \n he et al . \n fitc - labelled \n mf \n -albumin accumulated in the gut lumen of schistosomula after incubation for 24 h and 48 h. thus , \n mf \n -albumin must be transported intact and be metabolised slowly by schistosomula . \n we speculated that the anti - schistosomal activity would be accompanied by a series of proteins involved in signal transduction and protein trafficking ; these proteins might activate or strengthen the immune system and induce anti - schistosome effects during different stages of the schistosome life cycle in the vertebrate host . \n recent experimental evidence has shown that hormonal - like proteins can regulate a variety of cellular and physiological functions of schistosome in establishment , growth and reproduction , such as adrenal steroid hormones and testosterone ( \n barrabes et al . \n 2002 \n ) . in combination with the results of fitc - labelled \n mf \n -albumin accumulated in the intestines of schistosomula \n , it is possible that \n mf \n -albumin could influence the microenvironment or be indigested by the digestive tract excretions of schistosomula . \n thus , we presumed \n mf \n -albumin might play an important role in resistance to schistosome , by acting as a kind of a stressor . to explore further possible mechanisms of \n mf \n -albumin against \n s. japonicum \n , we collected digestive tract excretions of schistosomula to test their digestion differences from \n m. fortis \n and mice serum albumin in vitro . \n the results showed that \n mf \n -albumin was more difficult to be digested by the digestive tract excretions of schistosomula , compared with \n ms \n -albumin . when infected , the growth and development of \n s. japonicum \n were hindered and no adult worms or eggs could be found in either wild or laboratory - bred \n m. fortis \n ( \n he et al . \n therefore , we presumed that \n mf \n -albumin might be an important factor of the anti - schistosomula effects of \n m. fortis \n during \n s. japonicum \n development . in combination with the results of fitc - labelled \n mf \n -albumin accumulated in the intestines of \n s. japonicum \n , it also provided insight that digestive tract excretions could be used for rapid immunodiagnosis , even during early stages of infection . \n it is worthwhile to point out that when we studied the inhibitory effects of \n mf \n -albumin on schistosomula in vitro , we observed some large blebs appearing in the tegumental outer membrane . \n the appearance of these blebs was reported and was suggested to be the result of the action of complement ( \n mclaren et al . \n however , in our in vitro experimental system , there was no complement or the complement was inactivated . \n thus , we suggest that the presence of \n mf \n -albumin created an environment that was not suitable for the survival of schistosomula because of the obvious morphological changes to the tegument of the schistosomula . in summary , we screened proteins of \n m. fortis \n serum by molecular sieve chromatography , hplc and blue sepharose ff and identified \n mf \n -albumin as having anti - schistosomula effects , both in vitro and in vivo . \n we preliminarily explored the possible mechanism of \n mf \n -albumin anti - schistosomula and concluded that \n mf \n -albumin was an effective protein against schistosome .", "role": "user" }, { "content": "schistosomiasis is an endemic parasite disease and praziquantel is the only drug currently in use to control this disease . \n experimental and epidemiological evidence strongly suggests that \n microtus fortis \n ( \n mf \n ) is a naturally resistant vertebrate host of \n schistosoma japonicum \n . in the present study \n , we found that \n mf \n serum albumin ( \n mf \n -albumin ) and the conditioned medium of pcdna3.1- \n mf \n -albumin caused 46.2% and 38.7% schistosomula death rates in 96 h , respectively , which were significantly higher than that of the negative control ( p < 0.05 ) . \n we also found that mice injected with \n mf \n -albumin had a 43.5% reduction in worm burden and a 48.1% reduction in liver eggs per gram ( p < 0.05 ) in comparison to the control animals . to characterise the mechanisms involved in clearance \n , schistosomula were incubated with fluorescein isothiocyanate - labelled \n mf \n -albumin and fluorescent enrichment effects were found in the gut lumen of schistosomula after 48 h of incubation . \n next , digestive tract excretions from schistosomula were collected and the sensitivity of \n mf \n -albumin to digestive tract excretions was evaluated . \n the results indicated that schistosomula digestive tract excretions showed indigestibility of \n mf \n -albumin . \n the death of schistosomula could be partially attributed to the lack of digestion of \n mf \n -albumin by digestive tract excretions during the development of the schistosomula stage . \n therefore , these data indicate the potential of \n mf \n -albumin as one of the major selective forces for schistosomiasis .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: influenza a virus infects a wide variety of animals and , also , human hosts . among the avian influenza virus ( aiv ) subtypes , h9n2 virus has the potential to cause influenza pandemic and vaccination is a prevalent solution for this problem . \n the vaccine used for rapid interposition should be safe to use and highly effective , after administration ( 1 ) . \n aluminum salts and oil - based emulsions were previously used as adjuvant , to enhance the immunogenicity of inactivated influenza vaccines ( 2 ) . \n chitosan was introduced as an effective adjuvant for delivery of biological materials , such as drugs , and also , vaccines containing specially inactivated viral types , such as influenza , in several publications . \n chitosan adjuvant vaccines enhanced antibody titers against influenza , in comparison to vaccines without chitosan ( 3 , 4 ) . since the 1970s , the ecology of influenza viruses in birds has been better understood , when surveillance studies showed the enormous pool of viruses presenting in the feral bird population , particularly waterfowl , and the great variation in these viruses . at present , delivery - depot effect , or specific immune activation , \n are regarded as two mechanisms constituting the main core of all recently developed adjuvant systems . \n however , multiple kinds of adjuvant systems have been extended and approved , by preclinical methods , and several of them are useful for human beings . \n the first restrictions to the application of recent adjuvant systems for medicine concern the safety issues . however , investigation and research plans have decreased the toxicity of adjuvants , over the last 80 years . \n the safety barriers , presented by regulatory and liability issues , have continued to increase . in medicine , \n , the vaccines given to infants or children , today , heighten the safety concerns of vaccine adjuvants ( 1 ) . \n there are different methods by which adjuvants can improve the immune response against vaccines : a ) developing the immunogenicity of faint antigens ; b ) boosting the velocity and the length of the immune response ; c ) adjusting antibody avidity , specification , isotype , or subclass dissemination ; d ) stimulation of cytotoxic t lymphocyte response ; e ) increasing the induction of mucosal immunity ; f ) reducing the antigen volume in the vaccine , for lower costs . \n prophylaxis of influenza has been successfully used for more than 50 years for inactivated influenza vaccines \n . however , the results of presenting inactivated vaccines are less impressive in the aged population and are incapable to protect from influenza virus drift variants . \n chitosan is a polymer formed by the reaction between two different monomers , with units of more than one kind of glucosamine and n - acetyl glucosamine , taken from the sectional depolymerization and deacetylation of chitin . \n moreover , chitosan was found to represent immune adjuvant characteristics , by improving humoral and cell - mediated immune responses , followed by inducing vaccination ( 3 ) . \n this research is based on the optimization of chitosan concentration , for vaccine delivery , and , also , the evaluation of antigen loading capacity for vaccine preparation . \n chitosan nanoparticles ( cnp ) represent an interesting new platform for antigen delivery and a promising adjuvant candidate for h9n2 inactivated influenza vaccine . \n standard vaccine strain aiv ( a / chicken / iran/99/h9n2 ) was inoculated into 11-day - old specific pathogen - free ( spf ) eggs . \n the eggs were observed for 24 - 72 hours post inoculation , according to the international office of epizootics ( oie ) terrestrial manual ( 2008 ) . \n the amnion - allantois fluids of the inoculated eggs were collected and centrifuged at 1200 rpm , for 30 minutes ( 5 ) . \n the 50% of egg - infective dose ( eid50 ) was calculated by reed and munch method ( 1938 ) in 11-day - old embryonic spf eggs . \n hemagglutination assay ( ha ) was performed in v - bottom 96-well plates , with 1% chicken red blood cell , as described by burleson et al . \n the eid50 assay was carried out to measure the concentration of infectious influenza disease virus in a suspension , where every well is represented by an unique egg . \n each virus dilution is injected into replicate eggs , which are tested , after a period of incubation , for signs of virus growth . in case of influenza \n , we use the ha . the infectivity titer of a suspension of influenza disease virus is the number of infectious units of virus per unit volume , usually expressed per ml . \n an adaptation of the mathematical technique , devised by reed and muench ( 1938 ) , is used to calculate the dilution of the suspension of virus being tested that produced the end point . \n formaldehyde ( merck kgaa , darmstadt , germany ) solution was diluted in double - distilled water . \n the presence of virus will hold the red cells in a diffuse matrix and prevent them from settling out to the bottom of the well . \n the test is performed based on the inhibition of viral agglutination , by a specific antibody , and can be used for virus identification or antibodies measurement . \n louis , mo , usa ) were prepared according to a modified ionic gelation method . in this procedure , \n molecular weight of chitosan , in this study , was 220 kda , as specified by isometric methods . \n chitosan solution was prepared by dissolving 0.37 g of both low and medium molecular weights in 89.25 ml of acetic acid ( 0.1 m ) and heated at 37c , to dissolve . in the next step , \n the solution was filtered using pre - filter membranes under vacuum ( for excluding large particles ) . then \n , 160.78 ml of acetate sodium ( 0.1 m ) were added to the prepared solution . in this study , \n chitosan derivatives solution 0.5% and 1% ( deacetylated by 85% ) in 0.2 m glutamate buffer were used , for immune response activity . \n fleetingly , chitosan was dissolved in different concentrations ( 0.5% and 1% ) of acetic acid . for better dissolving and to form an unique complex with microorganisms , especially viruses , a ph 4.6 - 4.8 must be reached , by using naoh ( 7 ) . inactivated influenza virus was added to the prepared solution to reach a final concentration of 5 hemagglutination units ( hau ) ( suspension was gently stirred for 10 min ) . \n louis , mo , usa ) was added to suspension , in concentration of 0.01% , as an emulsifier . \n then , sodium tripolyphosphate ( tpp ) was separately dissolved in deionized water ( 80 ml ) , at a concentration of 0.1% w / v . \n nanoparticles were formed instantaneously , upon the drop - wise addition ( using pumping device ) of tpp solution to chitosan solution . \n the nanoparticle suspensions were gently stirred for 60 minutes , at room temperature , before characterization . \n the suspension was centrifuged at 10000 rpm , for 15 min , to remove the supernatant containing free aiv . \n the loading capacity of cnp were calculated by difference of supernatant ha titer and initial antigen loading . \n remaining pellets were dissolved in equal volume of citrate buffer [ 92 ml of sodium citrate ( 0.1 m ) in 8 ml of citric acid ( 0.1 m ) ] ( 3 , 7 ) . for vaccine - antibody response assay , \n different solutions of prepared vaccines ( 0.5% and 1% of chitosan ) were injected via intramuscular route in 21-day - old spf chickens . \n sera were collected every week after vaccination , for seven weeks , and measured by hemagglutination inhibition ( hi ) test . \n fifty spf broilers ( 21 days old ) were randomized into five groups of 10 chickens . \n before the chickens were introduced into the laboratory experimental facilities , they were tested for the major viral diseases , affecting chickens . \n the spf birds were maintained in air - filtered bio - security isolation units , with feed and water under controlled condition . \n group 1 ( with chitosan of low molecular weight and a concentration of 0.5% ) was vaccinated with cnp - aiv vaccine ( 0.2 ml / chicken ) ; group 2 ( with chitosan of low molecular weight and a concentration of 1% ) , group 3 ( with chitosan of medium molecular weight and a concentration of 0.5% ) and group 4 ( with chitosan of medium molecular weight and a concentration of 1% ) were vaccinated with the same volumes of vaccine , respectively . \n blood samples were collected on day 0 ( vaccination day ) and at the end of each week ( for 7 week ) post vaccination . \n , chicago , il , usa ) statistical package and correspondences evaluated using one - way anova tests . \n standard vaccine strain aiv ( a / chicken / iran/99/h9n2 ) was inoculated into 11-day - old specific pathogen - free ( spf ) eggs . \n the eggs were observed for 24 - 72 hours post inoculation , according to the international office of epizootics ( oie ) terrestrial manual ( 2008 ) . \n the amnion - allantois fluids of the inoculated eggs were collected and centrifuged at 1200 rpm , for 30 minutes ( 5 ) . \n the 50% of egg - infective dose ( eid50 ) was calculated by reed and munch method ( 1938 ) in 11-day - old embryonic spf eggs . \n hemagglutination assay ( ha ) was performed in v - bottom 96-well plates , with 1% chicken red blood cell , as described by burleson et al . \n the eid50 assay was carried out to measure the concentration of infectious influenza disease virus in a suspension , where every well is represented by an unique egg . \n each virus dilution is injected into replicate eggs , which are tested , after a period of incubation , for signs of virus growth . in case of influenza \n , we use the ha . the infectivity titer of a suspension of influenza disease virus is the number of infectious units of virus per unit volume , usually expressed per ml . \n an adaptation of the mathematical technique , devised by reed and muench ( 1938 ) , is used to calculate the dilution of the suspension of virus being tested that produced the end point . \n formaldehyde ( merck kgaa , darmstadt , germany ) solution was diluted in double - distilled water . \n the presence of virus will hold the red cells in a diffuse matrix and prevent them from settling out to the bottom of the well . \n the test is performed based on the inhibition of viral agglutination , by a specific antibody , and can be used for virus identification or antibodies measurement . \n louis , mo , usa ) were prepared according to a modified ionic gelation method . in this procedure , \n molecular weight of chitosan , in this study , was 220 kda , as specified by isometric methods . \n chitosan solution was prepared by dissolving 0.37 g of both low and medium molecular weights in 89.25 ml of acetic acid ( 0.1 m ) and heated at 37c , to dissolve . in the next step , \n the solution was filtered using pre - filter membranes under vacuum ( for excluding large particles ) . then \n , 160.78 ml of acetate sodium ( 0.1 m ) were added to the prepared solution . in this study , \n chitosan derivatives solution 0.5% and 1% ( deacetylated by 85% ) in 0.2 m glutamate buffer were used , for immune response activity . \n fleetingly , chitosan was dissolved in different concentrations ( 0.5% and 1% ) of acetic acid . for better dissolving and to form an unique complex with microorganisms , especially viruses , a ph 4.6 - 4.8 must be reached , by using naoh ( 7 ) . inactivated influenza virus was added to the prepared solution to reach a final concentration of 5 hemagglutination units ( hau ) ( suspension was gently stirred for 10 min ) . \n louis , mo , usa ) was added to suspension , in concentration of 0.01% , as an emulsifier . \n then , sodium tripolyphosphate ( tpp ) was separately dissolved in deionized water ( 80 ml ) , at a concentration of 0.1% w / v . \n nanoparticles were formed instantaneously , upon the drop - wise addition ( using pumping device ) of tpp solution to chitosan solution . \n the nanoparticle suspensions were gently stirred for 60 minutes , at room temperature , before characterization . \n the suspension was centrifuged at 10000 rpm , for 15 min , to remove the supernatant containing free aiv . \n the loading capacity of cnp were calculated by difference of supernatant ha titer and initial antigen loading . \n remaining pellets were dissolved in equal volume of citrate buffer [ 92 ml of sodium citrate ( 0.1 m ) in 8 ml of citric acid ( 0.1 m ) ] ( 3 , 7 ) . \n for vaccine - antibody response assay , different solutions of prepared vaccines ( 0.5% and 1% of chitosan ) were injected via intramuscular route in 21-day - old spf chickens . \n sera were collected every week after vaccination , for seven weeks , and measured by hemagglutination inhibition ( hi ) test . \n fifty spf broilers ( 21 days old ) were randomized into five groups of 10 chickens . before the chickens were introduced into the laboratory experimental facilities , they were tested for the major viral diseases , affecting chickens . \n the spf birds were maintained in air - filtered bio - security isolation units , with feed and water under controlled condition . \n group 1 ( with chitosan of low molecular weight and a concentration of 0.5% ) was vaccinated with cnp - aiv vaccine ( 0.2 ml / chicken ) ; group 2 ( with chitosan of low molecular weight and a concentration of 1% ) , group 3 ( with chitosan of medium molecular weight and a concentration of 0.5% ) and group 4 ( with chitosan of medium molecular weight and a concentration of 1% ) were vaccinated with the same volumes of vaccine , respectively . \n blood samples were collected on day 0 ( vaccination day ) and at the end of each week ( for 7 week ) post vaccination . \n all data were analyzed by using spss version 17 ( spss inc . , chicago , il , usa ) statistical package and correspondences evaluated using one - way anova tests . \n no respiratory or clinical complications were observed right after vaccination and in the weeks following vaccination . \n mean hi titers , in the different weeks of the study groups , are shown in figure 1 and table 1 . \n the mean hi titer for the group of chitosan with low molecular weight and a concentration of 1% was significantly higher than that of the other groups ( p < 0.05 ) \n . abbreviations : lcch , low molecular weight of chitosan ; mcch , medium molecular weight of chitosan . \n \n no respiratory or clinical complications were observed right after vaccination and in the weeks following vaccination . \n mean hi titers , in the different weeks of the study groups , are shown in figure 1 and table 1 . \n the mean hi titer for the group of chitosan with low molecular weight and a concentration of 1% was significantly higher than that of the other groups ( p < 0.05 ) \n . abbreviations : lcch , low molecular weight of chitosan ; mcch , medium molecular weight of chitosan . \n an efficient vaccine requires antigens for being presented to the immune system and it would be have maximal results if combined with suitable and premiere adjuvant , for long - term immunity , and also for increasing the immunogenicity . \n adjuvants that were utilized in boosting humoral and cellular immunity had a limited release from the injection site . on the other hand , the side effects of adjuvants , instead of swelling , \n tissue injury and also pain , are very important in histopathology for medicine development and vaccine protocol guidelines . \n meanwhile , residues of the adjuvant , especially mineral , at the injection site , caused multiple problems , like tissue damage and certain cases of necrosis , which could present carcinogenic transformation and decrease the standard commercial values ( 8 , 9 ) . \n loaded vaccines , the use of antigen was obviously reduced and the vaccine caused clearly much less inflammatory response , after inoculation . also , chitosan adjuvant has immune enhancing effects ( 10 , 11 ) . \n this research also paved the way for the introduction of the optimum concentration of chitosan for nanoparticle preparation and , also , realized an evaluation of the loading of aiv into cnp . \n although aluminum salts are used in the vaccine industry , for many years , however , research in this field , in human being , did not have high efficacy ( 12 ) . \n due to the ability of stimulating local factors , the mucosal affinity of adjuvant is the goal in vaccine preparation and administration . \n recently , scientists indicated the potential efficacy of cnp that were formulated by inactivated flu virus , for intranasal route administration ( 13 ) . \n ( 2008 ) ( 14 ) showed that chitosan - adjuvant formulations also significantly increased antibody titers against homologous and heterologous virus strains . \n chitosan , with an antigen increased proliferation of antigen - specific cd4 t lymphocytes in mice spleen , as well as the quantities of th-1 lymphocytes , amplifying t - helper function and inducing cytotoxic t - lymphocytes . \n ghendon et al . , ( 2008 ) ( 14 , 15 ) represented that using low and high molecular weights of chitosan , with inactivated influenza , enhanced the activity of cytotoxic natural killer ( nk ) lymphocytes \n . this process could be more effective when the nanoparticles used low molecular weights , by increasing cd3 , cd3/nk and cd25 t - lymphocytes ( 16 ) .", "role": "user" }, { "content": "background : influenza a is a virus that affects a wide range of animals and also human beings . \n avian influenza virus ( aiv ) subtype h9n2 has the potential to create influenza pandemic and vaccination is a common solution for this problem . \n the vaccine , used for rapid intervention , should be safe to use and highly effective , after a single administration . \n chitosan nanoparticles ( cnp ) have already been recommended as a new adjuvant for inactivated aiv h9n2 vaccine immunization.objectives:this study aimed at the evaluation and better understanding of optimum concentration of cnp preparations and also , assessment of loading capacity of aiv into cnp , as an adjuvant in specific pathogen - free ( spf ) chickens.materials and methods : for measurement of vaccine - antibody response , different types of cnp were injected intramuscularly , in a single dose , to 21-day - old specific pathogen - free chickens . \n chickens were monitored for the efficacy of the nanoparticles and , also , their immune response , during a follow up of 7 weeks , by using hemagglutination - inhibition ( hi ) test . \n the cnp were prepared according to modified ionic gelation method and inactivated antigen was loaded in four hemagglutinin units ( hau ) concentrations . \n loading capacity of nanoparticles was determined by hemagglutination ( ha ) method . inactivated a / h9n2 aiv was mixed with chitosan of low molecular weight.results:the cnp did not cause any mortality or side effects , when chickens were administered the prepared vaccine . \n the results strongly showed that this novel vaccine significantly enhances the immunogenicity of inactivated aiv , comparing with isa70 ( seppic , puteaux , france ) adjuvant that is used routinely in the razi serum and vaccine research and production institute , karaj , iran , to reduce isa70 s side effects.conclusions:the aiv loaded into cnp vaccines induce appropriate antibody titers , after a single immunization , while requiring a low dose of antigen . \n the cnp also represent an interesting new platform for antigen delivery and a promising adjuvant candidate for h9n2 inactivated influenza vaccine .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: homosexuality is rarely discussed publically as it is considered a taboo subject by both indian civil society and the government . \n since the decriminalization of homosexual intercourse between consenting adults , there has been an increased acceptance of homosexuality and msms have come out more openly with their problems . \n msms are one of the risk group targeted by national aids control organization ( naco ) to reduce human immunodeficiency virus ( hiv ) transmission . \n naco estimates that india is home to 2.5 million msms of which 100,000 are at high risk of contracting hiv due to multipartner and commercial sexual practices . \n their importance has also been stressed upon by the fact that former health minister dr . \n the minister also called for a relook into the impending bill to amend the immoral trafficking ( prevention ) act . interestingly , in most of the cases , it is bisexual behavior , which is commoner than isolated homosexual behavior . \n this bisexual behavior makes msm a bridge population for spread of sexually transmitted diseases ( stds ) and hiv . \n an interesting classification was given by humsafar trust , working for this community , where msm are classified into different categories based on identity , gender , behavior , and profession . \n based on identity they are categorized as gay , \" \" bisexuals , kothis , ( receptive partner ) or panthis \n transgender are those who are born males , but believe themselves to be females , also referred as \n there are msm who do not identify with queer labels but are homosexual or bisexual behaviorally . \n lastly , there are men who indulge in sex with men for cash or kind . \n apart from being important for hiv transmission , other issue is the pattern of stds in this group . \n . these are either community - based studies or from clinics catering specifically to high - risk population.[68 ] second , most of the present studies deal with their high - risk sexual behaviors and assessment of risks for developing stds due to such behavior . in contrast , large numbers of studies on std profile of msm are present from the western world . \n however , these can not be applied for indian population because of the difference in social and cultural behavior . \n henceforth , we carried out this study to delineate the clinical patterns of stds in msm coming to our suraksha clinic . \n the study was carried out in the suraksha clinic of a tertiary care hospital in a metropolitan city of india . \n we retrospectively analyzed the available data on msm who visited the suraksha clinic from 2004 to 2010 . \n the male patients were identified as msm either if they self reported to have homosexual or bisexual behavior or it was elicited after careful history taking . \n the diagnosis of the various types of stds was made clinically , substantiated by investigations . \n hiv screening and venereal disease research laboratory tests were carried out in all the patients . \n additional investigations , such as dark ground microscopy , tzanck smear , gram staining , and koh smears , were carried out wherever required . \n a total of 75 msm visited our suraksha clinic in six years from 2004 to 2010 . \n they constituted 11.4% of the total 660 patients visiting our suraksha clinic in these six years . \n the average age of the patients was 23 years , the youngest patient being 14 years of age and the oldest 49 years . \n the professions of the patients varied from manual laborers , salaried jobs , and businessmen . \n most of the patients ( 72% ) were unmarried and nearly 75% gave a history of promiscuous behavior . \n one - third msm admitted of indulging in only homosexual activities , whereas the other two - third indulged in bisexual acts . \n the type of predominant practice ( being an active or a passive partner ) and the prevalence of condom use could not be assessed in all the patients from the available records . \n of a total of 20 patients with syphilis infection , 8 had primary syphilis , 10 had secondary , and 2 patients were identified as latent syphilis during routine vdrl screening . \n condyloma acuminata ( ca ) was the next common std detected , being present in 26% ( 16 ) patients . \n of the 16 patients with ca , 8 patients are presented with penile warts , 6 are presented with perianal warts , and 2 are presented had both penile and perianal warts . \n all 6 patients with perianal warts gave history of being passive partners at some times , and thereby recipient of unprotected anal intercourse ( uai ) . \n 19% patients ( 14 out of 75 ) , presented with herpes genitalis , of which 8 patients had first episode of herpes genitalis , whereas six had recurrent herpes genitalis . \n gonococcal infection was present in 12% ( 9 of 75 ) patients , while five had non - gonococcal urethritis . \n three patients each had genital scabies and candidal balanoposthitis and two cases of chancroid were recorded . \n eight patients visited the clinic due to their fear of std and were found to have pearly penile papules , sentinel piles , and folliculitis . \n eight of 75 patients ( 11% ) had mixed infection at the time of presentation . \n two patients had ca along with primary syphilis and herpes genitalis , one patient had ca with primary syphilis , and one had ca with nongonococcal urethritis . \n one patient each had herpes genitalis with latent syphilis , herpes genitalis with scabies , and chancroid with scabies . \n all the patients with syphilis had reactive vdrl , the titres ranging from 1:8 to 1:256 . \n two patients with latent syphilis were identified during routine vdrl screening , one of them presented to us with herpes genitalis infection and the other was detected because of positivity in wife during routine antenatal check up . \n none of them identified themselves as sex worker , neither there were any transgenders in our study . \n the proportion of msm who were married was low as also reported by brahmam et al . \n a cross - sectional survey of 4597 self - identified msms from four high hiv prevalence states in south india reported 19.8% msm as bisexual , which constituted 15.9% of hiv seropositive population in that study group . \n thus , strategies focusing specifically on msm for control of stds and hiv are needed . \n the most common std recorded in our study was syphilis ( 27% ) followed closely by ca ( 21% ) and herpes genitalis ( 19% ) . \n studies from india carried out in clinics catering only msm have shown nearly 20% of msm to be affected by some stds , with syphilis , herpes , and condyloma acuminata as the most common stds affecting them.[68 ] brahmam et al . also reported a high prevalence of syphilis among \n kothis and hijras ( 15.8 and 13.6% , respectively ) but a low prevalence of gonorrhea ( < 1% ) and chlamydia ( 0.44% ) . \n studied the prevalence of stds among hiv seropositive indian msm and demonstrated a very low rate of syphilitic infections ( 5% ) . \n the proportion of cases with ca , herpes genitalis were high as in our study , nearing 30% . \n they reported a much higher number of cases with gonorrhea ( 23% ) as against our study . \n the results in western studies have been slightly different . in a study on msm attending the primary health clinic in boston , \n gonorrhea was diagnosed in 25% and syphilis seropositivity was found in 6.2% . in another study on male transvestites in jakarta , \n gonococcal test positivity was present in 17% and seropositivity to treponemal and nontreponemal test was present in 44% . \n similar rise in the prevalence of gonorrhea and syphilis has also been pointed out by truong et al . on comparison with the data on the std profiles of the heterosexual males who visited our clinic during the same 6 years period \n , it was seen that syphilis was four times more common in msm ( 6.25% in heterosexual males vs. 27% in msm ) and gonorrhea was three times more common in msm ( 4.1% in heterosexual males vs. 11% in msm ) . \n therefore , though the proportions of various stds among msm worldwide vary , a point common is the increased prevalence of gonorrhea and syphilis throughout as compared to stds in heterosexual males . to the best of our knowledge , \n comparative data of stds in msm and heterosexual males are not available in any previous indian studies . \n have reported significantly higher prevalence of hiv ( 14.5% in msw vs. 10.8% in non - msw ) and stds ( 17.5% in msw vs. 11.2% in non - msw ) in msm . \n the proportion of hiv seropositivity was not very high in our study , with only 3 patients ( 4% ) tested positive . \n it was less compared to 6.6% prevalence rate of hiv seropositivity among all the std patients visiting our suraksha clinic . \n the national prevalence rate of hiv seropositivity among msm is 7.4% and delhi is considered a high risk state with 11.7% msm showing hiv positivity . in a facility - based survey conducted in selected geographical areas of delhi , a hiv prevalence of 12.3% was reported . \n setia et al . have reported a combined estimate of hiv prevalence of 16.5% among msm in india from five different studies . \n gupta et al . have reported a significant association of hiv with genital ulcer disease among msm . \n hiv testing in the window period might be one of the reasons for low number of hiv positive cases in our study . \n second , we did not encounter any msm who was professionally homosexual , which could also explain low hiv positivity in our study . \n an interesting concept of seropositioning and serosorting has been put forward by truong et al . to explain the stabilization of hiv incidence over a period of 6 years . \n the concept of serosorting hypothesizes that msm are increasingly selecting seroconcordant partners when engaging in uai . \n seropositioning refers to the idea that as receptive uai has much higher risk of hiv acquisition compared to insertive uai ; hiv negative people are chosen as insertive partner . \n however , this does not apply to our population , as most of the patients are unaware of the hiv status of their partners . \n a careful history and examination is a very important tool in identifying an underlying std . \n most of the msm are socially shy in accepting their sexual behavior . but their identification is important as most of them are bisexual and promiscuous , thereby playing a role in spread of stds in vast number of partners . \n the profile of stds may also differ in msm , which makes it all the more important to identify them . \n therefore , it has been suggested that a rectal and pharyngeal swab should always be taken in all msms as these sites harbor silent infection . \n as this is a retrospective study of msm attending our suraksha clinic , proctoscopy , rectal , and pharyngeal swabs or other investigations were not done to detect various stds in asymptomatic patients , apart from screening done for hiv and syphilis . \n , the study highlights the profile of stds among male homosexuals or bisexuals attending the suraksha clinic of a tertiary care hospital and compared it with std profile of heterosexual males .", "role": "user" }, { "content": "background : men who have sex with men ( msms ) are a vulnerable population for spread of sexually transmitted diseases ( stds ) and human immunodeficiency virus ( hiv ) . \n apart from being important for hiv transmission , they tend to have a different distribution patterns of stds . \n few indian studies have looked into this aspect.objective:we retrospectively analyzed the available data on msm from suraksha clinic of a tertiary care hospital in a metropolitan city from 2004 to 2010.materials and methods : a total of 75 msm constituting 11.4% of the total 660 patients visited our suraksha clinic in these 6 years . \n the clinical and behavioral profile of each patient along with the pattern of stds was evaluated.observations and results:75% of the msms were promiscuous ; one - third of them indulging in only homosexual activities . \n syphilis was the most common std , followed by condyloma acuminata , herpes genitalis , and gonorrhea . \n on comparing the data on the std profiles of the heterosexual males , predisposition toward bacterial stds among msms was observed.conclusion:identification of msms is important as most of them are bisexual and promiscuous , thereby playing a role in spread of stds in vast number of partners . \n the profile of stds also differs in msm , which makes it all the more important to identify them .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: we begin by charting the history of the elucidation of the eukaryotic actin filament structure . \n actin , in its filamentous form ( f - actin ) , was first discovered by straub in the early 1940s as an integral component of muscle . \n twenty years later , hanson & lowy used the novel structural tool of the early 1960s - electron microscopy to show that f - actin , when polymerized in vitro , forms straight right - handed helices composed of 2 tightly intertwining strands . using myosin , which binds f - actin strongly in the absence of nucleotide ( corresponding to the rigor state of muscle ) , \n hugh huxley , who invented negative stain and spearheaded the early stages of structural electron microscopy , observed a unique arrow head pattern under the electron microscope . \n he concluded that the 2 strands forming the f - actin helical filament were in the same orientation and the filament therefore is polar . \n huxley 's myosin labeling technique for f - actin was exploited several years later to show that not only muscle cells but eukaryotic cells contain substantial amounts of f - actin , and pollard and colleagues in 1975 determined that the barbed and pointed ends of the filament , indicated by the arrow heads , related to the fast and slow growing ends of the filament . \n revolutionary work by toshio yanagida in the mid-1980s showed that actin filaments could be visualized with fluorescence microscopy by labeling with rhodamine - phalloidin , which binds tightly at the interface between 3 actin protomers . \n this methodology aided the characterization of a myriad of cellular proteins associated with actin , many of which regulate the ability of actin to polymerize or depolymerize . in 1990 , \n back - to - back publications of the actin monomer x - ray structure with the determination of its orientation within the filament , based on fitting of the monomer structure into x - ray fiber diffraction data , produced the \n , this model stood the test of time , with the near - atomic resolution details being slowly revealed as cryo - electron microscopy techniques improved . \n the current model was determined at 3.6 by the raunser group in 2016 . at the time of the holmes model \n , there was no reason to think that there would be more than a single filament architecture produced by actin - like sequences . \n the absolute conservation of eukaryotic actin filament structure likely arises from its role as an exceptionally highly connected hub where the universal - actin - pool is harnessed by several filament nucleating machineries to provide force and architecture to a wide variety of biological processes . \n essentially , once the eukaryotic actin filament was integrated into more than one biological process , a high degree of negative selection pressure restricted the actin sequence and structure to become frozen in time , since genetic drift favoring one biological process would have a negative impact on biological processes competing for actin . \n the concept of there being a single actin began to change in the early 1990s , when the homologous structure of the non - polymerizing 70-kda heat shock cognate protein was determined . \n subsequent bioinformatics analysis predicted that eukaryotic actin is related to a larger family of proteins including hexokinase , heat shock proteins and the bacterial proteins ftsa , mreb and stba / parm . \n similarity in these sequences was characterized by the conservation in amino acids participating in the atp - binding pocket , later known as the actin fold . \n yet it was not until 2001 that experimental evidence emerged that functional filament - forming actins exist in prokaryotes . \n three common classes of alps have been identified in bacteria , with a forth being the rare mamk forming the scaffold of the magnetosome in magnetobacteria . \n mrebs form single stranded protofilaments , which can be arranged as antiparallel pairs , are present in most rod - shaped prokaryotes and are involved establishing cell shape . \n ftsas contain a membrane inserting c - terminal amphipathic helix and recruit the cell division protein and tubulin homolog ftsz to the mid cell surface via ftsz 's c - terminal peptide . in concert both proteins \n lastly the plasmid segregating actins , the first characterized being parm from the r1 plasmid in e. coli ( ecparm ) . together these alps all contain the conserved actin fold , but often with very low overall sequence identities of below 20% . despite these large variations , all monomer structures solved to date by x - ray crystallography proved to be very similar . \n despite the structural similarity of the monomers , this did not prove true for the filament structures of parm 's , which turned out to have huge variations . in the initial paper describing ecparm \n , this filament was thought to be just a small variation of f - actin in its helical parameters . only after more extensive em reconstructions \n did it become apparent that the structure differed substantially , in being a left - handed helical filament as opposed to the right - handed f - actin ( fig . 1 ) . \n the evolutionary pressure that determined the handedness of the various biological filaments is not known at present . other parm 's investigated in the following years obtained by electron microscopy \n ( alfa from bacillus subtilis plasmid pbet131 ( bsparm ) , parm from staphylococcus aureus plasmid psk41 ( saparm ) ) showed that the helical parameters could differ even more substantially from f - actin . yet \n all these filaments ( ecparm , bsparm and saparm ) were still polar double stranded straight helices , like f - actin . \n figure 1.two views of the structures of filaments formed from the actin ( blue / orange ) and tubulin ( red / yellow ) folds . \n the actins are : the twisted single - stranded crenactin from the archaeon pyrobaculum calidifontis , \n caulobacter crescentus mreb filament formed from an antiparallel non - twisted pair of strands ( ccmreb ) , right - handed eukaryotic f - actin , left - handed escherichia coli parm from the r1 plasmid ( ecparm ) , \n clostridium tetani open nanotubules from the pe88 plasmid ( ctparm ) which are 2 antiparallel related copies of a parallel pair of strands and bacillus thuringiensis supercoiled antiparallel filaments and nanotubules from the pbmb67 plasmid ( btparm ) . \n the tubulins are : the eukaryotic microtubule , \n bacillus thuringiensis tubz from the pbtoxis plasmid ( bttubz ) and methanococcus jannaschii ftsz ( mjftsz ) . \n in addition the tubulin fold of btuba / b from prosthecobacter vanneervenii can form tubules comprised of 5 strands . \n two views of the structures of filaments formed from the actin ( blue / orange ) and tubulin ( red / yellow ) folds . \n the actins are : the twisted single - stranded crenactin from the archaeon pyrobaculum calidifontis , \n caulobacter crescentus mreb filament formed from an antiparallel non - twisted pair of strands ( ccmreb ) , right - handed eukaryotic f - actin , left - handed escherichia coli parm from the r1 plasmid ( ecparm ) , \n clostridium tetani open nanotubules from the pe88 plasmid ( ctparm ) which are 2 antiparallel related copies of a parallel pair of strands and bacillus thuringiensis supercoiled antiparallel filaments and nanotubules from the pbmb67 plasmid ( btparm ) . \n the tubulins are : the eukaryotic microtubule , \n bacillus thuringiensis tubz from the pbtoxis plasmid ( bttubz ) and methanococcus jannaschii ftsz ( mjftsz ) . \n in addition the tubulin fold of btuba / b from prosthecobacter vanneervenii can form tubules comprised of 5 strands . the first departure from the dogma that all parm 's formed \n only double stranded polar filaments came with the structure of alp12 ( ctparm ) from clostridium tetani , which segregates the pe88 plasmid encoding the lethal tetanus toxin . \n ctparm formed 4-stranded filaments ( 2 double strands ) arranged in an open cylinder separated by a wide cleft ( fig . 1 ) . \n subunits within a single - stranded protofilament associated through subdomain interactions that have parallels to all other known actin structures . at the macroscopic level , subdomain 3 to subdomain 4 inter - subunit connections are the constant feature that lead to relatively unidirectional protofilaments , despite the lack of conservation of inter - subunit interacting residues or their positions within the protein sequences . yet \n the 2 protofilaments formed a polar double - stranded filament through a completely different interface than observed for other actins . although each double strand is polar , the 2 double stranded filaments are paired in anti - parallel fashion , by 2 sheets of subdomain 3 to construct this novel open cylindrical architecture . more recently , \n parm from the pbmb67 plasmid in bacillus thuringiensis ( btparm ) was shown to form supercoiled , rather than straight filaments , which although double stranded , are anti - parallel rather than polar ( fig . 1 ) , a far departure from the construction of f - actin or any other parm . \n parm 's are plasmid segregation polymerizing actins that are linked to the parc regions of the plasmid dna via adaptor proteins ( parrs ) , which are specific to each individual parm / parc combination . within the parcmr system from e. coli ( ecparcmr ) , the parr / parc \n complex was shown to pair 2 or more filaments into randomly oriented bundles . in this system \n other forces , such as cellular crowding , also paired ecparm filaments in the absence of the associated molecules . \n the roles of the 2 competing effects ( direct binding of parr / parc versus osmotic pressure ) are difficult to separate , and hence their relative contribution is ambiguous . \n the parcmr system from bacillus thuringiensis ( btparcmr ) proved to be entirely different and more amenable to teasing apart each contribution . \n polymerization of btparm in the presence of btparr stimulated atp hydrolysis by btparm and formed a cylinder , comprised of 4 antiparallel strands , with inner and outer diameters of 57 and 145 , respectively , which is also formed in the presence of the btparr / parc complex ( fig . 1 ) . \n the structure of the btparm cylinder is composed of 2 interwoven supercoiled antiparallel filaments , which is the geometry of the btparm filament formed in the absence of parr and parc ( fig . 1 ) . here \n osmotic pressure originating from molecular crowding also paired filaments , but in this case they form rafts of individual supercoiled filaments arranged in parallel ( fig . 2 ) . \n thus in the case of btparcmr , the unique btparm nanotubule geometry requires a second component of the btparcmr system , parr or parr / parc . \n figure 2.in the presence of molecular crowding btparm formed rafts of filaments lying in parallel as observed by electron microscopy ( a ) . the filaments within the rafts were not paired into a cylinder , but were supercoiled single btparm filaments . \n ( b ) a projection image calculated from the supercoiled btparm filament model for comparison . \n pairing of filaments only occurs in the presence of parr or the parr / parc complex . \n in the presence of molecular crowding btparm formed rafts of filaments lying in parallel as observed by electron microscopy ( a ) . \n the filaments within the rafts were not paired into a cylinder , but were supercoiled single btparm filaments . \n ( b ) a projection image calculated from the supercoiled btparm filament model for comparison . \n pairing of filaments only occurs in the presence of parr or the parr / parc complex . \n eukaryotic actin filaments function as in a universal - pool - of - actin in which variety in actin - binding proteins is able to harness the same molecular polymerizing motor for many cellular processes . \n prokaryotic cells have developed different mechanisms for force generation . we have previously speculated that since bacteria have one - filament - one - function systems to create force through polymerization , they evolved distinct actin filaments to power specific functional processes within a single cell . \n this leads to several questions , firstly : are there more novel actin filament architectures out there ? given the vast numbers of uncharacterized parm 's that are known to exist from phylogenetic analyses , \n a more pertinent question is : how much further will these filaments differ from f - actin ? \n the answer to this question will likely be quickly revealed by cryo - electron microscopy , which in just a few years has become the structural biology tool of choice to study filament systems at near - atomic resolution , due to the improvements in electron microscopes , and in particular , direct electron detectors . \n finally , why did nature create so many different designs of actin - like filaments for plasmid segregation ? \n firstly , when 2 different plasmids reside in the same cell they will require different segregation systems to be faithfully inherited , hence , there may have been some positive evolutionary pressure for the parms to diversify . \n secondly , if evolution is truly deterministic , the filament structures of plasmid segregating actin - like filaments are likely related to the sizes of the individual plasmids to be segregated . \n that is a parm filament consisting of 4 strands ( like ctparm ) should be substantially stronger than a double stranded filament ( like ecparm ) , thus being able to push a bigger load . \n actin - like and tubulin - like filaments have transposable roles in biology . in prokaryotes , \n the tubulin - like protein ftsz assembles to form the cytokinetic ring , whereas this function is performed by f - actin in eukaryotes . in contrast , plasmid dna segregation is often orchestrated by parm actin - like proteins in bacteria , while chromosome dna segregation is choreographed by microtubules in eukaryotes . \n comparison of the known structures formed by the actin and tubulin folds , indicates that these basic building blocks are capable of forming extensive ranges of structures . \n both folds can form linear single stand filaments , twisted parallel pairs of filaments and tubules of various dimensions ( fig . 1 ) . \n the ability to alter a tubule 's dimensions , though changing the number of strands in the tubule , has been demonstrated through protein engineering to require relatively minor surface amino acid modifications in the case of barrels formed from -helices . \n thus , various diameters of tubules formed from the actin and tubulin folds are likely to have been sampled during evolution . \n whether a superior property is intrinsic to the actin fold for forming microfilament - like architectures , and to the tubulin fold for forming microtubule - like structures , or whether the use of these protein scaffolds were stochastic events that became entrenched in early eukaryotes remains open to debate . \n nevertheless , it is clear that the highly specialized filament geometries of microfilaments and microtubules are just 2 of many possibilities that are available in nature . \n the structures of f - actin and microtubules have been conserved over a billion years in eukaryotic cells . \n these structures are maintained through evolution by their interactions with large numbers of binding proteins , which have likely restricted their genetic drift and have allowed for the filament properties to be exploited by many cellular processes . in contrast , the one - filament - one - function design observed in many prokaryotic filaments has allowed for the adoption of a large variety of different filament structures . despite the many variances in filament structures , 2 features are preserved between all actins . \n firstly , the individual strands forming actin - like filaments ( protofilaments ) share grossly similar contacts , subdomains 1 and 3 from one monomer interact with subdomains 2 and subdomain 4 , respectively in the neighboring monomer in the protofilaments . \n secondly the nucleotide - binding site , which accepts gtp in some alps , acts as a conformational switch activated by polymerization controling atp / gtp hydrolysis and phosphate release . \n the atp / gtp switch ( converting the initially bound atp / gtp to adp - pi / gdp - pi and subsequently to adp / gdp ) acts as a timing mechanism , which coordinates the depolymerization of actin filaments via the conserved contacts in the protofilaments . \n the determinants as to whether filaments treadmill or are dynamically unstable largely remain unexplored for the alps . \n these activities will be impacted by : 1 ) off and on rates for each nucleotide - bound state of the monomers at both ends of the filament in its different nucleotide - bound forms ; 2 ) nucleotide - exchange rates in monomers and at filament ends ; 3 ) hydrolysis and phosphate - release rates on polymerization ; 4 ) concentrations of alps and nucleotides ; 5 ) higher order mechanisms for monomer association / dissociation with the filament ; and 6 ) binding partners . a simple case for an in vitro dynamically unstable \n filament occurs when the nucleotide - exchange rate for monomers is slow relative to the off rate for adp - bound protomers at the ends off the filament . on approaching steady state \n the filaments will depolymerize until the dissociated monomer pool has regenerated sufficient atp - bound monomers to support repolymerization , as is suggested for ecparm . in contrast , in vitro treadmilling for actin results from the 2 ends of the filament having different on and off rates for monomers . however , these are just 2 of the multitude of possible scenarios . \n mreb and ftsa filaments form non - helical protofilaments allowing them to present a consistent binding interface to the membrane , which would not be possible if they were twisted . \n parm 's , in contrast , form 2 to 4 ( and perhaps even more ) stranded helical filaments . as motor proteins , \n the helical design brings greater rigidity to the filaments and by increasing the number of strands expands the possibilities to segregate larger payloads . \n the functional requirement of any parm filament is to polymerize and interact with the specific plasmid through the complementary adaptor protein . \n this has allowed for far more latitude in exploring diverse filament architectures during evolution in comparison to filament systems that have more extensive interactions . \n thus , we speculate that the currently known actin filament structures are likely to be a fraction of the total diversity . \n ", "role": "user" }, { "content": "abstractfrom yeast to man , an evolutionary distance of 1.3 billion years , the f - actin filament structure has been conserved largely in line with the 94% sequence identity . \n the situation is entirely different in bacteria . in comparison to eukaryotic actins , \n the bacterial actin - like proteins ( alps ) show medium to low levels of sequence identity . \n this is extreme in the case of the parm family of proteins , which often display less than 20% identity . \n parms are plasmid segregation proteins that form the polymerizing motors that propel pairs of plasmids to the extremities of a cell prior to cell division , ensuring faithful inheritance of the plasmid . \n recently , exotic parm filament structures have been elucidated that show parm filament geometries are not limited to the standard polar pair of strands typified by actin . \n four - stranded non - polar parm filaments existing as open or closed nanotubules are found in clostridium tetani and bacillus thuringiensis , respectively . \n these diverse architectures indicate that the actin fold is capable of forming a large variety of filament morphologies , and that the conception of the actin filament has been heavily influenced by its conservation in eukaryotes . here , we review the history of the structure determination of the eukaryotic actin filament to give a sense of context for the discovery of the new parm filament structures . \n we describe the novel parm geometries and predict that even more complex actin - like filaments may exist in bacteria . finally , we compare the architectures of filaments arising from the actin and tubulin folds and conclude that the basic units possess similar properties that can each form a range of structures . \n thus , the use of the actin fold in microfilaments and the tubulin fold for microtubules likely arose from a wider range of filament possibilities , but became entrenched as those architectures in early eukaryotes .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: the environmental mold aspergillus fumigatus has become the most important airborne fungal pathogen of humans and is the second most frequent cause of systemic fungal infections.(1 ) in immunocompetent individuals , a. fumigatus can cause allergic reactions and noninvasive infections such as aspergilloma . \n however , in immunocompromised patients , this filamentous fungus is the main causative agent for life - threatening systemic infections named invasive aspergillosis ( ia ) . due to the improvements in transplant medicine and the therapy of hematological malignancies , \n the number of patients and thus the quantity of cases of ia has increased significantly in the last decades . \n specific diagnostics are still limited , as are the possibilities of therapeutic intervention , leading to a high mortality rate of 30 up to 90% for ia . \n infection is most commonly initiated by inhalation of airborne fungal spores ( conidia ) , which germinate in the lung alveoli and the formed hyphae start to grow invasively into the lung tissue . \n accumulating evidence suggests that a. fumigatus experiences hypoxia while growing in the lung . the oxygen partial pressure within the alveoli averages at 7.9 kpa ( 19.7 kpa in the atmosphere ) . in the surrounding tissue , \n it drops down to 24 kpa and in inflamed , necrotic areas even down to 0.13 kpa.(7 ) this can lead on one hand to a general reduction of phagocyte killing by the host , but on the other hand may promote the availability of free iron for the pathogen due to a decrease in redox potential.(8 ) additionally , nitric oxide ( no ) secreted by cells of the innate immune system , such as macrophages , acts as an inhibitor of cellular respiration and can cause so - called metabolic hypoxia.(9 ) thus , it is obvious that a. fumigatus has to adapt to low oxygen partial pressures during the infection . \n however , the fungus is considered to be an obligate aerobe and there is no clear evidence for either a fermentative metabolism or anaerobic respiration in a. fumigatus . \n nevertheless , some hints for ethanol fermentation during a. fumigatus infection were obtained in a murine model of ia(10 ) and a. fumigatus was shown to be capable of growing at low oxygen concentrations between 0.1% ( v / v ) and 0.5% ( v / v ) on agar plates.(11 ) under these conditions , the fungus faces the challenge to maintain the homeostasis of ergosterol and other cellular components , such as nad and heme , whose biosyntheses require molecular oxygen.(12 ) it is very likely that a. fumigatus has evolved mechanisms to cope with hypoxic growth conditions , but the details of the genetic and biochemical response of a. fumigatus to hypoxia has not been described in detail yet . \n recent discoveries in the nonpathogenic fission yeast schizosaccharomyces pombe(13 ) and the pathogenic encapsulated yeast cryptococcus neoformans(14,15 ) have led to the elucidation of a central regulatory pathway activated in response to hypoxia . here , o2 deprivation is indirectly sensed by measuring the level of ergosterol in the cell and is designated as the sterol response element binding protein ( srebp ) pathway . \n the er - bound transcription factor sre1p is activated upon hypoxia due to a decrease of the ergosterol content.(16 ) this allows sre1p to enter the nucleus and to activate genes essential for hypoxic growth.(17 ) the orthologue in a. fumigatus , srba was shown to be involved in hypoxic adaptation and virulence . \n an srba null mutant was unable to grow under hypoxic condition , was more sensitive to antifungal azoles , and was avirulent in a mouse infection model for ia.(6 ) target genes of the hypoxic regulator srba have not been studied in fully detail yet and global transcript profiling of the response of pathogenic fungi to low oxygen levels revealed a relative heterogeneous picture on hypoxia - responsive genes . in the pathogenic dimorphic \n yeast candida albicans , genes involved in fermentative metabolism were up - regulated , whereas genes regulating oxidative metabolism were down - regulated by hypoxia.(18 ) by contrast , the transcript levels of respiratory genes increased in c. neoformans during exposure to hypoxic growth conditions.(15 ) in a. fumigatus , transcriptional profiling of the srba null mutant revealed an srba - dependent regulation of genes involved in ergosterol biosynthesis , cell wall biosynthesis and transport processes.(6 ) in contrast , regulation of the hypoxic response on the level of proteins has not been elucidated in a. fumigatus or any other important human - pathogenic fungi yet . \n for this reason , we carried out global analysis of the change of the a. fumigatus proteome during hypoxia by two - dimensional gel fluorescence gel electrophoresis ( dige ) . because of the advantages in generating more reproducible , reliable and biologically homogeneous data sets , we used an oxygen - controlled chemostat(19 ) to cultivate a. fumigatus either under normoxic ( 21% o2 ) or under hypoxic ( 0.2% o2 ) conditions . by our proteome analysis \n we have shown that a. fumigatus compensates the depletion of molecular oxygen by the increased production of respiratory proteins and that a link exists between hypoxia and nitrosative stress . \n furthermore , for the first time we report that hypoxia is able to induce the activation of an otherwise silent secondary metabolite gene cluster in a. fumigatus . \n a. fumigatus was grown at 37 c in a glucose - limited oxystat ( chemostat with constant oxygen partial pressure [ po2 ] ) ( biostat b , braun , melsungen , germany ) in continuous culture ( 3 l working volume ) with constant stirring ( 550 rpm ) . \n the dissolved oxygen was measured with a high sensitive po2 electrode ( inpro6800/12/320 , mettler toledo , steinbach , germany ) connected to a data amplifier in a range of 0 10% po2 ( 8842698 braun , melsungen , germany ) . \n aerobic culture conditions were established ( 21% o2 atmospheric po2 ) by aerating with 0.8 l / min of air . under hypoxic culture conditions ( 0.2% o2 ) , \n the medium was continuously aerated with a mixture of nitrogen at a constant rate ( 0.65 l / min ) and air . \n the fungus was cultivated in aspergillus minimal medium ( amm ) as described in.(20 ) the medium contained 8.3 mm glucose as sole carbon source . \n the fermenter was inoculated with conidia to give a final concentration of 3 10 conidia / ml . \n fermentation started as a batch culture for 20 h under aerobic and 30 h under hypoxic ( 2% po2 for the initial batch cultivation ) conditions . \n subsequently , the dilution rate was set to 0.08 h ( correspond to a flow rate of 240 ml / h ) . \n samples were taken during fermentation to determine ph - value , glucose concentration ( biosen c - line , ekf diagnostic , barleben , germany ) and dry weight biomass ( hr73 halogen moisture analyzer , mettler toledo , steinbach , germany ) . \n after reaching the steady state ( constant biomass , constant po2 , constant ph , sixteen times exchange of culture volume , glucose - limitation ) the whole biomass was harvested , immediately rinsed with tap water , pressed to remove any liquid and the dried mycelium was immediately frozen in liquid nitrogen for further protein and rna extraction procedures or directly used for mitochondria isolation . \n proteins from 100 mg of this homogenate were extracted by the phenol extraction methanol / ammonium acetate precipitation method as described in(21 ) with slight modifications . \n after overnight precipitation the sample was centrifuged at 3200 g for 15 min at 4 c , the supernatant was removed and the pellet was rinsed three times with ice - cold acetone/0.3% ( wt / vol ) dithiothreitol . the pellet was air - dried and resuspended in 300 l lysis buffer ( 7 m urea , 2 m thiourea , 2% [ wt / vol ] chaps , 1% [ wt / vol ] zwittergent 310 and 20 mm tris ) , sonicated for 10 min in an ultrasonic bath and incubated for 1 h at room temperature.(22 ) the ph of the samples was adjusted to 8.5 by using 100 mm naoh . \n a. fumigatus mitochondria were isolated as described previously.(23 ) for 2-de , mitochondrial pellets ( 100 mg moist mass ) were resuspended in 400 l lysis buffer containing 2.8% ( wt / vol ) dodecylmaltoside . \n the protein concentration was determined according to the method of bradford(24 ) using the bio - rad protein assay ( bio - rad lab . , hertfordshire , u.k . ) . \n the dige ( difference in gel electrophoresis ) technique was used to analyze protein samples of mycelium cultivated under normoxic and hypoxic conditions and carried out as described.(25 ) samples from three independent normoxic and hypoxic cultivations were labeled with cydye minimal dyes according to the manufacturer s protocol with slight modifications ( ge healthcare bio - sciences , munich germany ) . \n 50 g of protein of each sample were labeled with 300 pmol of cydye dige flourophores ( dissolved in dimethyl formamide ) . \n samples obtained from normoxic ( 21% po2 ) and hypoxic ( 0.2% po2 ) conditions were labeled either with cy3 or cy5 . \n a pool of all six samples ( 3 normoxic and 3 hypoxic ) was prepared , labeled with cy2 , and used as a global internal standard . \n an equal volume of 4 sample buffer ( composition described above for the lysis buffer , plus 3.2% [ vol / vol ] servalyt ampholytes 310 [ serva electrophoresis , heidelberg , germany ] and 40 mm dtt ) was added . \n equal amounts of each of the three label preparations were combined and mixed with 50 g unlabeled protein extract of both growth conditions ( to increase the protein amount for subsequent mass spectrometry analysis ) and applied via anodic cup loading to ipgstrips ( ge healthcare bio - sciences ) . \n isoelectric focusing was carried out as described previously(22 ) using 24 cm ipg strips with a nonlinear ph range from ph 3 to 11 , which had been rehydrated overnight ( 7 m urea , 2 m thiourea , 2% [ wt / vol ] chaps ( 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonate ) , 1% [ wt / vol ] zwittergent 310 , \n 0.002% [ wt / vol ] bromophenol blue , 0.5% [ vol / vol ] ipg buffer , 1.2% [ vol / vol ] de - streak reagent [ ge healthcare bio - sciences ] ) . \n after separation by the first dimension the strips were equilibrated for 15 min in 10 ml of equilibration buffer ( 6 m urea , 30% [ vol / vol ] glycerol , 2% [ wt / vol ] sds ( sodium dodecyl sufate ) , 75 mm tris , 0.002% [ wt / vol ] bromophenol blue ) containing 1% ( wt / vol ) dithiothreitol and subsequently for 15 min in 10 ml of equilibration buffer containing 2.5% ( wt / vol ) iodoacetamide . for the separation of proteins in the second dimension , the ettan dalt system ( ge healthcare bio - sciences ) was used . \n sds polyacrylamide gels [ 1116% ( wt / vol ) ] of 1.0 mm thickness were casted with the a2deoptimizer ( biometra , gttingen , germany ) . \n separation conditions were as follows : 2 w / gel for 1 h followed by 15 w / gel for 4 h. proteins were visualized by analyzing the gels with a typhoon 9410 scanner ( ge healthcare bio - sciences ) using a resolution of 100 m . \n spot detection of cropped images was performed with the decyder software package ( version 6.5 ) . \n the following parameters were applied : detection sensitivity , 2500 spots ( excluding filter set ) ; slope , > 2.1 ; area , < 200 ; peak height , < 600 ; and area , < 10 000 . \n changes in the abundance of protein spots were regarded as significant with a threshold of 2-fold standard deviation difference . \n gels of six independent experiments were analyzed with the bva software , and average ratios as well as t test values for each spot were calculated . \n after scanning , the gels were stained with ruthenium ii tris ( bathophenanthroline disulfonate ) according to rabilloud(26 ) and lamanda(27 ) and protein spots were excised manually on a dark reader transilluminator ( dr-88 m , clare chemical research , dolores , co ) . \n protein spots were tryptically digested according to the protocol of shevchenko.(28 ) extracted peptides were identified on an ultraflex i maldi - tof / tof device using flexcontrol 3.0 for data collection and flexanalysis 3.0 spectra analysis / peak list generation ( bruker daltonics , germany ) as described.(23 ) peptide mass fingerprint ( pmf ) and peptide fragmentation fingerprint ( pff ) spectra were submitted to the mascot server ( mascot 2.1.03 , matrix science , u.k . ) , searching the ncbi database limited to the taxon fungi . with respect to the sample preparation , fixed modification of cysteine thiols to s - carbamidomethyl derivatives and variable methionine oxidation were defined for the database search \n further , up to one missed cleavage , and a peptide mass tolerance of 50 ppm was allowed . \n results were regarded as significant with an allowed likelihood for a random hit of p 0.05 , according to the mascot score . \n all proteome data were imported into our in - house data warehouse omnifung(29 ) ( http://www.omnifung.hki-jena.de ) , which is based on protecs ( decodon , germany ) . \n identified proteins were classified with the fungifun annotation tool as described in priebe et al.(30 ) a. fumigatus mitochondria were solubilized by a dodecylmaltosid solution ( 750 mm amino caproic acid , 50 mm bis(2-hydroxyethyl)-imino - tris(hydroxymethyl)-methane , 0.5 mm edta , ph 7.0 , 1% [ wt / vol ] dodecylmaltoside ) and the protein concentration was determined using the bradford assay . \n 170 g mitochondrial protein were separated on a two - dimensional blue - native / sds - page as described . \n quantification of the spot volume was achieved by using imagequant 5.2 ( ge healthcare ) . to 500 l of \n a. fumigatus culture grown in a chemostat under normoxic or hypoxic condition 5 l mito tracker red fm was added ( invitrogen , germany ) and was incubated for 1 h at 37 c . afterward the mycelium was washed three times with phosphate buffered saline ( pbs ) . \n fluorescence microscopic analysis was performed with an axiovert 200 m / lsm 5 live confocal laser scanning microscope ( zeiss , jena , germany ) . \n determination of the respiration rates of a. fumigatus hyphae were performed with an oxygen monitor ( ysi 5300 , ysi life sciences , yellow springs , oh ) equipped with polarographic clark - type electrodes . \n one ml of a. fumigatus culture grown in a chemostat under normoxic or hypoxic condition were added to 4 ml fresh amm and applied to the sample chamber . \n the depletion of dissolved oxygen in amm was measured for 20 min at 37 c under continuous stirring under atmospheric oxygen partial pressure . \n oxygen consumption rates were expressed as percent o2 consumed per minute per mg mycelium dry weight . \n northern blot analysis was performed as previously described.(33 ) the oligonucleotides used for amplifying dig - labeled probes are shown in supplementary table 1s ( supporting information ) . \n the culture filtrates were exhaustively extracted with ethyl acetate , the resulting extracts were dried with sodium sulfate and concentrated under reduced pressure . for hplc analysis the extracts were dissolved in 500 l of methanol . \n analytical hplc was performed on a shimadzu lc-10avp series hplc system consisting of an autosampler , high pressure pumps , column oven and pda . \n hplc conditions : c18 column ( eurospher 1005 250 4.6 mm ) and gradient elution ( mecn/0.1% ( v / v ) tfa 0.5/99.5 in 30 min to mecn/0.1% ( v / v ) tfa 100/0 , mecn 100% for 10 min ) , flow rate 1 ml min ; injection volume : 20 l . \n lcms measurements were performed using a surveyor hplc system ( thermo electron , bremen ) coupled to a finnigan lcq benchtop mass spectrometer with an electrospray ion source . \n hplc conditions : gradient elution ( mecn/0.1% ( v / v ) hcooh 2/98 in 30 min to mecn/0.1% ( v / v ) hcooh 98/2 ; flow rate 0.6 ml min ) . \n pseurotin a was identified by comparison with an authentic standard ( obtained from alexis biochemicals , lrrach , germany ) . \n eight week - old female cd-1 mice were infected with 1 10 conidia of the a. fumigatus wild - type strain cbs 144.8s . \n immunosuppression and infection was performed as described previously.(34 ) mice were monitored for weight loss and physical conditions and sacrificed when moribund . \n mice were cared for in accordance with the principles outlined by the european convention for the protection of vertebrate animals used for experimental and other scientific purposes ( european treaty series , no . \n lungs of two mice , which were sacrificed on day four and five were directly removed and stored in rnalater ( ambion europe , huntingdon , u.k . ) until further processing . \n rna isolation and cdna synthesis were performed as described.(35 ) harvested mycelium was directly frozen in liquid nitrogen as described above , and ground to a fine powder . \n one - hundred milligrams were used for rna isolation and subsequent cdna synthesis as described above . \n qrt - pcr was performed on a corbett rotor gene 6000 ( qiagen , hilden , germany ) real - time pcr system in triplicate for each sample . \n the a. fumigatus actin gene act1 ( afua_6g04740 ) was used as reference gene to calculate the expression levels of the genes of interest . \n the cdna samples from the in vitro and in vivo experiments were 250 times diluted and undiluted , respectively , used for amplification . to obtain sybrgreen - labeled pcr fragments ( dynamo flash sybr green qpcr kit , finnzymes , espoo , finland ) the gene - spezific primers afua_8g00540_qrt - pcr - for 5 gaa gac cac agt cat cga cc 3 , afua_8g00540_qrt - pcr - rev 5 tta aca ctt gcc cgt agc gg 3 , \n afua_8g00550_qrt - pcr - for 5 gca cct ggc ttc gca atg t 3 , afua_8g00550_qrt - pcr - rev 5 gca cgt tct tca agc cgg 3 , afua_6g04740_qrt - pcr - for 5 gct ttg \n cta cgt cgc tct cg 3 and afua_6g04740_qrt - pcr - rev 5 gga gga gca atg atc ttg acc 3 were used . \n the size of the pcr amplicons of afua_8g00540 , afua_8g00550 and afua_6g04740 were 252 , 416 and 353bp , respectively . \n the cycling parameters included an initial dna denaturation step at 95 c for 15 min , followed by 40 cycles with dna denaturation at 95 c for 15 s , primer annealing at 60 c for 30 s and extension at 72 c for 30 s. controls with no added template were included for each primer pair to exclude primer dimers interfering with amplification detection . \n qrt - pcr results were analyzed using rotor gene software 1.7 ( qiagen , hilden , germany ) . \n relative expression levels for each cdna sample were obtained via normalization to actin using the formula 2 [ 2 , 2 ] for all samples . \n porphyrins were quantified by hplc with uv and fluorescence detection and normalized to the sample protein content as described.(36 ) for determination of the total cellular iron , copper , zinc and manganese content the freeze - dried mycelia was decomposed in closed polytetrafluorethylene vessels containing 2 ml of hno3 and 0.5 ml of hydrogen peroxide using a high - performance microwave digestion unit ( mls-1200 mega ) as described.(37 ) the mentioned metals were determined by inductively coupled plasma - atomic emission spectroscopy ( icp - aes).(38 ) pyridine nucleotides were quantified using the amplite colorimetric nad(p)/nad(p)h assay kit ( biomol , hamburg , germany ) according to the manufacturer s instructions . \n fifty micrograms of ground mycelium were treated with 1 ml pbs buffer , incubated on ice and centrifuged . \n the supernatant was used to determine the ratio of nad(p)/nad(p)h in a plate reader ( fluostar omega , bmg labtech , offenburg , germany ) at 575 nm . \n to study the adaptation mechanism of a. fumigatus to hypoxia , we analyzed the proteomes of a. fumigatus wild - type strain grown either under normoxic ( 21% po2 ) or hypoxic ( 0.21% po2 ) conditions ( figure 1 ) . \n therefore , we established continuous cultivation for a. fumigatus in a fermenter system , which allowed to culture a. fumigatus at different defined oxygen partial pressures while keeping constant all other cultivation parameter ( ph , temperature , nutrition supply , dilution rate / growth rate , glucose limitation ) . to determine the optimal dilution rate ( equal to the growth rate in the chemostat at steady state ) for the continuous cultivation of a. fumigatus , we performed batch fermentations to define the maximal growth rate ( max ) of a. fumigatus under different oxygen concentrations ( supplementary figure 1 , supporting information ) . \n the max values of a. fumigatus were determined with 0.11 h and 0.25 h for cultivation under hypoxic and normoxic growth conditions , respectively . to ensure the same growth rate of a. fumigatus under normoxic and hypoxic culture conditions , we applied a dilution rate of 0.08 h during chemostat cultivation . \n a culture volume of 3 l and a dilution rate of 0.08 h led to a flow rate of 240 ml / h . the glucose concentration was set to 8 mm in the feed medium , which resulted in a biomass of around 750 mg dry weight / l . \n these settings turned out to be optimal for continuous cultivation of a. fumigatus and , at the same time , prevented clogging of the outlet tube . \n steady state ( constant biomass , ph value , and glucose depletion ) was reached after 7296 h. 2d gel electrophoresis of protein extracts of a. fumigatus grown under hypoxic ( cy5 , green ) and normoxic conditions ( cy3 , purple ) . \n the spot numbers refer to differentially synthesized proteins under hypoxic growth conditions ( see table 1 ) . \n the results of the proteome analyses are summarized in figure 1 , table 1 and table s2 ( supporting information ) . \n fifty - nine protein spots showed an increased while 31 protein spots showed a decreased standard abundance of > 1.5 fold ( 2-fold standard deviation ) in extracts of a. fumigatus mycelia grown under hypoxic conditions . \n several proteins appeared as multiple spots on the gels with the same apparent molecular mass , but with varying pi values and abundance , probably due to post - translational modifications or isoenzyme variation , for example , thiamine biosynthesis protein ( nmt1 ) and acetyl - coenzyme a synthetase ( faca ) . \n hence , the identified 90 protein spots represented 65 different proteins ( figure 1 and table 1 ) . \n the protein spot number was assigned to the protein spot with the highest change in intensity ( figure 1 ) . \n average ratios extracted from statistical analysis of dige gels by the decyder software programs dia and bva . \n exposure of a. fumigatus to hypoxic conditions resulted in an increased level of many proteins involved in glycolysis , pyruvate metabolism , respiration , the pentose phosphate pathway and amino acid metabolism . \n interestingly , the drastic reduction of the o2 concentration also led to an increased abundance of enzymes catalyzing reactions in secondary metabolite biosynthesis . \n 17 and 19 in figure 1 ) , are involved in the formation of pseurotin a , a known fungal mycotoxin with a highly substituted 1-oxa-7-azaspironon-2-ene-4,6-dione skeleton . \n most likely , because oxidative stress is typically low under hypoxia , also the hydrogen peroxide scavenging catalase cat1 and the redox - balancing protein thioredoxin showed reduced protein expression . \n a flavohemoprotein ( spot no.16 in figure 1 ) and an acetyl - coenzyme a synthetase ( spot no . 31 in figure 1 ) showed the highest up- and down - regulation , respectively , under hypoxic culture conditions . \n flavohemoproteins are known to be involved in scavenging reactive nitrogen intermediates as shown for a. oryzae and a. nidulans . \n because of the hypoxic upregulation of many proteins involved in respiration and electron transport , we examined the mitochondrial protein fraction in more detail . for this reason , \n mitochondria were isolated and protein samples were produced for two - dimensional gel electrophoresis as described previously.(23 ) the result of the comparative analysis of the mitochondrial proteome under hypoxic and normoxic growth conditions is shown in figure 2 , table 2 and table s3 ( supporting information ) . in total , 93 protein spots showed a different abundance upon hypoxia , in which 10 protein spots were down- and 83 protein spots were up - regulated . \n the 93 protein spots represented 50 different proteins ( table 2 ) . of the 50 differentially regulated proteins 35 proteins had not been detected in the mycelial fraction before ( table 2 footnote e ) comprising proteins involved in glycolysis , the tca cycle , respiration , protein folding and amino acid metabolism . \n obviously , mitochondrial fractionation did not completely avoid contamination by cytosolic proteins and , in addition , a small number of cell - wall associated proteins were detected as well , for example , gel proteins and ecm33 . \n 2d gel electrophoresis of mitochondrial protein extracts of a. fumigatus grown under hypoxic ( cy5 , green ) and normoxic conditions ( cy3 , purple ) . \n the spot numbers refer to differentially synthesized proteins under hypoxic growth conditions ( see table 2 ) . \n average ratios extracted from statistical analysis of dige gels by the decyder software programs dia and bva . \n mitochondrial import sequence according to mitoprotii - v1.101 . only found in the mitochondrial fraction as differentially regulated . \n 42 in figure 2 ) revealed the strongest induction ( 34-fold ) among the proteins of the mitochondrial fraction under hypoxic growth conditions . \n moreover , a drop in o2 levels gave rise to an increased ubiquinone biosynthetic activity , which is illustrated by the higher abundance of the two enzymes ubiquinone biosynthesis monooxygenase coq6 and methyltransferase coq5 ( table 2 ) . \n since the mitochondrial membrane complexes are difficult to resolve by conventional 2d - gel analysis , mitochondrial fractions ( 170 g of protein ) of a. fumigatus grown under normoxic or hypoxic conditions were applied to two - dimensional blue - native ( bn)/sds page after solubilization of the mitochondrial membrane proteins by using n - dodecylmaltoside as detergent . \n ( figure 3 , table 3 and table s4 ( supporting information ) ) . in the first separation , \n the respiration complexes with their high molecular masses were separated under native conditions according to their size , whereas the subunits of the native complexes are then separated by sds - page in the second separation . \n altogether , the bn / sds - page analysis revealed an increase in abundance of proteins of all five respiratory complexes in a. fumigatus mitochondria under hypoxic conditions ( figure 3 ) . \n however , proteins of complex iii and iv showed the highest , proteins of complex v the lowest rise in protein expression . due to this imbalance it is interesting to speculate that some electrons may not enter the respiratory chain via complex i or ii during hypoxic growth conditions . \n analyses of the protein complexes of the mitochondrial respiration chain of a. fumigatus by two - dimensional blue - native / sds page under hypoxic conditions . \n average ratios extracted from analysis of bn gels by the imagequant 5.2 . software . to investigate whether the up - regulation of proteins involved in the respiratory machinery is directly correlated with a higher respiratory capacity , we measured at atmospheric oxygen partial pressure the rate of oxygen consumption of a. fumigatus hyphae , which had been grown either under normoxic or hypoxic growth conditions ( figure 4a ) . \n adaptation to hypoxic compared to normoxic condition increased the oxygen consumption rate of a. fumigatus hyphae 1.7-fold ( 4.83% compared to 2.78% ( v / v ) o2 per minute and mg dry weight ) . \n visualization of mitochondria by staining with the mitochondrion - selective fluorescent mitotracker red probe indicated that hypoxia increases the number of mitochondria in comparison to normoxia ( figure 4b ) . \n physiological adaptation of a. fumigatus to hypoxic growth condition with regard to modifications in the respiratory activity and the amount of mitochondria in the hyphae . \n ( a ) oxygen consumption rate of a. fumigatus hyphae grown under normoxic ( 21% po2 ) and hypoxic ( 0.2% po2 ) conditions . \n the oxygen consumption rate of a. fumigatus grown in hypoxia is significantly increased compared to that of a. fumigatus cultivated under normoxic environments ( p < 0.01 ) . \n ( b ) determination of the amount of a. fumigatus mitochondria by mito tracker red fluorescence staining . \n clsm fluorescence ( fl ) , brightfield ( bf ) and overlay ( ov ) images showing a clearly elevated amount of mitochondria in the mycelium of hypoxic ( 0.2% po2 ) cultivated a. fumigatus in comparison to a. fumigatus grown under normoxia ( 21% po2 ) . \n the most striking difference in the growth morphotype of a. fumigatus at hypoxic growth conditions was a change in the mycelial color from whitish to reddish starting approximately after 2 days of cultivation ( figure 5a ) . to test the hypothesis that an increased heme or iron content may cause the red color formation during hypoxic chemostat cultivation \n , we quantified the heme and metal concentration of a. fumigatus mycelium by hplc and icp - aes , respectively ( figure 5b ) . \n the heme content increased 4-fold in a. fumigatus mycelium under hypoxic growth conditions and also the amount of iron and the trace elements zinc and copper were elevated . \n these changes most likely reflect the hypoxia - induced rise in mitochondrial respiratory enzyme complexes containing heme , copper and iron as cofactor . \n o2-dependent changes in the mycelial color and in the intracellular concentration of heme and metals in a. fumigatus . \n ( a ) color of the mycelium of a. fumigatus turned from whitish to reddish due to the reduction of the oxygen partial pressure from 21 to 0.2% in the medium . \n ( b ) differences in the metal ( left y - axis ) and heme ( right y - axis ) content of a. fumigatus mycelia grown under hypoxic ( dark gray bar ) in comparison to normoxic ( light gray bar ) conditions . \n the metals iron , zinc and copper and the prosthetic group heme are significantly ( p < 0.05 * or p < 0.01 * * ) enriched in a. fumigatus mycelia under hypoxia . \n error bars indicate standard deviations of the means . because of the up - regulation of several proteins involved in the pentose phosphate pathway and in the respiration machinery \n , we analyzed the intracellular concentrations of the pyridine nucleotides of a. fumigatus under both , hypoxic and normoxic conditions ( figure 6 ) . \n hypoxia increased the nadp and nadph content 47-fold and 8-fold , respectively , which changed the nadp / nadph ratio from 0.4:1 under normoxic to 2.4:1 under hypoxic growth conditions . \n the increase of the nadp / nadph ratio during hypoxia indicates that either reductive biosynthesis pathways or nadph - consuming mechanisms of oxidative or nitrosative stress defense are more active upon hypoxia . \n by contrast , no differences in the concentration of intracellular nad was detectable in hyphal cells of a. fumigatus cultivated under normoxia and hypoxia , while nadh was not detectable under neither of the cultivation conditions probably due to the low concentration of nadh in the cell . \n intracellular concentrations of the pyridine nucleotides of a. fumigatus under normoxia ( light gray bar ) and hypoxia ( dark gray bar ) . \n hypoxic growth conditions led to a significant ( p < 0.05 * or p < 0.01 * * ) increase of the nadp and nadph concentration ( left y - axis ) and to a complete conversion of the nadp / nadph ratio ( right y - axis ) in a. fumigatus . \n the proteome analysis of a. fumigatus grown under normoxic or hypoxic conditions revealed an increased abundance of enzymes involved in the biosynthesis of secondary metabolites ( table 1 and figure 1 ) , for example , an /-hydrolase and a sirn - like methyltransferase , whose corresponding genes are part of the already characterized pseurotin a biosynthesis gene cluster.(39 ) northern blot analysis confirmed the proteome data and revealed upregulation under hypoxic conditions of all genes of the pseurotin a biosynthesis gene cluster(39 ) at the transcriptional level ( figure 7 ) . \n moreover , northern blot analysis also revealed the induction of additional secondary metabolism genes during hypoxia : several genes of the fumitremorgin gene cluster ( afua_8g00190 ; afua_8g00200 and afua_8g0010 ) and genes of a predicted gene cluster of unknown function ( afua_8g00300 afua_8g00520 ) ( figure 7a and table s2 , supporting information ) . \n reversed phase - hplc confirmed induction of pseurotin a production under hypoxic conditions at the metabolite level ( figure 7a and b ) . \n induction of the pseurotin a biosynthesis gene cluster as well as the biosynthesis of pseurotin a under hypoxic , glucose - limited growth conditions in a. fumigatus . \n ( a ) northern blot analysis and organization of the pseurotin a biosynthesis gene cluster . \n mt : methyl transferase ; p450 : cytochrome p450 oxidoreductase ; gst : glutathione s - transferase . \n ( b ) hplc profiles of supernatant of the cultures of a. fumigatus grown under hypoxic and normoxic conditions . \n ( c ) northern blot analysis of the expression of srba in a. fumigatus grown under normoxic and hypoxic conditions . \n as shown in figure 7c , the mrna level of the hypoxic regulator - encoding srba , which was previously shown to be induced by hypoxia,(6 ) was increased during chemostat cultivation at 0.2% compared to 21% po2 , confirming the accuracy of the cultivation method . to elucidate the transcriptional activity of the pseurotin a biosynthesis gene cluster during infection , we applied qrt - pcr analysis by comparison gene expression of the hybrid polyketide synthasenonribosomal peptide synthetase ( pks / nrps ) gene afua_8g00540 and the methyl transferase gene afua_8g00550 under in vivo ( murine model of pulmonary aspergillosis in cortisone - treated mice ) and in vitro ( chemostat cultivation ) conditions ( figure 8a ) . \n transcript levels of both genes increased relative to the control gene act1 under hypoxic compared to normoxic in vitro conditions and during a. fumigatus infection in mouse lungs . \n the relative expression of the methyl transferase afua_8g00550 was similar during hypoxic in vitro and in vivo conditions during mice infection , i.e about 11-fold higher compared to normoxic in vitro conditions ( 12-fold in vitro , 8.3- and 13.4-fold in vivo ) . \n compared to normoxic in vitro conditions , the relative expression of the hybrid pks / nrps gene was about 8-fold upregulated during hypoxic in vitro conditions and even 64-fold and 28-fold , respectively , in the two mouse lungs tested . \n taken together , the qrt- pcr analysis revealed a clear induction of genes of the pseurotin a biosynthesis gene cluster in experimental aspergillosis . \n quantitative real time - pcr to determine the relative expression of the hybrid pks / nrps and methyl transferase genes of the pseurotin a biosynthesis gene cluster in comparison to the control gene act1 ( afua_6g04740 ) under normoxic and hypoxic conditions . \n ( a ) relative expression of the genes for the hybrid pks / nrps and methyl transferase of the pseurotin a biosynthesis gene cluster in vitro and in the lung of infected mice . for the in vitro tests a. fumigatus \n was grown under hypoxic and normoxic conditions and the total rna was subsequently isolated from the mycelium . for the in vivo data , \n mouse infection experiments were performed and the total rna was isolated from two infected lungs . \n ten g of total rna of a. fumigatus mycelium grown during hypoxia ( 0.2% po2 ) and normoxia ( 21% po2 ) was analyzed . \n to colonize the human lungs , the saprophytic fungus a. fumigatus , which is generally found in soil and decaying organic matter such as compost piles , has to adapt to the new environmental conditions . besides mechanisms to evade the attack of immune cells and an effective nutrient uptake system,(46)a . \n recently , it was shown that the ability to grow under hypoxia is a virulence trait for a. fumigatus and other pulmonary infection pathogens . despite \n the reported connection between hypoxia adaptation and virulence , comprehensive knowledge about the detailed molecular basis of stress adaptation in a. fumigatus to hypoxia is lacking . to close this gap \n , we carried out detailed proteome analyses of the total soluble and the mitochondrial fraction of a. fumigatus mycelium . to achieve a data set with a maximum in reproducibility and reliability we established a chemostat cultivation method for a. fumigatus to maintain the mycelium under strictly controlled environmental conditions . here , we identified 117 proteins that were differently regulated in this fungus under hypoxic versus normoxic growth conditions . \n the majority of these proteins belong to the funcat categories respiration / electron transport , carbon metabolism , amino acid metabolism , cell wall construction , glycolysis and vitamin and cofactor synthesis.(30 ) under hypoxia , a. fumigatus seems to enhance the flux through the glycolytic pathway . \n similar to c. neoformans(15 ) and c. albicans,(18 ) hypoxia up - regulated proteins of the glycolysis in a. fumigatus . \n consequently , for an effective conversion of the main glycolytic end product pyruvate to the entry molecule of the tca cycle , acetyl - coa , a. fumigatus increased the biosynthesis of proteins of the pyruvate dehydrogenase complex ( pdc ) . \n such a hypoxic dependent up - regulation of the pdc was also shown for a. nidulans,(48)trichoderma reesei(49 ) and s. cerevisiae . \n in addition to the increased glycolytic activity , a. fumigatus responded to hypoxia with the induction of several enzymes of the tca cycle like the aconitase , the -ketoglutarate dehydrogenase complex and the succinyl - coa synthetase , whereas the level of succinate dehydrogenase decreased . \n an increased tca cycle activity at low oxygen levels was also demonstrated for s. cerevisiae,(52 ) whereas the opposite , a repression of genes coding for enzymes of the tca cycle with reduced oxygen concentration , was reported for c. albicans.(18 ) the increased supply with precursor molecules for the biosynthesis of amino acids or tetrapyrrols or an increased flux through the tca cycle for the generation of nadh and fadh2 are possible explanations for the general induction of the tca cycle in a. fumigatus at reduced oxygen levels . altogether , the proteome data did not give any hint for the induction of fermentative pathways in a. fumigatus under hypoxia . \n in contrast to a. nidulans,(53 ) no enzymes involved in ethanol or lactate production were induced upon hypoxia . \n the level of the soluble fumarate reductase osm1 , which has been reported to be involved in the reoxidation of nadh and fadh2 during anaerobiosis in s. cerevisiae,(54 ) decreased at hypoxic growth conditions in a. fumigatus . \n furthermore , proteomic analysis of a. nidulans cultures under hypoxic conditions by shimizu et al.(48 ) revealed an increased activity of enzymes of the -aminobutyrate ( gaba)-shunt , which bypasses the nadh - generating activity of the -ketoglutarate dehydrogenase in the tca cycle . \n our proteome data do not support a similar hypoxic response in a. fumigatus , since levels of the gaba - shunt enzyme succinate semialdehyde dehydrogenase decreased at hypoxic growth conditions . \n however , the same group also reported an increased branched - chain amino acid biosynthesis under hypoxia , which contribute to nad and nadp regeneration.(55 ) indeed , also in a. fumigatus two enzymes of the valine , leucine and isoleucine biosynthesis pathway , namely dihydroxy acid dehydratase and acetolactate synthase , increased in abundance at low oxygen levels . \n but hypoxia may also affect in a general way the intracellular amino acid pools especially of amino acids that are partly synthesized in mitochondria.(50 ) overall , a. fumigatus presumably does not face a drastic metabolic imbalance during glucose - limited chemostat cultivations at hypoxic conditions . \n a drastic nadh accumulation is probably avoided by the upregulation of proteins of all five respiratory complexes , which is accompanied by an increased cellular content in heme , iron copper , and other metals as well as an augmentation of the mitochondrial mass and respiratory capacity . \n all these adaptation mechanism most likely lead to an increased efficiency in using the low amount of available oxygen . \n a similar hypoxic response was also observed for s. cerevisiae cultivated in glucose - limited chemostats.(52 ) by contrast , in a. niger a decreased activity of the respiratory chain , a lowered level of atp production and an accumulation of nadh in response to hypoxia was reported.(56 ) similar to c. neoformans,(15 ) proteins of the ubiquinone biosynthesis , an oxygen - dependent metabolic pathway , were more abundant under hypoxic environments . \n moreover , two mitochondrial - localized heat shock proteins , hsp70 and hsp60 that contribute to the biogenesis of fe / s cluster , mitochondrial protein biosynthesis and translocation processes of cytosolic proteins into mitochondria , were also more abundant under hypoxic environments . \n as in a. nidulans,(48 ) proteins of the biosynthesis of vitamins and cofactors were also found to be up - regulated in a. fumigatus under hypoxia , e.g. , the thiamine and the thiazole biosynthesis proteins , which are involved in the formation of thiamine that is subsequently converted into the active thiamine diphosphate ( tdp ) by the thiamine diphosphokinase.(59 ) tdp is an essential coenzyme of the transketolase , the pyruvate dehydrogenase and the -ketoglutarate dehydrogenase . \n all these enzymes were identified to be up - regulated in a. fumigatus under hypoxic growth conditions . \n the flavohemoprotein afua_4g03410 showed the highest upregulation in the mycelial fraction of a. fumigatus when cultured under hypoxia . in aspergilli , flavohemoproteins are involved in detoxifying reactive nitrogen species ( rns)(41 ) and are coregulated with nitrate assimilation.(42 ) an induction of this type of enzyme under hypoxia has not been reported for aspergilli yet . \n however , it was shown for s. cerevisiae that under hypoxic conditions cytochrom c oxidase catalyzes the production of no within mitochondria by reducing nitrite . \n castello et al.(62 ) suggested that in yeast the mitochondrially produced no is involved in hypoxic signaling , possibly via protein tyrosine nitration . \n therefore , the hypoxia - induced flavohemoprotein may protect cells of a. fumigatus against rns generated at low oxygen levels . in addition , the rns may also act as a signal molecule in the hypoxic response in a. fumigatus.(66 ) a hypoxia induced up - regulation of a flavohemoprotein was already demonstrated for c. neoformans.(15 ) the deletion of the corresponding genes of the no - detoxifying flavohemoproteins in the pathogenic yeasts c. neoformans and c. albicans led to attenuated virulence . \n to date , the role of rns in hypoxic signaling or pathogenicity of a. fumigatus has not been elucidated . \n the fact that some of the mitochondrially produced no is released from cells(69 ) raises the possibility that no has also an immuno - suppressive effect during later stages of infection . \n a further remarkable result of our work is the first identification of hypoxia as an inducing factor for secondary metabolism in a. fumigatus . \n two proteins , namely a sirn - like methyltransferase ( afua_8g00550 ) and an /-hydrolase ( afua_8g00530 ) of the pseurotin a biosynthesis gene cluster were found to be up regulated due to hypoxic condition.(39 ) the hypoxic induction of the entire pseurotin a biosynthesis gene cluster and the actual formation of pseurotin a under these conditions in a. fumigatus was proven by northern blot and hplc - ms analysis . \n qrt - pcr analysis clearly demonstrated a high expression level of two genes of the pseurotin a biosynthesis gene cluster under in vivo infection conditions as well . \n the pseurotin a hybrid pks / nrps gene in a. fumigatus during infection compared to hypoxic in vitro conditions indicating even lower o2 availability in the infected tissue or additional inducing stimuli besides hypoxia . \n it is conceivable that the production of pseurotin a could contribute to the survival of a. fumigatus in hypoxic microenvironments in the soil,(70 ) where it could act against competitors like nematodes and microbes . \n a demonstrated moderate cytotoxicity against lung fibroblasts(72 ) and the capacity to inhibit ige production(40 ) indicate a possible role of pseurotin a in the pathogenicity of a. fumigatus in mammals . \n our 2-d gel electrophoresis and blue native / sds - gel separation provided a global view on the proteome level changes during long - term adaptation of a. fumigatus to hypoxia at substrate limitation . \n we identified 117 different proteins from a. fumigatus which significantly changed their abundance at hypoxic in comparison to normoxic growth conditions . \n our data suggests that a. fumigatus compensates a drastic drop in oxygen levels by increasing the levels of respiratory proteins and thus increasing the respiratory capacity . \n however , it can not be ruled out that a short - term response of a. fumigatus to hypoxia at sufficient carbon source availability could induce other adaptation mechanisms . \n the high up - regulation of an no - detoxifying flavohemoprotein implies indirectly that hypoxia stimulates the formation of reactive nitrogen intermediates in a. fumigatus . \n furthermore , hypoxia activated the silent or poorly expressed secondary metabolite pseurotin a biosynthesis gene cluster . \n such an activation may have also an impact on the growth of a. fumigatus during the infection , when local obstruction of the airways and necrotic deaths of host s cells leads consequently to oxygen depletion . \n the characterization of the mechanisms of transcriptional and translational activation of hypoxia - induced genes and proteins , respectively , should form the focus of future studies .", "role": "user" }, { "content": "the mold aspergillus fumigatus is the most important airborne fungal pathogen . \n adaptation to hypoxia represents an important virulence attribute for a. fumigatus . \n therefore , we aimed at obtaining a comprehensive overview about this process on the proteome level . to ensure highly reproducible growth conditions , an oxygen - controlled , glucose - limited chemostat cultivation was established . \n two - dimensional gel electrophoresis analysis of mycelial and mitochondrial proteins as well as two - dimensional blue native / sds - gel separation of mitochondrial membrane proteins led to the identification of 117 proteins with an altered abundance under hypoxic in comparison to normoxic conditions . \n hypoxia induced an increased activity of glycolysis , the tca - cycle , respiration , and amino acid metabolism . \n consistently , the cellular contents in heme , iron , copper , and zinc increased . \n furthermore , hypoxia induced biosynthesis of the secondary metabolite pseurotin a as demonstrated at proteomic , transcriptional , and metabolite levels \n . the observed and so far not reported stimulation of the biosynthesis of a secondary metabolite by oxygen depletion may also affect the survival of a. fumigatus in hypoxic niches of the human host . among the proteins so far not implicated in hypoxia adaptation , an no - detoxifying flavohemoprotein was one of the most highly up - regulated proteins which indicates a link between hypoxia and the generation of nitrosative stress in a. fumigatus .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: adequate retention of the ocular prosthesis in the anophthalmic socket requires a well - formed inferior fornix , which in turn requires sufficient conjunctival length and a deep recess . \n obliteration of the inferior fornix might occur in contracted socket and in anophthalmic socket ( postenucleation socket ) syndrome . \n contracted socket is a condition characterized by fibrosis of the anophthalmic socket where shallow or obliterated fornix is a key finding in different stages of the disease and it occurs secondary to conjunctival shrinkage . \n the later occurs possibly as there is no globe and so the inferior rectus muscle is at a higher level in the socket with subsequent elevation of the lower lid retractors and their connections including the fornical conjunctiva . \n management of the shallow inferior fornix in contracted socket is usually done by deepening sutures following mucous membrane graft to avoid lower lid entropion . in anophthalmic socket syndrome , \n the shallow inferior fornix is reformed by deepening sutures that exit at the skin of the lower lid where they are tied over bolsters and left for 2 - 3 weeks . \n however skin erosion and infection necessitated early removal of the externalized sutures and increased the risk of recurrence . \n a transconjunctival inferior fornix fixation is another method of repair of shallow inferior fornix in anophthalmic socket syndrome where the edges of the conjunctival incision are directly sutured to the periosteum with no need for externalized sutures and stents . \n although this procedure yielded good results , it still carries the risk of triggering contracture of the socket as it involved dissection of the fornical conjunctiva and lower lid retractors with cauterization of the tissues and excessive sacrifice or destruction of conjunctiva . for the above reason \n , we designed this study to evaluate new surgical technique for deepening the inferior conjunctival fornix using fascia lata in contracted socket and anophthalmic socket syndrome . \n this was a prospective , nonrandomized interventional case series studythat was carried out in the oculoplastic unit at ophthalmology department , el minia university hospital , in the period from february 2009 to may 2012 . \n all procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 helsinki declaration and its later amendments or comparable ethical standards . \n all patients attending to the oculoplastic unit at el minia university hospital who were unable to wear and retain their ocular prosthesis due to shallow or obliterated inferior conjunctival fornix were examined and 24 male and female adult patients were considered for study enrollment after exclusion of patients with previous surgical correction of shallow inferior conjunctival fornix . \n the patient 's sockets were examined thoroughly and the patients were categorized into 2 subgroups according to the cause of the shallow inferior fornix as follows : anophthalmic group : 9 patients with anophthalmic socket syndrome.contracted group : 15 patients with moderate to severe contracted sockets . \n another 24 patients who underwent evisceration or enucleation with healthy sockets and can wear and retain their prosthesis comfortably were chosen as a control group . \n indication for evisceration or enucleation and time elapsed since that surgery were recorded for every patient in the control as well as patient groups . \n the depth of the inferior conjunctival fornix was measured in mm at the center of the lower eyelid in all participants in the control group and in patient subgroups . \n informed consents were obtained from all patients for the surgery under general anesthesia and after sterilization of the thigh with povidone iodine , a 2 cm vertical skin incision was made 10 cm above the lateral epicondyle of the knee over the iliotibial tract and then blunt dissection of the tissues was carried out down to the fascia lata site . \n a 3 mm wide free end of the fascia was fashioned using scissors and then the masson fascia lata stripper was engaged and advanced for 15 cm upwards towards the hip joint and then triggered several times till a fascia lata strip 3 mm wide and 1015 cm in length was obtained . \n ( 1 ) in case of anophthalmic socket with abundant conjunctiva but shallow inferior fornix ( figure 1 ) . \n a 2 mm conjunctival incision was made in each side of the center of the inferior fornix . \n another three horizontal cutaneous incisions were made in the lower lid over the inferior orbital margin , each of which was 5 mm in length and deepened down to the periosteum . \n the lateral incision was at the level of lateral canthus , the medial one was at the level of the lower punctum , and the middle one was in between ( figure 2 ) . \n one end of the fascia lata strip was secured to the periosteum using a nonabsorbable polyester 5/0 suture ( figure 2 ) and then wright 's fascia needle was used to deliver the free end of the fascia strip to lateral conjunctival incision ( figure 3 ) and then to the middle cutaneous incision . \n in the middle cutaneous incision . \n the free end of the fascia lata strip was passed under periosteal tunnel created to act as a pulley to change direction of pull and so facilitate intraoperative adjustment . \n again wright 's needle was used to pass the free end of the fascia strip to the medial conjunctival incision then and to the medial cutaneous incision . \n in the medial cutaneous incision . \n the free end of the fascia strip was pulled to achieve the desired tension to adequately reform the inferior fornix and then the free end was sutured to the periosteum by 5/0 polyester suture ( figures 4 and 5 ) . \n the skin incisions were closed by 6/0 nylon sutures while the conjunctival incisions were left without sutures . \n a proper sized conformer was placed to maintain the tissues in place for 3 weeks . \n a horizontal inferior fornical incision was performed and minimal dissection was done with excision of the fibrous bands if any to prepare the bed for the mucosal graft . \n the buccal mucosal graft was sutured at the bulbar side of the conjunctiva and then the fascia lata strip was manipulated as described above and the palpebral side of the mucosal graft was sutured to the palpebral conjunctiva before tightening the fascia strip . \n medications included systemic broad spectrum antibiotic for one week and topical mixed antibiotic / steroid eye drops and ointment for 4 weeks . \n skin sutures were removed after one week and temporary conformer after 3 weeks ( figure 6 ) and then the patients wore their ocular prostheses ( figure 7 ) . \n one month postoperatively the central inferior fornix depth ( cifd ) was measured in mm and recorded . \n after follow - up period , data were collected and analyzed using spss statistical program version 19 for windows and a statistically significant level was considered when p value was less than 0.05 . \n the follow - up period of all patients ranged from 9 to 26 months with a mean of 18.16 4.56 months . \n successful procedure was considered if the patient could wear and retain the prosthesis in place comfortably in all circumstances without lower eyelid entropion , ectropion , or retraction ( figure 7 ) . \n the ultimate goal of socket surgery is to fit the patient with a prosthesis that would simulate as much as possible to his or her normal fellow eye . \n the inability to retain an ocular prosthesis remains a psychologically devastating problem for patients and a healthy and deep inferior fornix is particularly important factor in accommodating and supporting the ocular prosthesis . \n obliterated or shallow lower fornix might occur either in contracted socket with conjunctival scarring and foreshortening or in anophthalmic socket or postenucleation socket syndrome where there is abundant conjunctiva with lacked inferior fornix fixation . \n management of the shallow or obliterated lower fornix usually is directed towards the underlying aetiology . in anophthalmic socket syndrome as there is no globe the inferior orbital fat migrated anteriorly and also the inferior rectus muscle is at a higher level in the socket with subsequent elevation of the lower lid retractors and their connections including the fornical conjunctiva . \n prolapse of the fornical conjunctiva may result in anterior rotation of the lower edge of the prosthesis . in these cases , a silastic stent ( e.g. , 240 retinal band ) \n can be positioned in the lowest aspect of the inferior fornix and anchored to the adjacent periosteum , with both arms of a double - armed suture passing through the stent , then through the deepest part of the fornix , through the periosteum of the inferior orbital rim , and through the full thickness of the eyelid . \n silicone or rubber bolsters are used when the sutures are tied on the skin surface , anchoring the stent securely in the inferior fornix ( figure 8) . \n these sutures may be removed in 2 - 3 weeks after adequate fibrosis has occurred between the inferior fornix and periosteum . \n but chronic mucoid conjunctival discharge and skin erosion with infection necessitated early removal of the externalized sutures and increased the risk of recurrence ( figure 9 ) . \n another method of repair consisted of a transconjunctival inferior fornix incision to gain direct exposure of the periosteum of the inferior orbital rim . \n direct suture fixation of the edges of the conjunctival incision to the periosteum is then achieved . \n contrary to anophthalmic socket syndrome where there is only poor lower fornix fixation , the contracted socket is characterized by inadequate conjunctiva in addition to cicatricial obliteration of the inferior fornix . \n so in management of contracted inferior fornix it is not enough to deepen the lower fornix but also additional tissue grafting is essentially needed . \n mucous membrane [ 2 , 9 ] and amniotic membrane [ 10 , 11 ] grafts are the most common tissue used in the reconstruction of the significantly contracted eye socket . \n after the tissue graft has been sutured to the conjunctival edges , maintaining the graft in its bed and deepening of the inferior fornix can be achieved by various techniques including deepening sutures , slightly larger conformer with temporary tarsorrhaphy or without tarsorrhaphy , or wire suturing of a custom conformer to the bone of the upper and lower orbital rim . in the current study we used fascia lata strips to pull down and hold the lower eyelid retractors for deepening of the shallow inferior fornix in 24 cases . \n the patients of the study had undergone evisceration or enucleation for different reasons ( table 2 ) and with different periods since these procedures ( table 1 ) . \n age of the patients ranged between 14 and 63 years with the mean age of 27.79 year ( table 1 ) and in comparison with the control group there was no statistically significant difference regarding the age ( p = 0.3633 ) . \n 41.7% of the patients were males while 58.3% of the patients had a left sided socket problem . \n regarding the preoperative cifd \n in the patient group , the depth ranged from 0 to 4 mm with a mean of 1.875 mm ( table 1 ) . \n the mean preoperative cifd was lower in anophthalmic subgroup ( table 3 ) and the difference between the 2 subgroups was statistically significant ( p = 0.0342 ) . in the control group , \n the range was 611 mm with a mean of 8.375 mm ( table 1 ) and the difference between the control and the patient groups was statistically highly significant ( p < 0.0001 ) . \n the postoperative cifd after 1 month in all patients was markedly increased and it ranged from 3 to 9 mm with an average of 6.833 mm ( table 3 ) and these changes were statistically highly significant when compared to the preoperative cifd ( p < 0.0001 ) . \n the improvement in mean postoperative cifd was higher in anophthalmic than in contracted subgroup ( table 3 ) and the difference between the 2 subgroups was significant ( p = 0.0028 ) . \n after a follow - up of average 18 months , 95.8% ( 23 out of 24 ) of patients had satisfactory results as they could wear and retain their prostheses all the time without significant problems . \n 93.3% of cases with contracted sockets achieved satisfactory postoperative results during the follow - up period ( table 2 ) . \n regarding the success rate , there was no significant statistical difference in both anophthalmic and contracted socket subgroups ( p = 0.4 ) . \n 16 cases ( 66.6% ) needed reconsultation of an ocularist for larger prosthesis to match the newly formed fornix and we did not encounter any case of lower eyelid entropion , ectropion , retraction , or lagophthalmos during the follow - up period . the only case that failed to retain the prosthesis was a 23-year - old female with severe left contracted socket and later on she required another 2 socket operations for buccal mucosal graft and finally the result was satisfactory for her . \n the 9 cases with anophthalmic socket syndrome had preoperative lax lower lids ( figure 1 ) which were spontaneously corrected after the fascia lata technique without need for lateral tarsal strip ( figure 6 ) . \n 6 out of 15 cases ( 40% ) with contracted socket had shallow superior conjunctival fornix and were managed simultaneously with buccal mucosal grafts that held in place with the conformer . \n all cases in our study reported pain at the lower outer side of the thigh for only one week and only 5 cases ( 21% ) reported limbing during the same period . \n none of the cases reported ugly scar , haematoma , or muscle herniation ; this might be due to the small skin incision ( 2 cm ) and narrow fascia lata harvested ( 3 mm ) . in comparison to deepening sutures , \n fascia lata is autogenous and it is buried in the tissues so no skin necrosis or infection was anticipated . \n also fascia lata technique is not a temporary procedure and so the desired pulling down tension on the lower lid retractors is expected to be maintained for long time as the nonelastic fascia is now fixed to the tough nonresilient periosteum . \n moreover intraoperatively the tension exerted by the fascia on the lower lid retractors can be judged and monitored before tying and suturing the medial end of the fascia strip and so postoperative lower lid entropion or retraction can be avoided . in the conjunctival fixation technique described by neuhaus and hawes and modified by ma'luf , \n all of the patients became able to retain a prosthetic eye postoperatively with two patients having minimal postoperative lower eyelid retraction and another two patients having minimal lower eyelid entropion but no secondary surgery was required . in spite of the above good results of the conjunctival fixation technique , it seems that the fascia lata technique is superior for the following reasons : the conjunctival fixation is a technique that can not be used in contracted socket due to the lack of sufficient conjunctiva to be fixed to the periosteum in these cases.fascia lata technique is more physiologic than conjunctival fixation to the periosteum as the former aimed at lowering and pulling down the abnormally elevated lower lid retractor to a near normal position and not to divide the lower lid retractors to secure the conjunctival edge to the periosteum.in anophthalmic socket , it is preferable not to manipulate or insult the precious conjunctiva too much to avoid triggering cicatrisation and the development of contracted socket . in fascia lata technique , the conjunctiva was minimally manipulated in contrary to the long conjunctival incision and deep tissue dissection in conjunctival fixation procedure . \n the conjunctival fixation is a technique that can not be used in contracted socket due to the lack of sufficient conjunctiva to be fixed to the periosteum in these cases . \n fascia lata technique is more physiologic than conjunctival fixation to the periosteum as the former aimed at lowering and pulling down the abnormally elevated lower lid retractor to a near normal position and not to divide the lower lid retractors to secure the conjunctival edge to the periosteum . in anophthalmic socket , it is preferable not to manipulate or insult the precious conjunctiva too much to avoid triggering cicatrisation and the development of contracted socket . in fascia lata technique , \n the conjunctiva was minimally manipulated in contrary to the long conjunctival incision and deep tissue dissection in conjunctival fixation procedure . \n in addition , harvesting fascia lata is not a simple procedure and it is time consuming with a second operation site in the thigh with its complications . \n the fascia lata technique is a new , alternative , and effective procedure to correct the shallow inferior fornix in anophthalmic socket syndrome where minimal socket dissection is required and complications of the deepening sutures are to be avoided also in moderate to severe contracted sockets fascia lata technique which is a second alternative to deepening sutures but avoids their complications .", "role": "user" }, { "content": "purpose . to evaluate a new surgical technique using fascia lata to deepen the shallow inferior conjunctival fornix in contracted socket and anophthalmic socket syndrome \n . methods . a prospective controlled study in which 24 sockets of 24 patients who were unable to wear and retain their ocular prosthesis due to shallow inferior fornix were enrolled and categorized into anophthalmic socket syndromes ( 9 patients ) and contracted sockets ( 15 patients ) . \n another 24 patients who underwent evisceration or enucleation with healthy sockets and can wear and retain their prosthesis comfortably were chosen as a control group . \n deepening of the fornix was performed using fascia lata strips under general anesthesia . \n central depth of the inferior fornix was measured preoperatively and postoperatively . \n results . \n a statistically significant improvement of the postoperative central inferior fornix depth was reported which was marked in anophthalmic subgroup . \n 100% of anophthalmic sockets and \n 93.3% of contracted sockets achieved satisfactory results during the follow - up period with no postoperative lower eyelid malposition or obvious skin scar . conclusion . \n fascia lata technique is a new alternative and effective procedure to deepen the shallow inferior fornix that can be used in moderate to severe contracted sockets or anophthalmic socket syndrome with minimal lower eyelid or socket complications .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: dna plasmids expression plasmids expressing cdna with a cmv \n promoter for myc - spar , ha - plk2 , and ha - plk2 were previously \n described ( 6 ) . \n pcmv - ha - plk2 was created from pcmv - ha - plk2 using pcr - based \n mutagenesis . \n cdna for full - length spar as well as spar fragments were \n amplified from pgw1-myc - spar \n ( 5 ) and cloned into pentr-6 , \n compatible with the gateway cloning system ( invitrogen ) . \n these were used for \n in vitro lr clonase reactions into pdest-53 ( for gfp - spar ) or \n pdest - n - myc ( for myc - spar fragments ) . \n point mutations in the -trcp \n phosphodegron were generated by pcr - based mutagenesis using pcmv - myc - spar or \n pdest - myc - act2 as templates . \n pcmv - myc - cki and pcmv - gsk3 were \n previously described ( 19 ) . \n f - box proteins previously cloned from cdna pools \n ( 14 ) were re - cloned into \n pentr-6 , and in vitro lr clonase reactions were performed with \n pdest-27 to generate gst - fused f - box proteins . \n cul1 ( residues \n 1 - 452 ) was previously described \n ( 14 ) ; cul3 and \n cul4 were prepared by cloning sequences encoding residues 1 - 418 \n of cul3 and 1 - 440 of cul4a into pcdna3 ( invitrogen ) , respectively . \n -trcp1f , fbw7f , and skp2f plasmids were previously \n described ( 14 , \n 20 , \n 21 ) , and -trcp2f \n was a gift from n. khidekel ( mit ) . \n pretrosuper ( prs)-sh-trcp and control \n shgfp plasmids were previously published \n ( 14 ) . \n t cells were grown in dulbecco 's modified eagle 's \n medium + 10% serum and seeded ( 1 10 cells / well of a 6-well \n dish ) 16 h before transfection with lipofectamine 2000 ( invitrogen ) . \n cells \n were typically harvested 24 - 30 h post - transfection , except for rnai \n experiments where cells were harvested 96 h post - transfection after a 48-h \n selection with puromycin ( 1 g / ml ) to enrich for a transfected population . \n treatment of transfected cells with cycloheximide ( 25 g / ml ) began 96 h \n post - transfection . for assays examining myc - spar abundance and turnover , \n cell pellets were \n lysed in 50 mm tris , ph 7.5 , 150 mm nacl , 1 \n mm edta , 0.5% deoxycholate , 1% nonidet p-40 , 0.1% sds , and cleared \n lysates were resolved on 4 - 12% gradient tris - glycine sds - page gels . \n resolved \n proteins were then transferred onto nitrocellulose ( 250 ma , 2 h ) and \n immunoblotted with c - myc 9e10 \n ( sc-40 , santa - cruz ) , ha f-7 ( sc-7392 , santa \n cruz ) , cdk2 m2 ( sc-163 , santa cruz ) , -trcp1 ( cell signaling ) , cul1 \n ( 71 - 8700 , invitrogen / zymed laboratories inc . ) , and cdc25a ab-3 ( ms-640-p0 , \n neomarkers ) , as indicated . \n promega horseradish peroxidase - conjugate anti - mouse \n igg ( w402b ) and anti - rabbit igg ( w401b ) were used for secondary detection . for single - cell immunofluorescence rnai experiments , \n hek293 t cells were \n seeded onto 18-mm glass coverslips coated with poly - d - lysine ( 30 \n g / ml ) and laminin ( 2 g / ml ) and transfected at 20 - 30% confluency using \n calcium phosphate . cells were fixed 96 h post - transfection for 10 min at room \n temperature using 4% paraformaldehyde and 4% sucrose in phosphate - buffered \n saline , and the gfp - spar signal was amplified using anti - gfp ( a-11122 , \n invitrogen ) at 1:1000 in gdb buffer ( 1% gelatin , 5% triton x-100 , 50 \n mm phosphate buffer , ph 7.4 , 2 m nacl ) \n ( 6 ) . binding and \n ubiquitination detection assays were performed with cleared \n whole cell lysates from transfected hek293 t cells in 50 mm tris , ph \n 7.5 , 150 mm nacl , 1 mm edta , 0.5% deoxycholate , and 1% \n nonidet p-40 . in the case of ubiquitination detection assays , cells were \n treated with 25 m mg-132 before harvesting , and 10 \n mm n - ethylmaleimide was added to the lysis buffer . \n gsh - sepharose or c - myc 9e10-agarose beads ( 10 l per condition ) were \n pre - washed in lysis buffer and incubated with 400 g of lysate for 2 h at 4 \n c while rocking . \n beads were then washed 3 - 4 times in lysis buffer , and \n bound protein was eluded in 2 laemmli protein loading buffer containing \n sds . \n both gsh - bound samples and crude extract were resolved on 4 - 12% gradient \n sds - page gels . \n myc - bound samples were resolved on 6% tris - glycine sds - page \n gels . \n samples were then transferred from the sds - page onto nitrocellulose and \n immunoblotted with anti - myc , anti - ha , anti - flag ( f3165 , sigma ) , or anti - gst \n ( 26h1 , cell signaling ) , as indicated . \n neuron cultures and immunostaining medium - density \n dissociated hippocampal cultures were prepared and cultured from e19 long \n evans rat hippocampi as previously described \n ( 5 ) . \n neurons were transfected \n at div16 , super - infected at div18 , and fixed 18 h \n post - infection ( div19 ) in 1% paraformaldehyde for 2 min at room temperature \n followed by -20 c methanol for 10 min . \n immunostaining was performed in \n gdb buffer ( 6 ) using rabbit \n spar polyclonal antibodies ( 5 ) \n and anti - flag m2 ( sigma ) . \n microscopy and quantification fixed neurons and hek293 t \n cells were imaged with an lsm510 confocal system ( zeiss ) . \n a 40 oil \n immersion lens was used for confocal microscopy , and each image was comprised \n of 0.5-m z - stacks projected into a single plane . \n spar immunostaining \n analysis was performed with metamorph software and carried out blinded with \n respect to the experimental conditions . \n quantification of spar puncta involved \n subjecting images stained for endogenous spar to threshold and was carried out \n from somatic and proximal dendritic regions from transfected , infected , or \n transfected plus superinfected cells . \n all spar intensity measurements \n represent integrated spar immunostaining intensity per area and are normalized \n to neighboring uninfected and untransfected cells \n plk2-dependent spar degradation requires a cul1-based e3 \n ub - ligase spar turnover in neurons is controlled through the upp in \n a manner that requires active plk2 protein kinase \n ( 6 ) . \n such regulation is \n reminiscent of the mechanism employed by most scf e3 ub - ligases , where \n substrate recognition depends upon upstream kinase signaling cascades . \n a \n central subunit of the scf complex is the cul1 scaffold protein . to explore \n the idea that spar turnover is regulated by an scf e3 ub - ligase , we made use \n of a previously reported dominant - negative version of cul1 \n ( cul1 ) . \n this c - terminal - truncated protein binds substrates but \n fails to associate with ubiquitin - charged e2 ubiquitin conjugating enzymes , \n thereby preventing substrate turnover \n ( 14 , \n 19 , \n 22 ) . \n a - c , dominant negative cul1 constructs block plk2-dependent loss of \n spar in hippocampal neurons . dissociated rat hippocampal neurons ( div16 ) were \n transfected with dominant negative cullin plasmids ( b and c ) \n or a control plasmid ( a ) and super - infected 2 days later with \n flag - tagged plk2 driven by sindbis virus ( sin - plk2 ) . \n neurons were \n fixed 18 h post - infection and immunostained for endogenous spar and \n infected plk2 . \n transfected cells were identified during image acquisition by \n the presence of a co - transfected fill protein ( gfp , seen in the \n first column of images ) . \n spar ( green ) and plk2 ( red ) were \n pseudo - colored for illustrative purposes after image analysis . \n arrows \n point to cells that are both transfected and infected ; arrowheads \n point to cells that are infected only . \n d and e , quantification of \n spar immunostaining in somatic and proximal dendritic regions as integrated \n immunofluorescence intensity per area in cells transfected with indicated \n plasmids and/or infected with plk2 sindbis virus ( sin - plk2 ) , \n normalized to nearby untransfected ( untr ) cells . \n , n > 17 cells for all conditions , \n , p < 0.001 , mann - whitney test ( e ) . \n a - c , dominant negative cul1 constructs block plk2-dependent loss of \n spar in hippocampal neurons . dissociated rat hippocampal neurons ( div16 ) were \n transfected with dominant negative cullin plasmids ( b and c ) \n or a control plasmid ( a ) and super - infected 2 days later with \n flag - tagged plk2 driven by sindbis virus ( sin - plk2 ) . \n neurons were \n fixed 18 h post - infection and immunostained for endogenous spar and \n infected plk2 . \n transfected cells were identified during image acquisition by \n the presence of a co - transfected fill protein ( gfp , seen in the \n first column of images ) . \n spar ( green ) and plk2 ( red ) were \n pseudo - colored for illustrative purposes after image analysis . \n arrows \n point to cells that are both transfected and infected ; arrowheads \n point to cells that are infected only . \n d and e , quantification of \n spar immunostaining in somatic and proximal dendritic regions as integrated \n immunofluorescence intensity per area in cells transfected with indicated \n plasmids and/or infected with plk2 sindbis virus ( sin - plk2 ) , \n normalized to nearby untransfected ( untr ) cells . \n , n > 17 cells for all conditions , \n , p < 0.001 , mann - whitney test ( e ) . cultured hippocampal neurons ( 16 days in vitro ( div16 ) ) were \n transfected with cul1 and then super - infected 2 days later with \n sindbis virus driving expression of flag - tagged plk2 for 18 h to promote \n degradation of endogenous spar \n ( 6 ) . \n we infected at a titer \n that resulted in 10% infection rate of cells already transfected with \n cul1 ( fig . 1 , \n a - c ) . \n the option of co - transfecting plasmids driving plk2 \n expression was precluded by the low plk2 expression achievable using this \n method.6 in the absence of plk2 infection , transfection of control empty vector \n ( pcdna3.1 ) or a control dominant - negative cullin 3 ( cul3 ) , which \n disrupts structurally related but functionally distinct cul3-based complexes \n ( 23 ) , had no effect on \n endogenous spar levels relative to nearby untransfected and uninfected cells \n as assessed by quantitative immunostaining \n ( fig . \n uninfected cells \n overexpressing cul1 displayed a trend toward increased spar \n levels compared with nearby untransfected cells , but did not reach statistical \n significance ( p = 0.13 ; fig . \n 1b , quantified in d ) . \n in plk2-infected but otherwise untransfected neurons , endogenous spar \n levels were close to undetectable ( fig . 1 , \n a - c , arrowheads ; quantified in e ) , in \n agreement with our previous findings \n ( 6 ) . in plk2-infected cells \n that had been previously transfected with control empty vector or \n cul3 \n , spar levels fell to the same extent as in cells only \n infected with plk2 ( fig . \n 1 , a and \n c , compare spar staining in cells marked by \n arrowheads ( infected ) and arrows ( transfected and infected ) ; \n quantified in e ) . however , in cells transfected with \n cul1 , infection with plk2 sindbis virus failed to reduce spar \n levels ( fig . \n 1b , \n notice the yellow color in cells marked by arrows , \n indicating the presence of both spar ( green ) and plk2 ( red ) ; \n quantified in e ) . \n thus , plk2-driven spar degradation in neurons \n depended upon a cul1-based scf complex but not any of the structurally related \n cul3-based ub - ligases . \n spar physically associates with the \n scf complex to further \n explore the idea that spar turnover is regulated by an scf e3 ub - ligase , we \n established a system in cultured hek293 t cells that recapitulates \n plk2-dependent spar degradation . \n this system facilitated biochemical studies \n that were otherwise limited by the physical properties of dendritic spines and \n allowed the use of molecular reagents previously developed for study of the \n human scf pathway ( 24 ) . \n we \n found that expression of spar alone ( as an myc - tagged fusion protein ) led to \n its accumulation in hek293 t cells ( fig . \n in contrast , co - expression with \n plk2 ( but not a catalytically inactive mutant plk2 ) promoted \n the degradation of myc - spar ( fig . \n 2 , \n a , lane 2 compared with lanes 4 - 6 , and \n b , lanes 1 and 2 ) . \n co - expression of spar with wild type plk2 \n correlated with the appearance of a slower mobility form of spar ( presumably \n phosphorylated ) that is sensitive to plk2-induced degradation \n ( fig . \n our recapitulation of plk2-dependent \n spar turnover in hek293 t cells provides a convenient system in which to search \n for components of the plk2-dependent degradation pathway . \n consistent with the idea that spar is a target of one of the scf complexes , \n overexpression of dominant - negative cul1 ( cul1 ) in hek293 t \n stabilized myc - spar despite cotransfection of active plk2 \n ( fig . \n 2b , lane \n 3 ) , in agreement with our findings in neurons ( see \n fig . 1 , b and \n e ) \n . moreover , these cells accumulated the slower \n migrating form of myc - spar that presumably corresponds to phosphorylated spar . \n in contrast , a dominant - negative form of the related cullin4 \n ( cul4 ) had no effect on spar turnover ( lane 4 ) , \n indicating that the effect of cul1 was specific . \n to uncover \n candidate f - box proteins for spar , we performed a cell - based interaction \n screen in hek293 t cells between spar and a panel of co - expressed gst - tagged \n f - box proteins . \n potential complexes between spar and gst - tagged f - box proteins \n were isolated by incubating cell lysates with gsh - sepharose beads . \n previous \n studies had indicated that the interaction of f - box proteins with substrates \n can be detected in tissue culture cells that co - express the f - box protein and \n substrate in the presence of cul1 , which blocks substrate \n degradation ( 14 , \n 19 ) . among a panel of f - box \n proteins individually co - transfected with myc - spar and ha - plk2 , we identified \n an interaction between myc - spar and both gst--trcp1 and gst--trcp2 \n ( fig . 2c , lanes \n 3 and 4 ) . \n our ability to assay for endogenous interactions was \n precluded by the unavailability of suitable antibodies for \n immunoprecipitation . \n although produced from different genes , -trcp1 and \n -trcp2 are 85% identical and are believed to have largely redundant \n functions ( 25 ) . \n importantly , \n the binding of -trcp1 and -trcp2 to myc - spar required \n co - transfection of active ( wild type ( wt ) ) plk2 and was undetectable in the \n presence of catalytically defective plk2 \n ( fig . 2c , lanes \n 5 and 6 ) . together with the plk2-dependent slower gel mobility \n of spar ( fig . \n 2 , a - c ) , \n this suggests that spar associates with -trcp in a \n phosphorylation - dependent manner , as is the case for all other -trcp \n substrates identified to date \n ( 13 ) . \n spar did not associate \n with any of 16 other f - box proteins tested \n ( fig . \n 2c , lanes \n 7 - 22 ) , including 5 f - box proteins , which like -trcp , bind \n substrates through their wd40 repeats ( fig . \n this indicates a high degree of \n specificity in the interaction between spar and -trcp . \n t cells were transfected with 1 g of \n pcmv - ha - plk2 ( lane 1 ) , 1 g of pcmv - myc - spar ( lane 3 ) , or \n 1 g of pcmv - myc - spar together with either 1 g of catalytically \n inactive pcmv - ha - plk2 ( lane 2 ) or increasing amounts \n of pcmv - ha - plk2 ( 0.3 , 1 , or 2 g ) ( lanes 4 - 6 ) . \n the \n total amount of transfected dna was kept constant among all conditions with \n use of empty vector . \n whole cell lysates were immunoblotted with myc antibody \n to assess myc - spar levels . \n pcmv - myc - spar ( 0.5 g ) and \n pcmv - ha - plk2 ( 1 g ) ( catalytically inactive , lane \n 1 ; wild type , lanes 2 - 9 ) were co - expressed in hek293 t cells with \n 2.5 g of either empty vector , dominant negative cullins , or dominant \n negative f - box proteins . \n changes in the abundance of myc - spar were determined \n by immunoblotting with anti - myc antibody . \n pcmv - myc - spar ( 0.6 g ) , pcmv - ha - plk2 \n ( 0.6 g ) , and pcmv - cul1 ( 2 g ) were co - expressed as shown \n with pcmv - gst ( lane 2 ) or the indicated f - box proteins as gst fusions \n ( 0.6 g ) ( lanes 3 - 22 ) in hek293 t cells seeded in 6-well plates . \n after 24 h , cell extracts were used for gsh - sepharose pull - down assays , and \n proteins were immunoblotted with anti - gst and anti - myc antibodies . \n extracts of hek293 t cells transfected with \n pcmv - cul1 and pcmv - myc - spar ( lane 1 ) , pcmv - ha - plk2 \n ( lane 2 ) , or both ( lane 3 ) were immunoprecipitated using \n anti - myc or anti - ha antibodies as indicated . \n . \n lysates from cells transfected with pcmv - myc - spar ( 0.5 g ) , \n pcmv - ha - plk2 ( 0.5 g ) , pcmv - gst--trcp ( 0.5 g ) , \n and pcmv - flag - cul1 ( 2 g ) , as indicated , were incubated \n with gsh - sepharose and immunoblotted with anti - myc , anti - ha , anti - gst , and \n anti - flag antibodies as shown . \n lanes 1 - 4 show lysates ( 6% of input of \n the gsh - sepharose binding reactions ) . \n t cells were transfected with 1 g of \n pcmv - ha - plk2 ( lane 1 ) , 1 g of pcmv - myc - spar ( lane 3 ) , or \n 1 g of pcmv - myc - spar together with either 1 g of catalytically \n inactive pcmv - ha - plk2 ( lane 2 ) or increasing amounts \n of pcmv - ha - plk2 ( 0.3 , 1 , or 2 g ) ( lanes 4 - 6 ) . the \n total amount of transfected dna was kept constant among all conditions with \n use of empty vector . \n whole cell lysates were immunoblotted with myc antibody \n to assess myc - spar levels . \n b , dominant negative versions of cul1 and \n -trcp stabilize spar . pcmv - myc - spar ( 0.5 g ) and \n pcmv - ha - plk2 ( 1 g ) ( catalytically inactive , lane \n 1 ; wild type , lanes 2 - 9 ) were co - expressed in hek293 t cells with \n 2.5 g of either empty vector , dominant negative cullins , or dominant \n negative f - box proteins . \n changes in the abundance of myc - spar were determined \n by immunoblotting with anti - myc antibody . \n pcmv - myc - spar ( 0.6 g ) , pcmv - ha - plk2 \n ( 0.6 g ) , and pcmv - cul1 ( 2 g ) were co - expressed as shown \n with pcmv - gst ( lane 2 ) or the indicated f - box proteins as gst fusions \n ( 0.6 g ) ( lanes 3 - 22 ) in hek293 t cells seeded in 6-well plates . \n after 24 h , cell extracts were used for gsh - sepharose pull - down assays , and \n proteins were immunoblotted with anti - gst and anti - myc antibodies . \n extracts of hek293 t cells transfected with \n pcmv - cul1 and pcmv - myc - spar ( lane 1 ) , pcmv - ha - plk2 \n ( lane 2 ) , or both ( lane 3 ) were immunoprecipitated using \n anti - myc or anti - ha antibodies as indicated . \n lysates from cells transfected with pcmv - myc - spar ( 0.5 g ) , \n pcmv - ha - plk2 ( 0.5 g ) , pcmv - gst--trcp ( 0.5 g ) , \n and pcmv - flag - cul1 ( 2 g ) , as indicated , were incubated \n with gsh - sepharose and immunoblotted with anti - myc , anti - ha , anti - gst , and \n anti - flag antibodies as shown . \n lanes 1 - 4 show lysates ( 6% of input of \n the gsh - sepharose binding reactions ) . \n consistent with previous studies that showed interaction between plk2 and \n spar ( 6 ) , we found that plk2 \n associated with spar in a coimmunoprecipitation assay upon co - expression of \n cul1 ( fig . \n interestingly , we also discovered that the associations of plk2 and \n -trcp with spar were not mutually exclusive , as we were able to detect a \n ternary interaction with plk2 that was not precluded by the -trcp - spar \n interaction ( fig . \n gst--trcp1 ( and gst--trcp2 ) \n associated with spar , plk2 , and the n terminus of cullin 1 \n ( cul1/cul1 ) , whereas catalytically inactive \n plk2 did not support assembly of the full complex \n ( fig . \n moreover , the plk2-dependent slower mobility form of spar was \n enriched in the complex compared with the faster mobility form of spar \n ( fig . \n a and \n b , depletion of -trcp by rnai protects gfp - spar from \n degradation in individual hek293 t cells . \n the indicated plasmids were \n transfected with pcmv - rfp as a co - transfection marker into hek293 t cells . \n after 96 h , cells were fixed and imaged for gfp - spar and rfp expression \n ( a ) and quantified as the percentage of rfp expressing cells that \n also expressed gfp - spar ( b ) . \n values represent the mean s.e . \n from three independent experiments , derived from analysis of 400 cells per \n experiment per condition ( n = 1200 cells per condition ) . \n , p < 0.01 ; ns , not significant ; one - way \n analysis of variance , compared with control condition \n ( prsp+plk2 ) . \n c and d , depletion of -trcp \n by rnai stabilizes spar abundance and turnover in the presence of plk2 \n activity . \n t cells were transfected with vectors expressing myc - spar ( 0.5 \n g ) , wt , or catalytically inactive ha - plk2 ( d / a ) ( 1 g ) , and \n the indicated shrna vector carrying a puromycin resistance selection marker \n ( 2.5 g ) . \n 36 h post - transfection , cells were incubated with media \n containing 1 g / ml puromycin to enrich for shrna expression . \n extracts were \n subsequently examined by immunoblotting 96 h post - transfection as indicated . \n endogenous cdc25a was probed to control for successful knockdown of \n -trcp . in panel d \n , hek293 t cells were transfected in an \n identical fashion to panel c , except that 24 h post - transfection the \n transfected cells were split among 5 wells in puromycin - containing media . \n after 48 h of puromycin selection , cells were treated with 25 g / ml \n cycloheximide ( chx ) and harvested at the indicated times before \n immunoblotting . \n t cells were transfected with myc - spar ( 1 g ) , \n his - ub ( 1 g ) , ha - plk2 ( 1.5 g , wild type or d201a ) , and gst or \n gst--trcp ( 4.5 g ) . \n twenty hours post - transfection cells were treated \n with 25 m mg-132 for 5 h and lysed in buffer containing 10 \n mm n - ethylmaleimide . \n spar was purified with c - myc \n 9e10-agarose , resolved on 6% tris - glycine sds - page gel , and immunoblotted \n ( ib ) with anti - myc antibodies . \n a and \n b , depletion of -trcp by rnai protects gfp - spar from \n degradation in individual hek293 t cells . \n the indicated plasmids were \n transfected with pcmv - rfp as a co - transfection marker into hek293 t cells . \n after 96 h , cells were fixed and imaged for gfp - spar and rfp expression \n ( a ) and quantified as the percentage of rfp expressing cells that \n also expressed gfp - spar ( b ) . \n values represent the mean s.e . \n from three independent experiments , derived from analysis of 400 cells per \n experiment per condition ( n = 1200 cells per condition ) . \n , \n p < 0.01 ; ns , not significant ; one - way \n analysis of variance , compared with control condition \n ( prsp+plk2 ) . \n c and d , depletion of -trcp \n by rnai stabilizes spar abundance and turnover in the presence of plk2 \n activity . \n t cells were transfected with vectors expressing myc - spar ( 0.5 \n g ) , wt , or catalytically inactive ha - plk2 ( d / a ) ( 1 g ) , and \n the indicated shrna vector carrying a puromycin resistance selection marker \n ( 2.5 g ) . \n 36 h post - transfection , cells were incubated with media \n containing 1 g / ml puromycin to enrich for shrna expression . \n extracts were \n subsequently examined by immunoblotting 96 h post - transfection as indicated . \n endogenous cdc25a was probed to control for successful knockdown of \n -trcp . in panel d \n , hek293 t cells were transfected in an \n identical fashion to panel c , except that 24 h post - transfection the \n transfected cells were split among 5 wells in puromycin - containing media . \n after 48 h of puromycin selection , cells were treated with 25 g / ml \n cycloheximide ( chx ) and harvested at the indicated times before \n immunoblotting . \n t cells were transfected with myc - spar ( 1 g ) , \n his - ub ( 1 g ) , ha - plk2 ( 1.5 g , wild type or d201a ) , and gst or \n gst--trcp ( 4.5 g ) . \n twenty hours post - transfection cells were treated \n with 25 m mg-132 for 5 h and lysed in buffer containing 10 \n mm n - ethylmaleimide . \n myc - spar was purified with c - myc \n 9e10-agarose , resolved on 6% tris - glycine sds - page gel , and immunoblotted \n ( ib ) with anti - myc antibodies . \n scf regulates spar abundance \n and turnover and promotes its ubiquitination to validate a role for \n the scf complex in plk2-dependent spar turnover , we next \n examined spar abundance in hek293 t cells after expression of dominant - negative \n -trcp ( -trcpf ) ( fig . \n the f - box construct is unable to assemble \n with cul1 due to absence of the f - box motif , but it maintains its ability to \n interact with substrates of both -trcp1 and -2 and can thereby sequester \n substrates and block their turnover \n ( 14 , \n 21 , \n 24 ) . \n expression of \n -trcpf resulted in increased levels of spar , especially of the \n slower migrating form of spar that is dependent upon plk2 activity \n ( fig . \n in contrast , f - box dominant - negative versions of other f - box \n proteins fbw7// ( containing wd40 repeats ) and skp2 \n ( containing leucine - rich repeats ) failed to promote an increase in the steady \n state abundance of spar ( fig . \n 2b , lanes 6 - 9 ) . to directly examine whether -trcp proteins are required for \n plk2-dependent spar turnover \n , we took advantage of a shrna vector \n ( sh-trcp ) that is capable of suppressing protein expression of both \n human -trcp1 and -trcp2 . this hairpin sequence and this particular \n shrna vector \n have been validated for numerous -trcp substrates \n ( 14 , \n 19 , \n 26 ) . \n cells were transfected \n with expression constructs for gfp - spar and red fluorescent protein ( rfp ) to \n mark transfected cells and simultaneously transfected with sh-trcp or \n control vector ( prsp ) in the presence of active plk2 or a catalytically \n inactive version , plk2 \n ( fig . \n we \n subsequently visualized cells for the presence of gfp - spar in rfp - positive \n cells ( fig . \n rfp - positive cells expressing plk2 , but not those expressing catalytically \n inactive plk2 , displayed very low levels of gfp - spar in the \n presence of the control shrna vector prsp \n ( fig . \n in contrast , rfp - positive cells transfected with sh-trcp \n contained high levels of gfp - spar despite the co - transfection of active plk2 \n ( fig . \n thus , depletion of -trcp by rnai substantially protected \n gfp - spar from plk2-dependent degradation . \n immunoblotting of transfected cell \n lysates confirmed that plk2-induced degradation of myc - spar requires \n -trcp ( fig . \n 3c ) . \n as expected , plk2 promoted loss of myc - spar in cells expressing a control \n shrna that targets gfp ( shgfp ) ( lanes 1 and 2 ) . \n in contrast , \n myc - spar was not efficiently degraded in cells depleted of -trcp despite \n the presence of active plk2 ( lane 4 ) . furthermore , depletion of \n -trcp led to accumulation of cdc25a , a known target of \n scf used here as a positive control \n ( 14 , \n 18 ) . \n figure 4.a candidate-trcp phosphodegron in spar . a , \n schematic representation of spar domains and spar fragments : actin - binding \n domains ( act1 and act2 ) , rapgap , pdz , and guanylate kinase \n binding ( gkbd ) domain . \n the candidate dsgidt phosphodegron motif \n ( residues 1304 - 1309 ) identified in the act2 domain of spar closely resembles \n the consensus -trcp phosphodegron ( inset ) . boundaries of \n generated c - terminal fragments of spar are depicted ; c-1 , c-2 , and act2 \n fragments contain the putative phosphodegron motif , whereas c-3 does not . \n hek293 t cells were transfected with vectors \n expressing spar fragments c-1 , c-2 , c-3 ( b ) or act2 ( c ) ( 0.6 \n g ) either alone or with pcmv - gst or pcmv - gst--trcp ( 0.6 g ) . in \n addition , pcmv - ha - plk2 ( 0.6 g ) and cul1 ( 2 \n g ) were co - transfected as indicated . \n after 24 h , cell extracts were used \n for gsh - sepharose pulldown assays , and proteins were immunoblotted with myc \n antibodies . \n constructs \n expressing point mutations ( s1305a , t1309a ) in full - length spar ( e ) \n and the act2 fragment ( d ) ( myc - spar or \n myc - act2 , 0.6 g ) were transfected into hek293 t cells along \n with pcmv - ha - plk2 ( 0.6 g ) , pcmv - cul1 ( 2 \n g ) , and pcmv - gst or pcmv - gst--trcp ( 0.6 g ) as indicated . \n cell \n lysates were incubated with gsh - sepharose and immunoblotted with myc \n antibodies . \n wild type \n constructs expressing myc - spar and myc - act2 were used for comparison as a \n positive control for interaction with gst--trcp . \n f , spar \n associates with endogenous scf complex in the presence \n of active plk2 and dependent upon its phosphodegron . \n constructs expressing \n full - length myc - spar ( wt or aa ) were co - expressed in hek293 t cells \n with active ( wt ) or catalytically inactive plk2 ( d201a ) . before lysis and \n immunoprecipitation with 9e10-agarose , \n cells were treated with proteasome \n inhibitor mg-132 ( 25 m ) for 5 h. proteins bound to 9e10-agarose \n were analyzed via immunoblotting using myc antibodies and antibodies that \n recognized endogenous -trcp1 and cul1 . \n myc - spar - c3 , a fragment of spar \n that does not contain the phosphodegron served as a negative control , and \n myc - cdc25a , a known target of scf , served as a positive \n control . \n myc - sparwt and myc - spar abundance was compared by \n immunoblotting with the indicated antibodies in hek293 t cells in the absence \n and presence of pcmv - ha - plk2 in the background of empty vector or \n -trcpf . a candidate-trcp phosphodegron in spar . a , \n schematic representation of spar domains and spar fragments : actin - binding \n domains ( act1 and act2 ) , rapgap , pdz , and guanylate kinase \n binding ( gkbd ) domain . \n the candidate dsgidt phosphodegron motif \n ( residues 1304 - 1309 ) identified in the act2 domain of spar closely resembles \n the consensus -trcp phosphodegron ( inset ) . boundaries of \n generated c - terminal fragments of spar are depicted ; c-1 , c-2 , and act2 \n fragments contain the putative phosphodegron motif , whereas c-3 does not . \n hek293 t cells were transfected with vectors \n expressing spar fragments c-1 , c-2 , c-3 ( b ) or act2 ( c ) ( 0.6 \n g ) either alone or with pcmv - gst or pcmv - gst--trcp ( 0.6 g ) . in \n addition , pcmv - ha - plk2 ( 0.6 g ) and cul1 ( 2 \n g ) were co - transfected as indicated . \n after 24 h , cell extracts were used \n for gsh - sepharose pulldown assays , and proteins were immunoblotted with myc \n antibodies . \n constructs \n expressing point mutations ( s1305a , t1309a ) in full - length spar ( e ) \n and the act2 fragment ( d ) ( myc - spar or \n myc - act2 , 0.6 g ) were transfected into hek293 t cells along \n with pcmv - ha - plk2 ( 0.6 g ) , pcmv - cul1 ( 2 \n g ) , and pcmv - gst or pcmv - gst--trcp ( 0.6 g ) as indicated . \n cell \n lysates were incubated with gsh - sepharose and immunoblotted with myc \n antibodies . \n wild type \n constructs expressing myc - spar and myc - act2 were used for comparison as a \n positive control for interaction with gst--trcp . \n f , spar \n associates with endogenous scf complex in the presence \n of active plk2 and dependent upon its phosphodegron . \n constructs expressing \n full - length myc - spar ( wt or aa ) were co - expressed in hek293 t cells \n with active ( wt ) or catalytically inactive plk2 ( d201a ) . before lysis and \n immunoprecipitation with 9e10-agarose , \n cells were treated with proteasome \n inhibitor mg-132 ( 25 m ) for 5 h. proteins bound to 9e10-agarose \n were analyzed via immunoblotting using myc antibodies and antibodies that \n recognized endogenous -trcp1 and cul1 . \n myc - spar - c3 , a fragment of spar \n that does not contain the phosphodegron served as a negative control , and \n myc - cdc25a , a known target of scf , served as a positive \n control . \n myc - sparwt and myc - spar abundance was compared by \n immunoblotting with the indicated antibodies in hek293 t cells in the absence \n and presence of pcmv - ha - plk2 in the background of empty vector or \n -trcpf . to directly examine whether -trcp is required for spar turnover \n , we \n performed a cycloheximide - chase experiment in cells expressing myc - spar and \n plk2 in the presence of shgfp or sh-trcp \n ( fig . \n spar \n levels persisted after 45 - 60 min of cycloheximide treatment under conditions \n of -trcp depletion ( sh-trcp ) but disappeared after the same time \n period in control cells ( shgfp ) ( fig . \n 3d , lanes 7 - 8 compared with 3 and \n 4 ) . taken together , \n these data indicate that \n scf is critical for plk2-dependent degradation of spar \n in heterologous cells . to explore whether the scf complex promoted \n ubiquitination of spar \n , we immunoprecipitated myc - spar from mg-132-treated \n hek293 t cells and immunoblotted for modified myc - spar \n ( fig . \n 3e ) . expression \n of active plk2 resulted in detection of ubiquitinated spar , and the \n ubiquitination reaction was further driven in cells by ectopic expression of \n gst--trcp ( lanes 2 and 3 ) . \n together , this suggests \n that the plk2-dependent turnover of myc - spar occurs through ubiquitination of \n spar . \n spar is a large protein of 1804 amino acids \n containing two actin binding domains ( act1 and act2 ) , a rapgap domain , a pdz \n domain , and a guanylate kinase binding domain ( gkbd ) \n ( fig . \n , we identified a candidate -trcp recognition motif ( dsgidt , \n residues 1304 - 1309 ) based upon the consensus -trcp recognition motif \n found in many of its targets ( dpsgx(ps / t ) ; = hydrophobic \n residue , x = any residue , ps or ps / t = phosphoserine or threonine ) \n ( fig . \n 4a ) . initially , \n we surveyed three fragments of spar spanning the c terminus ( c-1 , c-2 , c-3 , \n fig . \n 4a ) for their \n ability to interact with co - transfected gst--trcp1 in hek293 t cells in \n the presence of plk2 and cul1 . \n fragments c-1 and c-2 ( which \n contain the candidate phosphodegron ) bound to gst--trcp , whereas the c-3 \n construct lacking the phosphodegron failed to do so \n ( fig . \n n - terminal \n spar fragments that lack the dsgidt motif also could not bind to -trcp \n ( data not shown ) . \n a - c , \n dominant negative -trcp constructs block plk2-dependent loss of spar in \n hippocampal neurons . as in fig . \n 1 , div16 dissociated rat hippocampal neurons were transfected with \n dominant negative -trcp plasmids ( a and b ) or control \n skpf plasmid ( c ) and superinfected 2 days later with \n flag - tagged plk2 driven by sindbis virus ( sin - plk2 ) . \n arrows \n point to cells that are both transfected and infected ; arrowheads \n point to cells that are infected only . \n d and e , quantification of \n spar immunostaining in somatic and proximal dendritic regions , as in \n fig . \n n = 25 - 37 cells for all constructs in panel \n d ; , p < 0.05 indicates significant difference \n from theoretical mean of 100% , student 's t test ( d ) . for \n panel e , n = 19 - 37 cells ( infected only ) , n = 13 - 28 cells \n ( infected and transfected ) , , p < 0.001 , mann \n whitney test . \n a - c , \n dominant negative -trcp constructs block plk2-dependent loss of spar in \n hippocampal neurons . as in fig . \n 1 , div16 dissociated rat hippocampal neurons were transfected with \n dominant negative -trcp plasmids ( a and b ) or control \n skpf plasmid ( c ) and superinfected 2 days later with \n flag - tagged plk2 driven by sindbis virus ( sin - plk2 ) . \n arrows \n point to cells that are both transfected and infected ; arrowheads \n point to cells that are infected only . \n d and e , quantification of \n spar immunostaining in somatic and proximal dendritic regions , as in \n fig . \n n = 25 - 37 cells for all constructs in panel \n d ; , p < 0.05 indicates significant difference \n from theoretical mean of 100% , student 's t test ( d ) . for \n panel e , \n n = 19 - 37 cells ( infected only ) , n = 13 - 28 cells \n ( infected and transfected ) , , p < 0.001 , mann \n whitney test . \n a fourth fragment of spar spanning the act2 domain was sufficient to \n interact with -trcp , and its association depended upon expression of \n active plk2 ( fig . \n mutation of ser-1305 and thr-1309 to alanines within the act2 domain fragment \n alone ( act2 ) ( fig . \n 4e ) resulted in \n greatly diminished binding to -trcp compared with wild type spar \n constructs . \n moreover , only in the presence of catalytically active plk2 was \n full - length spar able to associate with endogenous -trcp and its \n associated cul1 protein ( fig . \n importantly , the c-3 fragment of spar that does not encompass the \n phosphodegron as well as full - length spar that carries point mutations of \n ser-1305 and thr-1309 did not support interaction with components of the \n endogenous scf complex \n ( fig . \n if this dsgidt motif in spar is important for spar \n turnover , then mutation of this candidate phosphodegron should render spar \n resistant to plk2-mediated turnover . \n indeed , plk2 efficiently promoted \n degradation of wild type spar but not the spar mutant \n ( fig . \n importantly , the accumulation of spar seen in the \n presence of plk2 was unchanged by the additional cotransfection of \n -trcpf ( compare lanes 4 and 8) , indicating that \n the mutation is specifically protecting turnover via the \n scf pathway . \n we further noted that spar \n continued to migrate as multiple bands upon sds - page analysis of extracts from \n cells expressing plk2 ( fig . \n this implies \n that the mobility shift seen with plk2 expression does not solely rely on \n phosphorylation of residues ser-1305 and thr-1309 . \n scf controls spar abundance \n in hippocampal neurons to investigate the role of the \n scf complex in post - mitotic neurons of the central \n nervous system , we first examined whether -trcp is present in the adult \n rat brain . \n mrnas encoding -trcp1 , -trcp2 , and cul1 are all present \n at high levels in rat brain , specifically in the ca1-ca3 regions of the \n hippocampus , the dentate gyrus , and cerebral cortex ( allen brain atlas ) . \n unlike for the human mrnas , we have yet to identify an shrna sequence \n that can target both -trcp1 and -trcp2 transcripts efficiently in \n rat neurons . \n co - transfection of different shrnas that individually target \n -trcp1 and -trcp2 has not yielded satisfactory knockdown of both \n proteins , possibly due to low efficiency of cotransfection of both shrna \n vectors in the same neuron . therefore , we opted to disrupt endogenous \n scf function by overexpression of dominant - negative \n -trcp constructs -trcp1f or -trcp2f . \n , div16 cultured \n hippocampal neurons were transfected with either -trcp1for \n -trcp2f and then super - infected 2 days later with sindbis virus \n to drive expression of flag - tagged plk2 and induce spar degradation \n ( fig . 5 ) . \n transfection of control dominant - negative skp2 f - box construct \n ( skp2f ) in the absence of plk2 infection had no effect on endogenous \n spar levels relative to nearby untransfected and uninfected cells \n ( fig . \n in contrast , uninfected cells overexpressing \n -trcp1f displayed higher levels of spar compared with nearby \n untransfected cells ( p < 0.05 , \n fig . \n spar levels trended upwards , but this did \n not reach statistical significance ( p = 0.11 , \n fig . \n these \n results suggest that there may be some scf \n complex - dependent turnover of endogenous spar in hippocampal neurons cultured \n under basal conditions . as expected , endogenous spar levels were close to undetectable in \n plk2-infected neurons that were otherwise untransfected \n ( fig . \n 5 , a - c , \n arrowheads , and e ) as well as in plk2-infected cells that \n had been previously transfected with skp2f \n ( fig . 5c , compare spar \n staining in cells marked by arrowheads ( infected ) and arrows \n ( transfected and infected ) ; quantified in e ) . \n in contrast , cells \n transfected with -trcp1f or -trcp2f showed no \n reduction in spar levels after infection with plk2 sindbis virus \n ( fig . 5 , b and \n c , quantified in e ) . \n these results are \n consistent with disruption of the scf complex through expression of \n cul1 , which also prevented plk2-dependent loss of spar ( see \n fig . \n 1 ) , and together they \n demonstrate that the scf pathway is required for \n plk2-induced spar degradation in neurons . because changes in spar expression can affect spine morphology \n ( 5 ) and synaptic strength \n ( 43 ) , we wondered if blocking \n the scf pathway might also affect these aspects of \n neuronal function . \n infection of div16 neurons with plk2 led to a significant \n decrease in the size and number of spines , consistent with previous results \n ( 5 ) . \n the additional \n overexpression of -trcpf in cells infected with plk2 , however , \n did not prevent the decrease in size and number of spines ( supplemental fig . \n similarly , blocking -trcp function had no effect on \n synaptic strength as assessed by recording of miniature excitatory \n postsynaptic currents ( mepscs ) ( supplemental fig . \n these results are perhaps not surprising given that \n scf is likely to regulate additional synaptic substrates \n besides spar . \n although we have clearly established the \n plk2-scf - spar degradative pathway , its function in \n neurons remains to be resolved . \n the upp has received increasing attention in neurobiology as an important \n regulator of synapse development , synaptic transmission and plasticity , and \n the dynamic turnover of psd proteins \n ( 27 - 31 ) . \n aberrant upp function has been implicated in the pathogenesis of certain \n neurodegenerative disorders , e.g. parkinson , alzheimer , huntington , \n and prion diseases , and amyotrophic lateral sclerosis \n ( 32 , \n 33 ) . \n to date most e3 substrate \n pairs in neurobiology have been identified in caenorhabditis elegans \n and drosophila melanogaster \n ( 34 - 42 ) , \n and relatively little is known about the specific e3s that regulate synaptic \n proteins in the mammalian brain . in this study we have uncovered a role for the e3 ub - ligase \n scf in the degradation of a key postsynaptic scaffolding \n protein and enzyme ( the rapgap spar ) that regulates the morphology of \n dendritic spines and the strength of synaptic transmission in hippocampal \n neurons ( 5 , \n 43 ) . \n mammalian \n scf has been implicated in the ubiquitin - dependent \n turnover of several cell cycle and other signaling proteins , including \n ib , -catenin , cdc25a , emi1 , and the period protein \n ( 14 - 17 , \n 19 ) . \n moreover , it has been \n connected to the differentiation of neural progenitors through its targeting \n of rest transcription repressor \n ( 44 , \n 45 ) . although we have defined \n the residues that are critical for scf - mediated \n degradation of spar , it is possible that spar degradation is also regulated by \n additional e3s , possibly in response to other signals and involving different \n determinants in the spar protein . \n previous studies have demonstrated that -trcp interacts with its \n substrates in a phosphorylation - dependent manner . \n the majority of -trcp \n substrates identified to date contain a dpsgxps sequence ( where \n ps is phosphoserine ) that functions as a phosphodegron and directly binds to \n the wd40 repeats of -trcp \n ( 46 ) . \n a variety of kinases \n have been implicated in the phosphorylation of -trcp phosphodegrons , and \n in many cases , multiple kinases collaborate to generate the phosphodegron , \n e.g. gsk3/cki ( for -catenin ) , chk1 and another unknown \n kinase ( for cdc25a ) , and cdc2/plk1 ( for emi1 ) \n ( 14 , \n 15 , \n 47 ) . \n interestingly , emi1 is \n one of several cell cycle regulators , in addition to wee1 , emi2 , and claspin , \n that is phosphorylated by plk family member plk1 for degradation by the \n scf complex \n ( 47 - 55 ) . \n thus , plk family members seem to collaborate with -trcp to regulate the \n degradation of a variety of substrates in different cellular contexts . \n we found that spar contains a canonical -trcp phosphodegron sequence \n ( dsgidt ) in its act2 domain . \n point mutations in ser-1305 and \n thr-1309 within spar inhibited plk2-dependent spar degradation and its binding \n to -trcp , providing evidence that this sequence functions as a \n phosphodegron . \n currently , the identity of the kinase that phosphorylates \n dsgidt in spar is not definitively established . a direct \n demonstration that plk2 phosphorylates ser-1305 and \n thr-1309 is currently \n lacking due , in part , to difficulty in purifying modified full - length spar . \n thus , further studies are required to definitively correlate that these \n candidate sites are phosphorylated by plk2 . \n nevertheless , because plk2 \n promotes the spar--trcp interaction and can phosphorylate spar in \n vitro ( 6 ) , we favor the \n idea that plk2 is the kinase responsible for phosphorylation of ser-1305 and \n thr-1309 in this phosphodegron . \n supporting this idea is that plk2 also binds \n to the act2 domain of spar ( 6 ) , \n so the kinase would be recruited to the vicinity of the phosphodegron . \n however , we can not rule out the possibilities that plk2 activity is required \n but not sufficient for phosphorylation of ser-1305 and thr-1309 , that \n additional kinases are involved , or that additional phosphodegrons exist in \n spar . \n plk2 is thus far the only kinase reported to promote spar degradation in \n neurons ( 6 ) , although it is \n possible that other kinases could contribute as well . during early \n xenopus and zebrafish development , \n casein kinase i has been \n implicated in the decrease of spar protein through wnt signaling \n ( 56 ) . \n spar levels influence spine shape and size \n ( 5 , \n 6 ) , and spar degradation is \n involved in activity - dependent synaptic scaling \n ( 43 ) . \n our results here show \n that the scf complex targets spar for turnover in \n response to plk2 activity \n . nevertheless , blocking plk2-dependent spar \n degradation through overexpression of the dominant - negative -trcp \n construct did not prevent plk2-dependent changes in spine morphology . \n this \n suggests that the exact role of the plk2-scf - spar \n pathway in neurons is still unclear . \n plk2 may regulate the function or \n expression of additional proteins besides spar that are unaffected by \n disruption of the -trcp degradation pathway . \n similarly , by globally \n disrupting the scf complex in the neuron , additional \n -trcp substrates may accumulate that offset the effects of accumulated \n spar protein . in c. elegans , the -trcp ortholog lin-23 targets \n -catenin / bar-1 for destruction in the ventral nerve cord , \n leading to altered glutamate receptor density \n ( 35 ) . \n endogenous plk2 expression is induced during periods of elevated neuronal \n activity and causes degradation of spar protein as well as thinning and \n elongation of spines ( 6 ) . \n the \n accumulation of spar in spines upon inhibition of the \n scf pathway as well as the recent report that \n proteasomes redistribute into spines with synaptic stimulation \n ( 11 ) is consistent with the \n possibility that phosphorylation of spar by plk2 leads to local interaction \n with -trcp and degradation of spar at postsynaptic sites . \n because spar \n is a large scaffolding protein in the psd that forms complexes with \n n - methyl - d - aspartate receptors and psd-95 \n ( 5 ) , regulation of spar \n degradation through the scf complex may be predicted to \n play a significant role in regulating the composition of proteins in the \n spine . given the critical role of the proteasome in turning over many psd \n proteins ( 10 ) , it is likely \n that additional specific e3s will be identified that regulate synaptic \n composition . \n ", "role": "user" }, { "content": "the ubiquitin - proteasome pathway ( upp ) regulates synaptic function , but \n little is known about specific upp targets and mechanisms in mammalian \n synapses . \n we report here that the scf-trcp complex , a \n multisubunit e3 ubiquitin ligase , targets the postsynaptic spine - associated \n rap gtpase activating protein ( spar ) for degradation in neurons . \n spar \n degradation by scf-trcp depended on the activity - inducible \n protein kinase polo - like kinase 2 ( plk2 ) . in the presence of plk2 , spar \n physically associated with the scf-trcp complex through a \n canonical phosphodegron . in hippocampal neurons , disruption of the \n scf-trcp complex by overexpression of dominant interfering \n -trcp or cul1 constructs prevented plk2-dependent degradation of spar . \n our results identify a specific e3 ubiquitin ligase that mediates degradation \n of a key postsynaptic regulator of synaptic morphology and function .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: this cross - sectional and case register study was conducted in isfahan which is a large province in iran , with a population of almost 4,559,256 in 2007 census ( 51.2% males).10 we reviewed documents of all patients with a pathologically confirmed diagnosis of crc who had been admitted to isfahan referral hospitals , either for surgery , chemotherapy , radiotherapy , or diagnostic evaluations , between 1995 and 2006 . in addition , we reviewed our crc registry at poursina hakim research institute which has been registering crc pa - tients in the province of isfahan in collaboration with vice chancellor of health of isfahan university of medical sciences , since 2003 . \n the results presented in this report are related to patients documents until the year 2008 . \n the study was approved by the ethics committee of isfahan university of medical sciences and informed consents were obtained from patients or their families participating in the study . \n we found hnpcc families based on the amsterdam ii criteria including at least three members in a family having a cancer asso - ciated with hnpcc ( colorectal , endometrial , small bowel , ureteral , renal , and pelvic can - cers ) in at least two consecutive generations , and one of these three members being a first - degree relative of the other two and at least one diagnosed before the age of 50 years.7 the fcc families were characterized by the pres - ence of at least one individual with crc whose diagnosis of cancer was under 50 years of age and another first / second degree relative with colonic / extracolonic tumors associated with the hnpcc . \n patients with fap or those with underlying inflammatory bowel disease ( ibd ) were excluded . \n hnpcc and fcc families ( patients or their siblings / parents in case the patient was dead or not available ) were interviewed and total number of members , number of patients with crc , number of patients with extracolonic tumors , primary sites of the extracolonic tu - mors , and ages at diagnosis were evaluated . \n diagnosis of extraco - lonic tumors was based on history taking from the family members . \n data was analyzed by spss for windows ( version 16.0 ) using chi - square and t - tests . \n during 1996 to 2008 , a total of 2580 crc cases have been diagnosed in isfahan . among thes cases , \n the total num - bers of individuals belonging to hnpcc and fcc families were 324 and 1132 members , re - spectively ( 755 males and 701 females ) . \n the most common age of cancer occurrence ranged from 40 to 55 years . in hnpcc families , the mean age of cancer oc \n - currence with colon origin was 48.0 14.6 years ( range : 23 - 90 years ) and for fcc families it was 49.0 13.9 years ( range 26 - 84 years ) ( p = 0.726 ) . \n age distribution of crc occurrence in hnpcc and fcc cases the total numbers of observed tumors were 70 ( 21.6% ) and 157 ( 13.8% ) in the hnpcc and fcc families , respectively . \n in addition , crc was found in 52 and 76 cases of the hnpcc and fcc families , respectively . \n the prevalence of hnpcc and fcc was therefore calculated as 2.0% ( 52/2580 ) and 2.9% ( 76/2580 ) , respec - tively . for extracolonic involvements , \n the mean age of cancer occurrence was 53.6 11.0 years ( range : 36 - 70 years ) in hnpcc families and 54.8 18.0 years ( range : 17 - 97 years ) for fcc families ( p = 0.801 ) . in both groups , can - cer with colon origin was the most observed tumor . \n in fact , it constituted 52 ( 74.2% ) and 76 ( 48.4% ) patients in the hnpcc and fcc fami - lies , respectively . among extracolonic cancers in fcc families , \n gastric cancer was the most frequent ( 12.1% ) while stomach and uterine were the most frequent sites for extracolonic cancers ( each one 7.1% ) among hnpcc fami - lies . \n as table 1 shows , multiple tumors were present in 7 patients , one patient had 3 tumors and 6 patients had 2 . \n the aim of the present study was to determine the frequency of hnpcc and fcc , as well as the frequency of extracolonic cancers in the family members in central part of iran ( isfahan ) . with the study of 2580 crc cases , we found the frequency of hnpcc and fcc as 2.0% and 2.9% , respectively . \n the first study in this regard has been reported by mahdavinia et al . from tehran . \n these investigators evaluated 449 crc patients with pathologically confirmed crc and found hnpcc in 21 ( 4.7% ) probands . \n they suggested that family history of crc was more frequently reported by early - onset ( 45 years ) than by late - onset ( > 45 years ) patients ( 29.5% vs. 12.8% ) . \n they also found family history of cancer in 53.6% of early - onset and 42.7% of late - onset crc patients with breast ( 9.8% and 5% ) , stomach ( 8% and 5% ) and lung cancers ( 4.5% and 7% ) to be the commonest extracolonic cancers among family members of patients with early - onset and late - onset crc , respectively.4 another recent report from northern iran which included 293 crc cases found hnpcc in 10.9% and 2.7% of the cases based on either amsterdam i or ii criteria . \n hnpcc cases in that study included 25.5% of those aged < 45 years and 3.6% of those aged \n 45 years.11 in our study , we evaluated nearly all families with hnpcc and families with dominant hereditary predisposition in the province of isfahan . \n the age of cancer occurrence in hnpcc and fcc families was within the range de - scribed in other studies.91213 we found that in hnpcc families , crc represents more than 70% of all tumors that is much higher than 16.9% reported by the other study in iran,4 and 50% reported from other countries.14 another study from iran which included 1138 cases of crc from 2000 to 2007 reported family history of cancer in 35.1% and crc history in 4.3% of the crc cases relatives . \n however , it did not report data on hnpcc or fcc.15 the variation in reported frequencies of hnpcc and fap could be due to different genetic back - grounds,16 and also to differences in the ability of diagnoses of crc among family members.11 while in other studies from iran subjects were from a limited geographic region , our study included a large province and the largest number of crc cases from iran . \n we also found a high frequency of uterine cancer in our population which was expected as a result of its previously demonstrated rela - tion with hnpcc . \n moreover , a high incidence of gastric cancer was revealed in our study population although there has not been a defi - nite finding in previous studies . despite the fact that studies in western countries did not show this incidence to be higher than normal population,14 asian studies confirmed it.1718 another cancer which is still a controversy to consider as \n a part of hnpcc tumor spectrum is the breast cancer.81920 it did not show a high incidence in comparison to other extracolonic tumors in our study . \n a number of recent sur - veys which suggest including this type of can - cer in hnpcc tumor spectrum are mainly based on molecular studies.1920 however , it is important to know that the high incidence of cancer among the families is more essential than presence of mlh1 mutation because this mutation is in relation with other types of can - cers which are not associated to hnpcc.21 some other studies found higher incidences of breast cancer maybe due to closer screening.14 another study on hnpcc in iran also found family history of breast cancer in 6.9% of crc patients.4 nevertheless , we believe that there is still not enough evidence to include breast can - cer as a part of hnpcc tumor spectrum . \n the occurrence of rare tumors ( a part of the hnpcc spectrum ) , such as renal , pelvic and small bowel tumors , in the fcc families sug - gests that some members of these families have a genetic predisposition similar to hnpcc but do not meet the clinical prerequisites of hnpcc syndrome . \n therefore , these families should be included in screening programs per - formed for hnpcc families . \n like every other cancer registry , our cancer registry in the province of isfahan did not include all cases of crc . \n however , to the best of our knowledge , this study included the largest number of crc cases report from iran . \n like the other study in iran by malekzadeh et al,1 we used the amsterdam ii criteria to find hnpcc and fcc cases . since amsterdam ii criteria have a relatively low sensitivity , the bethesda guidelines were developed and later revised in 2004 to help identify a larger proportion of families with hnpcc.22 bethesda guidelines , however , have low specificity , i.e. 80% or more of patients who meet the bethesda guidelines will not have lynch syndrome . \n indeed both of these guidelines have limitations and the results of our study need to be confirmed by genetic studies . in addition , another limitation to this study was that history taking was used as the basis of information regarding cancer in the family . \n although family members , in our society , are usually aware of serious diseases such as cancer in their relatives , a future national cancer registry program could provide more reliable information regarding cancer history in the families . \n based on the results of this study , clinical diagnosis of hnpcc and fcc was respectively observed in 2.0% and 2.9% of the studied cases in central iran ( isfahan ) . along with other studies from our society , \n these results show that hereditary and familial forms of crc are important types of cancer and need appropriate screening strategies to decrease the burden of the disease . \n high incidence of extracolonic tumors in hnpcc and fcc families in our society indicates that a detailed family history is mandatory when evaluating patients with crc . \n the results of this study need to be confirmed by further studies focusing on molecular basis in this field . \n moreover , a specific screening program for hnpcc and fcc families and a national registry program should be designed . \n all authors participated in data gathering , data analysis , and writing the draft of the manuscript and finally approved the final report .", "role": "user" }, { "content": "background : there is a lack of data on familial aggregation of colorectal cancer ( crc ) in iran . \n we aimed to determine the frequency of hereditary nonpolyposis colorectal cancer ( hnpcc ) and familial colorectal cancer ( fcc ) and to determine the frequency of extracolonic cancers in these families in isfahan.methods:we reviewed documents of all patients with a pathologically confirmed diagnosis of crc admitted to isfahan referral hospitals between 1995 and 2006 . \n we also studied our crc registry at poursina hakim research institute from 2003 to 2008 . \n we found hnpcc and fcc families based on the amsterdam ii criteria and interviewed them for family history of crc and extracolonic tumors . \n the family history was taken at least up to the second - degree relatives.results:during 1996 to 2008 , a total of 2580 crc cases have been diagnosed . \n we found 14 hnpcc and 53 fcc families . \n mean age of crc at diagnosis was 48.0 14.6 and 49.0 13.9 years in the hnpcc and fcc families , respectively ( p > 0.05 ) . \n the total numbers of observed extracolonic tumors were 70 ( 21.6% ; mean age = 53.6 11.0 years ) and 157 ( 13.8% ; mean age = 54.8 18.0 years ) in hnpcc and fcc families , respectively ( p > 0.05 ) . \n crc was respectively found in 52 and 76 members of the hnpcc and fcc families , revealing the frequency of hnpcc and fcc as 2.0% ( 52/2580 ) and 2.9% ( 76/2580 ) , respectively.conclusions:we found a relative high frequency of hnpcc ( 2.0% ) and fcc ( 2.9% ) among crc cases in our society and high incidence of extracolonic tumors in their families . \n further studies focusing on molecular basis in this field and designing a specific screening and national cancer registry program for hnpcc and fcc families should be conducted .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: posterior reversible encephalopathy syndrome ( pres ) is a reversible clinicoradiological syndrome characterized clinically by acute onset headache , altered alertness , seizures , vomiting , and abnormalities of visual perception , and neuroradiologically by edema predominantly involving the parieto - occipital regions . \n cerebellar involvement has been reported with the predominant involvement of posterior cerebral regions , but review of published english literature revealed only one case report with isolated cerebellar involvement in pres . \n we report here a case of pres with isolated cerebellar involvement in a child with acute lymphoblastic leukemia ( all ) during induction chemotherapy . \n a 7-years - old boy presented with slowly progressive bilateral cervical lymphadenopathy of few months duration . \n induction chemotherapy was initiated as per mcp 841 protocol which includes prednisolone , 40 mg / m for 29 days ; l - asparaginase , 6 000 iu / m on alternate days for 10 doses ; vincristine , 1.4 mg / m weekly for 5 doses ; daunorubicin , 30 mg / m at day 8 , 15 , and 29 ; and weekly intrathecal injections of methotrexate for 4 weeks . \n after documentation of febrile neutropenia on 14 day of induction , piperacillin - tazobactam and amikacin were initiated . \n he responded well and became afebrile within 24 hours of initiation of antibiotics . on 19 day of induction \n he was lethargic but well oriented and did not have signs of meningitis , abnormal movements , or visual disturbances . \n he was hypertensive initially ( blood pressure , 150/110 mmhg ; > 99 centile ) , but subsequent records were within normal range . \n a few hours later , the patient had two episodes of left focal tonic - clonic seizures at one - hour interval which were managed with midazolam and phenytoin . \n computed tomography ( ct ) of brain revealed diffuse hypodensity involving bilateral cerebellar hemispheres [ figure 1a ] . \n follow - up cranial magnetic resonance imaging ( mri ) examination performed 2 weeks after the episode showed almost complete resolution of cerebellar lesions [ figure 1b ] . \n he achieved remission after induction and received two cycles of high - dose cytarabine ( i2a ) and repeat induction ( ri1 ) cycle . \n he did not develop any neurological symptom during subsequent chemotherapy , but unfortunately succumbed to infective endocarditis after the third cycle of chemotherapy . \n ( a ) ct scan brain showing bilateral cerebellar edema ; ( b ) mri after 2 weeks showing complete resolution of cerebellar edema \n the reversible syndrome of altered mental status , headache , seizures , visual disturbances , and hypertension has been described in literature with different terminologies like hypertensive encephalopathy , reversible posterior leukoencephalopathy , and reversible occipitoparietal leukoencephalopathy . \n this syndrome has been recognized without hypertension or with only mild increase in blood pressure ; there is usually no accompanying destructive process of the white matter ; and neuroradiologically , the involvement is not strictly restricted to parieto - occipital regions . \n pres is seen exclusively in the setting of a significant systemic process or condition , including sudden hypertension , bone marrow or solid organ transplantation ( specially on cyclosporine immunosuppression ) , infection / sepsis / shock , toxemia of pregnancy , autoimmune disease , and during cancer chemotherapy ( e.g. , l - asparaginase , prednisolone , high - dose cytarabine , etc . ) . \n a case series from india revealed induction chemotherapy , hypertension , high - dose cytarabine , and cyclosporine as the triggers for pres in 13 leukemic patients . \n cerebral hypoperfusion and cytotoxic edema secondary to underlying systemic condition or drugs seems to be the principle pathophysiologic process leading to development of encephalopathy . \n an obvious risk factor in our patient was induction chemotherapy for all , but he also had one record of sudden onset hypertension at the time of neurotoxicity , which might have modulating effect . \n furthermore , despite receiving high - dose cytarabine and repeat induction cycle subsequently , pres did not recur , probably due to absence of hypertension during subsequent therapy . \n although mri is the most sensitive imaging test to diagnose pres , it is not necessary for the diagnosis if ct is suggestive . \n ct usually shows bilateral hypodensities in the posterior white matter , with some involvement of the overlying cortex . on mri , these lesions appear hyperintense on t2-weighted images , and are usually hypointense or isointense on diffusion - weighted images , with an increase of the apparent diffusion coefficient , indicating vasogenic edema . \n mckinney et al . reported two cases of completely unilateral involvement in their series , which is difficult to distinguish from infarction . \n the calcarine and paramedian occipital - lobe structures are usually spared , a fact that distinguishes pres from bilateral infarction of the posterior - cerebral - artery territory . \n the posterior part of the brain is more frequently involved due to poor sympathetic innervation of the vertebrobasilar system . \n fugate et al . found the parieto - occipital regions as the most commonly involved site ( 94% ) on mri in a large prospective study , followed by the frontal lobe ( 77% ) , temporal lobe ( 64% ) , and cerebellum ( 53% ) . \n cerebellar involvement was significantly more frequent in patients with a history of autoimmunity , and patients with sepsis were more likely to have cortical involvement . \n although parieto - occipital involvement was found in 99% of the cases , mckinney et al . noted higher incidence of atypical distributions ( posterior frontal , 78.9% ; temporal , 68.4% ; thalamus , 30.3% ; cerebellum , 34.2% ; brainstem , 18.4% ; and basal ganglia , 11.8% ) than commonly perceived . \n only one patient lacked parieto - occipital edema , but this patient had severe brainstem , thalamic , and deep white matter edema . \n atypical cases with only frontal , temporal , or brain stem lesions have been reported in literature . \n only one case of pres with isolated cerebellar involvement has been reported in the english literature , ours being the second . \n although the reversibility of the clinicoradiological features is most characteristic , it should be noted that prolonged seizures , hypertension , or both may result in permanent neurologic deficits and cerebral infarction . \n hence , pres must be managed with good supportive care including antihypertensive and antiepileptic medications to prevent any irreversible damage . a follow - up scan in a period of 1 to 2 weeks \n awareness of atypical patterns and variations in distribution of imaging abnormalities is important to recognize pres more accurately , to avoid expensive or potentially invasive work - ups for other primary cerebral diseases .", "role": "user" }, { "content": "parieto - occipital region is the most commonly involved site in posterior reversible encephalopathy syndrome ( pres ) . \n cerebellar involvement has been reported with the predominant involvement of posterior cerebral regions , but isolated cerebellar involvement in pres has been reported only once in english literature . \n we report here a 7-year - old boy with acute lymphoblastic leukemia who had pres with isolated cerebellar involvement during induction chemotherapy . \n he presented with sudden onset headache , vomiting and hypertension followed by seizures , unconsciousness , and involuntary movements . computed tomography scan revealed bilateral cerebellar hypodensities . \n he improved within few hours and reversibility of the lesions was documented on magnetic resonance imaging after 2 weeks . \n awareness of atypical patterns in distribution of imaging abnormalities is important to recognize pres more accurately and to avoid unnecessary diagnostic procedures and treatment .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: graves ' disease is the most common cause of hyperthyroidism in the pediatric population [ 1 , 2 ] . \n treatment options for graves ' disease include antithyroid medications , radioactive iodine , and surgery [ 1 , 2 ] . \n antithyroid medications used in children and adults include propylthiouracil ( ptu ) and methimazole ( mmi ) , and carbimazole , which is metabolized to mmi , and is available in europe but not the united states . \n recently , a significant safety concern related to hepatotoxicity risk associated with ptu use in children was brought to attention [ 4 , 5 ] . \n it is thus now recommended that ptu not be used in children , except in special circumstances [ 4 , 5 ] , and mmi should be used for antithyroid drug therapy in children . to date , the majority of publications related to medical therapy for graves ' disease have focused on children treated with ptu [ 614 ] . \n the description of the nature of adverse events that are associated with methimazole use in the pediatric population is modest , as well . at our center , \n we have routinely used mmi for graves ' disease therapy for many years . to provide insights into adverse events that can be associated with mmi use \n , we reviewed the adverse events associated with mmi use in our last one hundred consecutive pediatric patients treated with this medication . \n this review of treatment practice outcomes was conducted with the approval of the yale university human investigation committee . \n all adverse events were reported to the united states food and drug administration via the medwatch program . \n patients with the diagnosis of graves ' disease were identified from icd-9 coding ( 242.0 or 242.9 ) . \n the diagnosis of graves ' disease was made if there were elevated total and/or free thyroxine [ t4 ] and/or triiodothyronine [ t3 ] concentrations , subnormal thyrotropin levels , and evidence of thyroid autoimmunity not thought to be consistent with hashitoxicosis . \n the presence of goiter and ophthalmopathy supported the diagnosis of graves ' disease ; however eye disease was present in only 60% of the patients . in situations where there was a question of the diagnosis \n patients were diagnosed with graves ' disease in this setting only if the i uptake was elevated . \n the medical records of the last 100 consecutively treated patients with the diagnosis of graves ' disease were reviewed . \n medical records were reviewed to determine if adverse events occurred , and the length of time from initiating therapy until when adverse events developed . \n for those patients receiving treatment with either surgery or radioactive iodine , this was noted . \n initial thyroid function tests at diagnosis along with levels of thyroid stimulating immunoglobulin ( tsi ) or thyrotropin binding inhibitory immunoglobulin ( tbii ) were collected . \n 70% of patients were from the new haven ct area ; 30% of patients were from outside of the new haven area . \n 62% of the individuals were caucasian , 16% were hispanic , 16% were asian , and 6% were african american . at diagnosis , tsh levels were 0.01 0 \n the initial total t4 levels were 18.3 2.0 mcg / dl . \n the initial free t4 levels were 4.9 2.4 ng / dl . \n tsi levels were available in 73% of the individuals and were 180 70% . \n the patients were treated with average daily dose of mmi of 0.3 0.2 mg / kg / day . \n medication was given once a day in 60% of patients and twice daily in 40% of patients . \n adverse events attributed to the use of mmi , were seen in 19 patients ( table 1 ) . \n the most common side effects included pruritus and hives , which were seen in eight patients . \n two patients developed neutropenia with absolute neutrophil counts of 500 and 750 counts per cubic ml . \n three patients developed stevens - johnson syndrome with diffuse cutaneous eruption and mucous membrane involvement . \n u / l , the alanine aminotransferase ( alt ; sgpt ) was 379 u / l , the alkaline phosphatase was 355 \n u / l ; the bilirubin was 0.18 mg / dl ; and the gamma - glutamyl transpeptidase , ( ggt ) was 193 \n the age , methimazole dose , and length of time from initiation of treatment until development of adverse events for each patient are shown in table 1 . when clinical characteristics were compared in individuals who developed adverse events to methimazole with those who did not develop such reactions , no differences were detected as related to age , gender , dose , or ethnicity . \n there was no relationship to initial thyroid hormone levels , or circulating levels of immunoglobulins . \n of those individuals who developed adverse events to mmi , thyroidectomy was performed in three patients ( ages 34 years ) . \n radioactive iodine was administered to thirteen individuals ( age range 818 years ) , and three patients were changed to ptu , as treatment with radioactive iodine or surgery was refused by the families . \n adverse events to mmi occurred within one month of therapy in 20% of the patients , within three months of therapy in 50% of patients , and within six months of therapy in 90% of patients . in three patients , adverse events occurred after one and a half years of treatment . \n published reports related to the treatment of children with graves ' disease have generally involved cohorts of children treated with ptu [ 614 ] . \n these studies reveal an incidence of minor adverse events between 1% and 15% . within reports \n in which the use of mmi has been described , there has been little description of adverse events associated with this medication . \n our data suggest that methimazole can be associated with a risk of adverse events in up to 19% of individuals . \n if one excludes the eight patients with pruritus and hives , which are minor side effects , the more serious adverse events were found in 11% of patients . \n based on published reports describing outcomes for children treated with antithyroid medications for graves ' disease , up to 10 years ago , ptu was more widely used than mmi [ 1113 ] . more recent data however , suggest that two thirds of children in the united states treated with antithyroid medications are now being treated with mmi , and one - third are treated with ptu . \n recently , a concerning risk of hepatotoxicity resulting in liver failure in children and adults and in pregnant women treated with ptu has been brought to attention [ 4 , 16 ] . \n based on the incidence of reported cases of acute liver failure and liver transplantation associated with ptu , it is estimated that up to 1 in 2,000 children will sustain acute liver injury in response to ptu [ 4 , 16 ] . \n as the result , it is recommended that ptu not be used in children except in special circumstances , such as when an individual has had a toxic reaction to methimazole , and antithyroid medication is needed until definitive treatment either in the form of surgery or radioactive iodine can be performed . as such , mmi use in the pediatric population \n we observed one child who had cholestatic liver injury associated with methimazole . in the adult population , \n mmi associated liver injury is most typically seen in individuals who are older rather than younger , and in those who are treated with higher rather than lower mmi doses . \n in the individual who developed modest transaminase and alkaline phosphatase elevations , this condition reversed fully within one month after discontinuation of the medication . \n of concern was the development of stevens - johnson syndrome in three of the children , one of which required hospitalization . in each child , the condition reversed without long - term sequelae . of note , the three patients who developed stevens - johnson syndrome were receiving large doses of mmi ( 30 mg ) . at present \n we do not know , though , if the risk of stevens - johnson syndrome is dose - related . \n whereas most of the adverse events associated with mmi occurred within the first half year of the treatment onset , we observed adverse events after one and a half years of therapy in three children . \n these observations show that children treated with mmi warrant close follow - up for the development of potential toxic events . \n our observations raise the question about the utility of routine monitoring of hematological profiles or liver function tests or transaminase levels in patients on antithyroid medications . at present \n there is little evidence to support the notion that routine monitoring of these parameters is effective in minimizing the risk of antithyroid drug related adverse events [ 3 , 18 , 19 ] . \n if ptu is used , it is recommended that ptu should be stopped immediately and liver function and hepatocellular integrity be assessed in children who experience anorexia , pruritis rash , jaundice , light colored stool or dark urine , joint pain , right upper quadrant pain or abdominal bloating , nausea or fatigue [ 19 , 20 ] . \n in addition , ptu and mmi should be stopped immediately and white blood counts be measured in children who develop fever , mouth sores , pharyngitis , or feel ill . while routine monitoring of white blood counts may detect early agranulocytosis , it is not recommended because of the rarity of the condition and the lack of cost - effectiveness [ 3 , 18 ] . \n agranulocytosis has been reported in about 0.3% of adult patients taking mmi or ptu [ 3 , 18 , 19 ] . \n data of the incidence of agranulocytosis in children is not available , but is estimated to be very low . in adults , agranulocytosis is dose - dependent with mmi , and rarely occurs at low doses [ 3 , 18 ] . \n when it develops , agranulocytosis typically occurs within the first 100 days of therapy in 95% of individuals [ 3 , 18 ] . \n we recognized that a potential limitation of our study is that our referral patterns may bias our outcomes , as some of the patients coming for second opinions may have been treated beforehand with mmi doses higher than we typically use . \n the demographics of self - referred patients may also differ from that seen in a typical cross - section of children with graves ' disease . \n our patients are also not typically treated beyond two years with mmi , which influences our ability to observe long - term side effects . at present , \n ptu and mmi are the only antithyroid drugs available for graves ' disease in the united states . \n although mmi is less hepatotoxic than ptu , our data show that mmi use is indeed associated with potential adverse events , which can be serious . \n considering the hepatotoxicity risk associated with ptu , and the other minor and major adverse events associated with both ptu and mmi , strong consideration should be given to the development of less toxic antithyroid medications for use in children and adults .", "role": "user" }, { "content": "objective . graves ' disease is the most common cause of hyperthyroidism in the pediatric population . \n antithyroid medications used in children and adults include propylthiouracil ( ptu ) and methimazole ( mmi ) . at our center \n we have routinely used mmi for graves ' disease therapy . \n our goals are to provide insights into adverse events that can be associated with mmi use . methods . \n we reviewed the adverse events associated with mmi use in our last one hundred consecutive pediatric patients treated with this medication . \n results . \n the range in the patient age was 3.5 to 18 years . \n the patients were treated with an average daily dose of mmi of 0.30.2 mg / kg / day . \n adverse events attributed to the use of the medication were seen in 19 patients at 177 weeks of therapy . \n the most common side effects included pruritus and hives , which were seen in 8 patients . \n three patients developed diffuse arthralgia and joint pain . \n two patients developed neutropenia . \n three patients developed stevens - johnson syndrome , requiring hospitalization in 1 child . \n cholestatic jaundice was observed in 1 patient . \n no specific risk - factors for the development of adverse events were identified . \n conclusions . \n mmi use in children is associated with a low but real risk of minor and major side effects .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: ossification of the posterior longitudinal ligament ( opll ) , a result of heterotopic ossification , can induce spinal canal stenosis and lead to the development of severe myelopathy . \n there have been numerous studies of surgical techniques or clinical outcomes for opll of the cervical spine.1 in contrast , there are only a few reports regarding opll of the lumbar spine , and its clinical characteristics have not yet been well established because the number of patients who require surgical treatment is quite low.2 to our knowledge , there has been only six reports of surgically treated lumbar opll published in english.7 according to the data in the database for the spinal injuries center in fukuoka , japan , during the past 27 years , although 6,192 patients underwent operations for degenerative lumbar spine diseases , only ten underwent surgery for lumbar opll.2 thus , the frequency of lumbar opll requiring surgical treatment was remarkably low , indicating a lack of awareness of the disease even among orthopedic surgeons . \n we have recently witnessed that radiculopathy due to lumbar opll was found to be the cause of chronic right lower abdominal pain in a japanese man.8 after encountering this unusual case , we thought that lumbar opll should be considered to be a factor for not only orthopedic symptoms but also abdominal symptoms . because abdominal symptoms are common in the primary care setting,9 we tried to clarify the incidence of lumbar opll in an outpatient clinic in japan where primary care physicians are working . \n we analyzed consecutive outpatients making their first visit to the department of general medicine , asahikawa medical university hospital , between april 2009 and march 2012 . \n as we have recently demonstrated,913 the hospital consists of 602 beds in which approximately 250 doctors are working to address almost all medical problems . among them , four or five primary care physicians are working in the department of general medicine . during the period , we analyzed the patients ( n = 393 ) who received abdominal and pelvic computed tomography ( ct ) scan to clarify their several problems such as abdominal symptoms , origins of fever , high level of tumor markers , and so on . \n since it has been widely established that ct is a useful tool for detecting and accurately locating opll,14 lumbar opll was diagnosed by the ct scan according to a previous report.2 all data were drawn from medical records and the computerized physician order entry system in the hospital . \n each parameter such as age , sex , body mass index ( bmi ) , and clinical presentation was investigated from the source . \n analyzed comorbid conditions included diabetes mellitus , systemic hypertension , dyslipidemia , and cigarette smoking . \n statistical analysis was performed by fisher s exact test , and student s t - test for age and bmi . a level of p < 0.05 was considered to be statistically significant . \n statistical analysis was performed by fisher s exact test , and student s t - test for age and bmi . a level of p < 0.05 was considered to be statistically significant . \n lumbar opll was diagnosed by ct according to a previous report.2 representative images are shown in figure 1 . as shown in this figure , opll was diagnosed when an apparent ossification of the ligament causing more than 10% canal stenosis was revealed with ct . out of 393 patients who underwent abdominal and pelvic ct scan , 33 ( 8.4% ) were diagnosed with lumbar opll . \n all 33 patients diagnosed with lumbar opll detected by ct scan did not complain of main symptoms related to spinal stenosis , such as intermittent claudication . \n table 1 shows the clinical characteristics of 33 patients with lumbar opll in this study . \n when compared with patients without lumbar opll ( n = 360 ) , there was no significant difference in gender , bmi , presence of hypertension , diabetes mellitus or hyperlipidemia , and smoking habit , while a significant difference was identified in age in patients with lumbar opll . \n as clearly demonstrated in table 2 , the incidence of lumbar opll increased in the elderly . \n the highest incidence was observed in male patients aged 5069 years and female patients aged 7089 years . among 33 opll patients , \n seven patients complained of lower abdominal pain as their chief complaint . in the unique case mentioned earlier , \n the patient s pain was considered to be dependent on lumbar opll as reported in our recent publication;8 in another case , advanced colon cancer was considered to be the cause of abdominal pain . in the remaining five cases , the pain was diagnosed as functional gastrointestinal disorders after careful examination , but we could not completely exclude the possibility that the abdominal pain in the five patients was related to lumbar opll . \n , lumbar opll might be involved in the symptom , and was followed by consultation with the department of orthopedic surgery in our hospital . \n opll is more common in east asian populations , particularly in the japanese;15 the prevalence of opll is highest in japan , at a rate of 1.9%4.3%.16 although a number of authors have reported on opll of the cervical spine , few have discussed opll of the lumber spine.7 the number of reports regarding lumbar opll has been limited , and its prevalence and regional difference remain unclear . \n a lack of evidence may be a result of a large majority of lumbar opll possibly being asymptomatic , as described below . \n clinical presentations in patients with opll correspond to the level and magnitude of spinal cord compression . \n cervical and thoracic opll typically manifests with signs and symptoms of myelopathy , while the lumbar disease usually manifests with signs and symptoms related to spinal stenosis.3,17,18 matsunaga et al17 have demonstrated that after 30 years , myelopathy - free rates as high as 71% have been reported among patients who had no myelopathy when the cervical opll was first diagnosed . \n although we do not have any evidence that the time - course changes in lumbar opll are similar to cervical opll , these results led us to speculate that a large majority of lumbar opll patients are asymptomatic . \n in fact , all patients with lumbar opll detected in this study did not complain of spinal stenosis - mediated symptoms as described above . \n we do not know at this moment whether lumbar opll is indeed rare in the general population , or whether physicians are not aware of the presence of lumbar opll . in the present study , out of 393 patients who underwent abdominal and pelvic ct scans , 33 ( 8.4% ) \n were diagnosed as lumbar opll , indicating for the first time the incidence of lumbar opll in japan . according to previous data , \n the prevalence of cervical and thoracic opll in japanese and east asian countries has ranged from 1.9%4.3%.15 these results suggest that the frequency of lumbar opll is not much lower than that of cervical and thoracic opll.1 kobashi et al have demonstrated a higher frequency of diabetes mellitus among japanese men and women with opll,19 suggesting that diabetes mellitus may play a role in the development of opll . \n the present results demonstrated that there was no relationship between the presence of diabetes mellitus and lumbar opll . \n the discrepancy may be due to the difference in pathophysiology between cervical and lumbar opll . \n because of a lack of evidence whether pathogenesis is common in patients with cervical and lumbar opll , further studies on this issue are needed . \n the present study also showed a lack of association of hypertension , hyperlipidemia , and bmi with lumbar opll . \n the present study clearly demonstrated for the first time that lumbar opll is highly identified in elderly people . based on these findings \n , we should pay special attention in elderly patients and work to determine whether they may have lumbar opll . \n although the presence of lumbar opll by itself does not necessarily mean a clinical problem , we should keep in mind that lumbar opll is possibly involved in not only spinal stenosis - mediated symptoms , but also radiculopathy - related symptoms seen in the primary care setting , such as abdominal pain.8", "role": "user" }, { "content": "purposelittle is known about the prevalence and epidemiological characteristics of lumbar ossification of the posterior longitudinal ligament ( opll ) . \n we analyzed the rate of lumbar opll in an outpatient unit where primary care physicians are working in japan , to better understand the epidemiological characteristics of the disease.methodswe analyzed consecutive , first - time visiting outpatients who received abdominal and pelvic computed tomography ( ct ) scan at the department of general medicine , asahikawa medical university hospital , japan , between april 2009 and march 2012 . \n each parameter such as age , sex , and clinical presentation was investigated.resultsout of 393 patients who underwent abdominal and pelvic ct scan , 33 ( 8.4% ) were diagnosed as lumbar opll . \n when compared with patients without lumbar opll ( n = 360 ) , there was no significant difference in gender , body mass index ( bmi ) , presence of hypertension , diabetes mellitus or hyperlipidemia , and smoking habit , while the age in patients with lumbar opll was significantly higher.conclusionthese results suggest for the first time that lumbar opll is frequently observed in elderly people in the primary care setting , in japan .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: the annual incidence of myasthenia gravis ( mg ) is reported to be 0.254 patients per 100 000 population , with the first peak of onset around the second and third decades of life and the second peak around the fifth and sixth decades . in most cases , \n it is caused by pathogenic autoantibodies directed toward the skeletal muscle acetylcholine receptor ( achr ) , but in others , non - achr components of the postsynaptic muscle endplate , such as the muscle - specific receptor tyrosine kinase ( musk ) , might serve as targets of autoimmune attack . \n human leukocyte antigen ( hla ) loci are associated with all autoimmune diseases and are the strongest genetic factors for individual predisposition . \n of the non - hla susceptibility genes , variants of protein tyrosine phosphatase non - receptor type ( ptpn22 ) show the strongest associations with autoimmune diseases . \n a snp in ptpn22 has been reported to be associated with type 1 diabetes ( t1 dm ) , rheumatoid arthritis ( ra ) , systemic lupus erythematosus ( sle ) , hashimoto s thyroiditis . \n in addition , some studies also suggested that ptpn22 r620w polymorphism was associated with mg risk [ 1319 ] . \n however , the association between this polymorphism and the risk of mg was controversial and inconclusive . \n thus , we did a meta - analysis to determine if this polymorphism could influence mg risk . \n we did a comprehensive search in pubmed , medline , embase , cnki ( china national knowledge infrastructure ) and wanfang to retrieve relevant articles . \n we retrieved the relevant articles using the following terms : myasthenia gravis , \n ptpn22 , and polymorphism or variant or mutation as well as their combinations . \n the studies must meet the following criteria : 1 ) the paper should be case - control studies ; 2 ) evaluating the contribution of ptpn22 r620w polymorphism with mg risk ; 3 ) genotype distributions in the cases and controls were available to extract , and the results presented in odds ratio ( ors ) with its 95% confidence interval ( ci ) ; and 4 ) for a certain polymorphism , genotype distribution in the controls must be in hardy - weinberg equilibrium ( hwe ) . \n two investigators independently estimated the extracted data of the included studies to reach a consensus . \n the following information was extracted from each article : first author , year of publication , country , ethnicity , age , gender , type of mg , sample size . \n the newcastle - ottawa scale ( nos ) for case - control studies was used to assess the methodological quality of every included study . \n the highest study quality was nine stars with a maximum of four stars for selection , two stars for comparability , and three stars for exposure in the nos . \n the significance of the pooled ors was determined by the z test with a p value less than 0.05 considering statistically significant . \n statistical analyses were conducted in stata version 11.0 ( stata corporation , college station , tx , usa ) . \n we did a comprehensive search in pubmed , medline , embase , cnki ( china national knowledge infrastructure ) and wanfang to retrieve relevant articles . \n we retrieved the relevant articles using the following terms : myasthenia gravis , \n ptpn22 , and polymorphism or variant or mutation as well as their combinations . \n the studies must meet the following criteria : 1 ) the paper should be case - control studies ; 2 ) evaluating the contribution of ptpn22 r620w polymorphism with mg risk ; 3 ) genotype distributions in the cases and controls were available to extract , and the results presented in odds ratio ( ors ) with its 95% confidence interval ( ci ) ; and 4 ) for a certain polymorphism , genotype distribution in the controls must be in hardy - weinberg equilibrium ( hwe ) . \n two investigators independently estimated the extracted data of the included studies to reach a consensus . \n the following information was extracted from each article : first author , year of publication , country , ethnicity , age , gender , type of mg , sample size . \n the newcastle - ottawa scale ( nos ) for case - control studies was used to assess the methodological quality of every included study . \n the highest study quality was nine stars with a maximum of four stars for selection , two stars for comparability , and three stars for exposure in the nos . \n the significance of the pooled ors was determined by the z test with a p value less than 0.05 considering statistically significant . \n statistical analyses were conducted in stata version 11.0 ( stata corporation , college station , tx , usa ) . \n of the 46 references , 3 records excluded for duplication and 43 articles were judged potentially relevant . \n following titles and abstracts screened for relevance , 36 full - text articles comprehensively assessed against inclusion criteria . \n a total of 7 case - control studies with 2802 cases and 3730 controls were finally included in this review . \n ptpn22 r620w polymorphism was significantly associated with the risk of mg ( or=1.57 ; 95% ci , 1.341.82 ; i=31% ; figure 2 ) . \n in addition , thymoma patients was significantly associated with risk of mg ( or=1.59 ; 95% ci , 1.281.98 ; i=0% ) . however , non - thymoma patients with this polymorphism did not have increased mg risk ( or=1.36 ; 95% ci , 0.862.15 ; i=77% ) . furthermore , ptpn22 r620w polymorphism showed increased early - onset myasthenia gravis ( eomg ) risk ( or=2.38 ; 95% ci , 1.523.71 ; i=0% ) . \n the sensitivity analysis suggested that the corresponding pooled ors were not materially altered ( data not shown ) . \n of the 46 references , 3 records excluded for duplication and 43 articles were judged potentially relevant . \n following titles and abstracts screened for relevance , 36 full - text articles comprehensively assessed against inclusion criteria . \n a total of 7 case - control studies with 2802 cases and 3730 controls were finally included in this review . \n ptpn22 r620w polymorphism was significantly associated with the risk of mg ( or=1.57 ; 95% ci , 1.341.82 ; i=31% ; figure 2 ) . \n in addition , thymoma patients was significantly associated with risk of mg ( or=1.59 ; 95% ci , 1.281.98 ; i=0% ) . however , non - thymoma patients with this polymorphism did not have increased mg risk ( or=1.36 ; 95% ci , 0.862.15 ; i=77% ) . furthermore , ptpn22 r620w polymorphism showed increased early - onset myasthenia gravis ( eomg ) risk ( or=2.38 ; 95% ci , 1.523.71 ; i=0% ) . \n the sensitivity analysis suggested that the corresponding pooled ors were not materially altered ( data not shown ) . \n mg is an acquired organ - specific autoimmune disease , characterized by fluctuating weakness and fatigability of voluntary muscles due to the impairment of the neuromuscular transmission caused by autoantibodies . \n there is an urgent need to look for biomarkers evaluating the severity of the disease , predicting the prognosis of patients , and targeting the pathogenic genes . \n the present meta - analysis including 7 studies evaluated the association between ptpn22 r620w polymorphism and mg risk . \n we found that individuals with the ptpn22 r620w polymorphism showed an increased risk of mg in the overall population . \n furthermore , we found that ptpn22 r620w polymorphism might also be a risk factor of eomg . in the subgroup analysis , \n vang and coworkers investigated effects of r620 versus w620 in cell stimulation with anti - cd3 and anti - cd28 . \n they found that mutant phosphatase inhibited nfat / ap1 more efficient with w620 variant than r620 variant . \n thus , they suggested that w620 variant predisposed to mg because w620 variant may suppress t cell antigen receptor ( tcr ) signaling more potently during thymic development . \n firstly , the subgroups may have a relatively lower power based on a small number of studies . \n secondly , a more precise analysis should be conducted if individual information including other covariates such as age , sex and smoking condition becomes available . \n thirdly , the gene - gene interaction is important for the development of complex diseases including mg . \n this meta - analysis suggested a significant association between ptpn22 r620w polymorphism and mg risk .", "role": "user" }, { "content": "backroundthe association between ptpn22 r620w polymorphism and risk of myasthenia gravis ( mg ) remains controversial . \n therefore , we did this meta - analysis to investigate this association.material/methodswe did a comprehensive search in pubmed , medline , embase , cnki ( china national knowledge infrastructure ) , and wanfang electronic databases to retrieve relevant articles . \n the overall effect was measured by odds ratios ( ors ) with its 95% confidence intervals ( cis ) . \n statistical analyses were conducted with stata software.resultsoverall , a total of 7 case - control studies with 2802 cases and 3730 controls were finally included in this review . \n ptpn22 r620w polymorphism was significantly associated with an increased risk of mg ( or=1.57 ; 95% ci , 1.341.82 ; i2=31% ) . in the subgroup analysis , \n thymoma patients were significantly associated with risk of mg ( or=1.59 ; 95% ci , 1.281.98 ; i2=0% ) . however , non - thymoma patients with this polymorphism did not have increased mg risk ( or=1.36 ; 95% ci , 0.862.15 ; i2=77% ) . in addition , ptpn22 r620w polymorphism showed increased early - onset myasthenia gravis ( eomg ) risk ( or=2.38 ; 95% ci , 1.523.71 ; i2=0%).conclusionsthis meta - analysis shows a significant association between ptpn22 r620w polymorphism and mg risk .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: after the benefits of fast post - operative recovery and reduced morbidities without compromise in long - term oncologic outcomes were demonstrated by many studies [ 17 ] , laparoscopic colectomy revolutionized colorectal surgery and gained wide acceptance to become the preferred approach for colon resection . on the other hand , \n the application of enhanced recovery program ( erp ) in colorectal surgery has also been shown to accelerate post - operative recovery effectively and to reduce morbidities by many clinical studies and meta - analysis [ 815 ] . \n the concept of erp advocated strongly on pre - operative optimization of patient information and organ function , means for reduction of surgical stress , adequate anesthesia and pain control , and modification of peri - operative care pathway [ 1618 ] . \n practice of erp is not complicated but revolutionary and it soon becomes a new standard of peri - operative care in patients after major surgery . \n the application of erp in open colectomy has been reported to achieve a hospital stay of 25 days with no increase in complication or re - admission rate [ 8 , 11 , 19 , 20 ] . \n this favorable result has generated the interest that erp could also be an effective measure to optimize the short - term outcomes of laparoscopic colectomy further . \n erp is very suitable for laparoscopic surgery as the patients have less wound pain and morbidity ; hence , they can be ambulated and discharged early . \n erp for laparoscopic colectomy had been reported to successfully reduce post - operative stay to 23 days in even high - risk patients and make a 23-h stay be possible in selected patients \n . however , literature review found only three reports that compared results of laparoscopic colectomy with conventional care and enhanced recovery care [ 2325 ] . \n the purpose of this study was to evaluate the impact of erp on outcomes of laparoscopic colectomy in a center which specializes in laparoscopic colorectal surgery . \n this study compared the short - term outcomes of patients who underwent laparoscopic resection for treatment of colonic cancer before and after the application of erp in a university teaching hospital . \n comparison was made between consecutive patients who underwent laparoscopic resection for colon cancer during the two periods . \n the control group included patients who had operation during january 2005 to november 2006 . within this period , \n patient management was directed mainly by order of surgeon in - charge during the ward round . \n there was no consensus on time to resume diet , management of drains and urinary catheter , measure of rehabilitation , criteria for discharge between surgeons . \n the erp group included patients who were operated on from december 2006 to december 2007 . from december 2006 , an erp \n the erp for patients in the unit included the following elements : \n case manager the case manager , who was a nurse specialist , took an active part in the peri - operative management of the patients . before operation , the case manager interviewed the patients and their relatives to explain pre - operative preparation , expected recovery rate and also helped family members to prepare for the care of patient at home after discharge . \n the case manager had a telephone hotline to provide explanation and instruction to patients and their family members after discharge . \n peri - operative management peri - operative management the patient was admitted 1 day before the operation for bowel preparation and preoperative management . \n intravenous fluid was given at the discretion of anesthetist during operation but was restricted to 1.5 l / day in early post - operative period and was stopped as soon as the patient s oral fluid intake became adequate . unless there was contraindication , the medical and nursing care of patient after operation followed a standard clinical pathway , which aimed for accelerated recovery . \n the pathway emphasized on ( 1 ) education of patient and family in pain and wound management ; ( 2 ) routine chest physiotherapy and walking exercise ; ( 3 ) early resumption of nutrition without the need to wait for presence bowel sound or passage of flatus . \n the criteria for discharge of patient included ( 1 ) absence of complication except for mild wound infection ( 2 ) full ambulation or availability of care person at home and ( 3 ) tolerance of diet . \n table 1summary of clinical pathway for enhanced recovery programday 1assess for complicationresume fluid solid dietremove urinary catheterchest physiotherapyassisted ambulationeducate pain management and wound careday 2assess for complicationprogress to solid dietensure good pain controlwalking exercisediscuss with patient and family about discharge planday 3assess for complicationplan to dischargearrange community nursing service if indicatededucate patient / family on diet , wound care and pain management at homeday 4full ambulationdischarge from hospitalexclusion criteria summary of clinical pathway for enhanced recovery program patients with mid and low rectal cancer , who required low anterior resection or abdominoperineal resection with total mesorectal excision , were excluded because of the need for stoma care . \n patients who were not admitted to colorectal surgical ward because of administrative reason , who required synchronous second major procedure during the operation , who were anticipated to require intensive care unit ( icu ) or high dependency unit ( hdu ) admission after the operation were also excluded from care with erp . \n all patients in both groups had colonic or recto - sigmoid junction cancer and underwent laparoscopic colectomy with a standardized medial - to - lateral approach . \n patient controlled anesthesia ( pca ) was used for both groups of patients . to allow for fair comparison between two groups , \n the same exclusion criteria in the erp was applied to the control group , i.e. patients who met the exclusion criteria set for erp during january 2005 to november 2006 were excluded from the control group . \n data of patients demographics and pathology , operative details , complications and post - operative outcomes were collected prospectively . \n we defined high operative risk as patients who had an american society of anesthesiologist ( asa ) score 3 or higher and advanced cancer stage as patients who had stage 3 or 4 cancer . \n prolonged post - operative ileus was defined as absence of bowel movement and inability to tolerate diet for more than 5 days without evidence of intra - abdominal infection . \n re - admission was defined as any unplanned admission into any hospital within 30 days after discharge . to ensure accurate report of re - admission rate \n , a retrospective review was made through an electronic clinical management system , which contained admission records of patients in all public hospitals in hong kong . \n statistical analysis was performed with statistical package for social sciences ( spss 11.0 , spss inc . , \n categorical variables were compared by chi square test or fisher s exact test as appropriate . \n patient s age , operating time and blood loss were presented as mean with standard deviation and were compared by student s t test . \n hospital stay and time for resumption of gastrointestinal function were presented as median with range and were compared by mann whitney u test . \n this study compared the short - term outcomes of patients who underwent laparoscopic resection for treatment of colonic cancer before and after the application of erp in a university teaching hospital . \n comparison was made between consecutive patients who underwent laparoscopic resection for colon cancer during the two periods . \n the control group included patients who had operation during january 2005 to november 2006 . within this period , \n patient management was directed mainly by order of surgeon in - charge during the ward round . \n there was no consensus on time to resume diet , management of drains and urinary catheter , measure of rehabilitation , criteria for discharge between surgeons . \n the erp group included patients who were operated on from december 2006 to december 2007 . from december 2006 , an erp \n the erp for patients in the unit included the following elements : \n case manager the case manager , who was a nurse specialist , took an active part in the peri - operative management of the patients . before operation , the case manager interviewed the patients and their relatives to explain pre - operative preparation , expected recovery rate and also helped family members to prepare for the care of patient at home after discharge . \n the case manager had a telephone hotline to provide explanation and instruction to patients and their family members after discharge . \n peri - operative management peri - operative management the patient was admitted 1 day before the operation for bowel preparation and preoperative management . \n intravenous fluid was given at the discretion of anesthetist during operation but was restricted to 1.5 l / day in early post - operative period and was stopped as soon as the patient s oral fluid intake became adequate . unless there was contraindication , the medical and nursing care of patient after operation followed a standard clinical pathway , which aimed for accelerated recovery . \n the pathway emphasized on ( 1 ) education of patient and family in pain and wound management ; ( 2 ) routine chest physiotherapy and walking exercise ; ( 3 ) early resumption of nutrition without the need to wait for presence bowel sound or passage of flatus . \n the criteria for discharge of patient included ( 1 ) absence of complication except for mild wound infection ( 2 ) full ambulation or availability of care person at home and ( 3 ) tolerance of diet . \n table 1summary of clinical pathway for enhanced recovery programday 1assess for complicationresume fluid solid dietremove urinary catheterchest physiotherapyassisted ambulationeducate pain management and wound careday 2assess for complicationprogress to solid dietensure good pain controlwalking exercisediscuss with patient and family about discharge planday 3assess for complicationplan to dischargearrange community nursing service if indicatededucate patient / family on diet , wound care and pain management at homeday 4full ambulationdischarge from hospitalexclusion criteria summary of clinical pathway for enhanced recovery program patients with mid and low rectal cancer , who required low anterior resection or abdominoperineal resection with total mesorectal excision , were excluded because of the need for stoma care . \n patients who were not admitted to colorectal surgical ward because of administrative reason , who required synchronous second major procedure during the operation , who were anticipated to require intensive care unit ( icu ) or high dependency unit ( hdu ) admission after the operation were also excluded from care with erp . \n all patients in both groups had colonic or recto - sigmoid junction cancer and underwent laparoscopic colectomy with a standardized medial - to - lateral approach . \n patient controlled anesthesia ( pca ) was used for both groups of patients . to allow for fair comparison between two groups , \n the same exclusion criteria in the erp was applied to the control group , i.e. patients who met the exclusion criteria set for erp during january 2005 to november 2006 were excluded from the control group . \n data of patients demographics and pathology , operative details , complications and post - operative outcomes were collected prospectively . \n we defined high operative risk as patients who had an american society of anesthesiologist ( asa ) score 3 or higher and advanced cancer stage as patients who had stage 3 or 4 cancer . \n prolonged post - operative ileus was defined as absence of bowel movement and inability to tolerate diet for more than 5 days without evidence of intra - abdominal infection . \n re - admission was defined as any unplanned admission into any hospital within 30 days after discharge . to ensure accurate report of re - admission rate \n , a retrospective review was made through an electronic clinical management system , which contained admission records of patients in all public hospitals in hong kong . \n statistical analysis was performed with statistical package for social sciences ( spss 11.0 , spss inc . , \n categorical variables were compared by chi square test or fisher s exact test as appropriate . \n patient s age , operating time and blood loss were presented as mean with standard deviation and were compared by student s t test . \n hospital stay and time for resumption of gastrointestinal function were presented as median with range and were compared by mann whitney u test . \n after exclusion , there were 96 patients in the erp group and 84 patients in the control group . \n the number of patients excluded in each group and the reasons for exclusion were shown in table 2 . \n eight patients in the erp group and three patients in the control group were admitted to the hdu or icu admission after the operation . as a result , the eight patients in the erp group did not have post - operative care with erp . under the principle of intention to treat , patients who had unexpected hdu or icu admissions \n the age of patients were similar in both groups ( 69.6 13 versus 71.4 10 in erp and control groups respectively ; p = 0.30 ) . \n there were also no statistical difference in all other patient characteristics including gender , co - morbidities , asa score , stage of cancer , and incidence of previous abdominal surgery ( table 3 ) . \n the types of procedures performed were similar during the two periods ( p = 0.09 ) and were summarized in fig . 1 . between the erp and control groups \n , there were no difference in operating time ( 171.1 50 versus 160.5 41 min ; p = 0.12 ) , operative blood loss [ 100 ( range : 50150 ) versus 80 ( range : 50100 ) ml ; p = 0.91 ] and open conversion rate ( 6.3% versus 8.3% respectively ; p = 0.59 ) . \n table 2exclusion for enhanced recovery program and for comparison in control groupnumber of patientserpcontrolsynchronous second major resection52anticipated icu / hdu admission82not admitted to colorectal surgery ward9n.a.case manager not available3n.a.erp enhanced recovery program , icu intensive care unit , hdu high dependency unit , n.a . \n not applicabletable 3patient characteristicserp ( n = 96)control ( n = 84)p valueage72 ( 3194)72 ( 4692)0.30sex ( male / female)51/4550/340.39chronic pulmonary disease9 ( 9.4%)7 ( 8.3%)0.81chronic heart disease17 ( 17.7%)15 ( 17.6%)0.98previous stroke7 ( 7.3%)4 ( 4.8%)0.546diabetes mellitus15 ( 15.6%)18 ( 21.4%)0.32high anesthetic risk13 ( 15.6%)16 ( 19%)0.2previous abdominal surgery21 ( 21.9%)14 ( 16.7%)0.378advanced cancer stage42 ( 43.8%)41 ( 48.8%)0.50fig . \n 1types of laparoscopic colectomy in two groups of patients exclusion for enhanced recovery program and for comparison in control group erp enhanced recovery program , icu intensive care unit , hdu high dependency unit , n.a . \n not applicable patient characteristics types of laparoscopic colectomy in two groups of patients there was no operative mortality in both groups of patients . \n intra - operative complications occurred in four patients in the erp group ( two arrhythmia , bowel perforation , pre - sacral vein bleeding ) and three patients in the control group ( pre - sacral vein bleeding , iliac artery injury , bowel perforation ) . \n post - operative complications occurred in 12.5% and 10.7% of patients in the erp and control groups respectively ( p = 0.71 ) . \n there was no difference in the incidence of any type of post - operative complications between the two groups and the details are listed in table 4 . \n table 4post - operative complicationserp ( n = 96)control ( n = 84)p valuecardiac4 ( 4.1%)nil0.12pulmonary4 ( 4.1%)3 ( 3.6%)1.00intra - abdominal collection1 ( 1%)1 ( 1.2%)1.00anastomotic leakagenil2 ( 2.4%)0.22deep vein thrombosis1 ( 1%)nil1.0wound infection2 ( 2.1%)5 ( 6%)0.18 post - operative complications recovery of gastrointestinal function in patients was assessed by the resumption of diet , passage of flatus and the incidence of prolonged ileus . in the erp and control groups , \n the post - operative day of normal diet resumption diet was 1 ( range : 13 ) versus 1 ( range : 16 ) respectively ( p = 0.96 ) and the post - operative day of passing flatus was 2 ( range : 14 ) versus 2 ( range : 15 ) respectively ( p = 0.03 ) . \n no patient in the erp group and five ( 6% ) patients in the control group had prolong ileus after operation ( p = 0.02 ) . \n the median hospital stay after operation , however , had no difference in both groups [ 4 ( 223 ) days in erp group versus 4 ( 234 ) days in control group ; p = 0.4 ) ] . \n five patients in the erp group and six patients in the control group had re - admission and the re - admission rate were similar in both groups ( 5% and 7% respectively ; p = 0.59 ) . \n wound infection was the commonest reason for re - admission and the reasons of re - admission were summarized in table 5 . \n table 5reasons for re - admission of patientsenhanced recovery program group ( n = 5)1 . \n wound infection reasons for re - admission of patients factors affecting hospital stay of patients with laparoscopic colectomy were analyzed . while the median hospital stay for all patients in the study was 4 days , we arbitrarily defined hospital stay of 5 days or more as late discharge , which happened in 74 ( 40% ) patients . \n factors significantly associated with late discharge included age ( p < 0.01 ) , advanced cancer stage ( p = 0.04 ) , operative blood loss ( p = 0.04 ) , open conversion ( p = 0.01 ) , post - operation hdu / icu admission ( p < 0.01 ) , prolong ileus ( p < 0.01 ) and presence of complication ( p < 0.01 ) . among these factors , multivariate analysis with logistic regression found that age ( p < 0.01 ; 95% confidence interval , 1.031.12 ) , advanced cancer stage ( p = 0.01 ; 95% confidence interval , 0.170.82 ) and presence of complication ( p = 0.01 ; 95% confidence interval , 0.050.52 ) were independent factors associated with late discharge . \n after exclusion , there were 96 patients in the erp group and 84 patients in the control group . \n the number of patients excluded in each group and the reasons for exclusion were shown in table 2 . \n eight patients in the erp group and three patients in the control group were admitted to the hdu or icu admission after the operation . as a result , the eight patients in the erp group did not have post - operative care with erp . under the principle of intention to treat , patients who had unexpected hdu or icu admissions \n the age of patients were similar in both groups ( 69.6 13 versus 71.4 10 in erp and control groups respectively ; p = 0.30 ) . \n there were also no statistical difference in all other patient characteristics including gender , co - morbidities , asa score , stage of cancer , and incidence of previous abdominal surgery ( table 3 ) . \n the types of procedures performed were similar during the two periods ( p = 0.09 ) and were summarized in fig . 1 . between the erp and control groups \n , there were no difference in operating time ( 171.1 50 versus 160.5 41 min ; p = 0.12 ) , operative blood loss [ 100 ( range : 50150 ) versus 80 ( range : 50100 ) ml ; p = 0.91 ] and open conversion rate ( 6.3% versus 8.3% respectively ; p = 0.59 ) . \n table 2exclusion for enhanced recovery program and for comparison in control groupnumber of patientserpcontrolsynchronous second major resection52anticipated icu / hdu admission82not admitted to colorectal surgery ward9n.a.case manager not available3n.a.erp enhanced recovery program , icu intensive care unit , hdu high dependency unit , n.a . \n not applicabletable 3patient characteristicserp ( n = 96)control ( n = 84)p valueage72 ( 3194)72 ( 4692)0.30sex ( male / female)51/4550/340.39chronic pulmonary disease9 ( 9.4%)7 ( 8.3%)0.81chronic heart disease17 ( 17.7%)15 ( 17.6%)0.98previous stroke7 ( 7.3%)4 ( 4.8%)0.546diabetes mellitus15 ( 15.6%)18 ( 21.4%)0.32high anesthetic risk13 ( 15.6%)16 ( 19%)0.2previous abdominal surgery21 ( 21.9%)14 ( 16.7%)0.378advanced cancer stage42 ( 43.8%)41 ( 48.8%)0.50fig . \n 1types of laparoscopic colectomy in two groups of patients exclusion for enhanced recovery program and for comparison in control group erp enhanced recovery program , icu intensive care unit , hdu high dependency unit , n.a . \n intra - operative complications occurred in four patients in the erp group ( two arrhythmia , bowel perforation , pre - sacral vein bleeding ) and three patients in the control group ( pre - sacral vein bleeding , iliac artery injury , bowel perforation ) . \n post - operative complications occurred in 12.5% and 10.7% of patients in the erp and control groups respectively ( p = 0.71 ) . \n there was no difference in the incidence of any type of post - operative complications between the two groups and the details are listed in table 4 . \n table 4post - operative complicationserp ( n = 96)control ( n = 84)p valuecardiac4 ( 4.1%)nil0.12pulmonary4 ( 4.1%)3 ( 3.6%)1.00intra - abdominal collection1 ( 1%)1 ( 1.2%)1.00anastomotic leakagenil2 ( 2.4%)0.22deep vein thrombosis1 ( 1%)nil1.0wound infection2 ( 2.1%)5 ( 6%)0.18 post - operative complications \n recovery of gastrointestinal function in patients was assessed by the resumption of diet , passage of flatus and the incidence of prolonged ileus . in the erp and control groups , \n the post - operative day of normal diet resumption diet was 1 ( range : 13 ) versus 1 ( range : 16 ) respectively ( p = 0.96 ) and the post - operative day of passing flatus was 2 ( range : 14 ) versus 2 ( range : 15 ) respectively ( p = 0.03 ) . \n no patient in the erp group and five ( 6% ) patients in the control group had prolong ileus after operation ( p = 0.02 ) . \n the median hospital stay after operation , however , had no difference in both groups [ 4 ( 223 ) days in erp group versus 4 ( 234 ) days in control group ; p = 0.4 ) ] . \n five patients in the erp group and six patients in the control group had re - admission and the re - admission rate were similar in both groups ( 5% and 7% respectively ; p = 0.59 ) . \n wound infection was the commonest reason for re - admission and the reasons of re - admission were summarized in table 5 . \n table 5reasons for re - admission of patientsenhanced recovery program group ( n = 5)1 . \n wound infection reasons for re - admission of patients factors affecting hospital stay of patients with laparoscopic colectomy were analyzed . while the median hospital stay for all patients in the study was 4 days , we arbitrarily defined hospital stay of 5 days or more as late discharge , which happened in 74 ( 40% ) patients . \n factors significantly associated with late discharge included age ( p < 0.01 ) , advanced cancer stage ( p = 0.04 ) , operative blood loss ( p = 0.04 ) , open conversion ( p = 0.01 ) , post - operation hdu / icu admission ( p < 0.01 ) , prolong ileus ( p < 0.01 ) and presence of complication ( p < 0.01 ) . among these factors , \n multivariate analysis with logistic regression found that age ( p < 0.01 ; 95% confidence interval , 1.031.12 ) , advanced cancer stage ( p = 0.01 ; 95% confidence interval , 0.170.82 ) and presence of complication ( p = 0.01 ; 95% confidence interval , 0.050.52 ) were independent factors associated with late discharge . \n erp has been proven to improve short - term outcomes after operation . in colorectal surgery , erp was shown to have the benefits of early recovery of gastrointestinal function , reduced morbidities , reduced hospital stay and did not increase re - admission rate when applied for patients with open colectomy [ 912 , 19 ] . \n only a few studies have reported on implementation of erp in laparoscopic colectomy and the authors believe that investigation into this subject is important . in laparoscopic colectomy , it is unclear that if erp can hasten patient recovery further or actually add no further benefit to patients who already have prompt recovery \n . this information would be valuable in defining the role of erp in current practice of colorectal surgery . \n we included only cancer patients because they have a special need for nutritional and psychological care during peri - operative period when compared to patient with benign diseases and majority of our elective surgery is for colorectal cancer . \n an erp was started in the authors center since december 2006 and 80% of patients were suitable for erp during the study period . \n the main reason for excluding patients who were anticipated hdu or icu admission after the operation and patients not admitted to the primary ward of colorectal surgery because of administrative consideration ( table 2 ) . the implementation of erp was successful with no increase in complication or re - admission rate when compared to the control group . \n while the post - operative day of tolerating diet was the same in both erp and control groups , patients in the former group passed flatus significantly earlier [ 2 ( range : 14 ) versus 2 ( range : 15 ) respectively ; p = 0.03 ) ] . \n passage of flatus is unlikely to be altered by any external factor and it should be a more objective parameter to herald resumption of gastrointestinal function when compared with time to toleration of diet after operation . \n the incidence of prolonged ileus in the ft surgery group was also significantly lower than the control group ( 0 versus 6% respectively ; p = 0.02 ) . except for implementation of erp \n , there was no other difference in peri - operative patient management of two groups of patients . \n all operations in the study were performed by same operative approach ( medial - to - lateral mobilization ) and the procedures were done mostly by fellows in colorectal surgery under supervision of the senior author . \n the operating time , blood loss and complication rate did not show any difference in both groups . \n post - operative ileus is common after open abdominal surgery and the benefit of laparoscopic colectomy over open surgery is partly attributed by reduction of the duration of ileus . \n however , most studies on laparoscopic or erp for colectomy in the past did not report post - operative ileus and its actual incidence in current practice was unclear . \n the reduction of prolonged ileus in the erp group could be a result of enforced rehabilitation by effort of our nursing staff and physiotherapists in such a multi - modal care program . through motility study using indium scintigraphy , \n basse et al . demonstrated early normalization of gastrointestinal function when comprehensive erp was applied to patients with colonic resection . \n only three published reports [ 2325 ] had compared outcomes of laparoscopic colectomy with and without erp and they all reported reduction in hospital stay with the implementation of erp . \n reported more patients discharged within 8 days after laparoscopic colectomy by the implementation of clinical pathway , raue et al . and al chalabi et al . \n reported that erp resulted in reducing hospital stay from about 8 to 5 days among their patients . in the present study , there was no difference in length of hospital stay between the erp and control groups ( medial hospital stay = 4 days in both groups ) , despite a better gastrointestinal function in the erp group . \n multivariate analysis found that hospital stay was influenced by patient factors including age , cancer stage and presence of complication in the present study . in the erp of the present study , patients were targeted to be discharged on days 23 after operation and hence , pre - determined a hospital stay of about 4 days despite good recovery . as suggested by maessen et al . from the enhanced recovery after surgery ( eras ) group , \n the length of stay may be related to organization of care and not totally related to period of recovery . in view of the fact that patients managed with erp in the study had better post - operative bowel function and no increase in complication or re - admission rate for patients \n , the structure of our erp can be reviewed to target for earlier patient discharge on days 1 2 after operation and improve the patient outcomes further . \n most of previous studies in laparoscopic colectomy adopted traditional care pathway and reported hospital stay of 5 to 9 days , which was longer than what multi - modal rehabilitation program could achieve in open colectomy . \n two randomized control trials ( rct ) [ 12 , 19 ] had compared application of erp in open and laparoscopic colectomy and they showed that the overall short - term outcomes of laparoscopic surgery with erp were better or equal to open colectomy . however , a systematic review by vlug et al . in 2008 reported that due to the lack of data , no robust conclusion can be made on comparison of laparoscopic versus open surgery within application of erp . the present study provided further evidence that the benefit of structured multi - modal rehabilitation program is also suitable for laparoscopic colectomy . \n management of patients with enforced post - operative mobilization , pre - set time for resumption of nutrition and discharge according to pre - defined criteria was not associated with increase in patient morbidity , mortality or re - admission rate . \n the erp adopted in the present study is rather simple : it emphasized patient education , avoidance of tubes and drains , early resumption of nutrition and enforced rehabilitation through a structured post - operative care pathway . \n the design of this multi - disciplinary approach care system had made reference to the previous reports of successful experience in erp [ 8 , 12 , 16 , 18 , 21 ] and taken into account of the feasibility of co - operation by the paramedics as well as the patient acceptance in the light of our local culture . \n the present study did not employ epidural anesthesia which was an important element in many multi - modal rehabilitation programs [ 1113 , 19 ] . \n apart from better pain control and reduction of ileus , epidural anesthesia was shown to attenuate the endocrine - metabolic response through afferent neural blockade and to accelerate patient recovery through reduction of stress response . \n most of the evidence for benefit of epidural anesthesia is , however , from experience in open surgery [ 30 , 3234 ] . in the practice of minimally invasive surgery \n two randomized controlled trials [ 35 , 36 ] had compared results of pca versus epidural anesthesia for laparoscopic colectomy . \n reported that epidural anesthesia did not improve post - operative ileus and senagore et al . \n hence , most recent studies on use of erp for laparoscopic colectomy did not routinely use epidural anesthesia [ 23 , 37 ] . \n the role of epidural anesthesia for laparoscopic abdominal surgery shall be an important area for further evaluation . \n major limitation of this study is that it compared patients in two different periods and was not a randomized controlled study . \n the better gastrointestinal function in the erp group could theoretically be the results of better operative skill among surgeons and improvement in the surgical techniques . \n however , most of the laparoscopic colectomies in the two groups were done by different fellows in the unit . \n moreover , there was no difference in operating time , blood loss or complication rate between the control and the erp groups to suggest that the quality of operation was better in the erp group . \n no result from randomized controlled trial to compare laparoscopic colectomy with and without erp has been reported so far . \n the study protocol of a double blinded multi - center randomized controlled study for laparoscopic and open colectomy with or without fast track surgery in the netherlands was reported . \n the result of this study will provide stronger evidence on the value of erp in laparoscopic colectomy . \n in laparoscopic colectomy , application of erp was associated with significant improvement in gastrointestinal function including earlier passage of flatus and reduction of prolonged ileus after operation but no difference in complication rate , hospital stay and re - admission rate . \n erp could be an effective strategy to optimize the outcomes of laparoscopic colectomy and further investigation is warranted .", "role": "user" }, { "content": "introductionboth laparoscopic colectomy and application of enhanced recovery program ( erp ) in open colectomy have been demonstrated to enable early recovery and to shorten hospital stay . \n this study evaluated the impact of erp on results of laparoscopic colectomy and comparison was made with the outcomes of patients prior to the application of erp.methodsan erp was implemented in the authors center in december 2006 . \n short - term outcomes of consecutive 84 patients who underwent laparoscopic colonic cancer resection 23 months before ( control group ) and 96 patients who were operated within 13 months ; after application of erp ( erp group ) were compared.resultsbetween the erp and control groups , there was no statistical difference in patient characteristics , pathology , operating time , blood loss , conversion rate or complications . \n compared to the control group , patients in the erp group had earlier passage of flatus [ 2 ( range : 15 ) versus 2 ( range : 14 ) days after operation respectively ; p = 0.03 ) ] and a lower incidence of prolonged post - operative ileus ( 6% versus 0 respectively ; p = 0.02 ) . \n there was no difference in the hospital stay between the two groups [ 4 ( range : 234 ) days in control group and 4 ( range : 223 ) days in erp group ; p = 0.4 ) ] . \n the re - admission rate was also similar ( 7% in control group and 5% in erp group ; p = 0.59).conclusionsin laparoscopic colectomy for cancer , application of erp was associated with no increase in complication rate but significant improvement of gastrointestinal function . \n erp further hastened patient recovery but resulted in no difference in hospital stay .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: low back pain and hip and knee joints pain are major concerns in the elderly , whose population is increasing ( 1 - 3 ) . \n some clinical studies have found an alteration in spinal curves with increasing age . altered spinal curves \n may cause disk degeneration , radicular pain and vertebral fractures in the elderly ( 4,5 ) . \n a flattened spine exerts one - fifth the structural strength compared to a spine with thoraco - lumbar curves ( 4 ) . in addition , spinal mobility was proven as an important factor relating quality of life ( 6,7 ) . \n moreover , studies have shown hyperkyphosis is associated with decreased mobility , poor balance and greater body sway , which in turn is correlated with increased risk of falls , vertebral fractures , chronic back pain and functional decline ( 5,7 ) . \n reduced range of motion , as a natural consequence of the muscle - tendon unit and surrounding connective tissue stiffness with aging has been considered as a major cause of falls . \n some studies have indicated that reduced range of motion , especially in hip and ankle joints may increase the risk of falls ( 8,9 ) . \n gait studies of healthy elderly have shown decreased walking speed , less range of motion of the hip , knee , and ankle joints , and increased anterior pelvis tilt ( 9,10 ) . \n muscle tendon tightness that associated with aging process especially affect the bi - articular muscles . \n kerrigan et al ( 8) showed that elderly fallers are characterized by marked hip tightness . \n they also reported an increase in peak hip extension as a result of 10 week stretching exercises ( 11 ) . \n in addition , studies with older persons have shown an association between improved flexibility and increases in activities of daily living and quality of life ( 6 , 7,11 , 12 ) \n \n it seems that contractures of bi - articular muscles of hip , knee and ankle hip flexion contractures to occur with reduced of physical activity in elderly ( 8) . also , the standing posture depends not only on spinal alignment but also on hip , knee and ankle joints alignments ( 1,3 ) . therefore , it is important to explore this issue in the iranian elderly . the present study aimed to compare the thoracic and lumbar curves and hip extension and ankle dorsiflexion range of motions between old and young persons \n 30 healthy old persons over 60 years old ( age range , 63 - 78 yrs , 16 female , 14 male ) were recruited to participate in this study . \n the participants had been living in the facility care unit and were medically stable with no evidence of vestibular or neuromuscular conditions . \n the control group ( young group , n=30 ) with an average age of 23.37 2.31 years was height , weight and sex matched , with same exclusion criteria . before participating in the study , \n all subjects signed an informed consent form approved by the human subjects committee of university of social welfare & rehabilitation sciences . \n a standard goniometer was used for measuring passive range of motion of hip and ankle joints . \n the right and left lower extremities were assessed for each participant .the mean of the two measurements was considered as the joint motion value . for the measurement of passive hip extension \n relaxed hip extension angle was measured using a goniometer and defined as the angle between the horizontal and a line from the greater trochanter to the lateral femoral condyle ( 8) . \n length of plantar flexors was determined by measuring the amount of ankle joint dorsiflexion with the knee extended . \n the subject was positioned in the prone position with the foot hanging off the table and the subtalar joint maintained in the neutral position . \n dorsiflexion was measured with a standard goniometer as the angle formed by the lateral midline of the leg on a line from the head of the fibula to the tip of the lateral malleolus and the lateral midline of the foot in line with the border of calcaneus ( 13 ) . \n subject s spinal curves were assessed using flexible ruler and the measurement performed with the subject standing in relaxed posture . for the measurement of spinal curves \n , the flexible ruler is placed between two marked points ( t1 , t12 ) vertebrae for measurement of kyphosis and t12 , s2 vertebrae for measurement of lordosis ) and shaped as spinal curves . \n the flexible ruler was removed and placed on a piece of white paper so the spinal curves could be copied by a pencil along the flexible ruler . \n a vertical line was drawn to connect the t1 to t12 and t12 to s2 landmarks ( total length of curve / l line ) and the maximal width of spinal curvature measured in centimeter ( h line ) . then using the equation =4 [ arc tang ( 2h / l ) ] the degree of spinal curves were calculated ( 14 ) . \n the intra - class correlation coefficient ( icc ) , two way mixed effect model , and standard error of measurement ( sem values ) were used to assess intra - tester reliability of the measurement . \n we calculated the icc ( 3,1 ) as described by shrout and fleiss ( 15 ) , because only one judge evaluated the same population of subjects . \n independent t tests were performed to compare hip extension and ankle dorsiflexion range of motions and thoracic and lumbar curves between the young and elderly persons . \n a standard goniometer was used for measuring passive range of motion of hip and ankle joints . \n the right and left lower extremities were assessed for each participant .the mean of the two measurements was considered as the joint motion value . for the measurement of passive hip extension \n relaxed hip extension angle was measured using a goniometer and defined as the angle between the horizontal and a line from the greater trochanter to the lateral femoral condyle ( 8) . \n length of plantar flexors was determined by measuring the amount of ankle joint dorsiflexion with the knee extended . \n the subject was positioned in the prone position with the foot hanging off the table and the subtalar joint maintained in the neutral position . \n dorsiflexion was measured with a standard goniometer as the angle formed by the lateral midline of the leg on a line from the head of the fibula to the tip of the lateral malleolus and the lateral midline of the foot in line with the border of calcaneus ( 13 ) . \n subject s spinal curves were assessed using flexible ruler and the measurement performed with the subject standing in relaxed posture . for the measurement of spinal curves \n , the flexible ruler is placed between two marked points ( t1 , t12 ) vertebrae for measurement of kyphosis and t12 , s2 vertebrae for measurement of lordosis ) and shaped as spinal curves . \n the flexible ruler was removed and placed on a piece of white paper so the spinal curves could be copied by a pencil along the flexible ruler . \n a vertical line was drawn to connect the t1 to t12 and t12 to s2 landmarks ( total length of curve / l line ) and the maximal width of spinal curvature measured in centimeter ( h line ) . then using the equation =4 [ arc tang ( 2h / l ) ] the degree of spinal curves were calculated ( 14 ) . \n the intra - class correlation coefficient ( icc ) , two way mixed effect model , and standard error of measurement ( sem values ) were used to assess intra - tester reliability of the measurement . \n we calculated the icc ( 3,1 ) as described by shrout and fleiss ( 15 ) , because only one judge evaluated the same population of subjects . \n independent t tests were performed to compare hip extension and ankle dorsiflexion range of motions and thoracic and lumbar curves between the young and elderly persons . \n table 1 presents the icc and sem values for test- retest reliability of the each measurement taken in the pilot study . \n the result of independent t test showed that the mean of lumbar curve was higher in young group ( 31.29 6.37 ) than elderly ones ( 27.938.11 ) , however , no significant difference was found between two groups ( p=0.08 ) ( table 2 ) . \n the result also showed increased thoracic curvature with aging process ( young group=34.4313.27 , old group=36.198.97 ) , however , no significant difference was found between two groups(p=0.55 ) . \n the result of independent t test also showed that young person s demonstrated a significantly higher hip extension ( hip flexors length ) with ankle dorsiflexion ( plantar flexors length ) range of motions than elderly group ( p<0.01)(table 2 ) . \n due to increasing elderly population is worldwide , concern over health problems and medical and social costs of the elderly are growing . \n the data from this study demonstrated that reduced hip extension and ankle dorsiflexion occur in elderly ( p<0.01 ) . in agreement with previous studies ( 8,10,11,16 ) the present study showed that reduced hip extensors or hip flexion contractures \n there is a tendency with age to develop contractures of the hips , because the hip joint is less often fully stretched due to reduction in physical activity ( 10 ) . on the other hand , the only regular daily activity that extends the hip to its maximum and thereby stretching the hip flexors are walking and running ( 10,15 ) . thus a decline in these activities along with aging will contribute to hip flexors tightness and contracture ( 10 ) . \n it also suggested that hip flexion contractures or reduced hip extension may lead to an increase in anterior pelvic tilt and decrease in stride length and walking speed in elderly people ( 8,11 ) . \n a reduction in hip extension range and stride length may be particularly important in situations requiring rapid changes in stride length such as , attempting to change walking speeds rapidly , or when faced with uneven surfaces or obstacles ( 8,16 ) . \n the sagittal motion around the hip joint is achieved by the hip joint itself and by the support of the lumbar spine ( 17 ) . in our measurement of lumbar lordosis , \n the mean angle in the old and young group were 27.938.11 and 31.296.37respectively and the thoracic mean angle in the old and young group 36.198.97 and 34.4313.27 , respectively , however these differences were not significant . \n some studies showed that degenerative changes of the lumbar spine in middle - aged and elderly people can lead to decrease in lumbar lordosis and thoracic kyphosis ( 1 - 3,18 ) . \n berlemann et al ( 19 ) and tsuji et al ( 2 ) demonstrated that in elderly people , decreasing lumbar lordosis lead to increasing thigh muscle function and knee flexion while standing .offierski and macnab ( 3 ) and tsuji et al ( 2 ) reported a correlation between spine and hip joint pain and lumbar kyphosis and knee flexion position , respectively . \n they reported that concurrent diseases at hip - spine and knee - spine are common clinical problems in elderly ( 2,3 ) . \n it also suggested that kyphotic condition alters balance and increases the incidence of falling ( 17,20 ) . \n it increases the load at anterior part of vertebral body and increases fall risk ( 4 ) . \n our finding also showed that old people have decreased length of the calf muscles ( ankle dorsiflexion range ) than young people . according to previous studies , \n the old people had changes in the ankle joint capsule , associated ligaments fascia , the skin and decreased calf muscles length ( 21,22 ) . aging is known to bring about a loss of functional motor units , a decrease in the number and the size of both muscle fibers ( type i , ii ) , with the possibility of selective atrophy of type ii fibers . \n the reduction in the number of functional motor units and muscle fibers may account for the decreased muscle mass , strength and extensibility observed for the older people ( 21 ) . \n gajdosik also showed that the older women had decreased maximal isometric strength of the calf muscles ( 21 ) . changing calf muscle function due to \n decreased length and strength may have some impact on ambulation and static and dynamic standing balance that warrant further investigation ( 21,22 ) .", "role": "user" }, { "content": "background : falls have been strongly associated with decreased physical activity and impaired mobility . \n reduced range of motion , as a consequence of muscle stiffness , has been indicated to assume a positive relationship to fall incidence . \n also clinical observations suggest that maintaining the normal spinal curves is associated with the prevention of spinal , knee and hip disorders . \n thus , the aim of this study was to compare hip and ankle range of motions and thoracic and lumbar curves between young and old persons \n\n methods : using a nonprobability sampling 30 elderly persons at average of 68.14 4.03 years of age and 30 young people ( age 23.37 2.31 years ) through a case control design participated in the study . \n maximal hip extension and ankle dorsiflexion range of motions were measured by a standard goniometer . \n thoracic and lumbar curvatures were measured by a flexible ruler in both groups . \n independent t test were used to statistically analyze differences between groups . \n results : compared with the young group , the elderly group had decreased hip extension and ankle dorsiflexion motions ( p<0.01 ) . \n the result of independent t test showed that the mean of lumbar curve was higher in young group ( 31.29 6.37 ) than elderly subjects ( 27.938.11 ) , however , no significant difference was found between two groups ( p=0.08 ) . \n the result also showed increasing thoracic curvature with aging ( young group=34.4313.27 , old group= 36.198.97 ) , however , no significant difference was found between two groups \n\n conclusion : findings suggest decreased ankle and hip joint range of motions should be considered in rehabilitation of elderly people .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: cancer of cervix is the third most frequent cancer among women worldwide and is still a major concern especially in developing countries ( 1 ) . \n fortunately , due to hygiene measures and available screening methods , the mortality rate of the disease has been reduced , but still current estimates indicate that approximately 528,000 women are diagnosed with cervical cancer every year and about 267,000 die from the disease ( 1 ) . \n there are various risk factors involved in development of the disease . beside different environmental and behavioral factors like smoking , age at first intercourse , sexual behavior and partners , hygiene , and other sexually transmitted infections ( 2 - 5 ) , several epidemiological and experimental studies have shown a strong relationship between human papilloma viruses ( hpvs ) and cervical cancer . \n in fact , hpvs are known as the major risk factors for developing cervical cancer ( 6 - 8 ) . \n these nonenveloped viruses have an icosahedral capsid and a single double - stranded circular dna genome about 8 kbp in size ( 9 ) . \n it seems that two hpv proteins , e6 and e7 , are involved in the interfering with the cell cycle , leading to transformation of the host cell by inactivating p53 and retinoblastoma proteins ( 10 , 11 ) . \n depending on their association with cancerous or non - cancerous lesions , hpvs are categorized into two groups : high and low risk viruses ( 12 ) . \n low risk hpvs ( types 6 , 11 , 42 , 43 , and 44 ) are related to benign lesions and high risk hpvs ( types 16 , 18 , 31 , 33 , 34 , 35 , 39 , 45 , 51 , 52 , 56 , 58 , 59 , 66 , 68 , and 70 ) are associated with cervical cancer lesions . hpv-16 and hpv-18 are the most common high risk hpvs reported ( 6 , 13 - 16 ) . \n as hpv can not be propagated in tissue culture , its accurate identification relies on molecular biology techniques like pcr , real - time pcr , and nucleic acid - hybridization assays ( 17 , 18 ) . \n one of the mostly used pcr approaches is based on the my09/my11 pair primers which targets a 450 base pair ( bp ) fragment within the hpv l1 orf ( 19 ) . as various genotypes of hpv have different risk of tumorigenesis , determination of distributed genotypes in population is very important in the screening and prevention programs against the disease ( 20 ) . due to \n the limited published data on hpv prevalence rates , it is necessary to conduct more studies to know which hpv types need to be targeted by a prophylactic vaccine in each country . \n the present study was conducted to determine hpv genotypes which are circulating in women s population with cervical cancer and cervical dysplasia / metaplasia lesions using pcr and restriction fragment length polymorphism ( rflp ) molecular methods . \n the present study was retrospectively done and the samples were collected from pathology department of three different women hospitals in khuzestan province , iran ( southwest ) . \n these pathologically confirmed malignant samples were 72 paraffin fixed cervical biopsies which had been previously archived during 1998 to 2008 from women with cervical cancer and cervical dysplasia . \n the patients ages ranged from 32 to 90 years , with the mean age of 55 years . \n ten - micron sections were duplicately prepared from each of the paraffin - embedded blocks and these samples were subjected to dna extraction . \n first , the paraffin - embedded sections underwent a deparaffinization step using previously described method ( 21 ) . \n briefly , 500 l of universal buffer ( ph 9 ) was added to each microtube containing two ten - micron sections followed by heating at 120c for 20 minutes . \n the samples were then cooled ( paraffin forms a solid layer at the top of the liquid phase at this stage ) and the liquid phase was harvested . \n 500 l of phenol : chloroform : isoamyl alcohol ( 25:24:1 , v / v ) solution was added to each sample and the mixture was vortexed and centrifuged at 12000 g for 10 minutes . \n the supernatant was transferred to a clean microtube and equal volume of chloroform was added to the solution and the mixture was vortexed and centrifuged at 12000 g for 5 minutes . \n the supernatant was again transferred to a clean microtube and 0.1 volume of 3 m sodium acetate was added to the solution and the mixture was vortexed and one volume of isopropanol was added to the solution and microtubes were stored overnight at -20c . \n the microtubes were then centrifuged at 12000 g ( 4c ) for 10 minutes , the supernatant was discarded , and the pellet was once washed with 70% ethanol . finally , the microtubes were centrifuged , ethanol was discarded and 50 l of distilled water was added to the pellet ( 21 ) . \n first , quality of the extracted dna samples was investigated using specific primers ( ch20 and pc04 ; tag copenhagen co ) for human beta - globin gene . \n 7.5 l of extracted dna was added to 42.5 l of pcr mix containing 5 u of taq dna polymerase , 5 l of 10x pcr buffer , 1.5 l of 25 mm mgcl2 , 1 l of dntps mix , 0.5 l ( 50 pmol ) of each of ch20 and pc04 primers and 33.5 of l distilled water . \n the reaction was run under the following thermal cycling program : pre - denaturing at 95c for 2 minutes ; denaturing at 95c for 1 minute , annealing at 55c for 1 minute , extension at 72c for 1 minute , ( 40 repeats ) ; final extension at 72c for 5 minutes . \n finally , pcr products were analyzed by electrophoresis on 1% agarose gel in tae buffer containing ethidium bromide ( 0.5 g / ml ) . \n thereafter , positive samples in this pcr assay were subjected to second pcr assay with specific primers ( my09 , my11 ; tag copenhagen co ) for l1 region of hpv genome ( the sequences of the primers are presented in table 1 ) . \n therefore , 10 l of extracted dna was added to 40 l of pcr mix containing 5 u of taq dna polymerase , 5 l of 10x pcr buffer , 3 l of 25 mm mgcl2 , 1 l of dntps mix , 0.5 l ( 50 pmol ) of each of my09 and my11 primers and 29.5 l of distilled water . \n distilled water was also used as a negative control in each run of both pcr assays . \n pcr products of positive samples in hpv pcr assay were purified using gel extraction kit ( qiagen ) according to the manufacturer s protocol and 10 l of each of the purified dna samples were subjected to enzymatic digestion at 37c for 2 hours using rsai restriction endonuclease ( 22 ) . \n thus , digestion of the pcr products obtained by primers my09/11 can lead to the detection of the five most prevalent hpv types 6 ( 4 fragments of 67 , 72 , 149 , 159 bp ; indicator fragments are 149 and 159 bp ) , 11 ( 4 fragments of 26 , 72 , 135 , 216 bp ; indicator fragment is 216 bp ) , 16 ( 3 fragments of 70 , 72 , 310 bp ; indicator fragment is 310 bp ) , 18 ( 5 fragments of 38 , 85 , 72 , 125 , 135 bp ; indicator fragments are 125 and 135 bp ) , and 33 ( 4 fragments of 39 , 72 , 102 , 236 ; indicator fragments are 102 and 236 bp ) . \n afterward , the products of enzymatic digestion were analyzed by electrophoresis on 8% polyacrylamide gel ( page ) in tae buffer followed by staining using ethidium bromide solution to determine the number and size of digested fragments ( 22 ) . \n the present study was retrospectively done and the samples were collected from pathology department of three different women hospitals in khuzestan province , iran ( southwest ) . \n these pathologically confirmed malignant samples were 72 paraffin fixed cervical biopsies which had been previously archived during 1998 to 2008 from women with cervical cancer and cervical dysplasia . \n the patients ages ranged from 32 to 90 years , with the mean age of 55 years . \n ten - micron sections were duplicately prepared from each of the paraffin - embedded blocks and these samples were subjected to dna extraction . \n first , the paraffin - embedded sections underwent a deparaffinization step using previously described method ( 21 ) . \n briefly , 500 l of universal buffer ( ph 9 ) was added to each microtube containing two ten - micron sections followed by heating at 120c for 20 minutes . \n the samples were then cooled ( paraffin forms a solid layer at the top of the liquid phase at this stage ) and the liquid phase was harvested . \n 500 l of phenol : chloroform : isoamyl alcohol ( 25:24:1 , v / v ) solution was added to each sample and the mixture was vortexed and centrifuged at 12000 g for 10 minutes . \n the supernatant was transferred to a clean microtube and equal volume of chloroform was added to the solution and the mixture was vortexed and centrifuged at 12000 g for 5 minutes . \n the supernatant was again transferred to a clean microtube and 0.1 volume of 3 m sodium acetate was added to the solution and the mixture was vortexed and one volume of isopropanol was added to the solution and microtubes were stored overnight at -20c . \n the microtubes were then centrifuged at 12000 g ( 4c ) for 10 minutes , the supernatant was discarded , and the pellet was once washed with 70% ethanol . finally , \n the microtubes were centrifuged , ethanol was discarded and 50 l of distilled water was added to the pellet ( 21 ) . \n first , quality of the extracted dna samples was investigated using specific primers ( ch20 and pc04 ; tag copenhagen co ) for human beta - globin gene . \n 7.5 l of extracted dna was added to 42.5 l of pcr mix containing 5 u of taq dna polymerase , 5 l of 10x pcr buffer , 1.5 l of 25 mm mgcl2 , 1 l of dntps mix , 0.5 l ( 50 pmol ) of each of ch20 and pc04 primers and 33.5 of l distilled water . \n the reaction was run under the following thermal cycling program : pre - denaturing at 95c for 2 minutes ; denaturing at 95c for 1 minute , annealing at 55c for 1 minute , extension at 72c for 1 minute , ( 40 repeats ) ; final extension at 72c for 5 minutes . \n finally , pcr products were analyzed by electrophoresis on 1% agarose gel in tae buffer containing ethidium bromide ( 0.5 g / ml ) . \n thereafter , positive samples in this pcr assay were subjected to second pcr assay with specific primers ( my09 , my11 ; tag copenhagen co ) for l1 region of hpv genome ( the sequences of the primers are presented in table 1 ) . \n therefore , 10 l of extracted dna was added to 40 l of pcr mix containing 5 u of taq dna polymerase , 5 l of 10x pcr buffer , 3 l of 25 mm mgcl2 , 1 l of dntps mix , 0.5 l ( 50 pmol ) of each of my09 and my11 primers and 29.5 l of distilled water . \n distilled water was also used as a negative control in each run of both pcr assays . \n pcr products of positive samples in hpv pcr assay were purified using gel extraction kit ( qiagen ) according to the manufacturer s protocol and 10 l of each of the purified dna samples were subjected to enzymatic digestion at 37c for 2 hours using rsai restriction endonuclease ( 22 ) . \n thus , digestion of the pcr products obtained by primers my09/11 can lead to the detection of the five most prevalent hpv types 6 ( 4 fragments of 67 , 72 , 149 , 159 bp ; indicator fragments are 149 and 159 bp ) , 11 ( 4 fragments of 26 , 72 , 135 , 216 bp ; indicator fragment is 216 bp ) , 16 ( 3 fragments of 70 , 72 , 310 bp ; indicator fragment is 310 bp ) , 18 ( 5 fragments of 38 , 85 , 72 , 125 , 135 bp ; indicator fragments are 125 and 135 bp ) , and 33 ( 4 fragments of 39 , 72 , 102 , 236 ; indicator fragments are 102 and 236 bp ) . \n afterward , the products of enzymatic digestion were analyzed by electrophoresis on 8% polyacrylamide gel ( page ) in tae buffer followed by staining using ethidium bromide solution to determine the number and size of digested fragments ( 22 ) . \n although the collected sample had different statistical characteristics , chi - square test indicated that there was no significant difference between the various groups in all comparisons ( p value = 0.05 ) . \n 45 out of 60 samples ( 75% ) had cervical cancer , whereas , the rest showed cervical dysplasia / metaplasia ( 25% ) . \n these patients were of two groups : those who lived in downtown or rural areas and those who were residents in urban areas . \n fifteen patients ( 25% ) with cervical cancer and six patients ( 10% ) with cervical dysplasia were people who lived in downtown or rural areas , while thirty patients of urban people ( 50% ) had cervical cancer and nine patients ( 15% ) had cervical dysplasia . \n specific sequence of human -globin was successfully amplified in the first pcr assay from 60 out of 72 extracted dna samples . \n this pcr yielded a 268 bp dna fragment which confirmed efficient dna extraction from positive tissue samples . \n lane l , 100 bp dna ladder ( fermentas ) ; lane 11 , negative control contained no template dna . \n 268 bp amplified dna fragment can be seen in several lanes ( e.g. lanes 1 , 2 , 3 , 4 ) and not in lanes 5 , 6 , 7 . \n lane l , 100 bp dna ladder ( fermentas ) ; lanes 2 and 6 , hpv positive samples contained 450 bp amplicon from hpv genome . \n 17 cases of these hpv positive samples were from urban people and 9 cases were from rural people ( table 2 ) . \n moreover , the samples were classified into seven groups based on women s age from 32 to 95 years old . \n most hpv positive samples belonged to women aged from 48 to 55 and 55 to 63 years . \n however , 11 out of 45 samples with cervical cancer and 5 out of 15 cervical dysplasia / metaplasia samples belong to 48 - 55 years old group , while 55 - 63 age group only contained 8 samples with cervical cancer . \n meanwhile , the samples were categorized into four groups based on women s first - marriage age from 12 to over 23 years . \n 13 out of 26 hpv positive samples were from the group with the first - marriage age of 12 - 16 years . \n in addition , 15 and 41 cases of the study population were affected by dysplasia / metaplasia and squamous cell carcinoma , respectively . \n these results showed that percentage of hpv positive samples in scc samples were higher ( 48.8% ) than dysplasia / metaplasia samples ( 26.6% ) . \n digestion with this enzyme leads to the identification of five types of the virus including types 6 , 11 , 16 , 18 , and 33 ( 22 ) . \n polyacrylamide gel electrophoresis ( page ) showed that digestion of the products of hpv pcr assay led to the production of two 310 and 70 bp dna fragments ( although there were actually 3 fragments of 70 , 72 , and 310 bp , the difference between 70 and 72 bp fragments was not recognizable , and thus the electrophoretic pattern consisted of the two dna fragments ) . \n lane l , 100 bp dna ladder ( fermentas ) ; lanes 2 and 3 contain 310 and 70 bp dna fragments produced from enzymatic digestion ; lane 3 , undigested 450 bp product of hpv pcr as a control . \n specific sequence of human -globin was successfully amplified in the first pcr assay from 60 out of 72 extracted dna samples . \n this pcr yielded a 268 bp dna fragment which confirmed efficient dna extraction from positive tissue samples . \n lane l , 100 bp dna ladder ( fermentas ) ; lane 11 , negative control contained no template dna . \n 268 bp amplified dna fragment can be seen in several lanes ( e.g. lanes 1 , 2 , 3 , 4 ) and not in lanes 5 , 6 , 7 . \n lane l , 100 bp dna ladder ( fermentas ) ; lanes 2 and 6 , hpv positive samples contained 450 bp amplicon from hpv genome . \n 17 cases of these hpv positive samples were from urban people and 9 cases were from rural people ( table 2 ) . \n moreover , the samples were classified into seven groups based on women s age from 32 to 95 years old . \n most hpv positive samples belonged to women aged from 48 to 55 and 55 to 63 years . \n however , 11 out of 45 samples with cervical cancer and 5 out of 15 cervical dysplasia / metaplasia samples belong to 48 - 55 years old group , while 55 - 63 age group only contained 8 samples with cervical cancer . meanwhile \n , the samples were categorized into four groups based on women s first - marriage age from 12 to over 23 years . \n 13 out of 26 hpv positive samples were from the group with the first - marriage age of 12 - 16 years . \n in addition , 15 and 41 cases of the study population were affected by dysplasia / metaplasia and squamous cell carcinoma , respectively . \n these results showed that percentage of hpv positive samples in scc samples were higher ( 48.8% ) than dysplasia / metaplasia samples ( 26.6% ) . \n digestion with this enzyme leads to the identification of five types of the virus including types 6 , 11 , 16 , 18 , and 33 ( 22 ) . \n polyacrylamide gel electrophoresis ( page ) showed that digestion of the products of hpv pcr assay led to the production of two 310 and 70 bp dna fragments ( although there were actually 3 fragments of 70 , 72 , and 310 bp , the difference between 70 and 72 bp fragments was not recognizable , and thus the electrophoretic pattern consisted of the two dna fragments ) . \n lane l , 100 bp dna ladder ( fermentas ) ; lanes 2 and 3 contain 310 and 70 bp dna fragments produced from enzymatic digestion ; lane 3 , undigested 450 bp product of hpv pcr as a control . \n there are different risk factors involved in development of this lesion and it has been shown that one of the major risk factors is infection with human papillomavirus ( 6 - 8 ) . \n prevalence of this disease in developing countries is higher than in developed countries probably because there are no organized programs for screening and early diagnosis ( 23 ) . \n few studies have been conducted to determine the epidemiology and effective risk factors on hpv infected cases in the middle east countries and there are only sporadic available data about this cancer from iran . \n besides , there is no organized screening , vaccination , or eradication program to control cervical cancer . as a result , it is certainly difficult to take proper preventive measures against the disease without organized statistics . although there are some assays like pap smear test which are very applicable , but the results could be false negative ( 24 ) . in addition , knowing the prevalence of hpv infections provides invaluable data towards managing , controlling , and prevention of cervical cancer in women population . \n meanwhile , determination of the most prevalent hpv type is very important in preparing effective vaccines against viral infection in different regions and/or countries . \n although there are some approved vaccines being used against hpv infections ( 25 - 27 ) , type(s ) of prevalent papilloma viruses should first be identified to administer the most appropriate vaccine . \n several studies have been conducted to determine prevalence of hpv infections in different groups of patients , the most prevalent types of the virus with their geographic distributions , and risk factors involved in development of cervical cancer . in an extensive epidemiological study , bosch and colleagues tried to determine whether the association between hpv infection and cervical cancer is consistent worldwide and to investigate geographic variation in the distribution of hpv types . \n they tested over 1000 histologically confirmed specimens from patients with invasive cervical cancer at 32 hospitals in 22 countries by pcr - based assays , capable of detecting more than 25 different hpv types . \n their results showed that 93% of the tumors were infected with hpv dna with no significant variation in hpv positivity among countries . \n hpv 16 was present in 50% of the specimens , and were found to be the predominant virus type in all countries ( 6 ) . \n munoz and colleagues performed a study to broaden the knowledge of high risk hpv types . \n they pooled data from 11 case - control studies from nine countries involving 1918 women with histologically confirmed squamous - cell cervical cancer and 1928 control women . \n hpv dna was detected in 1739 of the 1918 patients with cervical cancer ( 90.7% ) and in 259 of the 1928 control women ( 13.4% ) . \n the most common hpv types in patients , in descending order of frequency , were types 16 , 18 , 45 , 31 , 33 , 52 , 58 , and 35 ( 16 ) . \n in another study performed by sadeghi et al . in 2008 , 52 paraffin - embedded blocks with cervical cancer and 52 paraffin - embedded cervical tissue blocks with normal histology were assayed and the results showed that 30.7% of samples with cervical cancer were infected by hpv , while there was not any positive sample in healthy specimens ( 28 ) . \n maleknejad et al . tried to delineate hpv infections in 64 paraffin - embedded samples from patients with cervical cancer . \n 59.4% of samples were infected with hpv dna , 34.4% of which were hpv 16 and the rest ( 25% ) were hpv 18 ( 29 ) . \n the results depicted that 84.5% of samples were infected by hpv 16 and hpv 18 , but the most prevalent virus was hpv 16 ( 30 ) . in a survey , yang and \n colleagues investigated correlation of viral factors with cervical cancer in taiwan . 27 cervical cancer biopsy specimens and 29 normal cervical scrapings \n were tested and they found that 18 ( 66.7% ) out of 27 cervical biopsies were infected by hpv dna . among those , \n hpv type 16 ( 9 cases ) was the most closely associated with cervical cancer and type 18 ranked second ( 31 ) . in another research , \n the two most common types were 16 ( 44% ) and 18 ( 39% ) , which also indicated high prevalence of hpv 16 ; however , there are some data which reported hpv 18 as the most common virus type in this region followed by hpv 16 ( 6 , 32 ) . \n considerably , hpv 16 was the most or one of the most prevalent types of the virus in many studies . \n the results of the present study were also in agreement with the previous findings . in this study , it was shown that 43.33% of patients with cervical cancer or cervical dysplasia were infected by hpv dna . \n unfortunately , as we did not have access to a reference hpv dna genome , we did not incorporate a positive control . \n however , it seems that my09/my11 , which is one of the mostly used pairs of primers , has been proved to be a highly sensitive and specific primer set for detection of hpv dna ( 33 ) . \n nevertheless , it would be better to use another pair of primers e.g. gp5+/gp6 + , to increase sensitivity for detection of hpv dna in the examined samples ( 34 ) . \n meanwhile , rflp analysis by rsai restriction enzyme showed that all ( 100% ) hpvs belonged to one of the most prevalent oncogenic hpvs , hpv type 16 . \n the results of this study confirmed that pcr could be a useful assay to detect hpv viral dna and subsequent rflp analysis may help to identify different hpv types circulating in a population . \n therefore , it seems that hpv 16 is one of the essential types of the virus which should be included in vaccines against the disease . \n meanwhile , although chi - square test did not show any significant difference between various groups of people , there are some points worthwhile to notice . \n most of cervical cancer and hpv positive samples belonged to the groups aged from 48 to 71 years , especially 48 - 55 years . \n in fact , the number of samples was not equal in different sample groups ; however , this may also imply on the higher frequency of patients who referred to hospitals in those age groups . \n it should be considered that previous studies reported that hpv infection is most common in sexually active young women , 18 to 30 years of age and there is also a sharp decrease in prevalence after 30 years of age . \n however , cervical cancer is more common in women older than 35 years , suggesting infection at a younger age and slow progression toward cancer ( 15 , 35 ) . \n this difference may be a reflection of social behaviors and traditions , and women s first sexual intercourse in the region which usually occurs in older ages compared to other regions . \n besides , most cases of cervical cancer ( 36.7% ) and hpv positive samples ( 21.7% ) belonged to women who got married before 17 . \n the correlation of marriage age and the incidence of the disease is noticed in some studies ( 36 , 37 ) . \n however , some researchers did not find any correlation between marriage age and the incidence of the disease ( 38 ) . \n these data show that there is a need to perform more studies to increase the knowledge about biology of different hpv types and involved risk factors in the development of the disease . \n the results of the present study can be guidance for future programs regarding screening and managing of this major disease and also for taking appropriate vaccination strategies against the prevalent types of the virus .", "role": "user" }, { "content": "background : cervical cancer is one of the important reasons of mortality among females . \n prevention , early diagnosis and immediate treatment can affect the rate of mortality in this cancer and several epidemiological studies have shown a strong relationship between human papilloma viruses ( hpvs ) and cervical cancer . \n objectives : the present study was conducted to survey hpv infections in a women population with cervical cancer and cervical dysplasia / metaplasia in southwest of iran.materials and methods:72 paraffin - embedded cervical biopsies which had been previously archived from women with cervical cancer and cervical dysplasia were examined by polymerase chain reaction ( pcr ) . \n afterward , the detected hpv strains were typed by restriction fragment length polymorphism ( rflp ) analysis of pcr amplicons.results:60 out of 72 samples had necessary requirements and hpv dna was detected in 43.3% of these samples . \n most hpv positive samples belonged to women aged from 48 to 63 years . on the other hand , \n hpv infection among patients with squamous cell carcinoma ( scc ) was 48.78% and in women with dysplasia / metaplasia was 26.66% . \n the most prevalent type of the human papilloma virus was hpv16 ( 100%).conclusions : knowing the most prevalent type of the human papilloma viruses circulating in the population ( hpv16 ) can be applied in the future screening and managing programs of this major disease and also in vaccination against the prevalent types of the virus . \n meanwhile , it seems that more studies should be performed to determine the role of different risk factors involved in development of the disease , especially those related with social behaviors and traditions with respect to different areas .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: the world health organization ( who ) defines overweight and obesity as body mass index ( bmi ) 25 and bmi 30 , respectively . around the world , obesity has become a worrying health and social issue , threatening lives of thousands of people . according to the who , over 1.9 billion adults ( 39% adults ) were overweight among which more than 600 million ( 13% adults ) were obese . \n childhood obesity is also common that 42 million children were overweight or obese in 2013 . \n considering its high prevalence , it is pressing to study the pathogenesis , manifestations , and prevention of obesity . \n obesity is related to a range of health - related problems , such as diabetes , heart disease , hypertension , and cancer . \n obese children show greater cardiovascular risk factors and persistence of obesity into their adulthood , which may be associated with higher likelihood of premature mortality [ 4 , 5 ] . \n in addition to health problems , obesity is associated with poorer cognition and motor control , and altered brain plasticity . in this review , \n we first look into the behavioral manifestations of obese individuals ' cognition and motor control capabilities . \n , the effects of physical exercise to combat obesity and obesity - related deficits in cognition and motor control will also be described . \n overweight and obesity are usually related to poorer cognition across lifespan [ 68 ] ; however , the association between bmi and cognitive function is weaker in old age [ 9 , 10 ] , partly due to inaccurate adiposity measurement in the aged people . \n indirect evidence has shown an association between western high fat diet and impaired cognitive functions . based on bmi data , individuals who are overweight or obese fall in the lowest quartile of global cognition , verbal fluency , delayed recall , immediate logical memory , and intelligence . \n other than bmi , other adiposity measures are also related to cognitive performance and brain changes . \n there is also a negative correlation between waist - to - hip ratio and hippocampal volume and a positive correlation between waist - to - hip ratio and white matter hyperintensities . \n compared to bmi , central adiposity has a stronger association with the risk of developing cognitive impairment and dementia in women . \n hence , studies using bmi as the only indicator of obesity may not be sensitive enough to capture obesity - induced cognitive dysfunctions . \n neuroimaging studies demonstrate atrophy in the frontal lobes , anterior cingulate gyrus , hippocampus , and thalamus in older obese individuals . \n bmi increase is associated with lower metabolic activity in the prefrontal cortex and cingulate gyrus , smaller gray matter volume in many brain regions ( particularly prefrontal cortex ) , and deficient white matter integrity in the uncinated fasciculus which is a structure connecting the frontal and temporal lobes [ 1822 ] . \n smaller gray matter volume in the left orbitofrontal region is related to poorer executive performance in obese women . \n childhood obesity is related to the reduced executive function , attention , mental rotation , mathematics , and reading achievement [ 2325 ] . \n obese adolescents have deficits in a range of cognitive functions , such as attention and executive functions [ 26 , 27 ] . \n an animal study shows that high fat diet induces similar morphometric and metabolic changes in juvenile and adult mice ; however , only early exposure to high fat diet hurts relational memory flexibility and decreases neurogenesis . \n people with higher midlife bmi have lower global cognition than their thinner counterparts and midlife obesity is related to the accelerated cognitive aging , but this association is weaker in older adulthood . \n both age and bmi contribute independently to decreased brain volume in middle and older adulthood . \n it is more likely for an older adult to have lower cognitive abilities if he / she was overweight or obese during middle age [ 32 , 33 ] . \n midlife obesity is related to an increased pace of deterioration in executive functions and an increase in waist - to - hip ratio is associated with substantial reduction in total brain volume . \n lower bmi and waist circumference and higher fat - free mass are associated with slower cognitive decline . \n midlife overweight / obesity , particularly with metabolic abnormality , is associated with higher dementia risk in older adulthood [ 33 , 3640 ] . moreover \n hence , midlife is a critical period in which the overweight / obese status can predict one 's cognitive functions and brain health in later life . \n besides cognition , obesity also affects motor control capabilities , degrading daily functions and health . \n children who are obese or overweight are poorer in gross and fine motor control and have delayed motor development [ 4450 ] . \n obese girls of 6th and 7th grades participate in less physical activity and have lower enjoyment of physical activity . \n children with high bmi have lower level of run which is a fundamental motor skill based on which complex motor skills are learned . \n observe that the prevalence of mastery of all fundamental motor skills is lower in overweight / obese children , especially for run , slide , hop , dribble , and kick . \n in addition to bmi , waist circumference is also related to children 's and adolescents ' ability to perform fundamental motor skills . \n there is an inverse relationship of bmi with fine motor precision , balance , running speed and agility , and strength in the 1st graders . \n obese children also have difficulty in postural coordination and a heightened dependency on vision during locomotion which is rather automatic in nonobese children [ 57 , 58 ] . \n adiposity is related to muscle quality ratio that is associated with motor conduction velocity and finger tapping speed . \n subcutaneous fatness can account for a significant variance of health - related and motor fitness . \n excessive fat mass is associated with poorer posture and walking . in middle and older adults , a combination of high bmi ( or waist circumference ) and high blood pressure is related to lower motor speed and manual dexterity . during postural control , \n obese individuals require greater attention resources to maintain balance during unipedal stance ; this implicates that obese people consume attention resources to compensate for their motor deficits . \n for example , obesity may affect brain structure , leptin and insulin dysregulation , oxidative stress , cerebrovascular function , blood - brain barrier , and inflammation [ 11 , 6571 ] . \n some also suggest that obesity - related changes in metabolism interact with age to impair brain functions . in terms of brain structure , \n obese individuals have lower cortical thickness in the left superior frontal and right medial orbitofrontal cortex . \n there is also a negative relationship between neuronal injury and gray matter density in hippocampus and cerebellum in overweight and obese individuals . \n it is suggested that the medial orbitofrontal cortex , hippocampus , and cerebellum are involved in reward - based learning , memory , and motor control and learning [ 7577 ] ; structural alterations in these regions may be associated with deficits in cognitive and motor domains . \n diet - induced obesity promotes reactive oxygen species in the brain which is associated with both body weight and adiposity [ 79 , 80 ] . in children , \n occurrence of 15-week obesity during childhood can induce permanent epigenetic changes in rat 's brain . in rats , triglycerides diminish the passage of insulin - like growth factors ( igfs ) into the brain through cerebrospinal fluid , impair hippocampal long - term potentiation , and impede leptin transportation across the blood - brain barrier [ 8385 ] . \n juvenile exposure to high fat diet impairs long - term spatial memory , but not short - term memory , suggesting a selective impairment of consolidation which is likely contributed by increased proinflammatory cytokine expression in the hippocampus . moreover , \n consumption of western diet is thought to degrade blood - brain barrier , which consequently damages hippocampus and leads to dementia . \n relative to those having normal diet , mice consuming high fat diet for 17 days develop insulin resistance in cerebral cortex tissues , degraded synaptic integrity , and poorer spatial memory . \n it can cross the blood - brain barrier and binds to presynaptic gabaergic neurons to produce its effects [ 89 , 90 ] . \n as leptin receptors are widespread in the brain ( e.g. , throughout the cortex and the hippocampus ) , leptin can modulate memory processes . \n obese mice with impaired leptin signaling have deficits of hippocampal - dependent memory and increased basal hippocampal inflammation [ 94 , 95 ] . \n leptin is related to neurogenesis , axonal growth , and synaptogenesis in addition to hypothalamic functions [ 9698 ] . for hippocampal neurons \n , leptin plays a role in long - term potentiation and depression and thus is important for synaptic plasticity [ 92 , 99 , 100 ] . \n compared to those with low leptin level , the elderly with high leptin level show less cognitive decline during aging . \n high leptin level in individuals with small waist circumference is related to less cognitive decline over 10 years . \n the presence of leptin may decrease the production of amyloid and speed up the removal of -amyloid . \n obese individuals usually develop leptin resistance which results in an increase in food intake and alteration of energy expenditure . \n the circulating levels of insulin and signaling pathway are altered in obesity ; this interacts with inflammatory processes to modulate cognition and behaviors . \n insulin receptors are widespread in hippocampal and cortical brain structures , insulin signaling can contribute to the formation of declarative memory . \n insulin concentrations vary with adiposity and there is a negative relationship between the amount of visceral fat and insulin sensitivity . \n insulin resistance can result from high fat consumption or obesity [ 110 , 111 ] . \n dysfunctional insulin signaling can induce inflammation and promote -amyloid and tau pathology , contributing to neurodegeneration [ 112 , 113 ] . \n insulin resistance can mediate cognitive impairment and neurodegeneration as insulin and igfs can regulate neuronal survival , metabolism , and brain plasticity [ 114 , 115 ] . during insulin resistance , there is a failure of cells to metabolize glucose , which consequently triggers an increase of insulin level . \n insulin signaling is related to tau phosphorylation , an early pathology of alzheimer 's disease [ 116 , 117 ] ; this is complementary to the fact that there are a large number of insulin - sensitive glucose transporters in the medial temporal lobe . \n thus , insulin dysregulation in the obese people likely confers a greater risk of dementia to them . \n the adipose tissue produces many substances for metabolism ( adipokines , such as bdnf ) and inflammation ( cytokines , such as leptin ) . \n many cytokines , such as interleukin-1 , produced by the adipose tissue can cross the blood - brain barrier and affect cognitive functions through neuroinflammation [ 95 , 119 ] . \n similar to leptin , it exerts its effects in the brain to bring about weight reduction . \n reduced hippocampal adiponectin levels are observed in aging animals , independent of high fat diet intake . \n neurotrophins , such as igf-1 and bdnf , can mediate obesity 's effects on cognition and behaviors . \n igf-1 is mainly produced in liver and binds to the igf-1 or insulin receptors to exert its effects to stimulate cell growth and proliferation and promote -amyloid clearance in the brain . \n obese individuals usually show igf-1 resistance , degrading their capability to prevent -amyloid deposition and neurodegeneration [ 114 , 123 ] . besides \n , bdnf can bind to many receptors , such as trkb and lngf receptors , to support neuronal survival and stimulate neurogenesis and synaptogenesis [ 124126 ] . \n bdnf promotes neuronal differentiation and survival , neurogenesis , and brain plasticity and is thus particularly crucial for learning and memory . \n high fat diet reduces bdnf level in the hippocampus , and the impaired hippocampal synaptic plasticity and cognition are possibly through bdnf 's effects on dendritic spines . \n diet - induced obesity reduces hippocampal expression of bdnf and presynaptic synaptophysin , which are related to an impairment of spatial learning in mice . \n although mounting evidence shows that obesity is associated with structural and functional brain changes , the causal link between them requires further investigations . \n the composition of gut microbiota appears to be causally related to obesity [ 132134 ] , playing a significant role in body weigh regulation since birth [ 135 , 136 ] . \n gut microbiota plays a key role in childhood obesity and brain development [ 137 , 138 ] . \n a comparison of germ - free mice and conventionally reared mice has demonstrated that germ - free mice are leaner and more resistant to diet - induced obesity . \n obese and nonobese individuals have different diversity and composition of gut microbiota [ 140 , 141 ] . as gut \n microbiota controls energy extraction and storage in the body , significant changes in gut microbiota can result in obesity and insulin resistance [ 139 , 140 , 142 ] . \n it has been suggested that diet can influence gut microbiota which in turn impacts the brain and behaviors through neural , hormonal , immune , and metabolic pathways [ 143 , 144 ] . \n transplantation of gut microbiota of diet - induced obese mice to lean mice is sufficient to bring about neurobehavioral changes through increasing neuroinflammation and disrupting cerebrovascular homeostasis [ 145 , 146 ] . \n mice consuming high energy diet containing higher percentage of clostridiales and lower expression of bacteroidales have poorer cognitive flexibility . in humans , the firmicutes \n gut microbiota can modulate a range of neurotrophins , such as bdnf and synaptophysin , to affect neural plasticity [ 149 , 150 ] . \n thus , diet changes gut microbiota which influences neurophysiology and neurotrophins , eventually impacting cognition and behaviors . \n previous results have shown that obesity - related brain plasticity alteration is a multifaceted issue , which can inflict permanent harm to individuals in their early ages . thus \n exercise can improve physical and cognitive performance , and quality of life in the elderly [ 151155 ] . in humans , those who are highly fit or aerobically trained have greater prefrontal and parietal activations for spatial selection and inhibitory functioning . \n there is a positive relationship between aerobic fitness and spatial memory which is mediated by hippocampus volume . \n aerobic training can increase hippocampal volume of the elderly ( with or without mild cognitive impairment ) and increases plasma bdnf level in both patients of alzheimer 's disease and healthy controls [ 158163 ] . \n regular physical activity is related to better cognition , less cognitive decline , and a lower risk of developing dementia [ 164 , 165 ] . \n as young as children , aerobic fitness can predict cognitive performance over time . besides cognition , higher level of physical activity \n is related to a reduced white matter hyperintensity burden on motor function in the aged people . \n lifelong exercise can preserve white matter microstructure related to motor control and coordination in the elderly . \n in addition , regular physical activity has long been suggested to be an effective way to improve obesity and related problems [ 170 , 171 ] . \n exercising 5 days per week for 15 weeks can improve executive functions in overweight children . \n high - intensity physical activity ( both aerobic and endurance training ) for 4 months improves cognition and oxygen extraction in obese individuals . \n the effectiveness of exercise may be moderated by exercise intensity and duration , and exerciser 's developmental stage [ 174 , 175 ] . \n exercise intensity can be related both to behavioral outcomes and to changes in brain structure and bdnf level . \n greater amount of physical activity in early life is associated with larger prefrontal and hippocampal volumes . \n individuals receiving low - intensity exercise , but not high - intensity , show increased bdnf expression . \n bdnf level depends on exercise intensity ; some observe that moderate - intensity exercise is the most effective to promote bdnf in the elderly . \n fail to observe any augmentation of task switching performance after a single bout of moderately intense exercise . in midlife mice , only 4-month ( but not 2-month ) \n running training can trigger activation of the antiamyloidogenic , prosurvival , synaptogenic , and neuroprotective pathways . \n wheel running for 14 days can increase cell proliferation in the dentate gyrus whereas wheel running for 56 days can additionally facilitate long - term potentiation in this region . \n four - week exercise can improve recognition memory in adult rats , but no such enhancement can be recorded 2 weeks after cessation of training . however , in adolescent rats , the enhancement of recognition memory is preserved . \n , physical activity can enhance neurogenesis , neuroadaptation , and neuroprotection though the actions of neurotrophic factors [ 184190 ] . \n hippocampal function is restored by physical activity through enhancing the expression of neurotrophic factors to promote neurogenesis , angiogenesis , and synaptic plasticity [ 191193 ] . \n for example , bdnf level increases with physical activity , particularly regular exercise [ 194 , 195 ] . \n it is found that bdnf can stimulate dna repair to protect cortical neurons against oxidative stress . \n short bout of mild exercise for 5 weeks improves both oxygen consumption and long - term spatial learning and memory in aged rats which is associated with hippocampal bdnf level . following physical activity , \n the elevated bdnf level in the dentate gyrus is sufficient to induce spatial memory improvement . \n a week of voluntary exercise is sufficient to increase the activity of tissue type plasminogen activator to facilitate the cleavage of probdnf into mbndf . \n a 7-day exercise intervention can increase gray matter volumes in the motor , somatosensory , association , and visual cortices in rats . \n exercising for 6 months reduces default mode network activity in the precuneus while one - year walking increases functional connectivity within the default mode network and frontal executive network . \n regular physical activity can reduce proinflammatory and increase anti - inflammatory signaling and reduce oxidative stress in aged animals [ 206 , 207 ] . \n exercise also reduces peripheral risk factors , such as diabetes and cardiovascular diseases which are associated with neurodegeneration . \n furthermore , vasculature is altered after exercise . in middle - aged rats , total length and surface area of cortical capillaries \n aerobic exercise at midlife can improve vascular dysfunctions , astrocyte hypertrophy , and myelin dysregulation associated with sedentary lifestyle [ 210 , 211 ] . \n exercise is associated with a range of improvements in the brain through a range of mechanisms in individuals of different weight statuses ( figure 1 ) . \n the effectiveness of exercise depends on the training parameters , such as intensity , duration , and developmental stage of exerciser . \n previous research results have consistently suggested that moderately intense exercise for a long enough period of time is especially beneficial for young exercisers . \n more and more people are becoming obese , producing aversive effects on their cognition , motor behaviors , and quality of life . \n previous research has suggested that altered brain structure and activation mediate obesity 's influences on cognition [ 1721 ] , whereas obesity influences the musculoskeletal system to degrade motor performance . as motor performance partly depends on cognitive ability , obesity may indirectly contribute to motor deficits through cognitive decline ( figure 1 ) . substantial research has shown that obesity affects our cognition and motor behaviors through different mechanisms , possibly through altering brain structure , leptin / insulin regulation , oxidative stress , cerebrovascular function , blood - brain barrier , and inflammation [ 11 , 6571 ] . \n it is suggested that moderately intense exercise for a long enough period of time seems favorable ; however , the training parameter for optimal outcomes remains to be determined . \n most of the previous research focuses on aerobic exercise ; the efficacy of anaerobic exercise to improve obesity and related dysfunctions is not well understood . \n more efforts should be devoted to investigate the efficacy of anaerobic exercise , in comparison with aerobic exercise . \n moreover , starting exercising in young age is particularly important to protect from neurodegeneration in old age . as childhood obesity is becoming more prevalent [ 2325 ] , introducing physical activity intervention in childhood may help children improve obesity and prevent age - related functional decline in old age . \n in addition to exercise , leptin replacement therapy , inhaled insulin therapy , and caloric restriction have also been proposed to improve obesity . \n leptin is responsible for energy balance and body weight and can affect neurogenesis and brain functions . \n it is observed that 18-month leptin replacement therapy increases gray matter concentration and activations in brain regions implicated in hunger and satiation neural circuits [ 213 , 214 ] . during weight loss , \n it is found that intranasal insulin exerts anorexic effects , promoting satiety and regulating food intake [ 219 , 220 ] . \n inhaled insulin reaches the brain through olfactory nerves and specific receptors in blood - brain barrier to exert its effects . \n caloric restriction also improves obesity and reverses deficits in leptin receptor protein and signaling associated with diet - induced obesity . \n after 3 months of caloric restriction , serum bdnf increases in overweight and obese individuals . \n diet - induced weight loss is related to a decrease in plasma free fatty acid and improvement in episodic memory . \n hitherto , the efficacy of leptin replacement therapy , inhaled insulin therapy , and caloric restriction on cognition and motor behaviors is poorly understood , which warrants further verification . \n it affects cognition mainly through altering the brain structures and functions [ 1721 ] , and motor performance through degrading musculoskeletal system . \n obesity can affect brain structure , leptin / insulin dysregulation , oxidative stress , cerebrovascular function , blood - brain barrier , and inflammation [ 11 , 6571 ] , which are involved in the deterioration of cognitive and motor functions . \n a host of previous research has suggested that exercise can improve both obesity - related cognitive and motor declines . as more and more people develop obesity in young age , introducing exercise intervention", "role": "user" }, { "content": "due to a sedentary lifestyle , more and more people are becoming obese nowadays . \n in addition to health - related problems , obesity can also impair cognition and motor performance . \n previous results have shown that obesity mainly affects cognition and motor behaviors through altering brain functions and musculoskeletal system , respectively . \n many factors , such as insulin / leptin dysregulation and inflammation , mediate the effect of obesity and cognition and motor behaviors . \n substantial evidence has suggested exercise to be an effective way to improve obesity and related cognitive and motor dysfunctions . \n this paper aims to discuss the association of obesity with cognition and motor behaviors and its underlying mechanisms . following this , \n mechanisms of exercise to improve obesity - related dysfunctions are described . \n finally , implications and future research direction are raised .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: the coronary artery disease(cad ) is the main cause of death in many european countries , including albania ( 1 ) . \n the conventional coronary angiography is the most used technique , the so called gold standard for the cad diagnosis . \n recently the coronary angiography has been challenged by a new noninvasive technique , the ctac ( ct angiography of coronaries ) , that has gained popularity in the last decade with the fast development of technology in the field of imaging , substituting increasingly the invasive coronary angiography in cad diagnosis ( 2 , 3 ) . \n the introduction of multi slice ct scans with 64 detectors brought a revolution in the imaging of the heart , increasing obviously the sensitivity and the specificity of this method in the diagnosis of the coronary atherosclerosis up to the values of 99% and 97% respectively ( 4 ) . \n many recent studies in this field strongly support the usage of ctac in the diagnosis of cad as an alternative of the coronary angiography ( 4 , 5 , 6 ) \n . cardiac risk assessment has been traditionally based on a combination of diabetes , obesity , history of elevated blood pressure , cigarette smoking , plasma level of total cholesterol and its fractions , and family history of premature coronary heart disease ( 1 , 7 , 8) . \n risk factor analysis was introduced to medicine by the framingham study in 1948 ( 9 ) . since \n then several risk factor algorithms have been developed with the goal of predicting major or fatal cardiovascular events ( cve ) and to help to find out the most appropriate individual diagnostic and treatment strategy . however , a significant number of cve occur either in the absence of risk factors or in presence of moderate risk when an aggressive treatment strategy would not be indicated ( 1 , 10 ) . computed tomography has emerged as a noninvasive , patient - friendly diagnostic modality to detect the presence of coronary atherosclerosis . the diagnostic potential of computed tomography coronary angiography is high because it allows not only the detection of significant coronary stenoses but also the presence of non obstructive coronary plaques ( cap ) ( 4 , 11 ) . \n early subclinical coronary artery disease diagnosed with this non - invasive tool might therefore have a role in refining risk on an individual basis beyond the conventional risk factors or algorithms to estimate risk of cve . \n there are several recent studies on the association of conventional risk factors with the prevalence and distribution of cap as detected by ctca in individuals with no history of coronary artery disease ( 1 , 12 , 13 ) and , to our knowledge , the relation between cap and algorithms to estimate risk still remains unclear . \n also the presence of calcium in the coronary arteries has been shown to be a marker for atherosclerotic disease ( 14 ) . \n this calcium is detectable on ct and is quantifiable using the agatston method ( 15 , 16 ) , which adds prognostic information to available demographic and serologic risk stratification ( 17 , 18 ) . \n however , ct performed for calcium scoring is not able to show non calcified atheroma or stenosis . \n many studies have shown that the ca score ( calcium score ) is a diagnostic test with a high negative predictive value to exclude cad but it has a low specificity especially in young ages because of the inability to diagnose the soft atherosclerotic plaques that have a high rupture risk ( 19 ) . \n the aim of our study was to ( 1 ) prospectively investigate the prevalence of cap as detected by ctac in albanian individuals with no history of coronary artery disease or acute coronary syndrome ; ( 2 ) to evaluate the correlation of traditional risk factors with the prevalence of coronary artery plaque and to the expected 10-year risk of fatal cve . \n the group of study : in this study we have included 456 patients without history of cad that applied for a cardiac check - up or referred by doctors to our hospital in order to rule - out coronary atherosclerosis from september 2009 to march 2013 . \n indications for performing a ctac were chest pain , shortness of breath , syncope or equivocal stress testing including exercise ecg , myocardial perfusion imaging or stress echocardiography unable to definitively rule - out / rule - in significant coronary artery disease . \n exclusion criteria for performing ctac were renal insufficiency ( serum creatinine 120 mol / l ) , contraindications to the administration of iodinated contrast , pregnancy , acute coronary syndromes , and ventricular and/or supraventricular arrhythmias . \n systolic and diastolic blood pressures were measured in the sitting position after 5 minutes of rest . \n an individual whose arterial blood pressure was .140/90 mm hg or was taking antihypertensive medications was classified as having hypertension ( 20 ) . \n an individual with a non - fasting plasma glucose concentration of at least 200 mg / dl ( 11.1 mmol / l ) , or fasting plasma glucose level of at least 126 mg / dl ( 7.0 mmol / l ) , or was being treated with anti - diabetic medication was considered to have diabetes ( 21 ) . \n an individual with a body mass index ( bmi ) ( calculated as weight divided by height squared ) of 30 kg / m2 or more was considered to be obese ( 22 ) . \n a smoker was defined as an individual who smoked at least one cigarette per day or had quit smoking during the previous year . \n hypercholesterolaemia was defined as a total serum cholesterol level of 240 mg / dl or more or a serum triglyceride level of 200 mg / dl or more ( or both ) or use of a lipid - lowering agent ( 23 ) . \n individuals were considered as having a positive family history , when they had first - degree or second - degree relatives with premature cardiovascular disease . \n the cardiovascular risk was calculated by using the method of score ( cardiology systematic coronary risk evaluation ) as recommended from esc ( european society of cardiology ) . \n cardiovascular risk charts based on gender , age , total cholesterol , systolic blood pressure , and smoking status have been developed by esc for estimating 10-year - risk of a fatal cve in different european countries ( 24 ) . \n albania is included in the high - risk regions of europe ( 24 , 25 ) . based on these risk charts \n , individuals were classified into high ( score > 10% ) , intermediate to high ( score between 4% and 10% ) and low ( score up to 3% ) risk groups . \n scan protocol and image reconstruction : data acquisition was performed with a 64-slice ct scanner ( somatom sensation , siemens , erlangen , germany ) , including ca score- a prospective low dosage examination without contrast agent and after that ct angiography using a retrospective ecg - gating protocol . the bolus tracking technique ( smartprep ) \n was used to trigger the acquisition , with a four - cavity view as the region of interest . \n a total of 70100 ml of iodinated , non ionic contrast agent ( optiray 350 , mallinckrodt ) was injected into the antecubital vein at a flow rate of 5.0 ml / s , followed by a 50-ml saline flush injected at a flow rate of 5.0 ml / s . \n scanning was initiated during a single breath hold for an acquisition time of 7 - 10 seconds . \n 65% of patients required pre - treatment with beta blockers to lower the heart rate . \n ctac analysis : reconstructed image data were transferred to a remote workstation ( leonardo , siemens ) for post - processing . \n in retrospect , ecg - gating protocol reconstruction of the image data was performed starting from early systole ( 10% of r \n image data sets were reconstructed immediately after the scan , then were analyzed by a specialized radiologist using a dedicated program ( circulation ) including mpr , mip , vrt reconstructions , based on the 16 separate segment model recommended by the american heart association ( 26 ) . \n any cap with a computed tomography attenuation below the contrast - enhanced coronary lumen < 130 hounsfield units ( hu ) , was judged as non - calcified . \n any cap with an attenuation more than 130 hu that could be visualized separately from the contrast coronary lumen was defined as calcified . \n coronary lumen narrowing was graded semi - quantitatively by visual assessment comparing the luminal diameter of the segment exhibiting the obstruction to the luminal diameter of the most normal appearing site immediately proximal to the plaque . \n the threshold for the evaluation of stenosis was 50% ; the atherosclerotic plaques that caused a stenosis up to 50% of coronary lumen diameter were called non stenotic , those with a stenosis of > 50% were classified as stenotic and were divided in two groups : moderate stenosis for atherosclerotic plaques with a stenosis of 51 - 70% and severe stenosis for atherosclerotic plaques causing a stenosis of > 70% . \n all the patients diagnosed with stenotic cad according to the above criteria were referred for coronary angiography . \n the amount of arterial calcification ( ca score ) was also calculated using agatston method where the coronary calcium was defined as a lesion over 130 hu in a 1 mm2(3 pixel ) surface , and according to this result the patients were divided in 4 groups : grade i : ca score of 0 , grade ii : ca score 1 - 10 , grade iii : ca score 11 - 100 , and grade iv : ca score 101 - 400 . the risk for having cad based on agatston score has been evaluated for the following 10 years ( 15 ) . statistical analysis : student t test was used to compare data between genders . \n logistic regression was also used to evaluate association of presence of cap with patients characteristics examined in our study . \n we also calculated odds ratio ( or ) and confidence interval ( ci ) 95% to measure the strength of the association . \n mean age of our study population was 56,7 years old ( range , 48 - 81 years ) . \n 60,5% of the total population had hypertension , 63,6% had hyperlipidemia and 32,5% were smoker , 18.4% were diabetic , 17.7% were obese , and 17.5% had positive family history for cad . \n table 1 shows the relationship between the presence of atherosclerotic plaque and the risk factors . \n all of the risk factors except hyperlipidemia were found to have strong statistical relationship with the development of atherosclerotic plaques . \n cap , coronary atherosclerotic plaque 61.4% of the patients were low risk and 32.9% were at intermediate risk according to score . \n though the presence and severity of cap increased significantly with the increase of score , it was found to be 44.1% in the low risk patients and 80% in the intermediate to high risk group , with a prevalence of 17% and 25% of stenotic plaques ( > 50% ) respectively ( table 2 , figure 1 , figure 2 , figure 3 ) . \n the prevalence dropped to 2% in the low risk group and 7% in the intermediate one when considering severe stenotic lesions ( figure 4 and figure 5 ) . \n furthermore there was no big difference in the presence of cap between intermediate and high risk group with a slight increase of stenotic lesions in the second one ( figure 3 ) . \n also 8.3% of patients with no risk factors ( score 0 ) had cap in ct angiography ( table 2 ) . \n summary of the cases with score , cap positive and stenosis score , systematic coronary risk evaluation ; cap , coronary atherosclerotic plaque distribution of cases of low risk , cap and stenosis according to the score group . \n distribution of cases of intermediate to high risk , cap and stenosis according to the score group . \n distribution of cases of high risk , cap and stenosis according to the score group a 46-year - old male classified as low risk according to score . \n ct angiography multiplanar reconstruction image showing multiple coronary atherosclerotic plaques on the proximal lad causing severe stenosis(arrows ) . \n a , b , c. a 56-year - old male classified in intermediate risk group according to score and as low risk according to ca score . a soft concentric stenotic plaque in mid - lad causing severe subocclusive stenosis ( arrows ) was diagnosed on ct angiography as shown in the multiplanar ( figure 5a ) and volume rendering images ( figure 5b ) . \n the patient was referred for coronary angiography where the lesion was confirmed ( figure 5c ) and treated with stenting . \n this demonstrates that ctac is more sensitive than ca score in early diagnosis of cad . in the cases that ca score \n is 0 , this technique is more reliable . in the cases that ca score has a moderated value , ctac is more sensitive in predicting the grade and severity of the disease . comparing ct angiography of coronaries with ca - score . \n to our knowledge this is the first large study performed with ctac in the albanian population for ruling out cad . \n even though albania is classified in the high risk regions of europe , large statistical data are lacking in this direction ( 24 , 25 ) . \n our study population group was classified mainly at low and intermediate risk to develop fatal cve according to score method . in these individuals , \n moreover , our data showed a clear relation between the presence of cap with an increased predicted risk of fatal cve . \n we had a similar prevalence of cap in the low risk group with a higher presence of stenotic lesions compared to previous studies from the literature(1 , 27 ) . \n also 8.3% of patients with no risk factors ( score 0 ) resulted with cap in ctac . \n these results support once more the theory that a considerable number of cve occur either in the absence of known risk factors or in the presence of low to moderate risk ( figure 4 , figure 5 ) when an aggressive treatment strategy would not be indicated ( 28 ) . \n also 18% of the patients with ca score 0 had cap in ct angiography , one of whom resulted with severe stenotic lesion(figure 5 a - c ) , supporting the theory that a considerable atheroma burden including significant stenoses may be present in patients with no coronary calcification suggested by previous studies(19 , 29 ) . \n the effective dosage of radiation taken by the patient during this examination varies from 6msv to 15msv that is approximately 1.5 - 3 times higher that the dosage of invasive coronarography . \n the new technological developments in this direction promise a considerable reduction of the dosage of radiation up to 5 msv through the different examination phases ( prospective method , ( 30 ) . \n the potential advantages of ctac over other invasive or non - invasive diagnostic procedures for coronary artery disease assessment have to be weighted against the potential hazards associated with long - term radiation risk . \n another limitation of our study is that the diagnosis of cap was based only on ctac , except cases with moderate - severe stenosis which were confirmed by conventional coronary angiography . \n though we must accept that more tailored analysis should have been done , this is an onging study and we would be glad to share our results modestly . \n in conclusion , despite its limitations , our study demonstrates once more that ct angiography is a reliable , very accurate noninvasive technique for the diagnosis of early cad , especially in the low - intermediate risk patients compared to the traditional evaluation schemes and ca score , thus should be considered in this group as a diagnostic guide for optimal therapy planning .", "role": "user" }, { "content": "objective : to evaluate the role of ct angiography of coronaries ( ctac ) in the diagnosis of subclinical atherosclerosis by detection of coronary artery plaques ( cap ) in a group of consecutive albanian individuals with no history of coronary artery disease ( cad ) or acute coronary syndrome and to investigate the relation between the prevalence of cap , traditional risk factors and the expected 10-year risk of fatal cardiovascular event ( cve ) based on our own experience.method and technique : this is a prospective study including 456 patients with no history of cad who underwent ctac in our hospital from september 2009 to march 2013 . risk estimation of fatal cve was assessed using systematic coronary risk evaluation ( score ) and then ct scan was performed with a 64 detector ct , including ca score and angiography of coronaries with iv contrast.results:from 456 patients 61.4% were low risk and 32.9% were at intermediate risk according to score . \n the prevalence of cap diagnosed by ctac was calculated as 55.7 % overall . \n though the presence and severity of cap increased significantly with the increase of score , it was found to be 44.1% in the low risk patients and 80% in the intermediate risk group , with a presence of 17% and 25% of stenotic plaques ( > 50% ) respectively . \n significant correlation was found between all traditional risk factors and cap.conclusion:although a direct relation between the prevalence of cap , risk factors and the related 10-year risk of fatal cve was found , there was a significant prevalence of cap in low intermediate risk group with a considerable presence of stenotic lesions . \n also 8.3% of patients with no risk factors and 18% of the patients with ca score 0 had cap in ct angiography , one resulting with severe stenosis . \n our results suggest once more that ct angiography is a reliable , very accurate noninvasive technique for the diagnosis of early cad , especially in the low - intermediate risk patients compared to the traditional evaluation schemes and ca score , thus should be considered in this group as a diagnostic guide for optimal therapy planning .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: dynamic nuclear cytoplasmic trafficking of macromolecules controls many eukaryotic cellular processes , such as gene expression , signal transduction , cell differentiation , and immune response . \n the karyopherin- family of transport factors recognizes targeting signals within cargo proteins for transport in and out of the nucleus . \n nuclear localization signals direct proteins into the nucleus , and nuclear export signals ( ness ) direct proteins into the cytoplasm ( reviewed in grlich and kutay , 1999 ; chook and blobel , 2001 ; conti and izaurralde , 2001 ; weis , 2003 ; kutay and gttinger , 2005 ; tran et al . , 2007 ; xu et al . , 2010 ) . \n the leucine - rich or classic nes is the only class of nuclear export signal that has been characterized . \n the name leucine - rich nes was coined because the first signals identified in the hiv-1 rev and pki proteins are enriched with leucine residues ( fischer et al . , 1994 ; meyer and malim , 1994 ; \n since then , many more nes - containing proteins have been identified , and mutagenesis and computational analyses have shown the nes sequences to be more diverse and conform to the loose consensus sequence -x2 - 3--x2 - 3--x- , where is l , v , i , f , or m and x is any amino acid ( bogerd et al . , 1996 ; henderson and eleftheriou , 2000 ; engelsma et al . , 2004 \n ; la cour et al . , 2004 ; kutay and gttinger , 2005 ) . \n the nes is recognized by the export karyopherin crm1 , which is also known as exportin 1 ( fornerod et al . , 1997 ; fukuda et al . \n , 1997 ; neville et al . , 1997 ; ossareh - nazari et al . , 1997 ; \n recently published crystal structures of crm1 bound to several ness showed that the signals adopt either combined -helix loop or all - loop structures that bind in a hydrophobic groove on the convex surface of crm1 ( dong et al . , \n leptomycin b ( lmb ) inhibits nuclear export by forming a covalent bond with cys528 of human crm1 , which is located in the nes - binding groove , thus blocking access of the nes to its binding site ( kudo et al . \n , 1999 ; dong et al . , 2009b ; monecke et al . , 2009 ) \n ness have been identified in > 300 proteins with diverse functions , such as transcription factors , cell cycle regulators , ribonucleoprotein complexes , translation factors , and viral proteins ( fischer et al . , 1994 ; wen et al . \n nuclear export of viral proteins by crm1 is important for replication of many viruses that cause human diseases . \n aberrant mislocalization of cellular crm1 cargoes also interrupts numerous cellular processes , often resulting in diseases . \n therefore controlling crm1nes interactions might be a potential therapeutic target for many disease conditions such as cancer and viral infections ( bogerd et al . , 1995 ; yi et al . \n , 2002 ; faustino et al . , 2007 ; noske et al . , \n a database of 80 ness named nesbase 1.0 was compiled in 2003 ( la cour et al . , 2003 ) . \n , we present nesdb , an up - to - date and substantially larger nes database with 221 experimentally identified entries . \n each entry is annotated with many detailed features related to the sequence , structure , and nuclear export activity of the ness and cargo proteins . \n nesdb is a valuable information resource for the biomedical research community to learn about nuclear export signals that have already been identified . \n analysis of the sequences and three - dimensional structures of ness in nesdb and false - positive ness generated from nesdb revealed some distinguishing features that might be important for the future development of accurate nes prediction algorithms ( xu et al . \n both the pubmed and uniprot databases were searched using keywords nuclear export signal , nes , and crm1 ( jain et al . , 2009 ; the uniprot consortium , 2011 ) . \n the returned literature was examined with the following criteria to identify the existence of an experimentally tested nes : 1 ) evidence of crm1-dependent nuclear export , such as binding to crm1 , inhibition by lmb , nuclear retention at nonpermissive temperature in crm1 temperature - sensitive yeast strains , or competition with other crm1 cargoes ; 2 ) the presence of a protein segment that matches the traditional nes consensus sequence -x2 - 3--x2 - 3--x- , which can target a reporter protein for nuclear export ; and 3 ) the presence of mutations within the tested nes segment that abolished nuclear export of the full - length protein . \n all proteins in nesdb meet the first criterion , and many meet all three criteria . \n the published literature is searched on a bimonthly basis and nesdb is updated with every 20 new entries . however , many sequences in the genome , especially those in amphipathic helices , match the nes consensus , thus making accurate nes identification difficult . \n it is likely that some published studies contain mistakenly identified ness . as a caution to the research community \n ness and contains experimentally identified ness with no contradicting experimental evidence . the second group is named \n ness in doubt and contains proteins that were initially reported as ness but with doubts on their validity cast by subsequent experiments . clicking the corresponding link on \n the list can be sorted alphabetically by protein names or numerically by protein i d numbers in nesdb . \n users are able to positively or negatively flag specific nes - containing proteins on their individual pages . \n a tally of flags for each protein is displayed next to its name on the list . \n an entry with many negative flags will be reevaluated and moved to the ness in doubt category or vice versa . \n the database is also equipped with a search button , which searches the full name , alternative names , and organism of proteins for the keywords . \n each entry contains 14 features related to the sequence , structure , and nuclear export activity of the ness and cargo proteins . a sample page for snurportin 1 ( snupn ) \n the nes features include the following : \n full name : the recommended name in uniprot database for the given protein , along with the link to its entry in uniprot ( jain et al . , 2009 ; the uniprot consortium , 2011).alternative names : other names of the protein that are commonly used in the literature.organism : the organism of the listed protein.experimental evidence for crm1-mediated nuclear export : reported experimental evidence for crm1-mediated nuclear export . reports on whether 1 ) the protein binds crm1 , 2 ) \n the protein is retained in the nucleus by lmb , 3 ) nuclear export of the protein is affected by another crm1 cargo such as the hiv - rev protein , or 4 ) crm1 is required for nuclear export in digitonin - permeabilization transport assays . \n all nesdb entries contain experimental evidence for crm1-mediated nuclear export.mutations that affect nuclear export : mutations that have been shown to disrupt nuclear export in cells.mutations that affect crm1 binding : mutations that have been shown to disrupt in vitro crm1 binding.functional export signals : the protein segment that resembles an nes and when fused to a reporter protein can independently target the reporter for nuclear export . \n such data define boundaries and sufficiency of the putative minimal ness.secondary structure of the export signal : the explicitly denoted , experimentally determined secondary structure for the reported nes.other residues important for export : residues known to contact crm1 or shown to affect crm1 binding or nuclear export but are located outside of the nes segment \n . this information may be useful since cargoes may bind crm1 in multipartite manner and contain additional binding epitopes.sequence : sequence of the full - length protein with the functional nes underlined . \n mutations that disrupt nuclear export in cells are highlighted in yellow , mutations that disrupt in vitro crm1 binding are in red , and other , non - nes residues reported to affect crm1 binding or nuclear export are in green . \n there are four tabs associated with the sequence : 1 ) the sequence in fasta format ( pearson and lipman , 1988 ) , 2 ) conserved domains of the protein obtained from the conserved domain database ( marchler - bauer et al . , 2009 ) , \n 3 ) predicted secondary structure of the protein by psipred ( mcguffin et al . , 2000 ) , and 4 ) conservation scores of the protein calculated by al2co ( pei and grishin , 2001).three - dimensional structures : links to three - dimensional structures in the protein data bank ( pdb ) , if available.comments : a short summary of protein functions and experiments related to the identification of its transport signals.references : the literature from which the features were extracted , with links to pubmed.user input : because ness are easily misidentified , a user input field at the bottom of each entry that allows users to positively or negatively flag the nes after submitting supporting comments / rationales . \n full name : the recommended name in uniprot database for the given protein , along with the link to its entry in uniprot ( jain et al . \n alternative names : other names of the protein that are commonly used in the literature . organism : the organism of the listed protein . \n reports on whether 1 ) the protein binds crm1 , 2 ) the protein is retained in the nucleus by lmb , 3 ) nuclear export of the protein is affected by another crm1 cargo such as the hiv - rev protein , or 4 ) crm1 is required for nuclear export in digitonin - permeabilization transport assays . \n mutations that affect nuclear export : mutations that have been shown to disrupt nuclear export in cells . \n mutations that affect crm1 binding : mutations that have been shown to disrupt in vitro crm1 binding . functional export signals : the protein segment that resembles an nes and when fused to a reporter protein can independently target the reporter for nuclear export . \n secondary structure of the export signal : the explicitly denoted , experimentally determined secondary structure for the reported nes . \n other residues important for export : residues known to contact crm1 or shown to affect crm1 binding or nuclear export but are located outside of the nes segment . \n this information may be useful since cargoes may bind crm1 in multipartite manner and contain additional binding epitopes . \n mutations that disrupt nuclear export in cells are highlighted in yellow , mutations that disrupt in vitro crm1 binding are in red , and other , non - nes residues reported to affect crm1 binding or nuclear export are in green . \n there are four tabs associated with the sequence : 1 ) the sequence in fasta format ( pearson and lipman , 1988 ) , 2 ) conserved domains of the protein obtained from the conserved domain database ( marchler - bauer et al . , 2009 ) , 3 ) predicted secondary structure of the protein by psipred ( mcguffin et al . , 2000 ) , and 4 ) conservation scores of the protein calculated by al2co ( pei and grishin , 2001 ) . \n three - dimensional structures : links to three - dimensional structures in the protein data bank ( pdb ) , if available . \n comments : a short summary of protein functions and experiments related to the identification of its transport signals . \n references : the literature from which the features were extracted , with links to pubmed . \n user input : because ness are easily misidentified , a user input field at the bottom of each entry that allows users to positively or negatively flag the nes after submitting supporting comments / rationales . \n a sample page from nesdb that shows 7 of the 14 illustrated features of the nes from snurportin 1 ( snupn ) . \n the features not shown include full name , alternative names , organism , three - dimensional structures , comments , references , and a user input form . \n nesdb will contribute to the understanding of how protein function is controlled by intracellular localization and will serve as a useful resource for the development of inhibitors that target crm1-mediated nuclear export . \n nesdb may be used to train and test new nes prediction algorithms to increase the reliability and accuracy of identifying vague and diverse ness in the genome .", "role": "user" }, { "content": "there are 221 experimentally validated , leucine - rich nuclear export signal ( nes)containing crm1 cargoes in a database named nesdb . \n entries in nesdb are annotated with sequence and structural information on both nes and cargo proteins , as well as with experimental evidence on nes - mapping and crm1-mediated nuclear export .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: type 1 diabetes ( t1 dm ) and obesity are main risk factors for cardiovascular events 1 , 2 . in particular , young adults with t1 dm have an increased risk of early asymptomatic atherosclerosis and consequent cardiovascular morbidity and mortality 3 - 5 . similarly , childhood obesity has been reported associated with biochemical and inflammatory factors that affect vascular endothelial function and that might confer a premature atherogenicity 6 . moreover , the insulin resistance , key feature of obesity , metabolic syndrome and type 2 diabetes , results in an array of metabolic and vascular events which finally promote the development of atherosclerosis 7 . \n the atherosclerotic process starts in childhood and proceeds silently over a long period of time before clinical manifestations 4 . \n carotid artery intima - media thickness ( cimt ) is considered a significant predictive marker of generalized atherosclerosis because of its correlation with coronary artery disease and it may predict future cardiovascular events in adults 4 . \n it is also recommended by the american heart association as a noninvasive imaging parameter for detecting atherosclerosis 8 , 9 . \n previous studies reported an increase of cimt in children with hypercholesterolemia as compared to controls 10 . \n however , data on this marker in obese children and in children and adolescents with t1 dm have yielded conflicting results 11 - 18 . this study aimed to evaluate the presence of early signs of atherosclerosis and \n seek for predisposing factors in a group of t1 dm children and adolescents treated with intensive insulin protocols and in a group of obese subjects . \n we enrolled 71 subjects ( mean age 12.86 2.38 years ) who attended our outpatients unit of pediatric endocrinology . \n out of them , 24 were obese ( 12 males , body mass index -bmi for age and sex > 95 centile ) , 26 subjects had t1 dm ( 12 males , mean hba1c 8.2 % ) , with onset of the disease at the age of 2.9 0.2 years , and 21 ( 11 males , bmi < 85 centile ) were healthy controls recruited from relatives of our medical staff . \n all diabetic patients were on treatment with insulin injections 4 times / day , including three meal doses of insulin lispro and one dose of insulin glargine at bedtime . \n the subjects with clinical , biochemical and instrumental signs of heart disease or family history of hypertension or dyslipidemia were excluded . \n all the procedures used were in accordance with the guidelines of the helsinki declaration on human experimentation . \n all patients underwent a general clinical examination , anthropometric measurements ( height in cm , weight in kg , bmi expressed in kg / m ) using italian growth charts 19 , and assessment of the pubertal and genital stage , according to tanner criteria 20 . both systolic ( sbp ) and diastolic blood pressure ( dbp ) were measured in all patients 21 , while waist circumference was assessed only in obese subjects 22 . \n blood glucose , insulin , total cholesterol ( tc ) , high ( hdl ) and low ( ldl ) density lipoprotein cholesterol , triglycerides ( tg ) and c - reactive protein ( crp ) were measured after overnight fasting in all subjects ; fructosamine and hba1c were assessed in diabetics . \n values of tc , ldl , hdl and tg were considered in the normal range if within the 5 and the 95 percentile 23 . \n an oral glucose ( 1.75 g / kg ) tolerance test ( ogtt ) was performed in obese subjects recording basal levels of blood glucose and insulin and after 120 min . \n b - mode high resolution ultrasonography ( us ) was performed with a philips hdi 5000 sono ct us scanner , equipped with a linear transducer l 12 - 5 , using a frequency of 18 mhz suitable to study vascular walls . \n participants were examined in the supine position with the head turning slightly to both sides . after identifying the bulb , longitudinal images of the common carotid artery and of the two traits that it generates ( internal and external carotid ) \n the far wall was scanned and the resolution box was used to magnify this segment , 10 - 20 mm proximally to the carotid bulb , where were focused all measurements . \n several images were acquired by using an anterior oblique angle ( 30 from midline ) and a lateral one ( 100 from midline ) 24 . \n the cimt of the far wall was measured during end diastole ( r - wave of the electrocardiogram ) . \n three scans on both sides were selected , and nine ( 33 ) measurements of maximum far wall imt were averaged ; thus , the conclusive mean cimt of each patient was calculated from a total of 18 measurements . \n we defined the cimt as thickened when 0.45 mm in children younger than 10 years or if it was 0.55 mm in children older than 10 years and younger than 18 years 25 - 28 . \n statistical analysis was performed using spss software for windows ( version 15 , chicago , il , usa ) . \n comparisons between groups or within the same group were made using the kruskal - wallis and the wilcoxon - mann - whitney u non parametric tests , respectively . \n spearman rho correlation analysis and multiple regression models were used to include cardiovascular risk factors . \n statistical analysis was performed using spss software for windows ( version 15 , chicago , il , usa ) . \n comparisons between groups or within the same group were made using the kruskal - wallis and the wilcoxon - mann - whitney u non parametric tests , respectively . \n spearman rho correlation analysis and multiple regression models were used to include cardiovascular risk factors . \n age , height and stage of pubertal development did not differ significantly within the three studied groups , while , as expected , obese subjects had increased weight and bmi values compared to t1 dm patients and controls . \n sbp and dbp were higher in obese and diabetics than in healthy subjects ( p<0.015 and p<0.002 , respectively ) . \n a positive correlation between sbp and bmi was found in obese individuals ( rho=0.26 ; p<0.02 ) ( data not shown ) . among t1 dm patients , 77% of them had blood glucose concentration greater than 180 mg / dl , 54% had hba1c greater or equal to 8% , and 73% had fructosamine levels greater than 300 \n mol / l ; tc , hdl , ldl cholesterol , tg and crp values were in the normal range . \n the levels of ldl cholesterol and tg were increased in 4.2% and 12.5% of the obese subjects , respectively . \n their tg values were significantly higher compared to both diabetics and controls ( p<0.001 ) . \n all obese patients had normal glucose tolerance with basal hyperinsulinemia ( 32.2 14.2 u / ml ) and after glucose loading ( 141.2 56.54 u / ml ) . \n cimt values resulted higher in obese subjects than in diabetics ( p<0.05 ) , and in both these groups compared to controls ( p<0.007 and p<0.02 , respectively ) ( fig.1 ) . \n diabetic subjects showed a positive correlation between cimt of the three studied segments and fasting blood glucose levels ( rho=0.47 , p<0.05 ) . \n cimt tended also to be negatively correlated with the units / kg / day of insulin administered in the same group of subjects ( rho=-0.29 , p=0.05 ) . \n the mean cimt values were also significantly higher among diabetic boys than age - matched healthy control subjects ( 0.51 0.05 vs 0.49 0.05 ; p<0.02 ) . \n we found a positive correlation between insulin levels ( basal and after oral glucose load ) and cimt of common , internal and external carotid artery in the obese ( rho=0.47 , p<0.03 ) . \n furthermore , the obese boys had higher mean cimt value than girls ( 0.52 0.03 vs 0.51 0.04 ; p<0.05 ) and the mean cimt was significantly higher among obese boys than age - matched healthy control subjects ( 0.52 0.03 vs 0.46 0.05 ; p<0.01 ) , but not among obese girls . \n cimt values were also higher in subjects during puberty than at the prepubertal age ( t1 dm vs obese vs controls : 0.51 0.05 vs 0.52 0.04 vs 0.46 0.05 ; p<0.005 ) . \n the overall measurement of cimt was also significantly different within studied groups stratifying the analysis by height : individuals less than 150 cm of height had higher values of c - imt , compared to controls , than taller subjects ( 0.53 0.03 vs 0.52 0.04 vs 0.47 0.05 ; p<0.04 ) . \n figure 2 shows the distribution of c - imt values within the study groups in pubertal age males with height less than 150 cm ( 0.52 0.02 vs 0.51 0.05 vs 0.46 0.03 ; p<0.02 ) . the multiple regression analysis performed in the obese group , selecting sbp , dbp , tc , hdl and tg as independent variables and cimt as the dependent variable , showed that hdl cholesterol represented a protective factor for the thickening of cimt ( or=0.58 with a 95% confidence interval of 0.31 - 0.73 , p=0.04 ) , while the other analyzed parameters did not seem to affect cimt in this group of subjects \n in this study we identified subclinical atherosclerosis in t1 dm patients and obese subjects as indicated by their significantly higher cimt ( mainly in boys ) compared to that of healthy controls . \n our results suggest that the common pathogenetic factor involved in endothelial damage in obese and t1 dm children and adolescents seems to be a reduced insulin function ( insulin deficiency in diabetics and insulin resistance in obese patients , respectively ) . \n insulin acts by modulating the release of vasodilator substances , such as nitric oxide and prostaglandins , from vascular endothelium , by both stimulating and inhibiting the sympathetic nervous system and by protecting smooth muscle cells in blood vessel from apoptosis induced by oxidative stress 29 . \n thus the vasodilatory and antioxidant effects of insulin are depressed in case of insulin deficiency ( i.e. type 1 diabetes ) and insulin resistance ( i.e. obesity ) conditions . \n it is well documented that cimt correlates with coronary atherosclerosis 4 , and that it represents an independent predictor of future cardiovascular events 9 . \n it also well correlates with parameters of invasive examination tools , such as coronary angiography and intravascular ultrasound 4 , 30 . \n studies on increased cimt in children and adolescents affected with obesity , t1 dm , dyslipidemia , hypertension and chronic renal failure , compared to healthy controls , have yielded conflicting results 31 , 32 . \n the different conclusions may be explained by the non - homogenous populations studied and by the variety of ultrasound methods used . \n although in our study obese individuals showed only slightly elevated mean cimt compared to subjects with t1 dm , a significant high number of diabetic and obese patients had cimt above the normal range , indicating that these patients showed early signs of atherosclerosis development compared with healthy controls . \n the degree and duration of hyperglycemia has been associated with macrovascular complications and increased cardiovascular risk in adults and preadolescent children with t1 dm 5 , 33 - 36 . in the present study no association between cimt and hba1c at the time of visit ( a marker of glycemic control ) was observed in diabetic subjects . \n however , the positive correlation between cimt ( especially of the left common carotid intima ) and fasting blood glucose suggests that accumulation of glycation end products , as a consequence of not optimal insulin therapy , increases oxidative stress and the subsequent cell damage . \n this hypothesis is also confirmed by the negative correlation , even if not statistically significant , between the values of cimt and the units of insulin daily administered . in obese individuals , the positive correlation between basal and after oral glucose load insulin levels and \n cimt shows that insulin resistance causes an imbalance of vascular homeostasis mechanisms responsible for the endothelial damage and dysfunction 29 , 37 . \n previous studies have shown that cimt is negatively correlated with hdl cholesterol levels and positively related with ldl levels , confirming that dyslipidemia has a prominent role in the pathogenesis of endothelial dysfunction with progression to atherosclerosis 6 , 38 , 39 . in line with data reported in literature , \n hdl cholesterol values represent a protective factor for cimt in our obese population . on the other hand , although increased cimt has been documented in children with daytime hypertension , as well as in those with nocturnal hypertension and t1 dm 25 , 40 , in our study population no association between conventional cardiovascular risk factors , such as blood pressure or ldl cholesterol and cimt was found . \n this could be explained by the short duration of the disease at the time of examination , and the young age of the study population . \n the gender - related difference in cimt measurements observed in our study ( higher prevalence of atherosclerosis and cardiovascular disease among males compared to females ) is in line with data reported in literature on the general population of children and adolescents 33 , 34 . \n furthermore , our obese subjects showed a higher value of cimt during puberty than that recorded in those at the prepubertal age . \n this could be explained by the change of insulin sensitivity for increased activity of the growth hormone axis ( gh)-igf1 , which worsens the condition of insulin resistance during puberty . \n furthermore , recently skilton and colleagues have positively correlated cimt with height in non - diabetic children aged 8-years 41 , while in this study we have observed that diabetic children with high risk for increased cimt are likely males at pubertal age less than 150 cm tall ( figure 2 ) . \n additional study in selected groups of children should be carried - out to get more conclusive results . in conclusion \n , we found that diabetic and obese children present ( as adults ) an endothelial damage , confirmed by increased cimt values . \n our results point out the importance of identifying children with the highest risk for cardiovascular heart disease using noninvasive studies , even if more studies on wider populations are warranted to confirm the findings . \n we also observed that there is a positive correlation between high levels of basal and after oral loading glucose insulin levels ( a sign of insulin resistance ) and increased cimt in obese , as well as between high fasting blood glucose levels ( a sign of relative insulin deficiency ) and cimt in t1 dm patients . \n thus it is necessary to normalize blood glucose values by optimizing insulin therapy in diabetic subjects , and to reduce hyperinsulinism by weight loss and physical activity in obese subjects , in order to prevent future cardiovascular risks .", "role": "user" }, { "content": "background . increased carotid intima - media thickness ( cimt ) is considered a marker of early - onset atherosclerosis and it seems to predict cardiovascular events both in obese and diabetic subjects . \n we aimed to evaluate early signs of atherosclerosis and investigate for predisposing factors in children and adolescents affected by type 1 diabetes ( t1 dm ) or obesity , comparing them with healthy controls.methods . \n out of 71 enrolled subjects ( mean age 12.8 2.3 years ) , 26 had t1 dm and 24 were obese , while 21 age- and sex - matched subjects acted as controls . \n cimt was measured using standardized methods . \n serum glucose , insulin , cholesterol , triglycerides and c - reactive protein levels were evaluated . \n an oral glucose tolerance test ( ogtt ) was performed in obese subjects.results . \n diabetic and obese individuals showed higher cimt mean values than healthy controls ( p<0.005 ) . \n cimt of the three examined segments correlated positively with fasting glucose levels and negatively with units of insulin / kg / day administered in t1 dm individuals . a positive correlation between insulin levels ( basal and after oral glucose load ) and cimt of common , internal and external carotid artery was found in obese subjects ( p<0.03 ) . \n high density cholesterol levels represented a protective factor for cimt in this latter group of the study population.conclusions . \n our findings show that cimt correlates with high insulin levels ( a sign of insulin resistance ) in obese patients and with high fasting glucose levels ( a sign of relative insulin deficiency ) in t1 dm subjects , confirming the need of reducing hyperinsulinism and monitoring blood glucose levels in these subjects to prevent atherosclerosis .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: a 58-year - old male presented with a lesion on the medial aspect of left lower eyelid near the punctum . \n he had a small elevated nodular swelling at the margin of the eyelid for the past two decades . \n there was a gradual increase in the size of the swelling for past 4 years , however , during past 6 months it became more visible associated with discomfort , itching , and a noticeable mass . \n physical examination revealed a solitary 2 cm 2 cm size brownish firm fleshy growth with hyperkeratotic surface covered with crusts in some areas , involving the skin of the left lower eyelid [ fig . 1 ] . \n it did not involve the underlying deeper tissues also and there was no regional lymphadenopathy . \n fleshy growth with hyperkeratotic surface covered with crusts in some areas , involving the skin of the left lower eyelid the mass was excised along with a 3 mm margin of healthy skin , and the resultant skin defect was repaired with a median frontoglabellar pedicle skin flap . \n the final pathologic diagnosis of excision biopsy specimen confirmed syringocystadenoma papilliferum . on gross examination , skin covered tissue measuring 1.3 cm 1.1 cm 1 cm . \n this showed a nodular lesion 0.7 cm in its greatest axis 0.3 cm away from the nearest peripheral surgical cut margins . on microscopic examination \n , section showed a nodular lesion lined by epidermis with varying degree of papillomatosis [ fig . 2 ] . \n these were lined by two rows of cells . the luminal row consisted of columnar cells with evidence of active decapitation secretion and outer row of cells composed of small cuboidal cells . \n on gross examination , skin covered tissue measuring 1.3 cm 1.1 cm 1 cm . \n this showed a nodular lesion 0.7 cm in its greatest axis 0.3 cm away from the nearest peripheral surgical cut margins . on microscopic examination \n , section showed a nodular lesion lined by epidermis with varying degree of papillomatosis [ fig . 2 ] . \n these were lined by two rows of cells . the luminal row consisted of columnar cells with evidence of active decapitation secretion and outer row of cells composed of small cuboidal cells . \n syringocystadenoma papilliferum is a rare benign tumor that is believed to be derived from the apocrine or the eccrine sweat glands . in a recent report \n , immunohistochemistry proved the apocrine nature of the benign , noncystic lesion by virtue of its nuclear androgen receptor and cytoplasmic gross - cystic disease fluid protein-15 positivity , along with its smooth muscle actin - positive myoepithelial layer . \n this is predominantly a childhood tumor ; however presentation at a later stage like our case has been described in literature . in half of those who are affected , it is present at birth , and in a further 1530% , the tumor develops before puberty . \n the lesion is frequently misdiagnosed as basal cell carcinoma or cyst or squamous cell carcinoma . in a series of 14 patients , \n this is unlike other benign eyelid lesions , where histopathological diagnosis confirms clinical diagnosis in 95.9% cases . \n tumor is usually described as a skin colored to pink , hairless , firm plaque of grouped nodules or as a solitary nodule . \n cauliflower like , verrucous , papillary , hyperkeratotic , or sometimes moist fleshy excrescences have also been described . \n most of the lesions develop and enlarge slowly , although a few can increase to significantly within a short period . \n the plaque type that presents a hairless area of the scalp is commonly associated with a sebaceous nevus of jadassohn . in about one - third of the case , \n appearance of the lesion in the face and neck region is seen in the linear type ; however , a solitary nodular type shows predilection for the trunk . \n syringocystadenocarcinoma papilliferum is a malignant counterpart of syringocystadenoma papilliferum . the diagnosis is clinically suspected and histologically confirmed . \n ulceration or a rapid enlargement of an existing tumor is indicative of a malignant transformation . \n we were suspicious in the index case because of the recent increase of size . in about one - tenth of cases of syringocystadenoma papilliferum \n squmaous cell carcinoma may also develop , but much less frequently . because of this , surgical excision is the treatment of choice . in our case , \n excision of the tumor was done followed by repair of skin defect with median frontoglabellar pedicle skin flap . \n \n ", "role": "user" }, { "content": "syringocystadenoma papilliferum can rarely affect eyelid skin . \n the lesion is frequently misdiagnosed as basal cell carcinoma or cyst or squamous cell carcinoma . we are presenting a case that was clinically diagnosed as basal cell carcinoma of eyelid but was later histologically diagnosed as syringocystadenoma papilliferum .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: fetal bovine serum was purchased from hyclone ( logan , utah ) , lipofectamine 2000 from invitrogen ( carlsbad , ca ) , tripure isolation reagent from roche molecular biochemicals ( mannheim , germany ) , gapdh ( glyceraldehyde phosphate dehydrogenase ) antibody from abmart ( shanghai , china ) , and phospho - stat3 ( tyr705 ) ( d3a7 ) xp rabbit monoclonal antibody . \n phospho - p44/42 mitogen - activated protein kinase ( mapk ) ( erk1/2 ) ( thr202/tyr204 ) ( 197g2 ) rabbit monoclonal antibody were all purchased from cell signaling technology ( beverly , ma ) . \n primer synthesis was performed by sangon biological engineering technology & service co ( shanghai , china ) . \n the cell cycle detection kit , cell lysis solution , and cell counting kit 8 ( cck-8 ) were from beyotime ( shanghai , china ) . \n dna marker , t4 dna ligase , and taq enzyme were all from takara ( dalian , china ) . \n rabbit immunoglobulin g antibody was purchased from abgent ( san diego , ca ) , dna rapid purification kit was from omega bio - tek ( norcross , ga ) . \n the primers were designed based on the coding regions of the arhi gene as follows : sense , 5-ccggaattcatgggtaacgccagctt-3 ; antisense , 5-cgcggatcctcacatgattatgcacttgt-3 ( the underlined gaattc and ggatcc sequences are the ecori and bamhi recognition sites , respectively ) . \n polymerase chain reaction ( pcr ) was performed using pcdna3.0-arhi as the template ( kindly donated by dr d. b. badgwell , at the university of texas m. d. anderson cancer center , houston , tx ) . \n the pcr products were separated by electrophoresis on a 1.5% agarose gel , followed by ethidium bromide staining . \n the fragments of interest were recovered from the agarose gel , purified , and ligated by t4 dna ligase to express the pires2-egfp - arhi plasmid . \n the positive recombinant products were selected on lb agar plates using 100 g / ml ampicillin and confirmed by pcr and dna sequencing . \n skov3 cells were purchased from shanghai institute of cell biology , chinese academy of sciences ( shanghai , china ) and were maintained in rpmi 1640 media supplemented with 10% fetal bovine serum , penicillin 100 u / ml , and streptomycin 100 u / ml at 37c in a humidified 5% co2 atmosphere . the cells were seeded in 6-well plates at a density of 34 10 cells per well in 2 ml of antibiotic - free medium and incubated for 1 day before transfection at 90% to 95% confluence . \n transfection was performed by first diluting 2 l plasmid and 5 l lipofectamine 2000 per well in serum - free medium to a final volume of 250 l , gently mixing , and incubating at room temperature for 5 min . \n this was followed by further gentle mixing and incubation at room temperature for 20 minutes . \n the cells were washed with serum - free medium before the diluted plasmid / lipofectamine complex was added to the 6-well plates for 4 to 6 hours , after which the complex was replaced with normal cell culture medium . \n the cells were visualized 24 hours after transfection under a nikon eclipse fluorescence microscope ( nikon , japan ) . \n the skov3 cells were divided into the following 4 groups : cells transfected with pires2-egfp - arhi plasmid ( treated group ) , cells transfected with pires2-egfp plasmid ( positive control group ) , cells without transfection ( negative control group ) , and a blank control without cells . \n cells were passaged at the logarithmic growth phase , trypsinized , and adjusted to 5 10 cells / ml . the cells were then seeded in 96-well plates in 100 l 10% fetal bovine serum and cultured for 24 , 48 , 72 , 96 , and 120 hours at 37c in a humidified 5% co2 atmosphere . \n after incubation for 4 hours under the same condition , 10 l cck-8 reagent was added to each well . \n the optical density value of each well was measured using a microculture plate reader at a wavelength of 450 nm . \n the growth inhibitory rate ( ir ) was calculated using the following formula : the skov3 cells were divided into the following 3 groups : pires2-egfp - arh transfected group ( treated ) , pires2-egfp transfected group ( positive control ) , and cells without transfection ( negative control ) . \n the cells were harvested after 48 and 72 hours , and cell apoptosis was detected using cell cycle and apoptosis analysis kits ( haimen , china ) , following the manufacturer s instructions . \n aliquots of cell lysates containing 50 g of proteins were separated by 10% sodium dodecyl sulfate \n the membranes were blocked with tbst ( tris - buffered saline and tween 20 ) buffer ( 10 mm tris - hcl , ph 8.0 , 0.15 m nacl , 0.05% tween 20 ) containing 5% skimmed milk ; incubated with rabbit polyclonal antibodies to phospho - stat3 , p44/42 mapk ( erk1/2 [ extracellular signal regulated protein kinase 1/2 ] ) , or phospho - p44/42 mapk ( erk1/2 ) at 4c overnight ; and followed by the addition of horseradish peroxidase linked anti \n rabbit immunoglobulin g. the bands were visualized by electrochemiluminescence , and the intensity of each western blot band was analyzed using quantity one software ( biorad ) . \n comparisons of growth irs and apoptotic rates were performed by test , and protein relative expression levels were compared by student t test . \n fetal bovine serum was purchased from hyclone ( logan , utah ) , lipofectamine 2000 from invitrogen ( carlsbad , ca ) , tripure isolation reagent from roche molecular biochemicals ( mannheim , germany ) , gapdh ( glyceraldehyde phosphate dehydrogenase ) antibody from abmart ( shanghai , china ) , and phospho - stat3 ( tyr705 ) ( d3a7 ) xp rabbit monoclonal antibody . \n phospho - p44/42 mitogen - activated protein kinase ( mapk ) ( erk1/2 ) ( thr202/tyr204 ) ( 197g2 ) rabbit monoclonal antibody were all purchased from cell signaling technology ( beverly , ma ) . \n primer synthesis was performed by sangon biological engineering technology & service co ( shanghai , china ) . \n the cell cycle detection kit , cell lysis solution , and cell counting kit 8 ( cck-8 ) were from beyotime ( shanghai , china ) . \n dna marker , t4 dna ligase , and taq enzyme were all from takara ( dalian , china ) . \n rabbit immunoglobulin g antibody was purchased from abgent ( san diego , ca ) , dna rapid purification kit was from omega bio - tek ( norcross , ga ) . \n the primers were designed based on the coding regions of the arhi gene as follows : sense , 5-ccggaattcatgggtaacgccagctt-3 ; antisense , 5-cgcggatcctcacatgattatgcacttgt-3 ( the underlined gaattc and ggatcc sequences are the ecori and bamhi recognition sites , respectively ) . \n polymerase chain reaction ( pcr ) was performed using pcdna3.0-arhi as the template ( kindly donated by dr d. b. badgwell , at the university of texas m. d. anderson cancer center , houston , tx ) . \n the pcr products were separated by electrophoresis on a 1.5% agarose gel , followed by ethidium bromide staining . \n the fragments of interest were recovered from the agarose gel , purified , and ligated by t4 dna ligase to express the pires2-egfp - arhi plasmid . \n the positive recombinant products were selected on lb agar plates using 100 g / ml ampicillin and confirmed by pcr and dna sequencing . \n skov3 cells were purchased from shanghai institute of cell biology , chinese academy of sciences ( shanghai , china ) and were maintained in rpmi 1640 media supplemented with 10% fetal bovine serum , penicillin 100 u / ml , and streptomycin 100 u / ml at 37c in a humidified 5% co2 atmosphere . the cells were seeded in 6-well plates at a density of 34 10 cells per well in 2 ml of antibiotic - free medium and incubated for 1 day before transfection at 90% to 95% confluence . \n transfection was performed by first diluting 2 l plasmid and 5 l lipofectamine 2000 per well in serum - free medium to a final volume of 250 l , gently mixing , and incubating at room temperature for 5 min . \n this was followed by further gentle mixing and incubation at room temperature for 20 minutes . \n the cells were washed with serum - free medium before the diluted plasmid / lipofectamine complex was added to the 6-well plates for 4 to 6 hours , after which the complex was replaced with normal cell culture medium . \n the cells were visualized 24 hours after transfection under a nikon eclipse fluorescence microscope ( nikon , japan ) . \n the skov3 cells were divided into the following 4 groups : cells transfected with pires2-egfp - arhi plasmid ( treated group ) , cells transfected with pires2-egfp plasmid ( positive control group ) , cells without transfection ( negative control group ) , and a blank control without cells . \n cells were passaged at the logarithmic growth phase , trypsinized , and adjusted to 5 10 cells / ml . the cells were then seeded in 96-well plates in 100 l 10% fetal bovine serum and cultured for 24 , 48 , 72 , 96 , and 120 hours at 37c in a humidified 5% co2 atmosphere . \n each experimental group included 6 wells , and all experiments were repeated in triplicate . after incubation for 4 hours under the same condition , 10 l cck-8 reagent was added to each well . \n the optical density value of each well was measured using a microculture plate reader at a wavelength of 450 nm . \n the skov3 cells were divided into the following 3 groups : pires2-egfp - arh transfected group ( treated ) , pires2-egfp transfected group ( positive control ) , and cells without transfection ( negative control ) . \n the cells were harvested after 48 and 72 hours , and cell apoptosis was detected using cell cycle and apoptosis analysis kits ( haimen , china ) , following the manufacturer s instructions . \n aliquots of cell lysates containing 50 g of proteins were separated by 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis and transferred to polyvinylidene fluoride membranes . \n the membranes were blocked with tbst ( tris - buffered saline and tween 20 ) buffer ( 10 mm tris - hcl , ph 8.0 , 0.15 m nacl , 0.05% tween 20 ) containing 5% skimmed milk ; incubated with rabbit polyclonal antibodies to phospho - stat3 , p44/42 mapk ( erk1/2 [ extracellular signal regulated protein kinase 1/2 ] ) , or phospho - p44/42 mapk ( erk1/2 ) at 4c overnight ; and followed by the addition of horseradish peroxidase linked anti \n rabbit immunoglobulin g. the bands were visualized by electrochemiluminescence , and the intensity of each western blot band was analyzed using quantity one software ( biorad ) . \n spss version 13.0 software was used for statistical analyses . comparisons of growth irs and apoptotic rates were performed by test , and protein relative expression levels were compared by student t test . \n 1a - c ) showed a high level of green fluorescence protein ( gfp ) in pires2-egfp - arhi - skov3 cells ( fig . \n the high level of gfp in the treated group compared with the control groups confirms successful transfection . \n green fluorescence protein expression 48 hours after transfection : ( a ) pires2-egfp - arhi - skov3treated group ; ( b ) pires2-egfp - skov3 plasmid control group ; ( c ) untransfected skov3 cells negative control group . \n the high level of gfp in the treated group compared with the control groups confirms successful transfection . \n after the cells had been cultured for 24 , 48 , 72 , 96 , and 120 hours , the growth irs were 64.69% , 70.17% , 67.01% , 66.87% , and 67.70% , respectively , in the test group , and were 27.20% , 31.10% , 29.80% , 30.73% , and 31.28% , respectively , in the plasmid control group ( fig . \n this showed that the growth ir was significantly increased in the test group compared with the plasmid control group ( p < 0.01 ) . \n the effect of arhi on cell proliferation in skov3 , measured by cck-8 assays , shows that arhi significantly inhibits growth of skov3 cells . \n the proportions of s - phase cells and apoptosis rates in skov3 cells were determined by flow cytometry . \n after 48 hours of culture , the mean proportions of s - phase skov3 cells in the test , plasmid control , and negative control groups were 64.18% , 38.43% , and 15.15% , respectively , and the mean apoptosis rates were 47.97% , 26.53% , and 9.33% , respectively . \n after 72 hours of culture , the mean proportions of s - phase skov3 cells in the test , plasmid control , and negative control groups were 43.29% , 10.37% , and 10.89% , respectively , and the apoptosis rates were 51.34% , 24.70% , and 4.39% , respectively ( table 1 ) . \n data show that the proportions of s - phase cells , and thereby s - phase arrest , were clearly higher in the pires2-egfp - arhi - skov3 cell group compared with the control group at both 48 and 72 hours ; the result at 72 hours was much more significant in the test group . \n in addition , the apoptosis rate was significantly increased in the test group compared with the control group ( p < 0.01 ) . \n influence of arhi on the cell cycle phase distribution and apoptosis rate the western blot images for p - stat3 ( phosphorylated signal transduction and activators of transcription 3 ) and p - erk protein expression for the control and treated groups : skov3 , pires2-egfp - skov3 , and pires2-egfp - arhi - skov3 , are shown in figure 3 . \n compared with gapdh reference protein , the relative expression levels of p - stat3 were 1.1473 0.0002 , 1.0913 0.0021 , and 0.7424 0.0006 , respectively , and the relative expression levels of p - erk were 1.2260 0.0011 , 1.1289 0.0018 , and 0.5866 0.0013 , respectively . in comparison to the 2 control groups , the expression levels of p - stat3 and p - erk proteins were significantly lower in the pires2-egfp - arhi - skov3 group ( p < 0.05 ) . \n changes in the expressions levels of p - erk1/2 and p - stat3 in the different skov3 groups : lane 1 , protein expression in the untransfected skov3 control group ; lane 2 , protein expression 48 hours after pires2-egfp was transfected into skov3 cells ; lane 3 , protein expression 48 hours after pires2-egfp - arhi was transfected into skov3 cells . \n the results show that arhi significantly reduces p - erk1/2 and p - stat3 expression levels . \n \n 1a - c ) showed a high level of green fluorescence protein ( gfp ) in pires2-egfp - arhi - skov3 cells ( fig . \n the high level of gfp in the treated group compared with the control groups confirms successful transfection . \n green fluorescence protein expression 48 hours after transfection : ( a ) pires2-egfp - arhi - skov3treated group ; ( b ) pires2-egfp - skov3 plasmid control group ; ( c ) untransfected skov3 cells negative control group . \n the high level of gfp in the treated group compared with the control groups confirms successful transfection . \n after the cells had been cultured for 24 , 48 , 72 , 96 , and 120 hours , the growth irs were 64.69% , 70.17% , 67.01% , 66.87% , and 67.70% , respectively , in the test group , and were 27.20% , 31.10% , 29.80% , 30.73% , and 31.28% , respectively , in the plasmid control group ( fig . \n this showed that the growth ir was significantly increased in the test group compared with the plasmid control group ( p < 0.01 ) . \n the effect of arhi on cell proliferation in skov3 , measured by cck-8 assays , shows that arhi significantly inhibits growth of skov3 cells . \n the proportions of s - phase cells and apoptosis rates in skov3 cells were determined by flow cytometry . after 48 hours of culture , the mean proportions of s - phase skov3 cells in the test , plasmid control , and negative control groups were 64.18% , 38.43% , and 15.15% , respectively , and the mean apoptosis rates were 47.97% , 26.53% , and 9.33% , respectively . \n after 72 hours of culture , the mean proportions of s - phase skov3 cells in the test , plasmid control , and negative control groups were 43.29% , 10.37% , and 10.89% , respectively , and the apoptosis rates were 51.34% , 24.70% , and 4.39% , respectively ( table 1 ) . \n data show that the proportions of s - phase cells , and thereby s - phase arrest , were clearly higher in the pires2-egfp - arhi - skov3 cell group compared with the control group at both 48 and 72 hours ; the result at 72 hours was much more significant in the test group . \n in addition , the apoptosis rate was significantly increased in the test group compared with the control group ( p < 0.01 ) . \n the western blot images for p - stat3 ( phosphorylated signal transduction and activators of transcription 3 ) and p - erk protein expression for the control and treated groups : skov3 , pires2-egfp - skov3 , and pires2-egfp - arhi - skov3 , are shown in figure 3 . \n compared with gapdh reference protein , the relative expression levels of p - stat3 were 1.1473 0.0002 , 1.0913 0.0021 , and 0.7424 0.0006 , respectively , and the relative expression levels of p - erk were 1.2260 0.0011 , 1.1289 0.0018 , and 0.5866 0.0013 , respectively . in comparison to the 2 control groups , \n the expression levels of p - stat3 and p - erk proteins were significantly lower in the pires2-egfp - arhi - skov3 group ( p < 0.05 ) . \n changes in the expressions levels of p - erk1/2 and p - stat3 in the different skov3 groups : lane 1 , protein expression in the untransfected skov3 control group ; lane 2 , protein expression 48 hours after pires2-egfp was transfected into skov3 cells ; lane 3 , protein expression 48 hours after pires2-egfp - arhi was transfected into skov3 cells . \n the results show that arhi significantly reduces p - erk1/2 and p - stat3 expression levels . \n cell cycle progression is regulated by multiple control points at different phases of the cell cycle ; the 3 principal ones being g1/s , g2/m , and at metaphase / anaphase transmission during mitosis . \n the g1/s check point is the most critical for control of cell proliferation via intracellular and extracellular signals related to transportation and integration of molecules into the nucleus.14 by reconstructing the pires2-egfp - arhi plasmid and successfully transfecting it into hoc skov3 cells with low arhi expression levels , as shown by high levels of green fluorescence from the plasmid observed by fluorescent microscopy , in addition , by using a cck-8 assay to analyze the growth irs , we observed significantly higher growth inhibition in pires2-egfp - arhi - skov3 cells compared with the control cells ; furthermore , flow cytometry showed the proportion of s - phase cells and the apoptosis rates were significantly higher after promoting expression of the arhi gene . \n these results suggested that arhi inhibited proliferation , arrested the cells at s - phase of the cell cycle , and induced apoptosis in skov3 cells . \n we therefore proposed that the arhi gene is involved in cell cycle regulation in hoc skov3 cells , preventing proliferation and inducing apoptosis . \n multiple signaling pathways are involved in cell growth and apoptosis , including the ras / raf / mek / erk1/2 signaling pathway , which is central for transporting the complex signals received by the cell surface receptors to transcription factors in the nucleus . \n extracellular signal regulated protein kinase 1/2 is a key physiological substrate of mek ; mek1 kinase can cause double phosphorylation of threonine and tyrosine at erk1/2 residues to activate erk1/2 ; activation of erk1/2 can subsequently lead to phosphorylation of downstream transcription factors , such as p90rsk kinase , creb , and c - myc . \n changes in cell proliferation , apoptosis , differentiation , and other critical physiological activities are a consequence of phosphorylation of these transcription factors.15 meloche and pouyssgur16 proposed that erk1/2 , which is a key regulatory factor in mitosis , is also involved in the evolutionary control of the cell cycle . \n a certain study showed that arhi plays roles in the egf - egfr - ras - raf - mapk / erk1/2 pathway.17 extracellular signals activate jak in the jak - stat signaling pathway through associated receptors , leading to phosphorylation of specific tyrosine residues on the receptors ; these phosphorylate stats , which are isolated from the receptors , transferred into the nucleus after dimerization , and combined with specific dna response elements . \n this initiates transcription at the target genes and promoting synthesis of a variety of proteins and their subsequent biological roles.18 studies have shown that inhibiting the jak - stat signaling pathway can significantly reduce p - jak2 , p - jak3 , and p - stat3 expression levels ; down - regulate cyclin - d1 , which is linked to cell cycle regulation ; and up - regulate p16 , p21 , and p27 expression , thereby inhibiting tumor cell proliferation and growth.19,20 stat3 , an important member of the stat family , is found in a variety of tumor tissues and cell lines and is now recognized as an oncogene.21,22 stat3 sirna down - regulates cyclin d1 , survivin , and vascular endothelial growth factor expression in hoc cells ; inhibits stat3 and its related genes ; inhibits hoc cell growth ; and induces apoptosis in vitro . \n as such , stat3 has recently become a target for hoc therapies.23 when the yeast 2-hybrid system was used to search for arhi - interactive proteins by screening the human mammary epithelial cell cdna gene pool , arhi was found to act with signal transduction proteins and the stat3 transcription activator , but not stat1 or stat5a . \n furthermore , when arhi and stat3 were coexpressed in skov3 cells , they did not form a complex in the cytoplasm . \n this suggests that by regulating stat3 phosphorylation , arhi can significantly reduce stat3 binding to dna , and thereby reduce the activity of the stat3-dependent promoter.24 our data showed that the expression levels of p - erk1/2 and p - stat3 proteins in the ras / raf / mek / erk1/2 and jak - stat3 signaling pathways were significantly reduced 48 hours after arhi transfection . in our study \n previously , we also measured the expression levels of stat3 and erk1/2 and did not find statistically significant . \n so we proposed that the mechanism by which arhi influences both the ras / raf / mek / erk1/2 and jak - stat3 signaling pathways to inhibit hoc skov3 cell proliferation is through regulating arhi gene expression by altering the phosphorylation levels of erk1/2 and stat3 protein kinases , thereby blocking the signaling pathway for tumor cell proliferation sufficiently to prevent cell proliferation and induce apoptosis . \n a study by lu et al25 revealed that the arhi gene maintains hoc cells in transplanted tumors in a resting state ; conversely , when the arhi level was decreased , the transplanted tumor grew rapidly , and when arhi expression subsequently recovered to normal levels , the hoc cells died through autophagy . \n the authors concluded that the arhi gene is a key factor in initiating autophagy in hoc cells ; a further study by mathias et al26 proposed that this may lead to cell death . \n an in - depth investigation is required to determine the molecular mechanism for autophagy initiation in the arhi gene , with the aim of developing an application based on arhi for the clinical treatment of hoc .", "role": "user" }, { "content": "objectiveaplasia ras homolog member i ( arhi ) is associated with human ovarian cancer ( hoc ) growth and proliferation ; however , the mechanisms are unclear . \n the purpose of this study was to investigate arhi effects in hoc skov3 cells.methodswe transfected skov3 cells with pires2-egfp - arhi and measured growth inhibition rates , cell cycle distribution , apoptosis rates , and expression of p - stat3 ( phosphorylated signal transduction and activators of transcription 3 ) and p - erk ( phosphorylated extracellular signal regulated protein kinase).resultsour data showed significant inhibition of growth , significantly increased s - phase arrest and apoptosis rates , and reduction of p - stat3 and p - erk1/2 expression levels.conclusionswe propose the mechanism may involve arhi - induced phosphorylation of erk1/2 and stat3 protein kinases , thereby blocking proliferation signaling pathways , to induce hoc skov3 apoptosis .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: from october 2006 to may 2008 , pta of the infrapopliteal arteries was attempted in 386 limbs ( 200 consecutive patients , 243 procedures ) of patients presenting with limb ischemia . in 143 patients , \n the patient ages ranged from 56 to 89 years ( mean 76 years ) and there were 118 women and 82 men . \n a total of 182 patients ( 91% ) were diabetics , 147 ( 74% ) had hypertension , 45 ( 23% ) had coronary artery disease , 16 ( 8% ) had cerebral vascular disease , and 41 ( 21% ) were smokers . \n clinical symptoms included rest pain in 69 ( 18% ) patients , intermittent claudication in 245 ( 64% ) , ulcers in 54 ( 14% ) , and gangrene in 18 ( 5% ) limbs . \n the mean ( se ) ankle brachial pressure index ( abi ) was 0.53 0.17 ( range 0.36 - 0.81 ) before the intervention . \n we generally defined arterial abnormalities as follows : foot pulse reduced or absent , abi < 1 ( in the absence of arterial calcification ) , or duplex scanning revealing stenosis > 50% of the vessel lumen . \n if two of the three criteria are confirmed , then a lower extremity mr angiography ( mra ) was performed to confirm these abnormalities and evaluate the vasculature and collateral compensation in all patients who were supposed to receive pta treatment . \n all mri examinations were performed on a 3.0 t mr system ( achieva 3.0 t , philips medical systems , the netherlands ) . \n automatic table movement bolus chase 3d contrast - enhanced mra was employed to perform a four - station protocol ( lower abdomen - thigh - lower leg - foot ) by using an integrated whole body coil with additional infragenual cuff - compression ( end pressure of 50 mmhg ) during the mri examination . \n a biphasic injection of gadodiamide was chosen at an initial volume of 20 ml and at a flow rate of 2.5 ml / s , and was immediately followed by a second volume of 20 ml at a flow rate of 2.0 ml / s . for all stations , \n the 3d mobiflex-3.0 t sequence in the coronal slice order was used with the following parameters : tr = 3.5 - 4.4 ms , te = 1.28 - 1.64 ms , flip angle = 20 , field of view = 375 mm , voxel size = 0.98 . \n antegrade or retrograde puncture of the femoral artery was decided according to the mra results and discussed between the two vascular interventional radiologists . \n digital subtraction angiography ( dsa ) was then performed , and a subsequent pta procedure was performed if necessary . \n the antegrade puncture of the femoral artery was used in all but 33 limbs . in those 33 limbs , \n pta was performed with a cross - over technique from the contralateral side because of the severe stenotic lesions in the ipsilateral puncture site or extreme obesity of the patient . \n all interventions were performed through a 6 fr access system ( terumo co. , tokyo , japan ) . \n a 4-fr vertebral catheter ( cook , bloomington , in ) was subsequently introduced and an angiography was performed . \n the tip of the catheter was placed at the proximal end of the occlusive vessel under the guidance of a road map . \n the outflow tract in the distal end of the occlusive lesion was an indication that an angioplasty was necessary . \n we used a low friction hydrophilic 0.035-inch guidewire ( terumo co. , tokyo , japan ) for the recanalization of the total occlusions . \n the tip of the guidewire was shaped into and allowed to form a ' u ' shape with the aid of a 4-fr catheter and collateral branch vessel in the proximal part of the occluded lesion . \n the ' u ' shape of the guidewire could ensure a wider profile , thus preventing the guidewire to exit the arterial wall or enter into the subintimal area . \n the 0.035-inch hydrophilic guidewire was replaced with a 0.014-inch skipper exchange guidewire ( skipper deep ; invatec , brescia , italy ) . \n hemi - compliance low - profile balloons ( amphirion deep ; invatec , brescia , italy ) with a balloon length of 80 or 120 mm and a diameter ranging from 2.0 to 3.5 mm were used for dilating the occluded vessel . \n the dilating pressure ranged from 6 to 10 atm and was maintained for three minutes . \n immediate postangioplasty angiograms were obtained in all cases . intravenous heparin ( a bolus of 4,000 iu followed by 1,000 iu ) \n was injected to maintain an activated clotting time of 250 to 300 seconds during the procedure . \n an additional 5,000 iu of low molecular weight heparin was administered subcutaneously every 12 hours for three days after the procedure . upon ambulation , \n the patients were to receive life - time antiplatelet ( aspirin , bayer co. , 100 mg daily ) treatment . \n technical success was defined as at least one anterior or posterior tibial artery with less than 50% residual stenosis that could be recanalized to the malleolar or dorsal foot along with no immediate severe complications . \n clinical success was defined in the wound group as complete healing of ulcers or dry and painless toe gangrene . in the intermittent claudication or rest pain group , \n success was defined as significant improvement in terms of distance walked or relief from rest pain for at least six months . \n a total of 417 occluded or severe stenotic lesions were treated by pta . among the 386 limbs \n treated , there were 305 lesions of the anterior tibial artery , 89 lesions of the posterior tibial artery , and 23 lesions of the peroneal artery ( table 1 ) . \n the lesion length varied from 2 to 32 cm , including 294 ( 71% ) lesions that were > 10 cm and 123 ( 29% ) lesions that were < 10 cm ( p < 0.0001 ) . \n altogether , 238 ( 57% ) lesions had an obvious outflow tract , whereas the rest did not . \n the patients were followed by clinical examinations at one , six and twelve months after the endovascular treatment . \n mras were supposed to be performed at six months in all patients . for those who were reluctant to undergo an mra follow - up , \n a doppler ultrasound examination was performed instead . at each follow - up examination , pain and wound healing \n were evaluated and palpated leg pulses were recorded . the visual analogue scale method was used to compare the pain levels before and after the procedure . \n pain relief was considered to have occurred if patients marked a 0.2-cm drop in pain level on a 0 to 10-cm scale . \n from october 2006 to may 2008 , pta of the infrapopliteal arteries was attempted in 386 limbs ( 200 consecutive patients , 243 procedures ) of patients presenting with limb ischemia . in 143 patients , \n the patient ages ranged from 56 to 89 years ( mean 76 years ) and there were 118 women and 82 men . \n a total of 182 patients ( 91% ) were diabetics , 147 ( 74% ) had hypertension , 45 ( 23% ) had coronary artery disease , 16 ( 8% ) had cerebral vascular disease , and 41 ( 21% ) were smokers . \n clinical symptoms included rest pain in 69 ( 18% ) patients , intermittent claudication in 245 ( 64% ) , ulcers in 54 ( 14% ) , and gangrene in 18 ( 5% ) limbs . \n the mean ( se ) ankle brachial pressure index ( abi ) was 0.53 0.17 ( range 0.36 - 0.81 ) before the intervention . \n we generally defined arterial abnormalities as follows : foot pulse reduced or absent , abi < 1 ( in the absence of arterial calcification ) , or duplex scanning revealing stenosis > 50% of the vessel lumen . \n if two of the three criteria are confirmed , then a lower extremity mr angiography ( mra ) was performed to confirm these abnormalities and evaluate the vasculature and collateral compensation in all patients who were supposed to receive pta treatment . \n all mri examinations were performed on a 3.0 t mr system ( achieva 3.0 t , philips medical systems , the netherlands ) . \n automatic table movement bolus chase 3d contrast - enhanced mra was employed to perform a four - station protocol ( lower abdomen - thigh - lower leg - foot ) by using an integrated whole body coil with additional infragenual cuff - compression ( end pressure of 50 mmhg ) during the mri examination . \n a biphasic injection of gadodiamide was chosen at an initial volume of 20 ml and at a flow rate of 2.5 ml / s , and was immediately followed by a second volume of 20 ml at a flow rate of 2.0 ml / s . for all stations , \n the 3d mobiflex-3.0 t sequence in the coronal slice order was used with the following parameters : tr = 3.5 - 4.4 ms , te = 1.28 - 1.64 ms , flip angle = 20 , field of view = 375 mm , voxel size = 0.98 . \n antegrade or retrograde puncture of the femoral artery was decided according to the mra results and discussed between the two vascular interventional radiologists . \n digital subtraction angiography ( dsa ) was then performed , and a subsequent pta procedure was performed if necessary . \n the antegrade puncture of the femoral artery was used in all but 33 limbs . in those 33 limbs , \n pta was performed with a cross - over technique from the contralateral side because of the severe stenotic lesions in the ipsilateral puncture site or extreme obesity of the patient . \n all interventions were performed through a 6 fr access system ( terumo co. , tokyo , japan ) . \n a 4-fr vertebral catheter ( cook , bloomington , in ) was subsequently introduced and an angiography was performed . \n the tip of the catheter was placed at the proximal end of the occlusive vessel under the guidance of a road map . \n the outflow tract in the distal end of the occlusive lesion was an indication that an angioplasty was necessary . \n we used a low friction hydrophilic 0.035-inch guidewire ( terumo co. , tokyo , japan ) for the recanalization of the total occlusions . \n the tip of the guidewire was shaped into and allowed to form a ' u ' shape with the aid of a 4-fr catheter and collateral branch vessel in the proximal part of the occluded lesion . \n the ' u ' shape of the guidewire could ensure a wider profile , thus preventing the guidewire to exit the arterial wall or enter into the subintimal area . \n the 0.035-inch hydrophilic guidewire was replaced with a 0.014-inch skipper exchange guidewire ( skipper deep ; invatec , brescia , italy ) . \n hemi - compliance low - profile balloons ( amphirion deep ; invatec , brescia , italy ) with a balloon length of 80 or 120 mm and a diameter ranging from 2.0 to 3.5 mm were used for dilating the occluded vessel . \n the dilating pressure ranged from 6 to 10 atm and was maintained for three minutes . \n immediate postangioplasty angiograms were obtained in all cases . intravenous heparin ( a bolus of 4,000 iu followed by 1,000 iu ) \n was injected to maintain an activated clotting time of 250 to 300 seconds during the procedure . \n an additional 5,000 iu of low molecular weight heparin was administered subcutaneously every 12 hours for three days after the procedure . upon ambulation , \n the patients were to receive life - time antiplatelet ( aspirin , bayer co. , 100 mg daily ) treatment . \n technical success was defined as at least one anterior or posterior tibial artery with less than 50% residual stenosis that could be recanalized to the malleolar or dorsal foot along with no immediate severe complications . \n clinical success was defined in the wound group as complete healing of ulcers or dry and painless toe gangrene . in the intermittent claudication or rest pain group , \n success was defined as significant improvement in terms of distance walked or relief from rest pain for at least six months . \n a total of 417 occluded or severe stenotic lesions were treated by pta . among the 386 limbs treated , \n there were 305 lesions of the anterior tibial artery , 89 lesions of the posterior tibial artery , and 23 lesions of the peroneal artery ( table 1 ) . \n the lesion length varied from 2 to 32 cm , including 294 ( 71% ) lesions that were > 10 cm and 123 ( 29% ) lesions that were < 10 cm ( p < 0.0001 ) . altogether , 238 ( 57% ) lesions had an obvious outflow tract , whereas the rest did not . \n the patients were followed by clinical examinations at one , six and twelve months after the endovascular treatment . \n mras were supposed to be performed at six months in all patients . for those who were reluctant to undergo an mra follow - up , \n a doppler ultrasound examination was performed instead . at each follow - up examination , pain and wound healing \n were evaluated and palpated leg pulses were recorded . the visual analogue scale method was used to compare the pain levels before and after the procedure . \n pain relief was considered to have occurred if patients marked a 0.2-cm drop in pain level on a 0 to 10-cm scale . \n in this study , a technically successful pta in at least one artery that could be recanalized directly to the malleolar or dorsal foot was achieved in 322 limbs ( 89% ) ( fig . \n 2 ) . altogether , 374 of 417 ( 90% ) infrapopliteal occluded or severely stenotic lesions were successfully recanalized . \n the loop technique was attempted in 393 of 417 lesions ( 94% ) in 361 limbs , and the technique success rate attained 90% ( 352 of 393 lesions ) . in certain cases , \n the loop guidewire could not cross over the occlusion segments because of severe calcification ; this situation occurred in 41 lesions longer than 10 cm . \n vessel rupture occurred in 19 segments and we treated 142 compound stenoses or occlusions in the iliac - femoral - popliteal axis arteries of 98 patients , which were subsequently followed by the implantation of 28 self - expanded stents . there were a total of 38 major immediate post - procedure complications that were noted . \n one patient suffered a retroperitoneal hematoma four hours after the procedure , which required an emergency surgical intervention . \n distal emboli were found in 24 vessels ( 6% ) , all of which were treated with urokinase and thrombosuction . \n ischemia did not improve in one patient , and he subsequently required a below - ankle amputation two weeks later ( fig . \n vessel rupture was identified in 19 of 393 ( 5% ) segments of 19 limbs because the guidewire perforated the wall of the vessel . for these patients , \n the heparin was immediately balanced out with protamine sulfate , and in 15 of these limbs , a successful intraluminal angioplasty was eventually performed . \n no other treatments were required for the remaining four limbs from the four patients , but their symptoms of limb ischemia did not improve . \n there were 46 ( 12% ) minor local dissections at sites after the dilatation , of which 21 were left untreated because no retrograde flap was detected by the antegrade angiography . \n the remaining serious detected intimal flaps were treated by re - dilation with a balloon of the same diameter , but at a higher pressure ( an additional 1 - 2 atm ) . \n the mean follow - up period for this study was 10.5 6.2 months ( range : 1 to 12 months ) . \n altogether , clinical success , including improvement of intermittent claudication or rest pain and wound healing , was achieved in 186 of the 200 ( 93% ) patients . \n after the procedure , rest pain was relieved in 58 of 69 ( 84% ) limbs and 207 of 245 ( 85% ) limbs showed improvement for intermittent claudication . \n complete wound healing was noted in 21 of 54 ( 39% ) limbs and 20 of 54 ( 37% ) limbs showed improvement in the wound size or depth . of these cases of improvement , \n a repeat pta on the same leg was performed on 43 limbs in 35 patients . in the healed / improved group , \n twenty - two patients had subsequent minor leg amputations , but the level of amputation was lower than previously estimated . \n the non - symptomatic patients did not undergo a diagnostic angiography during the follow - up period . \n mra data could be obtained from 144 patients , while 35 patients underwent a doppler ultrasound examination follow - up . \n the mra follow - up at six months found re - stenosis ( stenosis 50% ) in 34 of 264 segments , but the majority of these patients did not show any symptoms . \n in this study , a technically successful pta in at least one artery that could be recanalized directly to the malleolar or dorsal foot was achieved in 322 limbs ( 89% ) ( fig . \n 2 ) . altogether , 374 of 417 ( 90% ) infrapopliteal occluded or severely stenotic lesions were successfully recanalized . \n the loop technique was attempted in 393 of 417 lesions ( 94% ) in 361 limbs , and the technique success rate attained 90% ( 352 of 393 lesions ) . in certain cases , \n the loop guidewire could not cross over the occlusion segments because of severe calcification ; this situation occurred in 41 lesions longer than 10 cm . \n vessel rupture occurred in 19 segments and we treated 142 compound stenoses or occlusions in the iliac - femoral - popliteal axis arteries of 98 patients , which were subsequently followed by the implantation of 28 self - expanded stents . \n there were a total of 38 major immediate post - procedure complications that were noted . \n one patient suffered a retroperitoneal hematoma four hours after the procedure , which required an emergency surgical intervention . \n distal emboli were found in 24 vessels ( 6% ) , all of which were treated with urokinase and thrombosuction . \n ischemia did not improve in one patient , and he subsequently required a below - ankle amputation two weeks later ( fig . \n vessel rupture was identified in 19 of 393 ( 5% ) segments of 19 limbs because the guidewire perforated the wall of the vessel . for these patients , \n the heparin was immediately balanced out with protamine sulfate , and in 15 of these limbs , a successful intraluminal angioplasty was eventually performed . \n no other treatments were required for the remaining four limbs from the four patients , but their symptoms of limb ischemia did not improve . \n there were 46 ( 12% ) minor local dissections at sites after the dilatation , of which 21 were left untreated because no retrograde flap was detected by the antegrade angiography . \n the remaining serious detected intimal flaps were treated by re - dilation with a balloon of the same diameter , but at a higher pressure ( an additional 1 - 2 atm ) . \n the mean follow - up period for this study was 10.5 6.2 months ( range : 1 to 12 months ) . \n altogether , clinical success , including improvement of intermittent claudication or rest pain and wound healing , was achieved in 186 of the 200 ( 93% ) patients . \n after the procedure , rest pain was relieved in 58 of 69 ( 84% ) limbs and 207 of 245 ( 85% ) limbs showed improvement for intermittent claudication . \n complete wound healing was noted in 21 of 54 ( 39% ) limbs and 20 of 54 ( 37% ) limbs showed improvement in the wound size or depth . of these cases of improvement \n a repeat pta on the same leg was performed on 43 limbs in 35 patients . in \n twenty - two patients had subsequent minor leg amputations , but the level of amputation was lower than previously estimated . \n the non - symptomatic patients did not undergo a diagnostic angiography during the follow - up period . \n mra data could be obtained from 144 patients , while 35 patients underwent a doppler ultrasound examination follow - up . \n the mra follow - up at six months found re - stenosis ( stenosis 50% ) in 34 of 264 segments , but the majority of these patients did not show any symptoms . \n peripheral artery occlusive disease is estimated to occur in 3% of people aged 40 - 59 years and in 20% of people over 70 years , as documented by noninvasive testing ( 3 , 4 ) . \n percutaneous endovascular procedures are increasingly applied to treat symptomatic peripheral artery occlusive disease . despite the fact that advances in balloon catheters have made the infrapopliteal pta procedure easier than before for long or multi - segment occluded vessels , it is still difficult for the catheter to cross over occluded segment(s ) . \n if the occluded vessel is relatively short , it is easier for the guidewire to cross over to the distal true lumen simply with a straight head . \n , the straight tip guidewire could easily perforate into the intima or outside the arterial wall and eventually result in dissection or vascular rupture . \n thus we remolded a ' u ' shape loop out of a 0.035-inch hydrophilic guidewire and made the guidewire and a 4-fr catheter cross the occlusive segment . \n next , the guidewire was withdrawn and an angiography was performed to confirm that the catheter was in the true lumen . following this , \n the diameter of the compatible guidewire for the new infrapopliteal balloon catheter was 0.014 inches , which provided insufficient support compared to the commonly used 0.035-inch hydrophilic guidewire . in the early stages , we nevertheless tried to cross through the lesion with the skipper guidewire , but the tip was too soft to cross over . \n the first method involved the use of a remoldable guidewire ( terumo co. , japan ) . \n we can remold the tip to a ' j ' type in advance and then insert the catheter . \n this ' j ' type could recover in the larger diameter vessels such as the superficial femoral artery . \n this method was sometimes not effective because the remolded guidewire was inserted into the infrapopliteal target vessel with difficulty , especially into the anterior tibial artery . \n the other method relies on assistance from the catheter and vascular branch . under most circumstances \n , we can first insert the hydrophilic guidewire into a proximal branch and then make the ' u - shaped ' loop by continually rotating and advancing the guidewire . when using the 0.035-inch stiff guidewire to penetrate long occlusion lesions \n , we generally looped the head wire into a ' u ' shape by letting the head of the wire enter one of branches and then twisting the wire to form the loop . \n this technique helps to guarantee that the guidewire stays in the arterial lumen , prevent the guidewire from perforating the vessel , and hence reduce damage to the vessel wall during advancing . \n this technique also provides strength when penetrating an organized thrombus or calcified plaque as well as reduce the likelihood that the wire will enter the subintima as occurs when using the ' j - shaped ' guidewire head . \n this intraluminal angioplasty technique allows us to reduce the thrombosis occurrence and also provides a better long - term patency of the recanalized vessel compared to the subintimal angioplasty , according to our study ( 5 , 7 ) . \n however , this technique may have a high manipulation requirement and result in a somewhat higher vessel perforation rate ( 5% ) . \n careful advance must be maintained to prevent the guidewire from perforating the proximal vascular branch . \n sometimes we need to rotate the guidewire to better facilitate its passing through the lesion . \n the loop tip of the guidewire should be kept straight before it arrives at the dorsal artery of the foot or arteria plantaris , because its diameter is close to that of the guidewire ( fig . \n ( 8) report that a emergency surgical intervention was required after 31 of 1,377 procedures ( 2% ) with acute limb ischemia and hemorrhagic complications . in our group , only one patient with a retroperitoneal hematoma required emergency surgical treatment . \n the hemorrhage caused by infrapopliteal vessel perforation required no further treatment and no calf hematoma was found . \n distal embolic events may be inevitable in the procedure of recanalization of an occluded vessel . \n distal embolization of a plaque or thrombus may cause organ ischemia following percutaneous peripheral interventions , especially in the superficial femoral or iliac artery . \n this event may occur at various phases of intervention such as guidewire crossing , balloon angioplasty , stent deployment , and so on ( 9 ) . \n thrombolytic or thrombosuction therapy was usually effective and amputation was performed in only one patient after an angioplasty because of distal limb ischemia . \n ( 8) . to date , stent deployment below the knee is rare ; lam et al . \n ( 9 ) monitored the whole procedure during a superficial femoral artery intervention and found that embolic signals were noted in every patient during wire crossing , angioplasty , stent deployment , and atherectomy . \n however , patency was restored angiographically after thrombolysis . despite the frequency of these events , \n only one patient developed an angiographically and clinically significant embolization . in our patients , distal embolic events \n were noted angiographically in 24 vessels , however only one patient suffered from significant ischemia of a limb requiring below - the - knee amputation . an ipsilateral antegrade common femoral artery ( cfa ) puncture is usually preferred to a contralateral retrograde femoral access for the below - the - knee pta . in our group , procedures on all but \n antegrade puncture refers to the placement of an angiographic needle in the cfa in the direction of arterial flow . \n the puncture is made in the skin above the inguinal ligament , but the tip of the needle should always enter the cfa below the inguinal ligament . \n this procedure allows for the insertion of a vascular sheath for balloon angioplasty of the superficial femoral and/or popliteal arteries ( 10 ) . because of the frequent difficulty with providing sufficient manual pressure at the puncture site \n , an antegrade puncture is complicated by a high number of local bleeding complications , including retroperitoneal hematoma ( 11 ) . \n hemostasis of the puncture sites at the end of the procedure in all of our patients was achieved by manual compression . \n arterial access can be complicated by a ' hostile groin ' ( scarring , obesity , or previous failed cfa puncture ) ( 12 ) . \n it is a challenging procedure with a learning curve for the interventional radiologist ( 13 ) . \n a high puncture ( above the inguinal ligament ) is associated with increased incidence of groin hematoma and retroperitoneal hemorrhage , whereas a low puncture is associated with increased incidence of arteriovenous fistula and pseudoaneurysm formation ( 14 , 15 ) . \n a fluoroscopy is a helpful technique in guiding the arterial puncture towards the mid - femoral head . in obese patients , \n local bleeding may cause ecchymoma , which could be relieved via physical treatment . to date , \n femoral artery closure devices such as the angio - seal collagen plug and anchor device have been approved and shown to benefit patients after femoral artery catheterization . only a few studies in the literature \n have examined such closure devices of a post - antegrade puncture ( 11 , 16 - 18 ) . \n these vascular closure devices may be a safe , efficient and uncomplicated means of achieving arteriotomy closure of a post - antegrade puncture . \n in conclusion , in our experience , pta of infrapopliteal vessels is effective in treating infrapopliteal arterial occlusive disease , while also being associated with a low risk for major medical and surgical complications . based on our findings , \n the guidewire looping technique is safe and effective for the recanalization of occlusive segments of vessels . as far as we know \n , this paper is the first report on this technique applied to below - the - knee vessels .", "role": "user" }, { "content": "objectiveto determine the efficacy , safety and primary follow - up results of a guidewire looping technique for the treatment of infrapopliteal arteries.materials and methodsfrom october 2006 to may 2008 , an intraluminal angioplasty of the infrapopliteal arteries was attempted in 200 consecutive patients . \n altogether , 417 infrapopliteal lesions , with lengths varying from 2 cm to 32 cm , were treated as part of this study , including 305 lesions in the anterior tibial arteries , 89 in the posterior tibial arteries , and 23 in the peroneal arteries . \n the ' u'-shaped guidewire technique was attempted in 393 lesions from 361 limbs . \n the tip of a hydrophilic 0.035-inch guidewire was formed into a ' u ' shape with the aid of a 4-fr catheter and collateral branch vessel to recanalize the completely occluded long segment lesions.resultsa successful angioplasty with at least one artery recanalized directly to the malleolar or dorsal foot was achieved in 322 limbs ( 89% ) . \n the looping technique had a success rate of 90% ( 352 of 393 lesions ) . \n after the procedure , the rest pain was relieved in 58 of 69 patients , while 207 of 245 limbs ( 85% ) showed improvement for intermittent claudication . \n complete wound healing was noted in 21 of 54 patients , while 20 of 54 patients showed an improvement in the wound size or depth . \n a total of 38 major immediate procedure - related complications were noted , including retroperitoneal hematoma , distal emboli , and vessel rupture.conclusionthe results of this study suggests that the guidewire looping technique is a safe and effective method for the recanalization of the occluded lesions in infrapopliteal vessels .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: in recent years , plant derived polymers have evoked tremendous interest due to their diverse pharmaceutical applications such as diluents , binders , disintegrants in tablets , thickeners in oral liquids , protective colloids in suspensions , gelling agents in gels , and bases in suppository ; they are also used in cosmetics , textiles , paints , and paper - making . \n the plant based polymers have been studied for their application in different pharmaceutical dosage forms like matrix controlled system , film coating agents , buccal films , microspheres , nanoparticles , and viscous liquid formulations like ophthalmic solutions , suspensions , and implants and their applicability and efficacy has been proven [ 35 ] . \n these have also been utilized as viscosity enhancers , stabilizers , disintegrants , solubilizers , emulsifiers , suspending agents , gelling agents and bioadhesives , and binders in the above mentioned dosage forms . \n the term mucilage in plants means those substances which are soluble or at least swell very perceptibly in water and which , upon the addition of alcohol , are precipitated in a more or less amorphous or granular mass . \n mucilage originates in the plant either as a part of the contents of the cell or as a part of the wall thereof . \n these materials include guar gum , ispaghula husk , galactomannan from mimosa scabrella , gleditsia triacanthos linn ( honey locust gum ) , sesbania gum , mucilage from the pods of hibiscus esculenta , tamarind seed gum , gum copal and gum dammar , agar , konjac , and chitosan . \n industrial gums and mucilage , which , for the most part , are water - soluble polysaccharides , have enormously large and broad applications in both food and non - food industries . \n their use depends on the unique physicochemical properties that they provide , often at costs below those of synthetic polymers . \n manilkara zapota ( linn . ) p. royen syn . is a large , evergreen , forest tree more than 30 m in height a tree belonging to the family sapotaceae . \n p. royen syn was carried out so as to enable its use as a food additive . \n an understanding of the physicochemical properties and structural characterization of mucilage is essential in exploiting its potential as a food additive and for other industrial applications . in the present paper , \n we report morphological , physicochemical and structural aspects of mucilage from seeds of manilkara zapota ( linn . ) \n the methods employed for the analysis include ( i ) particle size analysis by microtrac , ( ii ) scanning electron microscopy ( sem ) , ( iii ) gel permeation chromatography ( gpc ) , ( iv ) differential scanning calorimetry ( dsc ) , ( v ) differential thermal analysis ( dta ) , ( vi ) thermogravimetry analysis ( tga ) , ( vii ) zeta potential ( zp ) , ( viii ) elemental analysis ( carbon ( c ) , hydrogen ( h ) , nitrogen ( n ) , and sulfur ( s ) ) , ( ix ) mineral analysis , ( x ) powder x - ray diffraction spectrometry ( pxrd ) , ( xi ) fourier transform infrared spectroscopy ( ft - ir ) , ( xii ) 1d nuclear magnetic resonance ( nmr ) , and ( xiii ) paper chromatography analysis . \n fruits of manilkara zapota were collected from the forest of rajgamar , district korba , chhattisgarh , in the months of april june and authenticated by professor h. b. singh , niscair , new delhi , india . \n mucilage was isolated from the seeds using maceration techniques in which seed powder ( 100 g ) was soaked in cold distilled water ( 500 ml ) and slurry was prepared . \n then slurry was kept aside for a day , then solution was heated on bunsen burner for 1 h ; after one day the mixture was filtered with the help of muslin cloth . \n the supernatant was collected after centrifugation then double volume of acetone was added in it to precipitate the mucilage . \n the mucilage was then dried at 40c45c in hot - air oven ( biotech ) and then passed through mesh number 120 and stored in desiccators until used for further studies in powder form [ 9 , 10 ] . \n dispersion of sample was prepared in suitable solvent ( water and 0.1 n nacl ) and the relevant information such as density , viscosity , and dielectric constant was added for the solvent in software ( microtrac flex operating software ) . about 3.0 ml sample dispersion was added in sample holder . \n sample holder made of optical probes which are paired with their opposite electrodes in an insulating sample cell . \n particle size distribution was determined from the velocity distribution of particles suspended in a dispersing medium , using the principles of dynamic light scattering . \n the tape was first firmly attached to the stub and the sample powder was scattered carefully over its surface . \n the stub with the sample was then coated with a thin layer of gold to make the sample conductive . \n gpc is a technique which enables polymer molecules of different sizes in solution to be separated from one another , such that the large molecules elute first and small molecules elute last . as they emerge from the bottom of the column , they are detected by a differential refractometer . since the molecular size in solution ( hydrodynamic volume ) \n is related to mw ( molecular weight ) , a picture of the entire mw distribution is obtained quickly and simply . \n gpc was carried out to estimate molecular weight of the mucilage relative to dextran polysaccharide as standard , using waters alliance model ( waters 2695 separation module with autoinjector ) coupled with waters 2414 refractive index detector ( ri ) . \n mobile phase was 0.2 m nano3 in water , flow rate was 1.0 ml / min , ultrahydrogel 500 and ultrahydrogel 120 ( 7.8 mm 30 cm 9 um ) were in series . \n detector and column were operated at 30c , which was started from mw : 5,200 ; 48,600 ; 2,03,000 ; 6,68,000 ; 14,00,000 spectra were processed using empower software . \n dsc analysis for mucilages was performed using a differential scanning calorimeter ( mettler toledo star system ) \n . accurately weighed ( 5 mg ) samples were placed into platinum cups and sealed . \n the temperature range was from 0c to 300c under nitrogen atmosphere at a heating rate of 10c / min . \n dta is a very popular thermal analysis technique ; it measures endothermic and exothermic transitions as a function of temperature . \n the analysis mucilage was performed on differential thermal analysis ( stapt , linseis ) in which accurately weighed ( 5 mg ) samples were placed into platinum cups and sealed . \n the temperature range was from 0c to 240c at a heating rate of 15c / min . \n tga is a thermal analysis technique for measuring the mass variation in a sample as a function of temperature under a controlled atmosphere . \n this variation of mass can be positive ( mass gain ) when the sample is subjected to oxidation or corrosion , or else negative ( mass loss ) when part of the sample has been transformed into vapor . \n thermal stability behavior can be investigated in both conditions : ( 1 ) dynamic , in which the temperature is increased at a linear rate and ( 2 ) isothermal , in which the temperature is kept constant . \n the analysis of mucilages was done using thermal analyzer ( stapt , linseis , germany ) in which accurately weighed ( 15 mg ) samples were placed into platinum cups and sealed . \n the temperature range was from 0 to 900c at a heating rate of 15c / min . \n zp is related to the charge present on the surface or near - surface of a suspended particle . \n zp was determined using zetatrac ( microtrac ) by measuring the response of charged particles to an electric field . in a constant electric field particles \n float at a constant velocity . through this velocity and charge zp can be determined . \n the brownian motion power spectrum is analyzed with the nanotrac controlled reference technique of particle sizing to determine the modulated power spectrum ( mps ) . \n zp was calculated for mucilage from the mps signal using formula in water and ph dependence of the zeta potential was investigated with the background electrolyte of 0.1 n nacl \n ( 1)= , \n where is zeta potential , is mobility , is viscosity , and is dielectric constant for water at 25c , zeta potential ( mv ) ~ 12.8 mobility ( /sec / volt / cm ) . \n elemental compositions ( chns ) were analyzed using elemental analyzer ( elemental , vario ) . accurately weighed 0.5 g of sample was heated to 1150c and corresponding element was determined by using elemental analyzer . \n the experiments were carried out at 25c under the following conditions : voltage 40 kv , current 30 ma , and 2 angle with scan step time of 10.33 s with specific length of 10 mm . \n ftir spectra of mucilage were recorded on a ft - ir spectrometer ( thermo scientific ) . \n nmr spectra of h and c of mucilage were recorded in an nmr ( 400 mhz ) spectrometer ( bruker avance ii 400 ) . \n the test mucoadhesive agent ( 100 mg ) was dissolved in d2o and chemical shifts were reported in ppm relative to an internal standard tsp ( 3-trimethylsilylpropionic-2,2,3,3,-d4 acid , sodium salt , 98% d ) for h nmr and 1,4-dioxane ( d 66.67 ppm ) for c spectra . \n proton nmr spectra were obtained at a base frequency of 400 mhz , with 16 transitions and delay time 1.5 s and for c , the base frequency was 100 mhz , with 3000 scans and delay time 2 s . \n carbohydrate or sugars occupy a central position in plant metabolism , so the method of their detection and estimation is very important . \n sugars are conveniently classified into three groups , on the basis of molecular size : the simple monosaccharide 's ( e.g. , glucose and fructose ) and their derivatives , the oligosaccharides , by the condensation of two or more monosaccharide 's units ( e.g. , sucrose ) , and the polysaccharides which consist of long chains of monosaccharide 's units , joined head to tail , either as straight chains or without branching . the samples were prepared in distilled water for paper chromatography . \n the paper chromatography analysis was performed using n - butanol : acetic acid : water ( 4 : 1 : 1 ) as solvent system with using arabinose , fructose , mannose , rhamnose , and xylose as reference standards . \n the average particle size as determined by zetatrac particle size analyzer indicated that obtained mucilages were in fine particle size and average range was obtained as 500551 nm diameters . \n the particles are mostly seen as aggregates of irregular shapes and dimensions which were fibrous in nature . \n the molecular weight of mucilages were determined by gel permeation chromatography and expressed as the dextran polysaccharide equivalent molecular weight . \n the computed average molecular weights , ( mw ) , number of average molecular weight , ( mn ) , and polydispersity ( mw / mn ) are tabulated in table 1 . \n dsc has emerged as powerful physical tools to monitor physical and chemical changes that occur in the gum during thermal processing and these methods yield curves that are unique for a given gum . \n the outcome of differential scanning calorimetry ( dsc ) ; analysis of mucilage revealed that the glass transition temperature is 138c . \n the major intense peak recorded in the dsc thermograms is an endothermic transition ( at around 200c ) followed by weaker exotherm(s ) . \n tga is a simple and accurate method for studying the decomposition pattern and the thermal stability of polymers . \n table 2 gives the details of thermal behavior according to the primary thermograms and derivative thermograms of mucilages . \n the representative plot results of thermogravimetric analysis carried out on the mucilages under lean oxygen ( 5% oxygen in nitrogen ) atmosphere are shown in figure 4 . \n the details of thermal behavior and thermal stability data according to the primary thermograms and derivative thermograms for the gum showed that heating at a rate of 10c per minute from 0c to a maximum of 900c resulted in two mass loss events \n . the results of elemental composition and heavy metal characterization are tabulated in table 3 . \n the elemental analysis study showed that the isolated mucilage had certain percentage of carbon , nitrogen , sulphur , and hydrogen , while toxic heavy metal like arsenic , lead , cadmium , and mercury were absent . \n the zp measurement was performed to collect information on the stability and charge behavior of the polymer . \n the zp of mucilage in aqueous medium ( water ) was recorded to be 18.05 mv and in 0.1 n nacl was recorded to be 5.15 mv , respectively . \n the result indicated that no characteristic peaks were observed in the spectrum , indicating that the mucilages were completely amorphous in nature . \n the spectra of mucilage showed band occurring at 3559.78 cm results from the presence of amide n h & c = o stretch and amine n h . the peak obtained at 3470.093418.01 cm results from the presence of hydroxyl ( oh and co ) groups . \n the peak obtained at 2884.672819.09 cm results from stretching modes of the alkyl c h stretch , carboxylic acid c = o & o h stretch , and methylene ( ch2 ) c h stretch . \n the peak obtained at 2167.121660.78 cm results from stretching mode of the alkenyl c h & c = c stretch and amide n h & c = o stretch . \n the interpretation of h and c nmr spectra analysis was done using chemix school software and was represented in figures 7 and 8 . \n the h and c nmr spectra of mucilage indicated presence of certain sugar composition such as signals of h nmr spectra between d 3.58d 3.40 ppm can be attributed to ch and oh group of rhamnose , mannose and arabinose . \n the signal at d 67.9 ppm , 72.2 ppm , and d 72.3 ppm of c nmr spectra can be attributed to ch group of rhamnose and mannose . \n the rf value of standards arabinose , fructose , mannose , rhamnose , and xylose matched with the obtained rf values of test substances ( mucilage ) . \n the traditional concept of the excipients as any component other than the active substance has undergone a substantial evolution from an inert and cheaper vehicle to an essential constituent of the formulation . \n excipients are any components other than the active substances intentionally added to formulation of a dosage form . \n micromeritics involve the study of small particles and the order of a few micron sizes . \n particle size analysis involves the characterization of individual particles , particles size distribution , and powders . \n particle size also previews the degree of densification , which could occur during tableting which was further also confirmed by sem analysis . \n microscopy is a major tool for the characterization of polymer material ultrastructure . in case of drug delivery system , \n such imaging technique is used to examine the detail shape , size , and distribution of polymeric micro- and nanoparticles , as well as interaction with biological environment . \n the sem results of the present study suggest that hydration capacity of mucilage depends on the surface property as presented in powder xrd studies . \n it was reported earlier that particle size and specific surface area influence the hydration behavior of gums , which in turn influence their intrinsic viscosity and molecular mass [ 2224 ] . \n further , wang experimentally reported that particle size influenced the hydration kinetics and molecular mass of guar gum which is a galactomannan - rich tree gum . \n polymer molecular weight determination is important because it determines many physical properties such as the temperatures for transitions from liquids to wax to rubber to solids and mechanical properties such as stiffness , strength , viscoelasticity , toughness , and viscosity . \n mw / mn ) is used as a convenient measure of the range of molecular weight present in a distribution and is in the range of 1.57.0 for natural polysaccharide gums . \n if the molecular weight is too low , the transition temperatures and the mechanical properties will generally be too low for the polymer material to have any useful commercial applications . for a polymer to be useful it must have transition temperatures to wax or liquids that are above room temperature and it must have mechanical properties sufficient to bear design loads . \n further , molecular weight is also a physical property of compound , which helps in standardization . whenever a polymer undergoes a phase transition such as melting or a glass transition from the rigid glass state to the soft rubber state , the temperature trends to remain constant while energy is taken in to the system . \n dsc and dta are essentially techniques that compare the difference between the energy acquired or released by a sample and a suitable reference as a function of temperature or time , while the sample and reference are subjected to a controlled temperature rise . \n the result of dsc and dta revealed that the isolated mucilage from seeds has good stability . \n dehydration , depolymerization , and pyrolytic decomposition are involved in dsc and dta at high temperature stages and resulted in the formation of h2o , co , and ch4 . however , because of the difference in structures and functional groups , either the degradation routes or the resulting fragments will be different . \n therefore , thermal scission of the carboxylate groups and evolution of co2 from the corresponding carbohydrate backbone may be a probable mechanism for the thermal transitions . \n the early minor weight loss in samples is attributed to the loss of adsorbed and structural water of biopolymers as related by other authors [ 25 , 26 ] or due to the desorption of moisture as hydrogen bound water to the saccharide structure . \n the second weight loss event may be attributed to the polysaccharide decomposition and is described by a weight loss . \n the weight loss onset ( representing the onset of oxidation or decomposition ) of polymer suggests that mucilages have good thermal stability . \n elemental analysis is a process where a sample of some materials ( e.g. , soil , minerals , and chemical compounds ) is analyzed for its elemental composition such as carbon , hydrogen , nitrogen , and sulphur . \n elemental analysis can be qualitative ( determining what elements are present ) , and even it can be quantitative . \n the elemental and heavy metal analysis showed presence of certain percentage of carbon , nitrogen , sulphur , and hydrogen and absence of heavy metals in mucilages . \n further study also suggested the presence of s - containing protein and amino acids and also confirms the formation of bond ( primary strong covalent bonds , weak secondary hydrogen bond , and vander waal 's forces ) of mucilage with mucosa in a short duration of time . \n electrokinetic measurement is one way to study complex surface chemical exchange processes at the clay mineral surface / liquid interface . \n all electrokinetic phenomena are related to the development of electrical double layer at the particle / electrolyte interface . \n the zeta potential is defined as the potential of shear plane of the particle when it moves in liquid . \n the results of zp studies suggested that the mucilages are ionic in nature and attributed to the presence of uronic acids as similar to what was previously reported by mohammadifar et al . for gum tragacanth which is uronic acid - rich gum . \n polarity of mucilage may be due to the presence of oh group , which is readily available for the formation of hydrogen bonds on the interface of reinforced systems . however , plant mucilages are covered with pectin and waxy substances , thus hindering the hydroxyl groups ( oh ) from reacting with polar matrices and deteriorate adhesion , but polar structure is only responsible for hydrophilic behavior of mucilage . \n for high adhesion forces , it is necessary to have polar functional groups , but swelling processes should be prevented . according to kanamaru , \n the extent of mucus adsorption is proportional to the absolute values of the positive zeta potential of mucilage and negative zp of mucus glycoprotein . \n shashikant reported that the significant difference in zp of a polymer may be obtained due to change in environment , that is , particles decomposition which yield small charge and surface properties of particles . \n it was concluded that conductivity of any solution is a function of electrolytes or ions and charge in ionic concentration is known to increase conductivity . the conductivity and dielectric constant often increase with thermal and electrical stress . \n this also finds support in the fact that there is concomitant shift in uv and ir spectra . \n taking account of earlier studies it can be believed that the zp caused huge impact on properties of mucilage . \n the powder x - ray diffraction is a useful method for investigating the arrangement of atom and molecules within the material . if there is an orderly arrangement of substructure within the material with repeat distances of a similar magnitude to the wavelength of light used , interference patterns are produced , and such patterns provide information on geometry of polymer structure . \n powder xrd analysis of mucilage sample showed no characteristic sharp peaks in the spectrum and indicated that the mucilage is completely amorphous in nature . \n natural gums , such as arabic , guar gum , and karaya gum , also show amorphous nature . \n ftir spectrometry has been extensively applied to characterize the polymer 's molecular and material structure . \n characterization using ftir spectroscopy often results in the identification of functional groups and the modes of their attachment to polymer backbone . \n the broad band occurring at 35003200 cm results from the presence of hydroxyl ( oh ) groups . \n natural gums usually contain fractions of sugar acid units which would usually impart a weakly anionic character to the gum macromolecule . \n absorption bands around 1618 and 1430 cm are typical of carboxylate groups of the galacturonic acid residues as reported by okafor et al . \n the region between 1500 and 1800 cm is typically used to detect presence of carboxylic groups . \n also , absorption peaks at 1740 cm and 1258 cm are typical of acetyl groups . \n the wave numbers between 800 and 1200 cm represents the finger print region for carbohydrates . \n nmr is the most powerful tool for the study of microstructure and chain configuration of polymers , both in solution and in solid state . \n the importance of nmr as a technique is from the fact that nmr signals can be assigned to specific atoms along the polymer backbone and side chains . \n the h nmr spectrum for mucilage showed few singlet at high field ( d 1.1785 ppm ( s ) , d 1.9559 ppm ( s ) ) , which is related to the environments of methyl groups of rhamnose and the protons linked to c-6 ( d 3.65 , d 3.70 ppm ) and c-4 of galactose ( d 3.98 , 4.28 ppm ) , respectively , and this suggests the existence of different galactose derivatives . \n the anomeric protons have been assigned to -sugar residues ( d 4.924.96 ppm ) and the -sugar residue ( d 5.15.3 ppm ) , as reported earlier by agrawal . \n the two closely neighbored signals observed in the h nmr spectrum of mucilage ( d 4.02 and d 3.84 ppm ) were assigned to h-1 of -glucose . \n the h nmr spectrum showed that crowded narrow region between 3 to 5 ppm is typical of polysaccharides and confirms the presence of many similar sugar residues . \n the signals between 3.1 and 4.3 ppm can be assigned to nonanomeric protons ( h2h6 ) , while signals between 4.3 and 4.8 , and 4.9 and 5.5 ppm arise from -anomeric and -anomeric protons , respectively . \n c nmr spectra of mucilages gave line widths which are typical of an amorphous natural polymer with broad band signals . \n there are resonance spectra due to the methyl group of rhamnose ( d 16.67 ppm ) as reported earlier by martnez et al . . \n plant gums and mucilages are an important group of plant constituents with pharmaceutical and technical uses . \n the separation and sensitive determination of various carbohydrates are currently of great interest in nutrition , medical cell biology , and biotechnology research . \n carbohydrates are difficult to analyze because they are very polar compounds , exhibit similar structural characteristics , and do not have a suitable chromophore . \n the paper chromatography analysis was performed in order to check the available carbohydrates in mucilage . \n the result obtained from 1d nmr , ftir , and paper chromatography studies indicated the presence of arabinose , fructose , mannose , rhamnose , xylose , and glucose . \n the result obtained from the study indicated similar rf values to the standard available data for sugars . \n according to lee , arabinose is reported to be a free , but minor , sugar component of onions , grapes , strawberries , commercial beer , corn , and alfalfa . \n the overall findings of the study indicated that mucilage isolated from the seeds have an inherent property which can be used in various dosage forms on modification . \n the pxrd and sem revealed that mucilage is amorphous in nature which indicated that wet granulation technology will be more suitable for dosage formulation . \n the thermal stability study by dsc , dta , and tga showed that mucilage had good thermal stability . \n ftir , solid 1d h , and c nmr and paper chromatography confirmed presence of nonreducing sugars . \n all these results demonstrate that mucilage is a useful pharmaceutical aid and can be used for effectively controlling the release of drugs from the designed matrix systems .", "role": "user" }, { "content": "mucilage isolated from seeds of manilkara zapota ( linn . ) p. royen syn . is a plant growing naturally in the forests of india . \n this mucilage is yet to be commercially exploited , and characterized as polymer . \n various physicochemical methods like particle size analysis , scanning electron microscopy , thermal analysis , gel permeation chromatography , x - ray diffraction spectrometry , zeta potential , fourier transform infrared spectroscopy , and nuclear magnetic resonance spectroscopy have been employed to characterize this gum in the present study . \n particle size analyses suggest that mucilage has particle size in nanometer . \n scanning electron microscopy analysis suggests that the mucilage has irregular particle size . \n the glass transition temperature of the gum was observed to be 138c and 136c by differential scanning calorimetry and differential thermal analysis , respectively . \n the thermogravimetric analysis suggested that mucilage had good thermal stability . \n the average molecular weight of mucilage was determined to be 379180 , by gel permeation chromatography , while the viscosity of mucilage was observed to be 219.1 cp . \n the x - ray diffraction spectrometry pattern of the mucilage indicates a completely amorphous structure . \n elemental analysis of the gum revealed the contents of carbon , hydrogen , nitrogen , and sulfur to be 80.9 ( % ) , 10.1 ( % ) , 1.58 ( % ) , and 512 ( mg / kg ) , respectively . \n mucilage had specific content of calcium , magnesium , potassium , lower concentrations of aluminum , cadmium , cobalt , lead , and nickel . \n the major functional groups identified from ft - ir spectrum include 3441 cm1 ( oh ) , 1660 cm1 ( alkenyl c h & c = c stretch ) , 1632 cm1 ( coo ) , 1414 cm1 ( coo ) , and 1219 cm1 ( ch3co ) . \n analysis of mucilage by paper chromatography and 1d nmr , indicated the presence of rhamnose , xylose , arabinose , mannose , and fructose .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: protein - protein interactions ( ppis ) are central to many if not all cellular processes . \n their importance has provoked broad interest in their analysis , which in turn has led to the construction of various large - scale interaction maps . \n the first ppi datasets were generated for model organisms such as saccharomyces cerevisiae , drosophila melanogaster and caenorhabditis elegans ( 15 ) . \n they are mostly based on review of literature ( 68 ) , extrapolation from interactions between orthologous proteins observed in other organisms ( 911 ) or application of high - throughput yeast two - hybrid ( y2h ) assays ( 12,13 ) . \n although these maps will certainly have profound impact on biological research , major limitations are lack of overlap , completeness and integration . \n scientists are required to interrogate numerous databases if they seek comprehensive information on potential interaction partners for specific human proteins . \n this generally involves time - consuming searches as various query formats and identifiers have to be used in different interaction databases . \n we have constructed the unihi database for the integration of large - scale human ppi maps . \n unihi offers a search platform that combines and gives access to ten different large - scale human ppi datasets . \n unihi is intended to reduce unnecessary duplication of data , while incorporating the strength of single databases regarding careful curation and annotation of ppis . \n the construction of unihi was motivated by the observation that human interaction maps tend to be highly divergent ( 14,15 ) . \n this is also the case for the interaction maps integrated in unihi ( table 1 ) . \n we observed that < 10% of all interactions occur in multiple maps , indicating a low degree of saturation ( figure 1b and supplementary data ) . \n the small number of shared interactions is remarkable considering the large number of proteins common to different datasets . \n more than 50% of all proteins are included in two or more maps ( figure 1a ) . \n thus , current ppi datasets are highly complementary sharing few interactions between many common proteins . \n numbers of proteins ( a ) and interactions ( b ) common to multiple maps . \n the histograms display frequency of proteins and interactions that are included in n different maps . \n ppi datasets currently integrated in unihi number of proteins and interactions in each dataset as well as construction approach are given . \n we have started to integrate available large - scale human ppi maps in unihi . in its initial version , unihi is based on the unification of the following interaction datasets recently generated : mdc - y2h , ccsb , hprd , dip , bind , cocit , reactome , ortho , homomint and ophid ( table 1 ) . \n these maps have been derived from manually curated databases ( 68,16 ) , computational approaches employing text - mining ( 13,17 ) , predictions based on orthology , ( 911 ) and from large y2h screenings ( 12,13 ) . \n matching of protein identifiers , which is essential for standardization , was performed using information from ensmart and hgnc ( 18,19 ) . \n for the combined map , we could assign 150 992 interactions between 17 064 unique proteins . for user friendliness , \n first , we wanted to indicate whether interactions are binary or complex . most of the included interactions are binary , while reactome comprises only complex interactions and hprd comprises both binary and complex ppis . to enable users to distinguish easily between the two types , we have split interaction data from hprd into two sets ( hprd - bin , hprd - comp ) . \n secondly , differentiation between ppis identified with different strategies was facilitated as choice of mapping approach has considerable impact on the ppis detected . \n ccsb data were divided into y2h- and literature - based interaction maps ( ccsb - y2h , ccsb - lit ) . \n the structure of the unihi database has been designed to integrate ppi data obtained from different sources . \n it consists of six key tables : protein , proteinaliases , proteindistribution , interactiondistribution , interactionproperties and interactionscore . \n it links the proteins with information about their properties , their interactions and their distribution and in the different ppi datasets ( supplementary figure s1 ) . \n a full description of the unihi database structure and its implementation can be found in the supplementary data . \n our aim was to provide easy and intuitive , but nevertheless efficient and comprehensive access to the integrated data . \n a search interface based on java programming language offers two different search options : in a single protein search , users input a single protein to query for its direct interaction partners . in a network - oriented multiple protein search users can supply a list of proteins . \n proteins can be entered by their corresponding gene symbol , entrez gene i d , uniprot i d , unigene i d , omim i d , ncbi geneinfo i d or ensembl i d . \n we utilized and extended a pre - existing java applet for graphical presentation of interaction networks ( 20 ) . \n retrieved interactions can be displayed either in textual ( figure 2 ) or graphical form ( figure 3 ) . \n for both types of views , interactions are directly hyperlinked to the maps from which they originate , with the exception of ophid , due to technical reasons , and ccbs , which is only available as a text file . to facilitate the interpretation of results , characteristic sets of colors \n textual representation of a query result for protein interactions in unihi . for each interaction partner found , a hyperlink is provided to the database from which the interaction originates . \n multiple links indicate inclusion in multiple maps . for easy discrimination between maps , specific colors have been assigned . \n shades of blue have been used for datasets derived by literature search , shades of green for orthology - based maps , shades of red for maps derived from y2h screens . \n after retrieval , users of unihi can visualize the interactions as graphs with interactions displayed as lines . \n ( b d ) output for a query with multiple proteins ( tp53 , cdc2 , e2f4 , hd , a2 m and gadd45a ) . \n multiple lines between proteins signify presence in several maps . queried proteins are symbolized by gray rectangles , their interacting partners by yellow rectangles . to assist users in the evaluation of results , \n several tools are offered to restrict interaction sets displayed according to selection of maps , multiple occurrences , common neighbourhood of proteins ( c ) , direct interactions between query proteins ( d ) . to permit users a highly targeted search , unihi offers several tools to specify the displayed interactions : ( i ) display only interactions from selected maps . \n ( ii ) display only proteins that are common interaction partners to multiple proteins in the query . \n such procedure can narrow down the context of a chosen set of proteins and can help to identify putative modifiers of physiological processes ( 12 ) . \n the aim of unihi is to provide a unified set of protein interactions included in the major human ppi maps that are publicly available . \n unihi has been designed with an open structure permitting future integration of further human interactome datasets . \n currently , perl scripts with integrated sql commands are used to preprocess and import interaction data after manual download from the corresponding web - pages . for future versions of unihi \n detailed information about the updating procedure can be found in the supplementary data . to examine the constitution of unihi \n , extensive statistical analysis was performed regarding network structure and functional annotation of integrated datasets . \n we also scrutinized the reliability of interaction maps using independent expression data and annotation ( see supplementary data ) . since the scope of unihi \n can be expected to be continuously expanding , these analyses will be regularly repeated and presented on the unihi webpage . \n this allows users a critical assessment of the single maps included in unihi as well as of unihi itself . to assess the quality of the interaction data , information on co - expression and co - annotation is presented for each interaction pair . \n additionally , unihi provides available links to the original pubmed articles that were used for curation in literature - based interactions maps . \n increasing numbers of human ppi datasets provide enormous amounts of valuable , but frequently unconnected information whose application in biology and medicine is still limited ( 2224 ) . \n lack of integration and overlap need to be addressed more strongly with experimental and bioinformatical strategies . \n unihi constitutes a highly practical integrated platform that allows simultaneous querying of the major human protein - protein interaction maps . \n it does not replace already available interaction maps , but facilitates single portal access to the larger part of the human interactome analyzed so far . \n unihi enables the assembly of comprehensive lists of protein interactions and flexible network - orientated searching . \n it allows identification of network structures which would not be detectable if single maps were analyzed separately . \n unihi is a flexible tool for the systematic utilization of human interactome data in biomedical research . \n ", "role": "user" }, { "content": "systematic mapping of protein protein interactions has become a central task of functional genomics . to map the human interactome , \n several strategies have recently been pursued . \n the generated interaction datasets are valuable resources for scientists in biology and medicine \n . however , comparison reveals limited overlap between different interaction networks . \n this divergence obstructs usability , as researchers have to interrogate numerous heterogeneous datasets to identify potential interaction partners for proteins of interest . to facilitate direct access through a single entry gate , \n we have started to integrate currently available human protein interaction data in an easily accessible online database . \n it is called unihi ( unified human interactome ) and is available at . at present , \n it is based on 10 major interaction maps derived by computational and experimental methods . \n it includes more than 150 000 distinct interactions between more than 17 000 unique human proteins . \n unihi provides researchers with a flexible integrated tool for finding and using comprehensive information about the human interactome .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: the recognition that plants host a diverse community of phytophagous organisms and mediate community interactions has led to a surge in studies investigating the interactions between herbivorous insects and plant parasitic nematodes ( mccarville et al . \n in particular , studies have begun to evaluate the plant - mediated interactions between insects and nematodes . \n insect feeding and nematode infection can induce various plant responses , namely plant defense compounds , and the resulting changes in a plant resource can alter nutrient availability , or the behavior and/or the fitness of the subsequent phytophagous organism ( bezemer et al . \n 2004 ; kaplan et al . 2009 ; manninen et al . 1998 ; van dam 2009 ) . \n a review of plant - mediated nematode and insect interactions showed that nematode - insect interactions may be positive , neutral , or negative but that feeding habits ( e.g. , piercing - sucking vs. chewing mouthparts ; migratory vs. sedentary endoparasitic ) in part mediate these interactions ( wondafrash et al . \n 2013 ) . in general , nematodes appear to neutrally and negatively affect piercing - sucking arthropods ( e.g. , reducing aphid fecundity and plant attractiveness to aphids ) , while chewing arthropods ( e.g. , caterpillars ) have neutral to positive interactions with nematode infected plants ( alston et al . \n the majority of plant parasitic nematodes feed belowground on roots , and understandably much of what we know comes from interactions between these root feeders and aboveground insects . \n studies on the interactions between aboveground feeding stem nematodes and insects are lacking and would contribute to the growing literature and knowledge of mechanisms driving nematode - insect interactions . \n alfalfa stem nematodes , ditylenchus dipsaci , are agriculturally important migratory endoparasites that harm medicago sativa ( alfalfa ) and other plant hosts ( duncan and moens 2006 ) . \n unlike many soil - dwelling plant parasitic nematodes , stem nematodes have free living stages in the soil that infect the crown buds of alfalfa but are carried in growing plant tissue aboveground and cause swollen nodes , and shortened internodes ( bohart et al . 1976 ) . \n furthermore , stem nematodes can migrate to apical regions of shoots and enter plant tissue through stomata , or penetrate directly into stems and leaf axils ( duncan and moens 2006 ) . \n similar to root - knot nematodes , stem nematodes have the ability to manipulate or suppress host plant responses when developing in the plant ( duncan and moens 2006 ; gheysen and mitchum 2011 ) . \n stem nematodes are pronounced in early spring when temperatures are cool ( < 20 c ) ( duncan and moens 2006 ) and interact with young plants before insect herbivores become active . \n the pea aphid , acyrthosiphon pisum , occurs from late - spring to mid - summer on shared host plants and is abundant , therefore , making it a good model herbivore to investigate nematode - insect interactions . \n induced plant responses toward phytophagous organisms rely on the plant hormones salicylic acid ( sa ) , jasmonic acid ( ja ) , and ethylene ( et ) ( pieterse et al . \n , sa mediates induction of systemic acquired resistance ( sar ) in plants toward plant pathogens , while the ja and et pathways are induced by insect feeding ( pieterse and van loon 1999 ) . \n current literature , however , finds that these responses can be tailored toward the type of herbivory . \n for example , necrotrophic pathogens and chewing insects tend to signal the ja pathway , while biotrophic pathogens and phloem - feeding insects tend to signal the sa pathway ( bari and jones 2009 ; smith and boyko 2006 ) . \n 2003 ) but there is a considerable amount of cross - talk among these signaling pathways ( bostock 2005 ; pieterse et al . \n 2012 ) . the complexity of this signaling leads to an array of defense responses against pathogens and insects , including the production of defensive proteins , toxins , and volatiles ( bezemer and van dam 2005 ; dicke et al . \n in particular , several proteins ( e.g. , chitinase , peroxidase , polyphenol oxidase , and trypsin inhibitor ) have been implicated in the toxicity , disruption of digestion , and decrease in nutritional value of plants toward nematodes and insects ( bi et al . \n 2006 ; kramer and muthukrishnan 1997 ; ryan 1990 ; zhu - salzman et al . 2008 ) . \n defense proteins can be linked with the sa ( chitinase and peroxidase ) and ja ( polyphenol oxidase and trypsin inhibitors ) pathways more specifically ( barto and cipollini 2005 ; and references therein ) . \n these responses may lead to trade - offs in defense when plants are attacked by pathogens vs. insects , and may help explain the outcomes of nematode - insect interactions . \n interestingly , however , an overlap of plant responses to pathogens and aphids has been observed . \n for example , aphid feeding can lead to the up - regulation of pathogen - response ( pr ) rna genes and production of defense proteins that lead to antixenosis ( girousse and bournoville 1994 ; smith and boyko 2006 ; walling 2000 ) . \n in addition , plants may respond to aphids with localized cell death at feeding sites , similar to the hypersensitive response of plants toward plant pathogens ( boyko et al . \n 2006 ; lyth 1985 ) . considering the general mechanisms of plant resistance for pathogens and insects ( e.g. , sa vs. ja signaling ) \n , tradeoffs may exist when selecting plants that are resistant to one phytophagous organism vs. another . \n alfalfa is a perennial , tetraploid legume with polysomic inheritance and broad genetic variation among commercial varieties ( maureira and osborn 2005 ) . breeding alfalfa and understanding the mechanisms directed at resistance \n is complex given that varieties are synthetic populations created by crossing a number of parents of selected genotypes ( maureira and osborn 2005 ) . \n saponins , a class of glycosides , are involved in herbivore suppression , and some alfalfa has been selected with elevated saponin content , but the effects on insects and pathogens are idiosyncratic and not consistent as resistance is in other systems . in tomato , for example , the mi-1 r protein requiring sa signaling leads to resistance toward root - knot nematodes and cross - resistance to aphids and whiteflies ( nombela et al . 2003 ; rossi et al . 1998 ; vos et al . \n plants without the mi-1 gene are more susceptible to aphids but only when ja signaling is blocked ( bhattarai et al . \n is geared toward particular phytophagous organism types , the possibility exists that host plants are silenced in particular defenses or that cross resistance is established . \n what complicates these interactions is that nematodes and insects have ways to evade various plant resistance traits and also induce plant responses . \n nematodes , for example , have enzymes that aid in cell wall degradation , protect against reactive oxygen species ( jones et al . \n 2004 ) , and effectors that suppress sa and ja production ( haegeman et al . 2012 ) . \n aphids avoid plant defenses with their saliva by plugging disrupted cells and creating a feeding tube and using watery saliva to prevent feeding site occlusion ( walling 2008 ) . \n in addition , some aphids reduce ja - regulated rnas by increasing sa pathways ( prado and tjallingii 2007 ; thompson and goggin 2006 ) . \n therefore , on a shared host plant , it is possible for one organism to prime plant responses in a way that affects the second herbivore . \n nematode infected plants , for example , have been shown to reduce caterpillar performance because they elevate levels of phenolics and glucosinolates in brassica ( van dam et al . \n 2005 ) and increase caterpillar performance in tobacco by interfering with nicotine synthesis ( kaplan et al . 2008 ) . \n field and greenhouse trials were conducted to examine how alfalfa varieties with varied resistance and stem nematode infection impact aphid abundance and plant defense responses . in particular , varieties were selected with resistance to nematodes only ( pgi-437 ) , aphids only ( rugged ) , both nematodes and aphids ( sutter ) , and a susceptible variety ( vernal ) . \n the objectives of this study were to determine ( 1 ) how selection of resistant varieties impact aphid populations , ( 2 ) how aboveground nematodes affect aphid abundance , and ( 3 ) to gain insight into the plant defense responses toward stem nematode and aphid among resistant varieties . \n a greenhouse trial was conducted at utah state university s research greenhouse complex in logan , ut , usa . in each of two years ( 20112012 ) \n , we evaluated the presence of nematodes on aphid abundance and plant defense protein concentration for each of four alfalfa varieties varying in resistance to nematodes and aphids . using the national alfalfa & forage alliance s pest resistance rating guides ( www.alfalfa.org ) \n , we chose varieties that crossed stem nematode resistance ( + , ) with pea aphid resistance ( + , ) . \n given the high level of genetic variation of m. sativa , resistance corresponds to % resistant plants , where highly resistant ( hr ) , resistant ( r ) , moderately resistant ( mr ) , and low resistant or susceptible ( s ) ratings are represented by > 50 % , 3150 % , 1530 % , and < 14 % resistant plants , respectively ( table 1 ) . here , we selected vernal ( susceptible to nematodes and aphids ) , pgi-437 ( resistant to nematodes but not aphids ) , rugged ( resistant to aphids but not nematodes ) , and sutter ( resistant to nematodes and aphids ) varieties.table 1resistance ratings for alfalfa varieties ( nafa 2014 ) , where highly resistant ( hr ) , resistant ( r ) , moderately resistant ( mr ) , and susceptible ( s ) ratings are represented by > 50 % , 3150 % , 1530 % , and < 14 % resistant plants , respectively . for this study , we lumped highly resistant and resistant varieties into a resistant category ( + ) and moderately resistant and susceptible varieties into a susceptible category ( )varietynematode resistanceaphid resistancepershing \n r(+)r(+)rugged \n mr()hr(+)vernal \n s()s()pgi-437 \n r(+)mr()sutter \n r(+)r(+)varieties examined in the greenhouse are indicated by ; field varieties are indicated by \n resistance ratings for alfalfa varieties ( nafa 2014 ) , where highly resistant ( hr ) , resistant ( r ) , moderately resistant ( mr ) , and susceptible ( s ) ratings are represented by > 50 % , 3150 % , 1530 % , and < 14 % resistant plants , respectively . for this study , we lumped highly resistant and resistant varieties into a resistant category ( + ) and moderately resistant and susceptible varieties into a susceptible category ( ) varieties examined in the greenhouse are indicated by ; field varieties are indicated by \n each of the four varieties was subjected to nematode treatments ( + , ) representing eight unique treatments each replicated 4 times for each of two trials ( n = 64 ) . \n our experimental unit was a microcosm , consisting of an 800-ml - capacity pot ( 8.9 8.9 8.9 cm ) filled with soil ( sunshine mix # 2 potting soil ) . \n each microcosm was seeded with 12 alfalfa seeds ( at standard field seeding rates ) , watered once a week , fertilized with 1 g of osmocote ( 10n-10p-10k ) , and maintained at 24/21 c day / night temperatures . \n after 2 wk of plant growth , each nematode treatment pot was inoculated with 30 field collected stem nematodes ( ditylenchus dipsaci ) . \n nematodes were extracted by chopping infested plant stems in water , and then collected with a pipette before transferring them to plants ( hanna and hawn 1965 ) . \n one week following nematode inoculation , each microcosm received 5 laboratory - reared ( 23 c ; 12 h l / d cycle ) aphids . \n aphids were confined using an inverted cup ( solo , 473-ml clear plastic cup , highland park , il , usa ) with mesh replacing the bottom and 40 cm sides ( vents on opposite sides ) of the cup . \n after 2 wk , aphids were counted , and alfalfa stems were clipped at the base from 13 cm tall vegetative stage plants and stored in the freezer to measure plant defense proteins at a later date ( see plant defense bioassays section below ) . \n a subsequent field experiment was conducted at utah state university s greenville research station in logan , ut , usa . we examined alfalfa plots in each of 3 yr ( 20112013 ) for the abundance of aphids and plant defense protein concentration on each of three alfalfa varieties . \n we selected vernal ( susceptible to nematodes and aphids ) , rugged ( resistant to aphids but not nematodes ) , and pershing ( resistant to nematodes and aphids ) . \n each of these varieties was planted in 5 replicate plots ( 1.5 3 m ; 15 plots total ) in a completely randomized design . \n plots within rows were separated by 1.2 m of fallow buffer , while adjacent plots were separated by 0.9 m of fallow buffer . \n a single variety was seeded in a plot at 9 kg seed per 4046.9 m. the option for including a nematode - only resistant variety in the field was limited because this variety had low winter dormancy , was a poor performer in our area , and therefore not included in our field experiment . \n it is important to note that stem nematodes have not been detected at this field site and are not prevalent in cache county , ut , usa . when alfalfa was ~30.5 cm tall , mesh sleeves ( tulle fabric ) were placed over four stems per plot and closed using a twist - tie . \n , plant stems from early - bud stage plants were clipped and immediately placed on ice to process aphid counts and for later analysis of plant defense protein concentration . \n leaves collected from stems of the field and greenhouse experiments were analyzed for levels of chitinase ( chi ) , peroxidase ( pod ) , polyphenol oxidase ( ppo ) , and trypsin inhibitor ( ti ) following modified methods from cipollini et al . \n pod , chi , and ppo activity were measured using a microplate reader ( biotek epoch , winooski , vt , usa ) . \n briefly , pod and ppo activity was determined following the oxidation of guaiacol and caffeic acid , respectively , for 1 min at 470 nm . \n chi activity in soluble protein extracts was examined by assessing the hydrolysis of p - nitrophenyl--n - acetylglucosaminide measured at 405 nm . \n ti activity was measured by examining the diffusion of protein extracts through a trypsin - containing agar followed by staining with n - acetylphenylalanine napthyl ester and o - dianisidine . \n ti concentration was determined using a standard curve made with soybean trypsin inhibitor , expressed as g trypsin inhibitor / g protein . \n briefly , foaming capacities of alfalfa plant tissues were measured by grinding 0.50 g ( greenhouse samples ) and 1.25 g ( field samples ) of alfalfa into 65 ml and 130 ml of distilled water , respectively , with a food blender for 2 min . \n the extract was poured into a graduated cylinder ( 100 ml for greenhouse samples and 250 ml for field samples ) . \n after the extract settled for 2 min , the cylinder was shaken vigorously for 10 sec to eliminate trapped air bubbles before the final amount of crude saponin level ( foam ) was recorded . \n the greenhouse microcosm experiments ( trial 1 and trial 2 ) were analyzed together within a 2 2 2 factorial design having aphid resistance ( + , ) , nematode resistance ( + , ) , and nematode application ( + , ) . \n aphid counts from the field experiment were analyzed with two factors , time ( for each of the 3 yr ) crossed with alfalfa variety ( vernal , rugged , and pershing ) . for greenhouse and field experiments , aphid counts were log transformed , and generalized linear model ( glm ) was used for analysis . \n glm was followed by tukey s post hoc test of variety given no significant interaction ( timevariety ) and a significant variety main effect . \n we also examined whether there was a correlation between aphid abundance and each plant defense protein concentration and nematode symptomatic plants . \n no relationship between aphid abundance and these factors was evident ( r < 0.11 ) and were not examined further . \n plant defense protein data from the greenhouse experiment were analyzed using a three - way anova with alfalfa variety , aphid treatment ( + , ) , and nematode treatment ( + , \n plant defense protein data from the field experiment were first analyzed using a three - way anova with time , alfalfa variety , and aphid treatment as factors . \n to better interpret time interactions for ppo , each year was analyzed separately using a two - way anova that included variety and aphid treatment as factors . \n significant differences in main effects were followed by tukey s post hoc tests and differences in interaction terms were examined with step - down bonferroni adjustments . \n analyses of aphid counts from the field study and greenhouse experiment were analyzed using systat ( version 13.0 ; spss , chicago , il , usa ) software ; plant defense protein data were analyzed using sas ( version 9.3 ; sas institute inc . , cary , nc , usa ) . \n at the start of the study , nematode treated pots showed symptoms of infection on 44 % of plants 9 se in the susceptible variety ( vernal ) , 33 % 6 in the nematode resistant variety ( pgi-437 ) , 48 % 8 se in the aphid resistant variety ( rugged ) , and 31 % 6 in the combined resistant variety ( sutter ) . in general , aphid abundance was lower when varieties had nematode resistance regardless of plants being resistant to aphids ( f1,55 = 25.669 , p < \n 0.001 ) ( fig . 1 ) . when plants were exposed to stem nematodes we found an increase in aphid abundance ( f1,55 = 8.234 , p = 0.006 ) . \n the effect of nematodes by variety was variable ; the percent increase of aphid abundance for the susceptible ( vernal ) , nematode resistant ( pgi-437 ) , aphid resistant ( rugged ) , and combined resistance ( sutter ) varieties were 84 % , 69 % , 88 % , and 20 % , respectively.fig . \n 1from the greenhouse experiment ( 20112012 ) , aphid counts on susceptible ( , ; vernal ) , nematode resistant ( + , ; pgi-437 ) , aphid resistant ( ,+ ; rugged ) , and combined aphid and nematode resistant ( + , + ; pershing ) varieties . \n values are means 1se from the greenhouse experiment ( 20112012 ) , aphid counts on susceptible ( , ; vernal ) , nematode resistant ( + , ; pgi-437 ) , aphid resistant ( ,+ ; rugged ) , and combined aphid and nematode resistant ( + , + ; pershing ) varieties . \n values are means 1se bioassays for each of the four defense proteins revealed variation among the varieties . \n the susceptible variety ( vernal ) had the highest level of chi ( variety : f3,45 = 15.03 , p < 0.001 ) and the variety ( rugged ) resistant to aphids had the highest level of ppo ( variety : f3,46 = 10.75 , p < 0.001 ; all tukey comparisons p < 0.02 ) . both of these varieties also had high levels of ti ( variety : f3,45 = 6.56 , p = 0.001 ) . \n alternatively , the variety ( sutter ) with combined resistance to nematodes and aphids had lower concentrations of ppo than the variety ( rugged ) resistant to aphids ( tukey p < 0.001 ) and chi ( fig . 2).fig . \n 2from the greenhouse experiment ( 20112012 ) , plant defense protein activity in susceptible ( , ; vernal ) , nematode resistant ( + , ; pgi-437 ) , aphid resistant ( ,+ ; rugged ) , and combined aphid and nematode resistant ( + , + ; sutter ) varieties . \n bar color indicates treatment type ( c = control , n = nematode application , a = aphids present , an = aphids and nematode application ) . \n capital letters that are different from each other represent significant ( p < 0.05 ) main effects of variety ( one letter per variety ) ; an asterisk indicates significant differences among treatments within a variety ; minus ( ) and plus ( + ) symbols indicate varieties that are different from the variety resistant to nematodes and aphids ( sutter ) when nematodes were not applied and when nematodes were applied , respectively . \n main effects of aphids ( for ppo , chi , and pod ) are not shown from the greenhouse experiment ( 20112012 ) , plant defense protein activity in susceptible ( , ; vernal ) , nematode resistant ( + , ; pgi-437 ) , aphid resistant ( ,+ ; rugged ) , and combined aphid and nematode resistant ( + , + ; sutter ) varieties . bar color indicates treatment type ( c = control , n = nematode application , a = aphids present , an = aphids and nematode application ) . \n capital letters that are different from each other represent significant ( p < 0.05 ) main effects of variety ( one letter per variety ) ; an asterisk indicates significant differences among treatments within a variety ; minus ( ) and plus ( + ) symbols indicate varieties that are different from the variety resistant to nematodes and aphids ( sutter ) when nematodes were not applied and when nematodes were applied , respectively . \n main effects of aphids ( for ppo , chi , and pod ) are not shown when plants were exposed to aphids , ppo concentrations decreased compared to plants without aphids ( aphid : f1,46 = 4.56 , p = 0.038 ) . \n similarly , we found a drastic decrease in chi for the susceptible variety ( vernal ) when exposed to aphids ( bonferroni p = 0.013 ) , but the magnitude of decrease by aphids for other varieties was small to non - existent ( variety : f3,45 = 15.03 , p < 0.001 ; aphid : f1,45 = 6.52 , p = 0.014 ; varietyaphid : f3,45 = 2.93 , p = 0.044 ) ( fig . 2 ) . \n the presence of aphids did , however , elevate ti in the combined resistant variety ( sutter ) ( bonferroni p = 0.056 ) , while aphids decreased ti concentrations in all other varieties , and apparently drove this interaction ( varietyaphid : f3,45 = 5.63 , p = 0.002 ) ( fig . 2 ) . \n nematodes did not alter ppo , ti , or chi levels among the varieties ( all main and interaction effects involving nematode : p > 0.05 ) . \n similarly pod concentration was not altered by nematodes for the aphid resistant variety ( rugged ) or combined resistant variety ( sutter ) . \n however , an interaction apparently driven by the presence of nematodes reduced pod in the susceptible variety ( vernal ) , and elevated pod concentrations in the nematode resistant variety ( pgi-437 ) ( variety : f3,45 = 6.14 , p = 0.001 ; aphid : f1,45 = 7.82 , p = 0.008 ; varietynematode : f3,45 = 3.07 , p = 0.037 ) ( fig . 2 ) . \n the susceptible variety had the lowest saponin levels ( variety : f3,4 = 7.13 , p = 0.044 ) but had similar levels to the combined resistant variety ( p = 0.159 ) , was not significantly different from the aphid resistant variety ( p = 0.082 ) , and was significantly different from the nematode resistant variety ( p = 0.039 ) ( fig . 3 ) . \n saponins were not affected by nematodes ( nematode : f1,12 = 2.40 , p = 0.147 ) or aphids ( aphid : f1,12 = 0.27 , p = 0.615).fig . \n 3from the greenhouse experiment ( 20112012 ) , crude saponin activity in susceptible ( , ; vernal ) , nematode resistant ( + , ; pgi-437 ) , aphid resistant ( ,+ ; rugged ) , and combined aphid and nematode resistant ( + , + ; sutter ) varieties . \n bar color indicates treatment type ( c = control , n = nematode application , a = aphids present , an = aphids and nematode application ) . \n different capital letters represent a significant ( p < 0.05 ) main effect of variety from the greenhouse experiment ( 20112012 ) , crude saponin activity in susceptible ( , ; vernal ) , nematode resistant ( + , ; pgi-437 ) , aphid resistant ( ,+ ; rugged ) , and combined aphid and nematode resistant ( + , + ; sutter ) varieties . \n bar color indicates treatment type ( c = control , n = nematode application , a = aphids present , an = aphids and nematode application ) . \n different capital letters represent a significant ( p < 0.05 ) main effect of variety changes in aphid abundance were dependent on the host variety , and these differences were constant over time ( time : f1,39 = 2.038 , p = 0.144 ; variety : f2,39 = 9.284 , p = 0.004 ; timevariety : f2,39 = 1.933 , p = 0.158 ) . \n a closer look revealed that the variety resistant to nematodes and aphids ( pershing ) had the lowest abundance of aphids in the field ( p = 0.049 ) ( fig . 4 ) . \n the aphid abundance was 25 % lower on the aphid resistant variety ( rugged ) than on the susceptible variety , however , this was not statistically significant ( p = 0.267).fig . \n 4from the field experiment ( 20112013 ) , aphid counts on susceptible ( white bar ; vernal ) , aphid resistant ( light grey bar ; rugged ) , and combined aphid and nematode resistant ( dark grey bar ; pershing ) varieties . \n different capital letters indicate significant differences ( p < 0.05 ) from the field experiment ( 20112013 ) , aphid counts on susceptible ( white bar ; vernal ) , aphid resistant ( light grey bar ; rugged ) , and combined aphid and nematode resistant ( dark grey bar ; pershing ) varieties . \n different capital letters indicate significant differences ( p < 0.05 ) a bioassay of ppo revealed that aphids induced elevated levels of ppo , but induction varied among varieties and years ( timevarietyaphids : f4,39 = 2.91 , p = 0.034 ) . \n this was contrary to the decreased ppo levels in the greenhouse experiment . to describe this interaction we analyzed each year separately . \n newly established alfalfa varieties ( 2011 ) with aphids showed consistent increases in ppo ( aphids : f1,8 = 0.06 , p = 0.065 ) ( fig . 5 ) . \n the response toward aphids , however , was not seen in the following year . in mature plants ( 2013 ) , \n an interaction between the presence of aphids and among varieties was apparently driven by the susceptible variety ( vernal ) having an elevated ppo concentration when aphids were present compared to the aphid resistant and combined resistant varieties where aphid feeding decreased ppo ( aphidsvariety : f2,12 = 10.44 , p = 0.002).fig . \n 5from the field experiment , plant defense activity in susceptible ( , ; vernal ) , aphid resistant ( ,+ ; rugged ) , and combined aphid and nematode resistant ( + , + ; pershing ) varieties . \n different capital letters represent a significant ( p < 0.05 ) main effect of variety ; different lowercase letters represent differences among treatments with the same shade bar ; an asterisk indicates a difference from the control within a variety . \n main effects of variety ( for pod ) and year ( for pod , chi , and ti ) are not shown from the field experiment , plant defense activity in susceptible ( , ; vernal ) , aphid resistant ( ,+ ; rugged ) , and combined aphid and nematode resistant ( + , + ; pershing ) varieties . \n different capital letters represent a significant ( p < 0.05 ) main effect of variety ; different lowercase letters represent differences among treatments with the same shade bar ; an asterisk indicates a difference from the control within a variety . \n main effects of variety ( for pod ) and year ( for pod , chi , and ti ) are not shown aphids did not appear to affect pod , chi , or ti concentrations in the field ( all main and interaction effects involving aphids : p > 0.05 ) . \n the susceptible variety ( vernal ) had higher concentrations of pod than the aphid resistant variety ( rugged ) ( variety : f2,12 = 3.91 , p = 0.049 ; tukey p = 0.045 ) . \n as varieties matured ( 3 year establishment ) , pod levels were elevated compared to newly established alfalfa plants ( time : f2,48 = 16.20 , p < 0.001 ; tukey comparisons p < 0.001 ) ( fig . 5 ) . \n induced responses to aphid were not evident , but in 2013 chi levels were at their highest compared to other years ( time : f2,44 = 37.47 , p \n in addition , in 2012 , ti was elevated in the combined resistant variety ( pershing ) compared to the other varieties ( time : f2,47 = 26.45 , p < 0.001 ; variety : f2,12 = 5.38 , p = 0.021 ; timevariety : f4,47 = 7.63 , p < 0.001 ; all bonferroni comparisons p < \n , the constitutive saponin defenses also were elevated in the resistant varieties compared to the susceptible variety , an effect we found in the greenhouse experiment . however , this was not witnessed in mature plants in 2013 ( time : f1,6 = 4.04 , p = 0.091 ; variety : f2,12 = 0.33 , p = 0.728 ; timevariety : f2,6 = 5.39 , p = 0.046 ) . upon further investigation of this interaction , in 2012 the saponin content was reduced in the presence of aphids , and this was not seen in mature plants in 2013 ( aphids : f1,12 = 2.78 , p = 0.121 ; timeaphids : f1,6 = 6.60 , p = 0.042 ; 2012 bonferroni comparisons p = 0.028 ) ( fig . 6).fig . \n 6from the field experiment , crude saponin activity in susceptible ( , ; vernal ) , aphid resistant ( ,+ ; rugged ) , and combined aphid and nematode resistant ( + , + ; pershing ) varieties . \n an asterisk indicates a difference from the control within a variety from the field experiment , crude saponin activity in susceptible ( , ; vernal ) , aphid resistant ( ,+ ; rugged ) , and combined aphid and nematode resistant ( + , + ; pershing ) varieties . \n nematodes can alter the performance of arthropods and other plant pathogens on shared host plants ( hawn and hanna 1967 ; wondafrash et al . \n this is in contrast to the studies that tested the effects of root - feeding nematodes on aphid abundance aboveground ( examples in mccarville et al . \n one of a few studies examining stem nematodes investigated the impact of stem nematodes on bacterial wilt ( corynebacterium insidiosum ) and found that nematodes also can increase the susceptibility of alfalfa varieties to this bacterium ( hawn and hanna 1967 ) . \n an example on melon plants shows that other plant pathogens also can enhance aphid abundance ( moran 1998 ) . \n what we know from root feeding nematodes is that their negative effect on aphids aboveground may be due to a decrease in the availability of amino acids in phloem sap ( bezemer et al . 2005 ) . in our study , we found that overall protein concentration increased in plants with stem nematodes ( mean 0.66 mg n / ml 0.09 se ) compared to the absence of nematodes ( mean 0.46 mg n / ml 0.09 se ) . \n we were careful not to confound protein from nematodes with plant protein by avoiding stem tissue where nematodes primarily reside and by selecting leaf tissue for assays . \n thus , our protein concentrations suggest that nitrogen based nutrients ( e.g. , amino acids ) play a role in nematode - insect interactions and may have contributed to increasing aphid abundances in this study . \n given the complexity of responses we found with the plant defense proteins , it is a worthy endeavor to evaluate how stem nematodes affect plant nutrition from root to shoot . \n plant parasitic nematodes have the ability to suppress sa and ja signaling pathways that suppress plant defenses ( haegeman et al . \n interestingly , isoflavonoids and other metabolites were not changed in shoots where stem nematodes reside but rather increased in concentration in the roots ( edwards et al . \n although root - lesion nematodes do not appear to affect isoflavonoids in alfalfa roots , they have been shown to increase phenylpropanoid and chitinase , among other mrnas important to plant defense in roots ( baldridge et al . \n we found an elevated pod concentration in leaves when a nematode resistant variety was exposed to nematodes . \n furthermore , nematodes were able to decrease the pod concentration in the leaves of the susceptible variety . \n the root accumulation of plant defenses as seen in these other studies may account for similar protein concentrations in leaves between nematode infected and non - infected alfalfa observed in our results . \n another factor to consider for the low detection of induction by nematodes is that the duration of plant defense changes by nematodes appears to be brief . \n plants infected with root - lesion nematodes had elevated concentrations of plant defenses within 12 h of infections , however , they were short lived , and the concentrations were decreased back to levels seen in susceptible varieties within 48 h ( baldridge et al . 1998 ) . \n alfalfa plants in our study were tested after 4 weeks of being exposed to nematodes . \n therefore , stem nematode effects on plant defense proteins may have occurred early on but were not captured because the effects diminish quickly . \n our selection of proteins , two of which are considered to be related to the sa ( chi and pod ) and ja ( ppo and ti ) pathways ( barto and cipollini 2005 ; and references therein ) , was to help tease apart competing sa and ja pathways and determine any tradeoffs related to defense toward pathogen and insect . \n although we did not see a decrease in plant defenses across varieties that would suggest the plant is more susceptible to aphids , nematodes may favorably alter nutrients and subsequent aphid feeding affected plant defense proteins . \n aphids divert nitrogen from the apical growth of alfalfa to feeding sites ( girousse et al . 2005 ) . \n only recently have we begun to understand the molecular bases of aphid resistance in plants . in medicago truncatula , for example , jasmonic acid has been implicated in r - gene mediated aphid resistance ( gao et al . \n more specifically , the akr gene imparts resistance toward the bluegreen aphid ( acyrthosiphon kondoi ) by elevating ja defenses ( gao et al . \n . however , there is much variation among aphid species , and distinctive medicago phenotypes are observed ( klingler et al . \n similarly , the tobacco aphid up - regulates ti in tobacco plants . across all alfalfa varieties , \n we found reduced ppo concentration with aphids , an effect encountered in sorghum where aphids down - regulate ppo ( smith and boyko 2006 ) . \n interestingly , we found this to be opposite when evaluating alfalfa plants in the field . \n pod was not affected by aphids in our study , but aphid feeding down - regulates pod in sorghum and arabodopsis , and is up - regulated in celery ( smith and boyko 2006 ) . \n given that aphids mostly decreased plant defense proteins in our study , aphids may be using various strategies to overcome alfalfa defenses ( see walling 2008 ) . in the variety resistant to nematodes and aphids and where we see aphid abundance to be lowest \n , we find increases in ti proteins when exposed to aphids , and this may in part aid in this variety being a less suitable host . \n the molecular mechanisms of plant defense for aphids are not fully understood but what is clear is that aphids in general activate multiple signaling pathways and that signaling is mediated by multiple compounds , as described above in several plant species ( smith and boyko 2006 and examples therein ) . \n general models of aphid - plant interactions according to host plant resistance have been described where aphid - resistant plants show gene - for - gene recognition and defense signaling early on , and resistant and susceptible plants recognize aphid - specific cell damage ( smith and boyko 2006 ) . \n sa signaling has been closely linked with aphids , but more specific loci have been identified in resistance of m. truncatula , a close relative of m. sativa , to the blue alfalfa aphid ( a. kondoi ) . in particular , a chromosome region \n neighboring resistance gene analogs has been mapped that encode the coiled - coil ( cc)-nbs - lrr resistance proteins ( klingler et al . \n more recently , jasmonate zim ( jaz ) was found to interact with jasmonate insensitive 1/myc2 and inhibit ja - responsive gene expression ( bari and jones 2009 ) . \n study of these specific genes and resistance regions is warranted to unravel the complexity of aphid - plant interactions and the inherent specificity among aphid species and their plant host . \n although saponin levels were not affected by nematodes or aphids , resistant varieties had higher saponin levels compared to the susceptible variety . \n there is evidence that high saponin levels negatively affect pea aphids by altering aphid probing behavior ( golawska 2007 ; golawska et al . \n high saponin levels however , were not important in the suppression of the spotted alfalfa aphid ( therioaphis maculate bruckton ) or root - knot ( meloidigyne hapla ) or stem nematodes ( d. dipsaci ) ( pedersen et al . 1976 ) . \n this may in part help explain the general resistance of pea aphid in our study toward both nematode and aphid resistant varieties . \n cross - resistance by other mechanisms has been seen in the soybean system where resistance toward the root nematode , heterodera glycines , reduces the performance of aphids , aphis glycines ( mccarville et al . \n increases in arthropod herbivore density in alfalfa have been shown to increase saponin levels ( pearson et al . \n saponins are a complex of many compounds and specific saponin compounds ( i.e. , zanhic acid tridesmoside and 3-glca , 28-ararhaxyl medicagenic acid glycoside ) appear to be more important in resistance than others ( golawska et al . \n it is possible that these particular saponins may be altered and may help explain the increased aphid abundance in the presence of nematodes but specialized bioassays for these compounds would be warranted . \n the mechanisms for pest resistance in alfalfa are complex given the level of genetic variation within and among varieties . in alfalfa roots , \n resistant varieties may be elevated in certain defenses compared to susceptible varieties , but other defense indicators such as chitinase mrnas were similar between resistant and susceptible varieties ( baldridge et al . \n indeed , we found elevated saponins in resistant varieties compared to the susceptible variety , but only particular varieties expressed elevated defensive proteins . \n the susceptible variety was elevated in chi and ti , and the aphid resistant variety was elevated in ppo and ti . \n overall , there appears to be a lack of consistency with any particular defense strategy , and perhaps unique combinations of defenses are expressed for each variety . \n for example , some species of aphids are susceptible to high saponin concentrations while other aphid species are not ( golawska , 2007 ; golawska et al . \n 1976 ) , yet the mechanism targets the mouthparts for which this is similar for aphids . \n alfalfa resiliency to respond to continual disturbances including routine harvest ( wounding ) periods in a season , herbivory , and environmental factors , may provide for more plasticity and perhaps unique methods for coping with these stressors in any given time . in our study , \n we found induced responses toward aphids for newly established plants and mature plants , but in 2012 induced responses were not evident . \n what we found , however , was that ti and saponin levels were elevated in this particular year . \n indeed , environmental factors ( e.g. , temperature , rainfall , drought , etc . ) can cause stresses in plants leading to induced defenses that may interact with responses to herbivores . \n we did not identify specific linkages with abiotic factors ( i.e. , temperature and relative humidity ) that would explain our results in the field but other environmental factors can not be ruled out . \n we were limited to only a single variety or two within a resistant variety type ( stem nematode resistant , pea aphid resistant , or resistance to both ) because of the lack of available commercial varieties with these particular resistance qualities . \n as we approach having a completed alfalfa genome available , we may better isolate particular genes and gene interactions involved in plant defense against pathogens and insects . \n it has long been known that alfalfa resistance is complex , and this also is evident in the classification of resistant varieties to particular phytophagous organisms . \n resistance is generalized at the population level where a highly resistant variety indicates that > 50 % of the population is resistant to the target organism . \n interestingly , in our study the aphid resistant variety maintained a high abundance of aphids in the field , yet the plant quality did not appear to be affected in the way the susceptible variety showed yellowing and other signs of damage ( pers . \n . therefore , some varieties classified as resistant may in fact be tolerant and still maintain high quality . \n resistance in alfalfa is unlike other plant systems ( e.g. , zea mays ) where resistance is more uniform within a variety . \n therefore , enhanced genomic resources will be critical tools for understanding resistance and the trade - offs to defend against multiple phytophagous organisms .", "role": "user" }, { "content": "plants are exploited by a diverse community of insect herbivores and phytopathogens that interact indirectly through plant - mediated interactions . \n generally , plants are thought to respond to insects and pathogens through different defensive signaling pathways . \n as plants are selected for resistance to one phytophagous organism type ( insect vs. pathogen ) in managed systems , it is not clear how this selection may affect community interactions . \n this study examined the effect of nematode - resistant varieties on aphid ( acyrthosiphon pisum ) suppression , and then determined how infection by the stem nematode , ditylenchus dipsaci , mediated ecological effects on aphids and on plant defense proteins . \n four alfalfa ( medicago sativa ) varieties were selected with resistance to nematodes only ( + , ) , aphids only ( ,+ ) , nematodes and aphids ( + , + ) , and susceptibility to nematodes and aphids ( , ) . field and greenhouse experiments were conducted to isolate the effect of nematode infection and aphid abundance on each variety . \n we found that varieties resistant to nematode , regardless of aphid resistance , had the lowest aphid counts , suggesting possible cross - resistance . \n aphid abundance , however , increased when plants were exposed to nematodes . \n resistant varieties were associated with elevated saponins but these compounds were not affected by insect or pathogen feeding . \n concentrations of peroxidases and trypsin inhibitors , however , were increased in nematode resistant varieties when exposed to nematodes and aphids , respectively . \n the patterns of plant defense were variable , and a combination of resistance traits and changes in nutrient availability may drive positive interactions between nematodes and aphids aboveground .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: skeletal muscle is the most abundant tissue in human body , except in obese patients , and is involved in several physiological functions . indeed , glucose uptake and metabolism take place primarily in the skeletal muscle , a tissue prone to adaptation in size by means of both hypertrophy and hyperplasia . \n the former relies on the regulation of protein synthesis and degradation rates , while the latter involves the myogenic process that is in charge of regulating myocyte turnover as well as of supporting the rapid regeneration following injury . \n the counterpart of muscle hypertrophy is muscle atrophy , even defined as sarcopenia , that naturally occurs in physiological conditions , such as aging . beyond aging , \n muscle wasting is a feature associated with several pathological states and chronic diseases such as immobilization following fractures or bed rest , malnutrition , cancer , chf , ckd , copd , burns , muscular dystrophies , aids , sepsis , and immune disorders . \n muscle depletion has important implications , exercise intolerance and inability to manage daily activities that eventually translate into poor quality of life . \n most of the above mentioned pathological conditions are associated with variable degrees of local and/or systemic chronic inflammation , an element that could play a relevant role in the onset of muscle wasting . \n indeed , inflammation is considered one of the diagnostic hallmarks of cachexia , a wasting condition that often occurs in chronic diseases . \n the aim of this review is to summarize the evidences supporting the role of inflammation , associated with several illnesses , in impairing muscle homeostasis and myogenesis , leading to muscle atrophy . \n skeletal muscle mass represents a determinant of physical performance , and muscle size varies according to physiological stimuli and pathological conditions that , in turn , modulate the activation state of signaling pathways involved in the control of protein turnover . \n muscle atrophy occurs when the balance between protein degradation and protein synthesis is poised towards degradation , leading to the loss of myofibrillar proteins and , consequently , to reduced fiber cross section area , finally resulting in impaired contraction ability and low force generation . \n muscle nitrogen balance is finely modulated by distinct agents , both intrinsic ( nutrient and energy availability , mechanical stress ) and extrinsic ( humoral mediators : hormones and cytokines ) . \n moreover , muscle wasting , beyond the loss of muscle mass , often determines a reduction of muscle quality , that is , specific force , as reported in patients with chf or cancer or admitted to the intensive care unit . \n protein breakdown in the skeletal muscle is mediated by two main degradation systems , the ubiquitin - proteasome and the autophagy pathways . \n the proteasome system preferentially targets short - lived proteins , and several reports considered the proteasome as the degradation machinery mostly involved in wasting processes of distinct origin . \n the autophagy system is in charge of degrading long - lived proteins and organelles ( mitophagy , pexophagy , etc . ) , and recent observations suggest that , beyond the proteasome , even autophagy plays a crucial role in muscle wasting . \n in addition to proteasome and autophagy dependent proteolysis , intact myofilaments were postulated to undergo a preliminary cleavage in order to be released from the myofibrils for the subsequent ubiquitin dependent degradation , and such activity was proposed to be carried out by calpains or by caspase-3 . \n the observations reported above prompted the idea that protein breakdown inhibition could be the right way to prevent disease - associated muscle wasting . \n indeed , proteasome inhibition proved effective only in experimental muscle unloading , while several reports show the detrimental effects of autophagy suppression [ 1214 ] . reasonably , \n both defective and excessive autophagy are deleterious by opposite mechanisms , namely , the lack of damaged protein / organelle removal and the exaggerated degradation , respectively . a distinct strategy in order to reduce muscle protein breakdown would be to target the muscle - specific ubiquitin ligases , since their activity represents the limiting step in determining both substrate - specificity and degradation rate . since the beginning of the 2000s , the discovery of muscle - specific e3 enzymes and the characterization of their substrates have emerged and are still growing . \n the first ones , mafbx / atrogin-1 and murf1/trim63 [ 15 , 16 ] , are actually the most commonly used read - out measurements for the molecular assessment of muscle protein catabolism , even though obvious limitations are implied . \n few years later , trim32 was shown to target myofibrillar components , but its role seems to be primarily related with muscular dystrophies rather than wasting processes . \n the role of fbxo40 , firstly characterized in denervation - induced atrophy in 2007 , was then clearly established by the identification of irs-1 as substrate , thus defining a negative feedback on the anabolic pi3k / akt axis . \n finally , fbxo30/musa1 was described as a bmp - regulated gene required for denervation- and fasting - mediated muscle loss . \n however , investigations in humans are still lacking and no evidence is actually available in order to validate the use of ubiquitin ligases as therapeutic targets for muscle wasting . \n muscle protein degradation systems are modulated by a coordinated network of signaling pathways activated or suppressed by hormones and cytokines ( figure 1 ) . on one hand , \n anabolic signals are activated by insulin , igf-1 , gh , and androgens , while catabolism is stimulated by a variety of proinflammatory cytokines as well as glucocorticoids and ros . \n igf-1 promotes muscle hypertrophy , while low igf-1 circulating levels have been associated with several muscle atrophy conditions . \n igf-1 , and similarly insulin , activates the pi3k / akt pathway , which promotes protein synthesis through mtor and its downstream effectors , mtorc1/2 complexes , not to mention that mtor activation in the skeletal muscle results in autophagy inhibition , thus determining at the same time both protein synthesis activation and inhibition of protein degradation . \n another igf-1-mediated anticatabolic action is due to akt phosphorylation / inhibition of the foxo transcription factor family , determining their inability to translocate to the nucleus and promote the expression of the ubiquitin ligases atrogin-1 and murf1 and autophagy genes . \n opposite to the igf-1 pathway , one of the most relevant inducer of muscle atrophy is myostatin , a member of the tgf- family . \n it signals through actriib , and recruiting the transcription factors smad2/3 leads to increased atrogin-1 and murf1 mrna levels . \n the negative regulation of muscle mass exerted by myostatin likely relies on the suppression of akt signaling . in the complex network of signals relevant to muscle homeostasis , \n the bmp pathway has been recently characterized , showing that the downstream activation of the transcription factors smad1/5/8 regulates a fundamental anabolic signal . in the same paper , \n the alternative activation of either the myostatin or the bmp pathway , both competing for smad4 recruitment , was demonstrated . \n mice , suggesting that the balance between these signal cascades is crucial for the modulation of muscle mass . \n finally , few signaling pathways , less characterized in the skeletal muscle , displayed their relevance to muscle mass regulation during wasting conditions . \n histone deacetylases 4 and 5 were shown to abrogate dach2 expression that in turn lead to myogenin increase and ubiquitin ligase accumulation . \n overexpression of atf4 was sufficient to induce muscle atrophy , regulating the expression of genes mainly related to cell growth suppression . \n similar to atf4 action , the transcription factor and tumor suppressor p53 was able to trigger muscle atrophy through the induction of the cyclin - dependent kinase inhibitor p21 . \n loss of muscle mass , a common feature of chronic diseases , is frequently associated with increased production of proinflammatory cytokines such as tnf- , il-1 , il-6 , and ifn-. for example , patients affected by chronic renal or heart failure show increased circulating levels of tnf- and tnf soluble receptors [ 30 , 31 ] . \n similarly , several proinflammatory cytokines are increased in cachectic cancer patients ( reviewed by ) . \n chronic inflammation may depend on increased expression of proinflammatory mediators but also on reduced levels of anti - inflammatory factors ; consistently , mice k.o . for il-10 , one of the best known anti - inflammatory cytokines , display weakness and accelerated muscle loss that can be improved by treatment with the anti - inflammatory , resveratrol rich , grape seed extract . \n in addition to cytokines , other factors are produced during the inflammatory response , such as the so - called acute phase proteins , markers of systemic inflammation . in this \n regard , cancer cachexia and the acute phase response appear to be correlated : the enhanced synthesis of acute phase reactants in the liver has been proposed to drive muscle protein hypercatabolism contributing to increase of the resting energy expenditure . of interest , \n acute phase proteins have been shown to be produced even by the skeletal muscle itself . \n proinflammatory cytokines , together with altered homeostasis of classical hormones , put on a complex network that results in inhibition of anabolic and/or anticatabolic signals ( see above ) , in favor of lipolysis and proteolysis . \n in particular , proinflammatory cytokines are well known to impinge on muscle protein metabolism . in this regard \n , data obtained in both experimental models and human pathology have demonstrated that systemic inflammation is associated with reduced rates of protein synthesis paralleled by enhanced protein breakdown , both accounting for the loss of muscle mass . however , the precise mechanism by which inflammation modulates protein turnover rates is still poorly investigated . \n data obtained in clinical studies show a variegated situation : both rates are markedly increased in severely burned patients , with the balance remaining in favor of degradation ; synthesis rates are maintained in face of enhanced degradation rates in critically ill septic subjects , while in cancer patients with cachexia , protein turnover rates have been described as increased , decreased , or unchanged ( reviewed in ) . \n the regulation of muscle protein metabolism by humoral factors is widely accepted . in this regard , the activity of both proteasome and lysosomes , the two proteolytic systems mainly involved in muscle depletion , is known to be affected by the hormonal and cytokine milieu . as an example , healthy animals exposed to proinflammatory cytokines such as tnf- , il-1 , or il-6 develop muscle wasting associated with increase of both ubiquitin expression and proteasome enzymatic activity ( reviewed in ) . \n consistently , few studies demonstrated in the past that cytokines play a crucial role in the onset of muscle wasting . indeed , \n muscle depletion , enhanced protein breakdown , and increased ubiquitin can be prevented by treating tumor - bearing animals with antibodies directed against il-6 , tnf- , or ifn- [ 4143 ] . \n proinflammatory cytokines contribute to muscle wasting also in chronic diseases of noncancer origin . indeed , \n increased circulating levels of tnf- , il-1 , and il-6 in sepsis appear to be correlated with disease severity and lethality . \n similarly , the proinflammatory shift occurring in aids patients likely accounts for muscle protein hypercatabolism , a feature frequently reported in these patients before the adoption of combined antiretroviral therapy . \n a recent report shows that muscle wasting in diabetic rats is associated with enhanced expression of tnf- , il-1 , and il-6 in the skeletal muscle and that such increase can be corrected by exercise training . \n finally , also sarcopenia and the loss of muscle quality that characterize aging are associated with high levels of proinflammatory mediators [ 46 , 47 ] . \n the effects exerted by proinflammatory cytokines on muscle mass are mediated , partially at least , by activating the transcription factor nf-b . \n the transcriptional activity is regulated by the phosphorylation and consequent degradation of the inhibitor i-b , allowing the positive regulation of murf1 and other atrophy related genes , including the inducible nitric oxide synthase . \n studies performed on experimental models suggest that the nf-b signaling is activated in skeletal muscle during cancer cachexia , and recently modulations of this transcription factor have been observed in gastric and lung cancer patients [ 50 , 51 ] , as well as in experimental models . \n another protein related to the tnf- cascade , traf6 , was shown to coordinate the activation of both proteasome and autophagy . among proinflammatory cytokines , tweak has been shown to induce muscle wasting mainly by stimulating proteasome - dependent proteolysis . \n tweak - induced downregulation of both pgc-1 expression and mitochondrial biogenesis has been proposed to mediate the effects on muscle mass . indeed , pgc1 hyperexpression protects against tweak - induced effects such as nf-b activation , increased ubiquitin ligase levels , and muscle wasting . \n in addition , several cytokines act on the jak / stat pathway , and muscle stat3 was demonstrated to induce atrogin-1 and to block autophagy , leading to muscle degeneration . \n proinflammatory cytokines act on muscle protein metabolism not only by activating catabolic pathways , but also by downregulating the anabolic ones . as an example \n , lipopolysaccharide - induced muscle wasting is associated with increased circulating levels of tnf- and il-1 that lead to inhibition of the akt / mtor signal transduction pathway [ 56 , 57 ] . in this \n regard , treatments able to restore akt physiological activity appear to counteract tnf- induced wasting [ 58 , 59 ] . \n recent observations show that cancer cachexia occurring in the apc ( min/+ ) mice also depends on the inhibition of mtor activation due to the high circulating il-6 levels . \n the antianabolic action of proinflammatory cytokines is partially exerted through interactions with the igf-1-dependent signaling pathway . \n indeed , tnf- leads to serine phosphorylation of irs-1 , inhibiting its recruitment to the insulin / igf-1 receptor . \n tnf- can impinge on the insulin / igf-1 signaling via direct interaction between the ikk complexes and irs-1 . alternatively , \n tnf-induced activation of jnk may play a role , as shown by the observation that the downregulation of the igf-1-dependent signaling exerted by the cytokine does not occur in the presence of a jnk inhibitor ( reviewed in ) . \n finally , proinflammatory cytokines may modulate anabolism also by inducing leucine - resistance , resulting in decreased mtor phosphorylation and reduced protein synthesis . \n in addition to modulations of protein synthesis and breakdown rates , several reports in the last years suggest that also alterations of the myogenic response may play a role in the maintenance of skeletal muscle mass in the adult , in both physiological and pathological states . \n myogenesis is the process that guarantees the generation of myoblasts to give rise to skeletal muscle tissue . \n embryonic myogenesis is definitely better understood thanks to the extensive work of developmental biologists that generate important genetic tools to establish the exact hierarchical activations of skeletal muscle transcription factors triggering the early embryonic process . in the adulthood , \n the situation is more complicated since those genetic tools are not sufficient to identify the major key players involved in adult myogenesis . \n skeletal muscle injuries are extremely common and mainly caused by intensive muscle exercise , trauma , laceration , burns , freezing , and toxin exposure ( the latter are also commonly used in experimental models of muscle regeneration ) . \n these insults result in muscle injury that determines a diffuse degeneration followed by the induction of regeneration . \n however , the characteristics of regeneration have been shown to differ according to the type of injury ; thus a direct comparison of the results obtained in various studies is extremely difficult . \n when skeletal muscle is damaged , muscular fiber degeneration is compensated by the regeneration of new fibers formed at the expense of resident myogenic satellite cells localized underneath the basal lamina of muscle fibers . \n skeletal muscle regeneration is mainly sustained by scs [ 64 , 65 ] and this is critical for chronic muscle diseases including muscular dystrophies . in mds , dystrophic scs share the same molecular defect and the newly formed fibers during regeneration cycle are susceptible to degeneration . with time , the reservoir of satellite cells is totally consumed and the muscle tissue is progressively replaced by connective tissue . in the adulthood , skeletal muscle represents half of the body weight , and scs are able to maintain their functionality thanks to the generation of muscle precursor cells able to proliferate and , upon fusion , to generate new fibers . all trunk and limb muscle source cells are originated from embryonic somite source , with exception of the head muscle . \n pax3 precursors were identified in the embryonic dorsal aorta ; they are able to give rise to both smooth and skeletal muscle cells , suggesting a common origin in the two muscle lineages . \n oxidative - slow muscles contain a relatively high number of scs , up to six times more than fast - glycolytic muscles . \n scs can be easily isolated after enzymatic digestion and/or physical trituration or by facs for specific surface markers including cxcr4 , 1-integrin , sca-1 , m - cadherin , syndecan-4 , notch-1 , and ncam / cd56 [ 7072 ] . \n it is relevant to note that sc myogenic potential seems to be diverse according to the markers considered for cell sorting , thus revealing heterogeneity in sc populations . \n pax7 is a transcription factor considered as a biomarker for quiescent and proliferating scs , while jagged-1 is considered a marker of activated scs . \n the expression of pax7 , ncam , and c - met has been shown also in human scs . \n numb is an inhibitor of the notch signaling but also a cell fate determinant and was found asymmetrically distributed in a sc subpopulation , suggesting that only a small subset of scs retains the stem cell characteristic and undergoes asymmetric division . there are still pending questions regarding sc isolation and characterization . \n although encouraging results have been obtained in preclinical [ 76 , 77 ] and clinical studies , the use of scs for the treatment of muscle degeneration is hampered by the inability of scs to pass the endothelial barrier when injected systemically . \n further work is needed to confirm and improve the therapeutic efficiency of sc autologous injection for skeletal muscle degeneration . \n bone marrow cells , including mscs , blood , and muscle - derived cd133 and sp cells , have been also implicated in skeletal muscle regeneration . \n however , in some cases , engrafted cells failed to express skeletal muscle proteins , suggesting that under standard conditions they fuse rather than differentiate to skeletal muscle potency . \n in other cases , results have been more encouraging . in bone marrow and peripheral blood , stem cells characterized by the expression of the cd133 antigen \n are present and have been shown to give rise to dystrophin - positive fibers following their intramuscular transplantation . \n sp cells are referred to as a small subpopulation of stem cells able to exclude hoechst 33342 dye and participate to adult myogenesis . \n related to the sp populations are the cd34/cd45 cells , known as sk-34 cells that are apparently derived from cd34/cd45 , named sk - dn cells . \n being different from the other myogenic stem cells , they still retain the ability to differentiate into vascular cells , including pericyte , endothelial cell , and smooth muscle cells and peripheral nerve cells as schwann and perineurial cells . \n interstitial cajal - like cells or telocytes are recently discovered c - kit cells type populating the muscle interstitium . \n these cells own a small body ( 915 um ) and a certain number of telopodes organized in network to maintain tissue homeostasis and renewal through exosome delivery . in the interstitium among the fibers are usually present several cell types that were also showed to contribute to adult myogenesis . whether these cells are missing or altered in pathological muscles and their origins are still heavily debated . \n emerging evidences highlight that intramuscular adipocytes and fibrocytes are the differentiated stages of faps [ 8486 ] . \n faps are isolated as cd34/sca-1 or as sca-1/cd140a and they are able to differentiate into myoblasts . \n these cells mediate the ability of hdac inhibitors to promote skeletal muscle regeneration in mdx mice , animal model for dmd . \n the key cytokines and growth factors responsible for their paracrine positive effects are still under investigation , and more information regarding the human counterparts is necessary for translational clinical implications . in murine models were identified a particular class of fibroblasts expressing the transcription factor 7-like 2 ( tcf / l2 or tcf4 , ) . \n these tcf4 fibroblasts present in the connective tissue seem also to regulate muscle fiber generation and as such they could be directly related to faps . \n pw1 is a zinc - finger - containing transcription factor expressed in myoblasts , and it seems as an important marker for myogenic progenitor cells since postnatal muscle growth is severely impaired in mice lacking pw1 in myogenic lineages . \n muscle - derived stem cells are also identified in the skeletal muscle interstitium based on the expression of flk1 , sca-1 , and desmin . \n since they are able to differentiate into myogenic , adipogenic , osteogenic , chondrogenic , and even hematopoietic lineages , they are strictly associated with mesoangioblasts and pw1 positive cells although comparative studies are still missing . \n noncanonical progenitors of mesodermal tissues were originally isolated from murine dorsal aorta and for their multipotent characteristic to give rise to mesodermal cell types in vitro and in vivo they were mesoangioblasts . \n mabs express cd34/c - kit / flk-1 but are negative for nkx2.5/myf5/oct4 and are able to give rise to multiple mesodermal lineages in vitro and in vivo . \n in the adult muscles , mabs are usually isolated and cloned using their pericyte markers , alkaline phosphatase , sca-1 , ng2 proteoglycan , cd140a , and cd140b [ 9499 ] . \n mabs are able to differentiate into myogenic , osteogenic , chondrogenic , and adipogenic lineages [ 97 , 98 ] . \n also human mabs display pericyte markers , as cd146/cd140b1/ng2 , but are negative for hematopoietic or sc markers cd45/cd34/cd56/cd144/pax7 [ 96 , 100 ] . since a few markers are shared between human pericytes and mscs ( cd10/cd13/cd44/cd73/cd90 ) , the origin and the interaction between those multipotent stem cells are still matter of debate [ 100 , 101 ] . \n interestingly , a comparison among mscs , mabs , and multipotent adult progenitor cells resulted in specific differention / functional properties that can be partially converted in vitro by culture conditions . \n in addition , notch signaling seems to have a primary role in modulating the myogenic potential of mscs and mabs . \n stem cell biology in adult myogenesis is a very active and fast moving field , and probably some redundancies in stem cell type and function observed here will be explained with further comparative studies . \n there is a clear need for more basic research to better understand the interconnected roles of stem cells in skeletal muscle regeneration and to explore the integration of signaling pathways such as notch and bmp . \n this new information not only will improve our understanding of the adult myogenesis process but also in principle could reveal potential therapies for the enhancement of tissue repair in acute and chronic skeletal muscle degenerations . \n changes in cellular composition of muscle microenvironment are crucial for metabolic modifications occurring during both acute damage with consequent muscle regeneration and chronic degeneration . \n overall , the pathophysiological alterations occurring during the onset and the progression of muscle diseases highlight the complexity of the possible interactions among muscle resident / recruited cells and immunological mediators ( figure 2 ) . \n the modulation that takes place during acute events , such as muscle trauma inducing regeneration after temporary atrophic conditions , is different from the one occurring during chronic events , such as long lasting inflammatory processes affecting muscle during genetic diseases ( mds or myopathies ) , cancer - induced muscle atrophy , and sarcopenia . \n the inflammatory process during trauma or fractures is a controlled and finely regulated event that through different and defined stages can ensure a complete and efficient reconstruction of muscle fibers . \n the process begins with the release of chemoattractant molecules like desmin but also heat shock proteins and hmgb1 among many others , responsible for local activation of the innate immune response and comprehensively recognized as damage associated molecular patterns . \n muscle injury is usually followed by a local increase in myeloperoxidase activity that reflects neutrophils activation . \n their contribution to skeletal muscle regeneration and myofiber remodeling relies on the oxidative and proteolytic modification of damaged tissue , to allow phagocytosis of cellular and matrix debris . \n interestingly , targeted ablation of cd11b cells ( including neutrophils and monocytes / macrophages ) reduces muscle fiber repair after muscle injury . \n moreover , neutrophils release proinflammatory cytokines such as tnf- , il-1 , ifn- , tgf-1 , and il-12 that reach a peak of concentration 24 h after injury and can modulate the regenerative process in skeletal muscle . \n recent findings show that il-4 production mainly due to eosinophils could play a role during early stages of muscle regeneration . \n myogenic cells attract monocytes from the blood stream to their location next to capillaries . indeed \n , this privileged position could be the reason for easy access and activation through chemotactic molecules such as mcp-1 , macrophage - derived chemokine , fractalkine ( cxcl1 ) , vegf , and the urokinase system . \n however , in vivo studies have reported that monocytes / macrophages recruited into skeletal muscle after injury could come from the connective tissue surrounding muscle / epimysium , where these cells accumulate before invading muscle tissue in the site of damage . \n notably , already twenty years ago , some studies reported that the activation of m1 macrophages ( cd68/cd163 ) at days 1 - 2 after injury was concomitant to activation and proliferation of scs . \n by contrast , m2 macrophages ( cd68/cd163 ) reach the peak of concentration closely to the surface of regenerating myofibers just 4 days after injury , when muscle differentiation is starting . \n consistently , different in vitro studies showed that scs increase their proliferation rates when cocultured with m1 macrophages , while they enhance both their fusion index and myogenin expression in the presence of m2 cells . \n muscle regeneration in mice induced by cardiotoxin injection was impaired after treatment with neutralizing antibodies against macrophage colony stimulating factor , a cytokine able to activate macrophages . \n m1 to m2 transition , timing , and correct sequence have been demonstrated to be essential for both resolution of inflammation and myofiber repair . indeed , il-10 administration after muscle injury leads to m1 disruption and m2 promotion , resulting in reduced myofiber growth and regeneration . \n ablation of il-10 in vivo can disturb the transition from m1 to m2 macrophages and decrease fiber growth . when cocultures of muscle cells with macrophages activated through il-10 to the m2 phenotype are stimulated to proliferate , no effect on myod or myogenin expression \n is observed , showing that m2 macrophages promote the early , proliferative stage of myogenesis . \n anyway , the window where il-10 can promote regeneration is short and if not respected can induce a delay in myogenic program [ 116 , 118 ] . \n moreover , macrophage - derived il-10 seems as well able to promote skeletal muscle differentiation of mabs , vessel derived myogenic precursors , and il-10 antibodies can effectively decrease the capabilities of these cells to participate in myofiber commitment . as il-10 , \n other mediators such as tnf- and ifn- are produced in the muscle microenvironment and play a role during muscle myogenesis . as an example , tnf- can impair regeneration , keeping scs in the proliferation stage and inhibiting differentiation , mainly by acting on nf-b and myod [ 119 , 120 ] . however , a second important role for tnf- is related to myofiber release of activin - a through both nf-b and p38 mapk , activating the phosphorylation of smad2/3 to inhibit sc differentiation and fusion . \n m1 macrophages produce high levels of activin - a that , in vitro , can act both on the further polarization of m1 macrophages and on the inhibition of anti - inflammatory il-10 release from m2 cells . \n suppression of p38 mapk signaling is usually followed by th2 cytokine elevation ( il-10 and tgf- ) , suggesting the switch in macrophage phenotype during skeletal muscles regeneration . \n the mechanisms through which ifn- can directly influence muscle regeneration are still poorly understood and , so far , the knowledge is limited to some in vitro data , reporting a delay in proliferation , impaired fusion , and low expression of terminally differentiation markers when this cytokine is added to myoblasts cultures [ 123125 ] . \n all of these data confirm the vision of skeletal muscle as an endocrine organ able to secrete specific myokines endowed with both paracrine and endocrine functions . indeed , \n leukemia inhibitory factor and il-15 are usually mentioned among the paracrine effectors , while myokines , such as myonectin , il-6 , irisin , calprotectin , and oncostatin m , once secreted from muscle tissue , especially during contraction but also in pathologic conditions , are able to induce anti - inflammatory cytokines ( il-1 receptor antagonist and il-10 ) . despite a transient and \n finely tuned activation of the inflammatory system required to sustain muscle repair after injury , chronic inflammation can be deleterious , driving uncontrolled wound healing and fibrosis , as well as triggering muscle wasting . \n for these reasons , systemic administration of immune - modulators can reveal beneficial effects , particularly in the case of anti - inflammatory drugs able to reduce m1-macrophages , as shown , for example , in dmd patients . \n interestingly , treatment with cyclosporin a , well - known immunosuppressant drug , was found to have multiple beneficial effects on the myopathic and mitochondrial phenotype of collagen vi a1-null mice , recovering muscle strength and , recently also , increasing pax7 pool and stimulating the formation of new myofibers [ 136138 ] . \n this effect may be exerted through an indirect regulation of the inflammatory state that occurs during muscle regeneration . \n all of these findings have been as well supported by the increased muscle regeneration observed in ullrich patients undergoing csa treatment , pointing on one side to immunosuppressive drugs as potent inducer of myogenesis and on the other side underlining the involvement of immune system with myogenesis , at least for these myopathies . \n administration of neutralizing antibodies against tnf- to mdx mice reduced p38-dependent inflammation , increased pax7 cells , and impaired the growth of regenerating fibers , suggesting the occurrence of an epigenetic link among inflammation , activated p38 mapk , and pax7 expression from scs during regeneration . \n further studies are needed to understand the function of scs and their possible induction to differentiate as a means to counteract muscle atrophy . \n increased pax7 and decreased myogenin levels have been reported also in the cachectic muscle of mice bearing the c26 carcinoma and in cancer patients , opening the possibility that cancer - driven inflammation induces muscle atrophy , dysregulating sc differentiation program [ 52 , 141 , 142 ] . \n sc accumulation was demonstrated to be due to an increase in the erk mapk signaling able to maintain the cells in an undifferentiated state . \n treatment with a chemical inhibitor of erk phosphorylation can indeed rescue the levels of pax7 and myogenin . \n lately , this process of sc accumulation has been pointed as contributing to muscle atrophy and has been related to nf-b , known transcription factor able to be activated by many proinflammatory cytokines . in the skeletal muscle of cachectic mice , nf-b is able to sustain pax7 expression , and moreover , in the same study , the authors revealed that other muscle precursors , such as mscs , participate in muscle wasting since they can not complete their myogenic commitment . \n chronic injury and proinflammatory cytokines due to tumor progression could be responsible for differentiation program failure . when mesenchymal progenitors are missing or are unable to display their prodifferentiation effects on scs , these latter ones actually contribute to fibrosis in mdx mice , leading to pathogenic effects in mds . in the last years \n , cells expressing cd34/sca1/pdgfra and not related with other lineages , such as hematopoietic , endothelial , or skeletal muscle , have received more and more attention . \n indeed , these cells can both differentiate in vitro and in vivo towards fibroblast and adipocytes producing asma and perilipin , respectively [ 84 , 143 ] . \n recently , suppression of fibroadipogenic phenotype of mesenchymal cells through hdac inhibitors has been demonstrated to induce the myogenic transcriptional activity in young mice by upregulating myod . on the contrary , \n faps taken from old mice fail in the activation of promyogenic phenotype mainly because of hdac inhibitor resistant . \n these last results already suppose a different behavior of the microenvironment that is actually acting on the myogenic program of scs in the elderly . aging of skeletal muscle \n indeed , geriatric scs show reduced proliferation and differentiation potential and can easily switch from a quiescent to a senescent state . \n some studies have already focused on the importance of the microenvironment where scs are studied , since if old murine scs are exposed to a young environment or growth factors , their ability to proliferate and differentiate is partly restored , suggesting the extreme importance the environment has on the single cells and on the other side the plasticity cells can have . during aging , \n extrinsic factors can alter sc functions , starting from the niche where they lay , since fibrous connective tissue is usually increasing [ 147 , 148 ] . \n in addition , scs in the elderly were shown to convert to a fibrogenic lineage mainly due to humoral factors . in particular \n , this lineage conversion seems to depend on wnt signaling . lately , a comparison between old and young scs identified an increased expression in jak - stat pathway with aging that could avoid their differentiation in old conditions . \n this pathway is of particular importance in chronic inflammatory conditions , since infiltration of inflammatory cells and increased circulating levels of proinflammatory cytokines ( such as tnf- ) can have together a detrimental effect on skeletal muscle regeneration . \n moreover , this pathway has usually a pivotal role in transduction of extracellular signals from cytokines and growth factors with particular importance for inflammatory cells . \n interestingly , in a recent work , murine and human scs were demonstrated to progress in their differentiation thanks to stat3 and a fine regulation of the activation of this protein could interfere with myogenesis . \n the authors indeed speculate that chronic degenerative stimuli could favor this prodifferentiative pathway leading to exhaustion of the sc pool . indeed , \n in chronic regenerative conditions , a pharmacologic inhibition of stat3 could have a therapeutic relevance . \n inflammation is a common trait of several pathological conditions characterized by the loss of muscle mass . during the past \n , a direct link was established among proinflammatory cytokines , modulation of intracellular signaling pathways , and protein breakdown . in recent years , \n an additional hypothesis , suggesting the impairment of the myogenic program as underlying cause of muscle atrophy , is becoming popular . in this line , \n the importance of a finely orchestrated balance between pro- and anti - inflammatory cytokines in regulating physiological myogenesis is a well - established concept , while data suggesting the relevance of inflammation in impaired myogenesis are growing . \n inflammation likely contributes to muscle depletion by both enhancing protein breakdown and impairing myogenesis in parallel and no priority between the two processes can actually be identified . \n the other way round , an effective strategy aimed at counteracting muscle wasting should take into consideration not only anabolic / catabolic aspects but analogously the continuous involvement of the myogenic counterpart . \n in addition , the role of interstitial and circulating progenitors involved in myogenesis and paracrine effects is also critical to modulate inflammatory responses in muscle wasting conditions . \n the adoption of anti - inflammatory agents for the treatment of chronic wasting diseases has been widely described and is not the topic of the present review ; however , the modulatory effect on inflammation exerted by exercise training deserves a short consideration . \n indeed , combined endurance and resistance training in elder subjects resulted in decrease of proinflammatory cd14/cd16 monocytes and low levels of tnf- production in vitro . \n moreover , circulating il-10 and regulatory t - cells ( cd4/cd25/cd127 ) increase in well - trained athletes with respect to a sedentary age matched population , even in resting conditions . keeping in mind the above - mentioned anti - inflammatory effect and considering \n that exercise is the most physiological stimulus able to coordinate both myogenesis and muscle hypertrophy , the adoption of patient - tailored exercise protocols will potentially have impact on muscle wasting associated with distinct pathologies .", "role": "user" }, { "content": "skeletal muscle mass is subject to rapid changes according to growth stimuli inducing both hypertrophy , through increased protein synthesis , and hyperplasia , activating the myogenic program . \n muscle wasting , characteristic of several pathological states associated with local or systemic inflammation , has been for long considered to rely on the alteration of myofiber intracellular pathways regulated by both hormones and cytokines , eventually leading to impaired anabolism and increased protein breakdown . \n however , there are increasing evidences that even alterations of the myogenic / regenerative program play a role in the onset of muscle wasting , even though the precise mechanisms involved are far from being fully elucidated . \n the comprehension of the links potentially occurring between impaired myogenesis and increased catabolism would allow the definition of effective strategies aimed at counteracting muscle wasting . \n the first part of this review gives an overview of skeletal muscle intracellular pathways determining fiber size , while the second part considers the cells and the regulatory pathways involved in the myogenic program . in both parts are discussed the evidences supporting the role of inflammation in impairing muscle homeostasis and myogenesis , potentially determining muscle atrophy .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: three transgenic lines that overexpress nf - l ( line 58 ) , nf - m - c50 ( line 42 ) and nf - h ( line 35 ) , have been produced and characterized in detail previously ( monteiro et al . \n , 1990 ; xu et al . , 1993 , 1996 ; wong et al . , 1995 ; marszalek et al . , 1996 ) . \n all three lines express their transgene products at the highest level in spinal cord motor neurons . to produce mice overexpressing two nf subunits simultaneously , the three lines were intercrossed and transgenic mice carrying two transgenes : nf - l and nf - m , nf - l and nf - h , or nf - m and nf - h were determined by southern blot as described previously ( xu et al . \n mice were killed at 17 d after birth and spinal cords were dissected and frozen immediately on dry ice . \n a piece of tail was saved from each mouse and used for transgene typing as described above . \n mice were fixed by transcardial perfusion using 4% paraformaldehyde in 0.1 m phosphate buffer ( ph 7.4 ) . \n a 0.3-cm segment of the lumbar enlargement of the spinal cord , centered at the attachment site of l5 ventral roots , was dissected out , immersed in 3% potassium bichromate and 1% osmium tetroxide for 3 d , and then stained with silver nitrate for 48 h. 70-m transverse sections from the stained tissue was cut using a vibratome ( tpi inc . \n spinal cords that contained more than seven clearly stained motor neurons were further analyzed . \n motor neurons located within the anterior horn , with their cell bodies intact and larger than 30 m in diameter , and with all the neurites clearly traceable , were drawn using a nikon drawing tube at 300 magnification . \n an equal number of neurons were then randomly selected from all the neurons drawn from each animal and measured using metamorph software ( universal imaging co. , west chester , pa ) . \n the number of neurons measured are 33 from 3 wild - type mice , 32 from 4 h mice , 21 from 3 m mice , 33 from 3 l mice , 20 from 2 hm mice , 20 from 2 lh mice , and 22 from 2 lm mice . \n dendritic segment : the length of a dendrite between two branch points , between the cell body and the first branch point ( primary dendrite ) , or the terminal segment that no longer branches . \n the number of dendritic trees in a neuron equals the number of primary dendrites . \n segment ratio : the ratio between the number of dendritic segments in the n + one order and that in the n order per neuron . in these measurements , axons were not excluded because it was not possible to differentiate the axon from the dendrites in every neuron with full confidence . \n however , the inclusion of axons in the measurements of neurons with numerous and long dendrites was not expected to influence the results significantly because each neuron contains only one axon and the length of the axon is limited by sectioning . for neurons with significantly shortened dendrites , \n the contribution of an axon could be significant , and therefore , the measured dendritic length in these neurons would bias towards an overestimation . \n this would tend to diminish the difference between the wild - type and transgenic neurons . despite this tendency \n , statistically significant differences were found between the wild - type and several transgenic lines ( see results ) . to measure dendritic diameter and tapering , \n five neurons randomly selected from the above motor neuron pool were further drawn using 1,000 magnification . \n the diameters at both ends of each dendritic segment and the length of the dendritic segments were measured from the drawings . \n the proximal end of the primary dendrite was determined using the following criteria : the transitions from cell body membrane to dendrite membrane were located at points at which the membrane changed from convex out to concave out . \n a smooth convex arc consistent with the rest of the soma outline was drawn between the points and the midpoint of the arc was defined as the origin of primary dendrite ( see chen and wolpaw , 1994 ) . \n the average diameter of a segment was calculated as the average of the proximal and distal diameters . \n dendritic tapering was the difference between the initial diameter and the end diameter of a dendritic segment per 100 m of segment length . \n rall 's ratio was calculated from ( [ddaughter])/ ( dparent ) using the average diameter of dendritic segments . \n all statistical comparisons were carried out using wilcoxon two - sample rank sum test . \n for em , mouse tissues were initially fixed by transcardial perfusion with a solution of 0.1 m sodium phosphate ( ph 7.4 ) , 4% paraformaldehyde , 2.5% glutaraldehyde , followed by further fixation in the same fixative for 24 h at 4c . \n tissues were then dissected out , postfixed with 2% osmium tetroxide in 0.1 m phosphate buffer , dehydrated in a graded ethanol series , and embedded in epon - araldite resin . \n thin sections were stained with uranyl acetate and lead citrate , examined and photographed in an electron microscope ( model em301 ; philips electron optics , mahwah , nj ) . \n animals were perfused as described above using 4% paraformaldehyde in 0.1 m phosphate buffer . \n sections were stained overnight at 4c using a monoclonal antibody against map2 ( a gift from dr . \n r. vallee ; de camilli et al . , 1984 ) and a rabbit polyclonal antibody against nf - l ( xu et al . , 1993 ) . \n mouse igg ( jackson immunoresearch laboratories inc . , west grove , pa ) were used as secondary antibodies . \n the doubly stained sections were examined with a scanning laser confocal microscope ( model 1024 ; bio - rad laboratories , hercules , ca ) . \n three transgenic lines that overexpress nf - l ( line 58 ) , nf - m - c50 ( line 42 ) and nf - h ( line 35 ) , have been produced and characterized in detail previously ( monteiro et al . \n , 1990 ; xu et al . , 1993 , 1996 ; wong et al . , 1995 ; marszalek et al . , 1996 ) . \n all three lines express their transgene products at the highest level in spinal cord motor neurons . to produce mice overexpressing two nf subunits simultaneously , the three lines were intercrossed and transgenic mice carrying two transgenes : nf - l and nf - m , nf - l and nf - h , or nf - m and nf - h were determined by southern blot as described previously ( xu et al . \n mice were killed at 17 d after birth and spinal cords were dissected and frozen immediately on dry ice . a piece of tail was saved from each mouse and used for transgene typing as described above . \n mice were fixed by transcardial perfusion using 4% paraformaldehyde in 0.1 m phosphate buffer ( ph 7.4 ) . \n a 0.3-cm segment of the lumbar enlargement of the spinal cord , centered at the attachment site of l5 ventral roots , was dissected out , immersed in 3% potassium bichromate and 1% osmium tetroxide for 3 d , and then stained with silver nitrate for 48 h. 70-m transverse sections from the stained tissue was cut using a vibratome ( tpi inc . , st . \n spinal cords that contained more than seven clearly stained motor neurons were further analyzed . \n motor neurons located within the anterior horn , with their cell bodies intact and larger than 30 m in diameter , and with all the neurites clearly traceable , were drawn using a nikon drawing tube at 300 magnification . \n an equal number of neurons were then randomly selected from all the neurons drawn from each animal and measured using metamorph software ( universal imaging co. , west chester , pa ) . \n the number of neurons measured are 33 from 3 wild - type mice , 32 from 4 h mice , 21 from 3 m mice , 33 from 3 l mice , 20 from 2 hm mice , 20 from 2 lh mice , and 22 from 2 lm mice . \n dendritic segment : the length of a dendrite between two branch points , between the cell body and the first branch point ( primary dendrite ) , or the terminal segment that no longer branches . \n . segment ratio : the ratio between the number of dendritic segments in the n + one order and that in the n order per neuron . in these measurements , axons were not excluded because it was not possible to differentiate the axon from the dendrites in every neuron with full confidence . \n however , the inclusion of axons in the measurements of neurons with numerous and long dendrites was not expected to influence the results significantly because each neuron contains only one axon and the length of the axon is limited by sectioning . for neurons with significantly shortened dendrites , \n the contribution of an axon could be significant , and therefore , the measured dendritic length in these neurons would bias towards an overestimation . \n this would tend to diminish the difference between the wild - type and transgenic neurons . despite this tendency \n , statistically significant differences were found between the wild - type and several transgenic lines ( see results ) . to measure dendritic diameter and tapering , \n five neurons randomly selected from the above motor neuron pool were further drawn using 1,000 magnification . \n the diameters at both ends of each dendritic segment and the length of the dendritic segments were measured from the drawings . \n the proximal end of the primary dendrite was determined using the following criteria : the transitions from cell body membrane to dendrite membrane were located at points at which the membrane changed from convex out to concave out . \n a smooth convex arc consistent with the rest of the soma outline was drawn between the points and the midpoint of the arc was defined as the origin of primary dendrite ( see chen and wolpaw , 1994 ) . \n the average diameter of a segment was calculated as the average of the proximal and distal diameters . \n dendritic tapering was the difference between the initial diameter and the end diameter of a dendritic segment per 100 m of segment length . \n rall 's ratio was calculated from ( [ddaughter])/ ( dparent ) using the average diameter of dendritic segments . \n all statistical comparisons were carried out using wilcoxon two - sample rank sum test . \n for em , mouse tissues were initially fixed by transcardial perfusion with a solution of 0.1 m sodium phosphate ( ph 7.4 ) , 4% paraformaldehyde , 2.5% glutaraldehyde , followed by further fixation in the same fixative for 24 h at 4c . \n tissues were then dissected out , postfixed with 2% osmium tetroxide in 0.1 m phosphate buffer , dehydrated in a graded ethanol series , and embedded in epon - araldite resin . \n thin sections were stained with uranyl acetate and lead citrate , examined and photographed in an electron microscope ( model em301 ; philips electron optics , mahwah , nj ) . \n animals were perfused as described above using 4% paraformaldehyde in 0.1 m phosphate buffer . \n sections were stained overnight at 4c using a monoclonal antibody against map2 ( a gift from dr . \n r. vallee ; de camilli et al . , 1984 ) and a rabbit polyclonal antibody against nf - l ( xu et al . , 1993 ) . \n the doubly stained sections were examined with a scanning laser confocal microscope ( model 1024 ; bio - rad laboratories , hercules , ca ) . \n previous studies have shown that the three mouse lines ( see materials and methods ) overexpress each of the three nf subunits predominantly in spinal motor neurons ( wong et al . , 1995 ; marszalek et al . , 1996 ; xu et al . , \n measurements from 6-wk - old mice indicate an increase of two- to threefold of the endogenous levels ( wong et al . , 1995 ; marszalek et al . , 1996 ; xu et al . , \n we measured the nf subunit levels in spinal cords of the 17-d - old animals . in each transgenic line , \n the respective nf subunit was increased ( table i ) , consistent with the previous measurements in sciatic nerves and ventral roots . however , some differences are noticeable in hm mice . in addition to large increases in the levels of nf - h and nf - m , nf - l level is also significantly increased ( table i ) , suggesting that increasing both nf - h and nf - m causes nf accumulation in motor neurons by hindering nf transport ( see below ) . to investigate the role of nfs in determining dendritic architecture , we stained spinal cord motor neurons using the golgi method . \n overexpression of each of the three nf subunits produced visually different patterns of dendritic arborization compared with the wild type ( fig . \n 1 ) . overexpression of either nf - h ( h mice ) or nf - m ( m mice ) shortened dendrites by 60 and 30% , respectively , whereas overexpression of nf - l ( l mice ) did not cause a significant change ( figs . 1 and 2 ) \n . the shortening caused by overexpression of either nf - h or nf - m was partially or completely alleviated by a simultaneous increase in the level of nf - l ( compare h mice with lh mice and m mice with lm mice in figs . 1 and 2 ) \n . a simultaneous increase of both nf - h and nf - m nearly eliminated dendrites ( figs . 1 and 2 ) . to determine changes in dendritic branch pattern , we measured the number of dendritic trees , the length of each dendritic tree , the number of dendritic segments , and the length of dendritic segments in each neuron . \n 3 ) . increasing nf - m also reduced , to a less extent , three out of four parameters , with the number of dendritic segments per dendritic tree unaffected ( fig . \n 3 ) . a combined increase of both nf - m and nf - h reduced all the parameters severely ( fig . \n although increasing nf - l alone did not affect any of these parameters ( fig . \n 3 ) , when increased together with either nf - h or nf - m , nf - l was able to reverse partially or completely the reduction caused by increasing either subunit alone ( fig . \n 3 ) . interestingly , in the case of increasing nf - l together with nf - h , the reduction in the number of dendritic trees and dendritic segments ( caused by increasing nf - h alone ) was completely reversed . \n but this had no effect on the reduction of the length of dendritic tree and dendritic segments ( compare h with lh in fig . \n 3 ) . to further examine changes in dendritic branch patterns , we calculated the branch ratio at the successive dendritic branch points . increasing either nf - h or nf - m reduced the third and fourth branches , while the second branches were unaffected ( fig . \n the reduction in the third branch number caused by increasing nf - h was not reversed by a simultaneous increase of nf - l ( fig . \n 4 ) , despite the recovery of total number of dendritic segments per neuron to the normal level ( fig . \n whereas the third and fourth dendritic branches were not different from the wild type in l and lm mice , there was a significant increase in the second order dendritic branches , indicating that the proximal dendritic branches are increased . \n increasing both nf - h and nf - m caused severe reduction in both proximal and distal dendritic branches . \n to determine how dendritic diameters were affected by the altered neurofilament subunit composition , we measured segmental diameter , tapering and rall 's ratio . increasing nf - h or both nf - h and nf - m caused a significant decrease in the diameter of the proximal dendrites but did not affect the diameter of the distal dendrites ( fig . 5 ) . increasing either nf - l or nf - m did not affect dendritic diameter significantly ( fig . \n 5 ) . but increasing nf - l restored the reduced diameters of the proximal dendrites caused by increasing nf - h to normal ( fig . 5 ; compare hl with h ) . altering nf subunit composition \n however , alteration of subunit composition by increasing any of the three subunits , with the exception of increasing nf - l , led to a reduction in rall 's ratio between the second and first order dendrites ( fig . 5 ) . to investigate how changes in nf subunit composition alter dendritic arborization , we examined the global organization of nfs and microtubules ( mts ) by double immunofluorescence using an antibody against nf - l ( xu et al . , 1993 ) , in combination with antibodies against either microtubule - associated protein 2 ( map2 ) , which is distributed in cell bodies and dendrites ( de camilli et al . \n both combinations revealed the same pattern in wild - type motor neurons : both nfs and microtubules were organized into elaborate networks , which extended well into the dendrites ( fig . \n a close alignment between nf and mt networks was also evident , as indicated by a substantial overlap between the two networks ( fig . \n the close alignment between the nf and mt networks was altered by an increase in nf - m . \n 6 d , arrowheads ) , with some of these bundles excluding microtubules ( fig . \n although the extension of both nfs and microtubules into the proximal dendrites was not affected ( fig . \n f , large arrows ) , the nf content in distal dendrites appeared decreased , as indicated by the reduced nf staining in small dendritic segments ( fig . 6 , d f , small arrows ) . \n both aggregation of nfs and dissociation of nf and mt networks were further aggravated by the elevated expression of nf - h ( fig . \n 6 , g i , arrowheads ) , resulting in a depletion of nfs from distal ( fig . 6 , g i , small arrows ) and proximal dendrites ( fig . 6 , g i , large arrows ) and dendritic attrition ( fig . 6 , g i , large arrows ) . \n in contrast to these effects , increasing nf - l did not cause aggregation of nfs or dissociation between nf and mt networks , nor did it lead to nf depletion from proximal dendrites or dendritic atrophy ( fig . 6 , j \n these results suggest that nf - l plays a role antagonistic to nf - h and nf - m and that a balanced ratio among these subunits is crucial to maintaining the cytoskeletal organization in neurons . to confirm this point \n increasing nf - l together with either nf - m or nf - h prevented the formation of nf aggregates and the dissociation of nf and mt networks ( fig . \n most notably , it prevented the depletion of nfs from dendrites ( fig . \n 7 , d f , arrows ) that was caused by increasing nf - h alone ( fig . 6 , g i , arrows ) . \n thus , increasing nf - l reverses the alterations in cytoskeletal organization caused by increasing nf - h and nf - m despite a further increase in the total nf protein content ( table i ) . \n in contrast to the role of increasing nf - l , increasing both nf - m and nf - h caused the most severe aggregation of nfs ( fig . 7 , g i , arrowheads ) and depletion of nfs from dendrites and dendritic attrition ( fig \n thus , a high ratio of nf - h and nf - m to nf - l , rather than an increase in the total amount of nf polypeptides , leads to the alteration in cytoskeletal organization and dendritic arborization . to determine the nature of the nf aggregates , \n motor neurons were examined using em . in neurons from the wild - type mice , various cellular organelles , including nfs and mts , were interspersed throughout the cytoplasm ( fig . 8 a ) . \n this cytoplasmic organization was altered in neurons that express high levels of nf - h , nf - m , or both nf - h and nf - m . \n all of these neurons had nf aggregates that were composed of large parallel filament bundles ( fig . \n these bundles were separated from the cell and nuclear membrane by a layer of other cytoplasmic organelles ( fig . \n these nonfilamentous cytoplasmic layers were connected by several tracks , which contain mts and various other organelles ( fig . \n 8 , b , c , and e , between arrowheads ) that correlate with the regions stained positively with map2 and tubulin antibodies , including the cell periphery , the perinuclear region and the thin tracks connecting these two regions ( figs . \n in contrast to increasing nf - h and nf - m , increasing nf - l led to an increase in nf bundles that traverse among small islands of rer and other cellular organelles , but did not cause large filament aggregation ( fig . \n nfs were also distributed in close proximity of cell and nuclear membrane ( fig . \n when nf - l was increased together with nf - h , the formation of nf aggregates caused by increasing nf - h alone was prevented ( compare fig . 8 , b with f ) . the same results were also observed when nf - l was increased together with nf - m ( data not shown ) . to determine how changes in nf subunit composition affected nf distribution in dendrites , and in particular , to find out how neurofilament network was disrupted abruptly in the proximal dendrites \n , we further examined dendritic initiation zone using em . in wild - type mice there were abundant neurofilaments in dendrites ( fig . \n they formed parallel bundles and were a continuous extension of the nf network in the cell body ( fig . \n 9 , a and b ) . in sharp contrast , the nfs forming aggregates in the cell body in h ( fig . 9 , c and d ) or hm mice ( fig . \n 9 , g and h ) failed to extend into the proximal dendrites , leaving mostly microtubules in the proximal dendrites . \n the disrupted extension of nfs into the proximal dendrites caused by an increase in nf - h was reversed by increasing nf - l ( fig . \n f ) , and lm ( not shown ) mice the number of nfs was increased in the proximal dendrites and the continuous extension of the nf network from the cell body to the proximal dendrites was not affected . \n this study demonstrates the distinct effects of different nf subunits on cytoskeletal organization and dendritic structure in spinal motor neurons . \n increasing nf - h , nf - m , or both subunits led to nf aggregation and a segregation of the nf network from the mt network ( figs . \n these effects were more severe in the case of increasing nf - h or both nf - h and nf - m , leading to a drastic reduction of nfs in dendrites ( figs . \n 6 , 7 , 9 ) . increasing nf - l , however , reversed these changes and led to a close association between the nf and mt networks and an extension of nfs into dendrites ( figs . \n correlating with these changes , increasing nf - h and/or nf - m inhibited dendritic arborization , whereas increasing nf - l alleviated this reduction ( figs . \n these results demonstrate that nf - h and nf - m antagonize nf - l in organizing the nf network and dendritic growth and that a lower ratio of nf - h and/ or nf - m to nf - l is required for the growth of elaborate dendritic trees . \n this finding sheds light on the significance of the developmental regulation of nf subunit expression ( see below ) . \n previous experiments have shown that a high ratio of nf - h and nf - m to nf - l inhibited radial axonal growth ( wong et al . , 1995 ; marszalek et al . , 1996 ; xu et al . , \n our current efforts demonstrate that the high ratio is also inhibitory to dendritic growth , thus proving that a high ratio of nf - h and nf - m to nf - l is generally inhibitory to neuronal process growth . \n our observations suggest that this inhibition is derived from interference with the interaction between nf and mt networks by the increased ratio of nf - h and/or nf - m to nf - l that leads to a dissociation of the nf and mt networks ( figs . \n because long distance intracellular organelle transport is known to be microtubule dependent ( vallee and bloom , 1991 ) , such a dissociation could lead to an inhibition of nf transport , resulting in a decreased nf content in both axons and dendrites ( wong et al . \n , 1995 ; marszalek et al . , 1996 ; xu et al . , 1996 ; figs . \n 6 , 7 , 8) and an accumulation of nfs in cell bodies ( fig . \n a close association of intermediate filaments and mts in cultured cells has been widely observed and disruption of this interaction leads to a collapse of intermediate filament network into aggregates ( blose et al . , 1984 ; gyoeva and gelfand , 1991 ) . \n we show that nfs are closely associated with mts in neurons in vivo . the dissociation of nfs from mts and the formation of nf aggregates after the increased ratio of nf - h and/or nf - m to nf - l suggest that nf - h and nf - m weaken the association of nfs with mts and promote nf self association . \n the reversal of this dissociation and nf aggregation by an increase in nf - l suggests that nf - l plays an interactive role between nfs and mts . \n candidates for mediating this interaction include microtubule associated proteins , such as map2 and tau . \n both have been shown to bind to microtubules as well as nf - l in vitro ( heimann et al . , 1985 ; miyata et al . , 1986 ) . \n other potential mediators include microtubule - based motors ( gyoeva and gelfand , 1991 ) and the intermediate filament - associated protein plectin ( svitkina et al . , 1996 ) . \n the mechanism for the inhibition of the interaction between nfs and mts by the high ratio of nf - h and/or nf - m to nf - l is not clear . \n one possibility is that an increased ratio of nf - h and/or nf - m to nf - l may increase the density of their cooh - terminal tails projecting from the filament core where nf - l resides ( hirokawa et al . , 1984 ; hirokawa , 1991 ) . because nf - l may play an interactive role with mts , a high density of nf - h and nf - m tails could block this interaction due to stereo hindrance . concomitant with inhibiting the interaction between nfs and mts , \n nf - h and nf - m may also promote self - association among nfs ( xu et al . , 1996 ) , mediated by their cooh - terminal tails of these molecules ( hirokawa et al . , 1984 ; hirokawa , 1991 ) . \n thus , both interference with the interaction between nfs and mts and a tendency for self - association may contribute to the formation of nf aggregates when the ratio of nf - h and/or nf - m to nf - l is increased . by the same token \n , increasing nf - l lowers the density of nf - h and nf - m tails , which in turn reduces the stereo hindrance , therefore allowing the interaction between nfs and mts to occur . \n the reduced nf content in dendrites as a consequence of an increased ratio of nf - h and nf - m to nf - l is consistent with a reduction in transport of nfs into neuronal processes and is a likely cause that leads to structural changes in dendrites . \n 3 a ) , distal dendritic branching ( fig . 4 ) and the length of dendritic segments ( fig . \n although increasing nf - l alone does not affect most of these parameters , when increased together with nf - h or nf - m , it partially or completely alleviated the reduction in these dendritic parameters ( figs . \n the most dramatic effects are in the recovery of proximal dendritic diameter and the number of primary dendrites in lh mice ( figs . \n coincident with these recoveries is the recovery of nf content in the proximal dendrites ( figs . \n 3 , a and d ) is not observed and the recovery of distal dendritic branching is incomplete in lh mice ( fig . \n 4 ) , suggesting that the effect of increasing nf - l is limited to the proximal dendrites . in a mathematical simplification of the electrical properties of complex dendritic arborization , rall used a single element or equivalent cylinder to describe a dendritic tree ( rall , 1995 ) . \n rall 's ratio is a constraint in this model that requires ( [ddaughter])/(dparent ) = 1 ( d represents diameter of dendritic segments ) . \n our results show a rall 's ratio being close to one in wild - type mice , agreeing with other studies ( ulfhake and kellerth , 1981 ; brnnstrm , 1993 ; chen and wolpaw , 1994 ) . \n changes in nf subunit composition do not lead to a significant deviation from the rall 's ratio in the overall dendritic structure . \n only the ratio between the second and the first order dendrites is reduced in all except l mice , whereas the ratios in distal dendrites are unaffected ( fig . \n the expression of nf - l and nf - m closely follows neuronal differentiation ( carden et al . , 1987 ; \n 1992 ) and is associated with the earliest stages of neuronal process growth ( cochard and paulin , 1984 ; landmesser and swain , 1992 ; crino and eberwine , 1996 ) . \n however , the expression of nf - h does not increase significantly until > 10 d after birth ( shaw and weber , 1982 ; pachter and liem , 1984 ) . whereas the coexpression of nf - l and nf - m during the early stages of development is consistent with the finding that at least two subunits , composed of nf - l and either nf - m or nf - h , are required for nf assembly in cells ( ching and liem , 1993 ; lee et al . , 1993 ) , maintaining a low level of nf - h may be important for dendritic growth . on this note , it is interesting to notice that at age 17 d , the ratio of nf - h to nf - l in wild - type mice is 0.06 ( table i ) , approximately threefold lower than the ratio in 6-wk - old mice ( xu et . \n increasing nf - h in h mice brings the ratio to 0.16 ( table i ) . \n al . , 1996 ) , yet it produces a remarkable inhibition of dendritic arborization ( figs . \n thus , our current results indicate that the delayed expression of nf - h enables dendritic growth . \n the outcome of this study raises the question of why the ratio of nf - h to nf - l increases as neurons develop to maturity . \n the early appearance and enrichment of nf - l and nf - m in neuronal processes suggests that nfs play a role in the early development of these processes ( cochard and paulin , 1984 ; landmesser and swain , 1992 ; crino and eberwine , 1996 ) . \n on the other hand , neuronal maturation correlates with an increase in nf - h and slowing of axonal transport ( willard and simon , 1983 ) . \n therefore , the increase in the level of nf - h may increase the stability of cytoskeleton and thereby stabilize neuronal structure in mature neurons ( willard and simon , 1983 ) . \n collectively , the existing experimental data suggests the following sequence of events during development : as nf - h increases , the interaction between nfs and microtubules weakens , and the interaction among nfs themselves strengthens . \n this leads to an extensive cross - bridge formation among nfs ( hirokawa et al . , 1984 ) and slower transport of cytoskeletal components ( willard and simon , 1983 ) . \n as these changes proceed , a stable nf matrix within axons and dendrites ( particularly proximal dendrites ) is formed and neuronal processes stabilize . \n thus , neurons may regulate the ratio of nf - h to nf - l in order to maintain a balance between plasticity and stability of neuronal processes during different stages of development . \n such a process is perhaps most relevant in large nf - rich neurons with long axons and long dendrites radiating from their cell bodies ( such as motor neurons and betz cells ) . \n the role of neurofilaments in the development of dendritic trees with short branches may be less significant . \n consistent with this view , small neurons that do not have long dendritic trees and large neurons with numerous but short segmental dendritic branches ( e.g. , purkinje cells ) express little neurofilaments . \n taken together , our results show that nfs play an important role in the development of large dendritic trees with long segmental branches . \n nf subunit content in spinal cord nf subunit levels are expressed as pmol/100 g total spinal cord protein . \n the levels of nf - h are the sum of both phosphorylated and unphosphorylated forms . \n camera lucida drawings of representative spinal motor neurons from mice transgenic for different nf subunits . \n w , wild type ; h , nf - h transgenic ; m , nf - m transgenic ; l , nf - l transgenic ; hm , lh , and lm , represent the mice that are doubly transgenic for two subunits . \n changes in the average total dendritic length per motor neuron . the length of dendrites was measured for each neuron and the average length per neuron was calculated for each transgenic line . \n error bars represent sem single and double stars represent the values that were significantly different ( p < 0.05 and p < 0.01 , respectively ) from that of the wild type . \n the definitions for dendritic tree and dendritic segments are described in materials and methods . \n dendritic branches are evaluated by calculating the ratio of dendritic segments in successive dendritic order ( see materials and methods ) . \n the ratio beyond the fourth order were not calculated because of low segment numbers . \n 1 and 2 . changes in dendritic diameter , tapering , and rall 's ratio ( see materials and methods ) . \n 1 and 2 . different effects of overexpressing each of the three nf subunits on cytoskeletal organization . spinal motor neurons were doubly stained with antibodies against nf - l ( red ) and map2 ( green ) . \n ( a c ) wild type ; ( d f ) m mice ; ( g i ) h mice ; ( j l ) l mice . \n overexpression of nf - l prevents whereas overexpression of nf - h and nf - m induces nf aggregation . \n spinal motor neurons were doubly stained with antibodies against nf - l ( red ) and map2 ( green ) . \n ( a c ) lm mice ; ( d f ) lh mice ; ( g i ) hm mice . \n changes in nf network and cytoplasmic organization in motor neurons overexpressing different nf subunits . \n spinal motor neurons were photographed with em and displayed at the same magnification . \n ( a ) wild type , ( b ) h mice , ( c ) m mice , ( d ) l mice , ( e ) hm mice , and ( f ) lh mice . \n nf marks nf aggregates ; arrows point to cytoplasmic membrane ; arrowheads point to tracks connecting the perinuclear and the peripheral cytoplasmic regions . \n ( a and b ) wild type ; ( c and d ) h mice ; ( e ) m mice ; ( f ) l mice ; ( g and h ) mh mice ; ( i and j ) lh mice .", "role": "user" }, { "content": "dendrites play important roles in neuronal function . \n however , the cellular mechanism for the growth and maintenance of dendritic arborization is unclear . neurofilaments ( nfs ) , a major component of the neuronal cytoskeleton , are composed of three polypeptide subunits , nf - h , nf - m , and nf - l , and are abundant in large dendritic trees . by overexpressing each of the three nf subunits in transgenic mice , we altered subunit composition and found that increasing nf - h and/or nf - m inhibited dendritic arborization , whereas increasing nf - l alleviated this inhibition . \n examination of cytoskeletal organization revealed that increasing nf - h and/or nf - m caused nf aggregation and dissociation of the nf network from the microtubule ( mt ) network . \n increasing nf - h or nf - h together with nf - m further reduced nfs from dendrites . \n however , these changes were reversed by elevating the level of nf - l with either nf - h or nf - m . \n thus , nf - l antagonizes nf - h and nf - m in organizing the nf network and maintaining a lower ratio of nf - h and nf - m to nf - l is critical for the growth of complex dendritic trees in motor neurons .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: non - hodgkin lymphomas ( nhls ) account for approximately 60% of all lymphomas in children and adolescents . \n childhood nhls are subdivided into burkitt 's lymphoma , diffuse large b - cell lymphoma ( dlbcl ) , lymphoblastic lymphoma , and anaplastic large - cell lymphoma . \n dlbcl is characterized by relatively more frequent extranodal presentation , seen in upto 40% of the cases . \n primary involvement of the lymphoma of the middle ear is rare , with only about 18 cases being reported in literature so far . here \n , we report a case of dlbcl , presented with features of facial palsy and otitis , who received initial symptomatic treatment and later chemotherapy after diagnosing dlbcl . \n a 2 years 8 months old boy visited our tertiary care hospital with complaints of ear ache ( left side ) and left facial palsy of 4 weeks duration . after initial 2 weeks of these symptoms \n , there was whitish serous discharge from the left ear . before referral to our hospital , he was treated for otitis media with antibiotics , details of which were not available . \n his complete blood picture and biochemistry investigations done in our hospital were within normal limits . \n as the symptoms persisted for 4 weeks , computed tomography of head and neck was done which was suggestive of solid mass lesion of 2 cm 2 cm size in the left mastoid with destruction of mastoid bone [ figure 1 ] . \n the disease was in stage 1 ( as per murphy 's staging ) and was completely resected . \n histopathological examination showed large cells of lymphoid cell proliferation immune histochemistry was positive for cd20 , bcl-2 and negative for cd3 , with low mib-1 , which confirmed dlbcl . \n his positron emission tomography for staging , bone marrow and cerebrospinal fluid revealed no abnormality . computed tomography head and neck suggestive of solid mass lesion of 2 cm 2 cm size in the left mastoid with destruction of mastoid bone computed tomography head and neck of right side showing no abnormality his initial clinical symptoms of earache and facial nerve palsy followed by ear discharge resolved after starting chemotherapy as per b - cell lymphoma protocol for 6 months . \n currently , he is 37 months off treatment , and no disease recurrence is seen clinically as well as radiologically . \n it is an aggressive form of lymphoma , usually curable with appropriate treatment and has high survival rate . \n rapid disease progression of dlbcl calls for an early , accurate diagnosis and appropriate treatment . \n however , unusual presentation can mislead the physician resulting in wrong diagnosis , which delays the treatment , thus promoting disease progression . \n extranodal presentations of childhood dlbcl are relatively uncommon in clinical practice , and much rarer are those primarily involving middle ear and mastoid . involvement of middle ear and mastoid can resemble the features of otitis media and unusual facial palsy mimicking mastoiditis . \n although these initial symptoms were suggestive of middle ear infection , the distinguishing factor was unresponsiveness to antibiotics . \n there have been very few reports of nhl with facial nerve involvement ; ogawa et al . \n mccabe et al . , reported a case of 2-year - old , an immunocompetent boy with spontaneous regression of an epstein - barr - virus - associated monoclonal lymphoid proliferation who presented with acute otitis media and facial palsy . \n have described a case of dlbcl with features of otitis media , mastoiditis , and facial palsy . \n as symptoms persisted even after 4 weeks of treatment , the patient was started on chemotherapy , to which he promptly responded . \n extranodal nhls of middle ear which is not a common clinical presentation may present as facial palsy , and misdiagnosed as otomastoiditis . \n there should be a high index of suspicion for primary neoplasms of the middle ear in patients with chronic otomastoiditis refractory to appropriate initial antibiotic therapy ; early diagnosis and appropriate treatment results in good therapeutic outcome and minimizes further complications . \n \n ", "role": "user" }, { "content": "extra nodal presentation of non hodgkins lymphoma ( nhl ) is a rare entity , and data available about the nhl that primarily involves of middle ear and mastoid is limited . \n we report a case of diffuse large b cell lymphoma ( dlbcl ) , in a 2 year 8 month old boy , who developed otalgia and facial palsy . computed tomography revealed a mass in the left mastoid . \n mastoid exploration and histopathological examination revealed dlbcl . \n this case highlights the importance of considering malignant lymphoma as one of the differential diagnosis in persistent otitis media and / facial palsy .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: literature describes several case reports of retained intracardiac foreign bodies especially in left sided chambers . our aim was to report a case of an unusual foreign body that extended from the right atrium to the tricuspid valve , thus causing tricuspid regurgitation . \n minimally invasive approach was utilized to excise the foreign body mass with an embedded plastic cannula in it . \n a 42 year - old caucasian female without significant past medical history , other than a history of status post ablation left leg varicosities , presented to the ed with the complaint of ongoing chest pain . \n she described the unbearable pain as sharp , pleuritic in nature , originating under her left breast and radiating to her back and neck . \n even when taking a deep breath , the patient defines the pain as , it is like somebody sticking a knife into her chest . \n she denied any frank shortness of breath , fever , nausea or vomiting and had not been diaphoretic . \n subsequent chest x - ray and v / q scans performed demonstrated no acute cardiopulmonary abnormality and a low probability for a pulmonary embolus respectively . on further exploring her medical history , it was noticed that post ablation of her left leg varicosities , she developed deep vein thrombosis of the right lower extremity and hence was commenced on anticoagulants . \n given that , she underwent a contrast magnetic resonance imaging ( mri ) of chest for cardiac morphology and function . \n mri demonstrated a mobile mass in the right atrium with echo - dense linear structure extending from the right atrium to the tricuspid valve causing tricuspid regurgitation ( tr fig . \n . the dilemma of her excruciating pain symptoms still remained unclear and the right atrial mass causing tr was thought to be plausible endocarditis in nature . \n she was taken to operating theatre for removal of the right atrial mass via minimally invasive approach . \n the femoral artery and veins were dissected and used for cardiopulmonary bypass ( cpb ) cannulation source . \n it was found that there was a large mass assembled around a piece of plastic cannula extending from the right atrial appendage onto the tricuspid valve ( fig . \n this was also reconfirmed by no evidence of tr on intraoperative transesophageal echocardiogram ( tee ) post - removal of the mass . \n once satisfaction to our intracardiac inspection was achieved , the right atriotomy was closed in 2 layers , and cardiopulmonary bypass was weaned off successfully . \n 24-french blake drain was placed in the pericardial cavity for drainage and the right anterior thoracotomy wound was closed in routine fashion . \n there have been several case reports describing retained intracardiac foreign bodies especially in left sided chambers \n we report a case of a young woman who presented to us with repeated episodes of debilitating stabbing chest pain . \n the differential diagnosis at the time was perplexed and ranged from recurrent polyserositis , familial mediterranean fever , amyloidosis , idiopathic pericarditis and tuberculosis associated with polyserositis . nevertheless , we found during the surgery that there was a retained iatrogenic piece of plastic cannula embedded in the inflamed right atrial mass causing the symptoms of the patient . \n earlier review conducted in 5000 patients who received intracardiac catheters inserted for treatment modalities suggested iatrogenic retention incidence of 0.07% . \n such retained foreign bodies in the heart could lead to number of complications , that may include thrombus formation , tricuspid valve insufficiency , neurological manifestations secondary to cardio - embolic stroke , infections , and arrhythmias . in this regard , actis dato et al . \n evaluated fourteen cases of foreign bodies in the heart and provided some procedural guidelines about the management of foreign bodies in the heart . \n the authors suggested that if the patient is symptomatic , surgical removal of the foreign body is indicated ; asymptomatic with associated risk of infection , embolization , or erosion , surgery is indicated ; and if patient is asymptomatic without the associated symptoms , surgery is indicated depending on the nature and location of the foreign body within the heart . \n these include new generation computer tomography scans , echocardiography , and mri . in our case \n we relied on mri yet the scans remained inconclusive in delineating between the mass or foreign body from the surrounding muscle tissues and/or organs . \n it is interesting to note that patients may remain asymptomatic for a period of time before constitutional signs and symptoms begin to appear as in our case . \n tsutsui and coworkers in 2003 suggested the identification of chemotherapy and dialysis retained catheter fragments during routine chest x - rays , though the patients in their report remained primarily asymptomatic . \n the exact aetiology of such retained plastic cannula fragment in our patient is still not clear , however our patient did recall to have a past history of cardiac catheterization several months prior to the surgery . \n henceforth , in view of catheters being widely used and necessary for a variety of procedures and treatments in this current age and era , it is of prime importance for physicians and other medical staff to be aware of the proper insertion and removal of plastic cannulas whenever being used on a patient . \n it is of utmost importance that the removal of the plastic cannulas be confirmed by two different attending personnel , a nurse and by the attending physician . \n we emphasize that every department should be equipped with the proper protocols for managing catheter based vascular access . \n the patient presented in this report was very fortunate that the plastic cannula fragment did not immediately pierce the right atrial wall , which could have led to severe internal bleeding into the pericardial cavity and ultimately progressing to further complications . \n \n \n we have obtained written consent from the patient and that is available to be provided , should the editor ask to see it . \n subbareddy konda helped in case identification , refining the literature search and writing and others .", "role": "user" }, { "content": "introductionwe report a case of a retained foreign body in the right atrium and the review of the literature discussing several cases where the poor attention and management of medical staff has led to worsening consequences to patient 's health.presentation of casein our case the mass demonstrated on mri scan turned out to be an inflammatory process and organized clotted blood built around a broken piece of a plastic cannula protruding out of the right atrium . \n this caused debilitating pleuritic pain to the patient on presentation.discussionthe cause of this iatrogenic retained piece of cannula may well be from the patients prior diagnostic investigations.conclusionalgorithm managed indications for surgical removal of such foreign bodies in symptomatic patients lead to better patient 's outcomes and decreases the chances of infection , embolization , or erosions within the heart . keeping this in view , we managed our patient with success .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: note , however , that this method has many disadvantages , including insufficient bone volume , donor site morbidity , bone resorption , and discomfort . \n still , recent studies based on multipotent stem cells have shown new possibilities for the treatment of bony defects . \n hematopoietic stem cells ( hscs ) have been reported to be able to make skeletal muscle1,2 , cardiac muscle3,4 , and liver5 because they can differentiate into adipocytes1 , condrocytes6,7 , myocytes1,8 and osteoblast7,9 , but there are ethical and surgical limitations6,10 . \n recently , mscs in the human maxillary sinus membrane have been known to differentiate to osteoblasts under some conditions . \n gruber et al.11 reported that porcine sinus mucosa cells were positive for stro-1 and alkaline phosphatase ( alp ) activity . \n they concluded that the sinus mucosa holds mesenchymal progenitor cells committed to the osteogenic lineage by responding to bone morphogenic protein ( bmp)-6 and bmp-7 . according to kim et al.13 , mscs in the human maxillary sinus membrane \n note , however , that there are controversies regarding mscs ' biological safety and effectiveness . \n statin is an inhibitor in the reduction of 3-hydroxy-3-methylglutaryl coenzyme a ( hmg co - a ) and is used for lowering serum cholesterol14 . \n recently , statin has been reported to be able to enhance bone metabolism and new bone formation . \n it also increased cancellous bone mass and cancellous bone compressive strength15 , and these results were related to the increased rate of new bone formation14,16 . \n as a protein that regulates the differentiation and function of osteoblasts and chondrocytes , bmp has a major role in bone formation17 . \n after the adipose - derived stromal cells modified with the bmp-2 gene were implanted into ulnar defects in the canine model , these cells produced significant new bone18 . \n bmp-2 and vitamin d(3 ) had synergistic effect on the osteogenic differentiation of adipose stem cells19 . \n for these reasons , osteogenic differentiation of human mscs has been evaluated with bmp-2 mrna expression levels20,21 . \n we hypothesized that simvastatin wields a beneficial effect on the osteogenic potential of human maxillary sinus membrane - derived stem cells ( hsmscs ) . \n this study sought to evaluate the osteoinductive potential of simvastatin and osteogenic potential of mscs derived from the human maxillary sinus membrane . \n the subjects of this study were five adults , and approval was obtained from the institutional review board of college of medicine , the catholic university of korea . \n informed consent was provided to patients as per the ' act on legal codes for biomedical ethics and safety ' and the declaration of helsinki . human maxillary sinus mucosal membrane ( hmsm)-derived stem cells were isolated and cultivated in accordance with the author 's previous study13 . \n maxillary sinus tissue was collected and contained in phosphate - buffered saline ( pbs ; gibco brl , grand island , ny , usa ) including 1% antibiotic and antimycotic ( 10 antibiotic & antimycotic , gibco brl ) and transported to the laboratory under aseptic condition . \n it was then washed with pbs ( gibco brl ) including antibiotic and antimycotic , cut into small pieces with a mesh under aseptic condition , treated with 0.06% collagenase type ii ( invitrogen , carlsbad , ca , usa ) , shaken with an incubator ( co2 incubator ; forma scientific , marietta , oh , usa ) containing 5% co2 at 37 for 4 hours , and centrifuged at 1,000 rpm for 10 minutes ( large - capacity table - top centrifuge ; hanil science industrial , gangneung , korea ) . \n louis , mo , usa ) containing 10% fetal bovine serum ( hyclone , logan , ut , usa ) and 1% penicillin - streptomycin ( gibco brl ) and filtered with 40 m cell strainer ( bd bioscience , bedford , ma , usa ) . tissues except hmsm - derived cells were filtered , with the filtered out solution cultured in the incubator . \n the daily morphological characteristics were observed with inverted microscope , and the culture solution was replaced every other day . \n when the medium was changed , cells not adhering to the culture plate were removed ; only the adhering cells were cultured . \n when culture dishes became near - confluent , cells were separated with trypsin - edta ( gibco brl ) and subsequently repeated for continued passaging . \n colony - forming unit - fibroblast assay was used to evaluate msc presence in hmsm . \n when the initially cultured cells became near confluent , cells were separated with trypsin - edta , plated at 110 cells / ml onto the 100 mm culture plate , stained with hematoxilin ( sigma - aldrich ) after 24 hours , and observed with inverted microscope . \n flow cytometry on cell surface markers stro-1 , cd105 , and cd34 for the separation of mscs from hmsm was conducted . \n the hmsm - derived cells at passage 3 were plated into a test tube ( bectom dickinson , franklin larks , nj , usa ) by 110 cells / ml . the antibodies of cd34 ( bd bioscience ) and cd105 ( serotec , oxford , uk ) - to which fluorescein isothicyanate ( fitc ) and phycoerythrin were adhered - were treated for 1 hour and washed with wash buffer 3 times , and the origin of stem cell was observed with flow cytometry ( bd bioscience ) . for stro-1 ( human anti - mouse monoclonal antibody , igm subclass ; r&d system , minneapolis , ma , usa ) , antibody was treated for 1 hour and washed 3 times with wash buffer ; the secondary antibody to which fitc was attached was treated for 30 minutes , washed 3 times with wash buffer , and observed with flow cytometry . \n sinus mucosa - derived stem cells were washed with pbs 72 hours after treatment on normal media and simvastatin ( 0.1 , 0.2 , 0.5 , 1.0 , 2.0 m)-treated media . \n afterward , the cells were collected with cell scraper ( sarsted , newton , nc , usa ) . \n after the total rna was separated by qiagen rna extraction kit ( qiagen , hilden , germany ) , it underwent reverse transcription reaction with the separated rna , sense primer , antisense primer , and reverse transcriptase - polymerase chain reaction ( rt - pcr ) premix ( bioneer , daejeon , korea ) for 60 minutes at 42 and 5 minutes at 94 and denatured for 5 minutes at 94 and 1 minute at 94 , and genes were annealed for 90 seconds at 52 , elongated for 1 minute at 72 , and then annealed and elongated repeatedly 30 times and incubated for 5 minutes at 72.(table 1 ) sinus mucosa - derived stem cells were washed with pbs 72 hours after treatment on normal media and simvastatin ( 0.1 , 0.2 , 0.5 , 1.0 , 2.0 m)-treated media . \n the cells were then collected with cell scraper ( sarsted ) and spun at 12,000 rpm for 20 minutes at 4 after being rinsed with ripa - b buffer ( 0.5% nonidet p-40 , 20 mm tris , ph 8.0 , 50 mm nacl , 50 mm naf , 100 m na3vo4 , 1 mm dtt , 50 g / ml pmsf ) for 1 hour at 4. the supernatant was transferred to the nitrocellulose membrane , blocked with ttbs ( 1 m tris , 1.5 m nacl , 500 l tween 20 ) containing 5% skim milk , and treated by bmp-2 primary antibody and secondary antibody . \n sinus mucosa - derived stem cells were cultured 3 , 6 , 9 , and 12 days after treatment on normal media and determined concentrated simvastatin - treated media . \n after the total rna was separated with qiagen rna extraction kit , it underwent reverse transcription reaction with the separated rna , oc sense primer , antisense primer , and rt - pcr premix for 60 minutes at 42 and 5 minutes at 94 and denatured for 5 minutes at 94 and 1 minute at 94 ; genes were annealed for 90 seconds at 52 , elongated for 1 minute at 72 , and then annealed and elongated repeatedly 30 times and incubated for 5 minutes at 72.(table 1 ) the control and simvastatin - treated group were each washed 3 times with sterile triple - distilled water 3 , 6 , 9 , and 12 days after the treatment date , fixed with citrate - acetone - formaldehyde fixer solution ( sigma - aldrich ) , and washed 3 times with sterile triple - distilled water . \n they were stained with alkaline - dye mix ( nitrite , frv - alkaline , naphthol asbi alkaline solution ; sigma - aldrich ) with light isolated for 15 minutes at normal temperature , washed 3 times with sterile triple - distilled water , counterstained with hematoxylin , washed again 3 times with sterile triple - distilled water , and observed with inverted microscope . \n all groups were washed 3 times with sterile triple - distilled water 3 , 6 , 9 , and 12 days after the treatment date . \n , darmstadt , germany ) at normal temperature for 15 minutes and washed again 3 times with sterile triple - distilled water . \n they were stained for 30 minutes with light isolated with 1% silver nitrate ( merck ) solution , washed 3 times with sterile triple - distilled water , left for 1 hour under ultraviolet , counterstained with 0.1% eosin ( sigma - aldrich ) , and observed with inverted microscope . \n human maxillary sinus mucosal membrane ( hmsm)-derived stem cells were isolated and cultivated in accordance with the author 's previous study13 . \n maxillary sinus tissue was collected and contained in phosphate - buffered saline ( pbs ; gibco brl , grand island , ny , usa ) including 1% antibiotic and antimycotic ( 10 antibiotic & antimycotic , gibco brl ) and transported to the laboratory under aseptic condition . \n it was then washed with pbs ( gibco brl ) including antibiotic and antimycotic , cut into small pieces with a mesh under aseptic condition , treated with 0.06% collagenase type ii ( invitrogen , carlsbad , ca , usa ) , shaken with an incubator ( co2 incubator ; forma scientific , marietta , oh , usa ) containing 5% co2 at 37 for 4 hours , and centrifuged at 1,000 rpm for 10 minutes ( large - capacity table - top centrifuge ; hanil science industrial , gangneung , korea ) . \n louis , mo , usa ) containing 10% fetal bovine serum ( hyclone , logan , ut , usa ) and 1% penicillin - streptomycin ( gibco brl ) and filtered with 40 m cell strainer ( bd bioscience , bedford , ma , usa ) . tissues except hmsm - derived cells were filtered , with the filtered out solution cultured in the incubator . \n the daily morphological characteristics were observed with inverted microscope , and the culture solution was replaced every other day . \n when the medium was changed , cells not adhering to the culture plate were removed ; only the adhering cells were cultured . \n when culture dishes became near - confluent , cells were separated with trypsin - edta ( gibco brl ) and subsequently repeated for continued passaging . \n colony - forming unit - fibroblast assay was used to evaluate msc presence in hmsm . \n when the initially cultured cells became near confluent , cells were separated with trypsin - edta , plated at 110 cells / ml onto the 100 mm culture plate , stained with hematoxilin ( sigma - aldrich ) after 24 hours , and observed with inverted microscope . \n flow cytometry on cell surface markers stro-1 , cd105 , and cd34 for the separation of mscs from hmsm was conducted . \n the hmsm - derived cells at passage 3 were plated into a test tube ( bectom dickinson , franklin larks , nj , usa ) by 110 cells / ml . the antibodies of cd34 ( bd bioscience ) and cd105 ( serotec , oxford , uk ) - to which fluorescein isothicyanate ( fitc ) and phycoerythrin were adhered - were treated for 1 hour and washed with wash buffer 3 times , and the origin of stem cell was observed with flow cytometry ( bd bioscience ) . for stro-1 ( human anti - mouse monoclonal antibody , igm subclass ; r&d system , minneapolis , ma , usa ) , antibody was treated for 1 hour and washed 3 times with wash buffer ; the secondary antibody to which fitc was attached was treated for 30 minutes , washed 3 times with wash buffer , and observed with flow cytometry . \n sinus mucosa - derived stem cells were washed with pbs 72 hours after treatment on normal media and simvastatin ( 0.1 , 0.2 , 0.5 , 1.0 , 2.0 m)-treated media . \n afterward , the cells were collected with cell scraper ( sarsted , newton , nc , usa ) . \n after the total rna was separated by qiagen rna extraction kit ( qiagen , hilden , germany ) , it underwent reverse transcription reaction with the separated rna , sense primer , antisense primer , and reverse transcriptase - polymerase chain reaction ( rt - pcr ) premix ( bioneer , daejeon , korea ) for 60 minutes at 42 and 5 minutes at 94 and denatured for 5 minutes at 94 and 1 minute at 94 , and genes were annealed for 90 seconds at 52 , elongated for 1 minute at 72 , and then annealed and elongated repeatedly 30 times and incubated for 5 minutes at 72.(table 1 ) \n sinus mucosa - derived stem cells were washed with pbs 72 hours after treatment on normal media and simvastatin ( 0.1 , 0.2 , 0.5 , 1.0 , 2.0 m)-treated media . \n the cells were then collected with cell scraper ( sarsted ) and spun at 12,000 rpm for 20 minutes at 4 after being rinsed with ripa - b buffer ( 0.5% nonidet p-40 , 20 mm tris , ph 8.0 , 50 mm nacl , 50 mm naf , 100 m na3vo4 , 1 mm dtt , 50 g / ml pmsf ) for 1 hour at 4. the supernatant was transferred to the nitrocellulose membrane , blocked with ttbs ( 1 m tris , 1.5 m nacl , 500 l tween 20 ) containing 5% skim milk , and treated by bmp-2 primary antibody and secondary antibody . \n sinus mucosa - derived stem cells were cultured 3 , 6 , 9 , and 12 days after treatment on normal media and determined concentrated simvastatin - treated media . \n after the total rna was separated with qiagen rna extraction kit , it underwent reverse transcription reaction with the separated rna , oc sense primer , antisense primer , and rt - pcr premix for 60 minutes at 42 and 5 minutes at 94 and denatured for 5 minutes at 94 and 1 minute at 94 ; genes were annealed for 90 seconds at 52 , elongated for 1 minute at 72 , and then annealed and elongated repeatedly 30 times and incubated for 5 minutes at 72.(table 1 ) \n the control and simvastatin - treated group were each washed 3 times with sterile triple - distilled water 3 , 6 , 9 , and 12 days after the treatment date , fixed with citrate - acetone - formaldehyde fixer solution ( sigma - aldrich ) , and washed 3 times with sterile triple - distilled water . \n they were stained with alkaline - dye mix ( nitrite , frv - alkaline , naphthol asbi alkaline solution ; sigma - aldrich ) with light isolated for 15 minutes at normal temperature , washed 3 times with sterile triple - distilled water , counterstained with hematoxylin , washed again 3 times with sterile triple - distilled water , and observed with inverted microscope . \n all groups were washed 3 times with sterile triple - distilled water 3 , 6 , 9 , and 12 days after the treatment date . \n , darmstadt , germany ) at normal temperature for 15 minutes and washed again 3 times with sterile triple - distilled water . \n they were stained for 30 minutes with light isolated with 1% silver nitrate ( merck ) solution , washed 3 times with sterile triple - distilled water , left for 1 hour under ultraviolet , counterstained with 0.1% eosin ( sigma - aldrich ) , and observed with inverted microscope . \n the subjects of this study were five adults ( mean age : 20 years old ) . \n bmp-2 mrna was expressed on the simvastatin group after 72 hours , with the highest expression on the 1.0 and 2.0 m groups.(fig . \n 1 ) bmp-2 protein was also expressed on the 0.5 , 1.0 , and 2.0 m simvastatin groups.(fig . \n 2 ) bmp-2 mrna and protein were not expressed on the control and smscs only group . \n 4 ) von kossa positive staining appeared on the 6th day , increasing with time.(fig . \n still , it has to be modified because of unfavorable environments such as excessive alveolar bone loss and sinus pneumatization . to improve such unfavorable environment , bone graft is considered to be the treatment of choice \n especially , maxillary sinus grafting is a standard procedure with dental implant in the posterior maxilla22 , and autogenous and various artificial bone substitutes are used in this procedure . \n autogenous bone has many disadvantages , e.g. , insufficient bone volume , donor site morbidity , bone resorption , and discomfort , even as it is an ideal graft material . \n artificial bone graft materials are cell - free , requiring more time for bone healing despite being more convenient to use than autogenous bone22 . \n recent studies show that multi - potent stem cells can replace bone graft for the treatment of bony defects23 - 26 . \n embryonic stem cells may have greater potential27,28 , but these cells have some limitations such as bioethical consideration and uncontrolled differentiation . \n mscs are adult stem cells with self - renewal capacity and multi - potentiality of being differentiated into multiple mesenchymal cell lineages29 - 31 . \n different types of mscs and combination methods with various osteoconductive scaffolds have been examined for their potential application in bone regeneration22 . \n most of mscs are generally isolated from bone marrow aspirates harvested from the iliac bone , periosteum of the mandible , or maxillary tuberosity and adipose tissue22 . \n note , however , that mscs in normal media did not show any calcified nodule and oc mrna . \n these results suggest that mscs itself may have not osteogenic potential , and that combination methods with mscs and osteoconductive scaffold induce new bone formation . \n song et al.32 investigated the effect of simvastatin , a widely used statin , on osteoblastic and adipocytic differentiation in primary cultured mouse bone marrow stromal cells . according to them , \n simvastatin enhanced the expression level of mrna for oc and protein for oc and osteopontin , increased alkaline phosphatase activity significantly , and inhibited adipocytic differentiation . \n it was suggested that statin induced the apoptosis of osteoclast and enhanced the differentiation of osteoblast through the inhibition of the mevalonate pathway in cholesterol metabolism and blockade of guanosine-5'-triphosphatase and prenylation32 . \n the oc mrna and alp activity of human alveolar osteoblasts and periodontal ligament cells was also increased by simvastatin33 . \n some clinical studies reported that the long - term administration of simvastatin is correlated with increasing bone mineral density and with decreasing bone fracture35,36 . \n this study has some limitations since quantitative experiments were not performed because of the preliminary study ; hence the need for quantitative analysis and evaluation of relative osteogenic potential of simvastatin . \n in this study , the authors tested the effect of simvastatin on maxillary sinus mucosal stem cells . \n the osteogenic potential of stem cells and simvastatin was evaluated with rt - pcr , and the mineralization of extra cellular matrix was verified by alp expression and von kossa staining . \n we found that bmp-2 was much more expressed in 1 m simvastatin than the other concentrations . 1 m simvastatin enhanced the expression of oc mrna and mineralization . \n the authors ' previous study showed that mscs and osteogenic media expressed oc mrna after 1 week , and that mineralization was detected after 2 weeks13 . \n these results suggest that the combination of stem cells and simvastatin enhance new bone formation , and that this method may be a useful treatment of bony defects .", "role": "user" }, { "content": "objectivesthe objective of this study is to determine the adequate concentration and to evaluate the osteogenic potential of simvastatin in human maxillary sinus membrane - derived stem cells ( hsmsc).materials and methodsmesenchymal stem cells derived from the human maxillary sinus membrane were treated with various concentrations of simvastatin . \n the adequate concentration of simvastatin for osteogenic induction was determined using bone morphogenetic protein ( bmp-2 ) . \n the efficacy of osteogenic differentiation of simavastatin was verified using osteocalcin mrna , and the mineralization efficacy of hsmscs and simvastatin treatment was compared with alkaline phosphatase and von kossa staining.resultsexpression of bmp-2 mrna and protein was observed after three days and was dependent on the concentration of simvastatin . \n expression of osteocalcin mrna was observed after three days in the 1.0 m simvastatin - treated group . \n mineralization was observed after three days in the simvastatin - treated group.conclusionthese results suggest that simvastatin induces the osteogenic potential of mesenchymal stem cells derived from the human maxillary sinus membrane mucosa .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: this was a population - based cross - sectional study , carried out in bavi district , a rural community located 60 km west of hanoi , the capital , within the epidemiological field laboratory of bavi ( filabavi ) . \n the filabavi health and demographic surveillance system ( hdss ) , supported by sida / sarec , was established in 1999 with a sample of around 50,000 individuals from the bavi district . \n people aged 50 and over accounted for about 17% of the total population under surveillance . \n the surveyed population includes three distinct groups : those in mountainous areas , highlands , and riverside or island dwellers ( 10 ) . \n face - to - face household interviews were planned for all people aged 50 years and over who lived in the filabavi area between the end of 2006 and the beginning of 2007 . \n the interviews were done by trained surveyors from filabavi using a summary version of the sage questionnaire ( available as a supplementary file to this paper ) . \n the questionnaire was translated into the local language and pre - tested before official use . \n spot - checks and re - checks on sample data were conducted by supervisors for quality control . \n health status scores were calculated based on self - reported health levels in eight health domains covering : affect , cognition , interpersonal activities and relationships , mobility , pain , self - care , sleep / energy , and vision . \n each domain included at least two questions . asking more than one question about difficulties in a given domain \n provides more robust assessments of individual health levels and reduces measurement error for any single self - reported item . \n health status scores were computed by using item response theory ( irt ) parameter estimates in winsteps , a rasch measurement software package ( http://www.winsteps.com ) . \n respondents who had health status scores below the median were categorised as having poor health status . \n qol was assessed by using the eight - item version of the world health organization quality of life instrument ( whoqol ) . \n results from the eight items were summed to get an overall whoqol score which was then transformed into a 0100 scale . \n the higher the whoqol score , the better the qol . respondents who had whoqol scores less than the median were considered as having poor qol . more details on how scores for this study were derived are given elsewhere ( 9 ) . \n explanatory variables included sex ( male , female ) , age ( grouped as 5059 , 6069 , 7079 , 80 + years ) , educational level ( no formal education , up to 6 years of formal education , more than 6 years of education ) , marital status ( in current partnership , never married , separated , divorced and widowed ) , wealth index ( quintiles ) , whether respondent stays alone ( yes or no ) and proportion of people aged 50 years and over in the same household ( < 25% , 2549% , 5074% , > 75% + ) . \n variables used included household income , the area of land owned , type of house , materials of roof and floor , toilet facilities , electricity and water supplies , and ownership of a range of durable assets for each household . \n the data analysis began with calculation of frequencies and percentages of the variables of interest . \n multivariable logistic regressions were then carried out to examine the association between health status and qol with the selected explanatory variables . \n all subjects in the study were asked for their written informed consent before collecting data , and they had complete right to withdraw from the study at any time without disadvantage . \n this was a population - based cross - sectional study , carried out in bavi district , a rural community located 60 km west of hanoi , the capital , within the epidemiological field laboratory of bavi ( filabavi ) . \n the filabavi health and demographic surveillance system ( hdss ) , supported by sida / sarec , was established in 1999 with a sample of around 50,000 individuals from the bavi district . \n people aged 50 and over accounted for about 17% of the total population under surveillance . \n the surveyed population includes three distinct groups : those in mountainous areas , highlands , and riverside or island dwellers ( 10 ) . \n face - to - face household interviews were planned for all people aged 50 years and over who lived in the filabavi area between the end of 2006 and the beginning of 2007 . \n the interviews were done by trained surveyors from filabavi using a summary version of the sage questionnaire ( available as a supplementary file to this paper ) . \n the questionnaire was translated into the local language and pre - tested before official use . \n spot - checks and re - checks on sample data were conducted by supervisors for quality control . \n health status scores were calculated based on self - reported health levels in eight health domains covering : affect , cognition , interpersonal activities and relationships , mobility , pain , self - care , sleep / energy , and vision . \n asking more than one question about difficulties in a given domain provides more robust assessments of individual health levels and reduces measurement error for any single self - reported item . \n health status scores were computed by using item response theory ( irt ) parameter estimates in winsteps , a rasch measurement software package ( http://www.winsteps.com ) . \n respondents who had health status scores below the median were categorised as having poor health status . \n qol was assessed by using the eight - item version of the world health organization quality of life instrument ( whoqol ) . \n results from the eight items were summed to get an overall whoqol score which was then transformed into a 0100 scale . the higher the whoqol score , the better the qol . respondents who had whoqol scores less than the median \n were considered as having poor qol . more details on how scores for this study were derived are given elsewhere ( 9 ) . \n explanatory variables included sex ( male , female ) , age ( grouped as 5059 , 6069 , 7079 , 80 + years ) , educational level ( no formal education , up to 6 years of formal education , more than 6 years of education ) , marital status ( in current partnership , never married , separated , divorced and widowed ) , wealth index ( quintiles ) , whether respondent stays alone ( yes or no ) and proportion of people aged 50 years and over in the same household ( < 25% , 2549% , 5074% , > 75% + ) . \n variables used included household income , the area of land owned , type of house , materials of roof and floor , toilet facilities , electricity and water supplies , and ownership of a range of durable assets for each household . \n health status scores were calculated based on self - reported health levels in eight health domains covering : affect , cognition , interpersonal activities and relationships , mobility , pain , self - care , sleep / energy , and vision . \n asking more than one question about difficulties in a given domain provides more robust assessments of individual health levels and reduces measurement error for any single self - reported item . \n health status scores were computed by using item response theory ( irt ) parameter estimates in winsteps , a rasch measurement software package ( http://www.winsteps.com ) . \n respondents who had health status scores below the median were categorised as having poor health status . \n qol was assessed by using the eight - item version of the world health organization quality of life instrument ( whoqol ) . \n results from the eight items were summed to get an overall whoqol score which was then transformed into a 0100 scale . the higher the whoqol score , the better the qol . respondents who had whoqol scores less than the median \n were considered as having poor qol . more details on how scores for this study were derived are given elsewhere ( 9 ) . \n explanatory variables included sex ( male , female ) , age ( grouped as 5059 , 6069 , 7079 , 80 + years ) , educational level ( no formal education , up to 6 years of formal education , more than 6 years of education ) , marital status ( in current partnership , never married , separated , divorced and widowed ) , wealth index ( quintiles ) , whether respondent stays alone ( yes or no ) and proportion of people aged 50 years and over in the same household ( < 25% , 2549% , 5074% , > 75% + ) . \n variables used included household income , the area of land owned , type of house , materials of roof and floor , toilet facilities , electricity and water supplies , and ownership of a range of durable assets for each household . \n the data analysis began with calculation of frequencies and percentages of the variables of interest . \n multivariable logistic regressions were then carried out to examine the association between health status and qol with the selected explanatory variables . \n the protocol of this study was approved by the scientific board of filabavi . all subjects in the study \n were asked for their written informed consent before collecting data , and they had complete right to withdraw from the study at any time without disadvantage . \n of the total 8,874 people aged 50 and over living in the study setting at the time of the survey , there were 8,535 who participated in the study ( amounting to 96% ) . \n four percent of subjects were unable to participate as they were away ( 2.3% ) or were not healthy enough to take part in the survey ( 1.7% ) . \n the background characteristics of potential study subjects ( respondents and non - respondents ) are described in table 1 . \n there were no significant differences in socio - economic characteristics between the respondents and the non - respondents . \n background characteristics of study subjects ( respondents and non - respondents ) \n table 2 presents the distribution of health status scores of the study population by age and sex . \n the overall mean health status score was 66.2 and median 65.0 . in both sexes , the average health status scores decreased with age . \n overall , the proportion of respondents with below - median health status among men and women was 39.1 and 58.3% , respectively . \n distribution of health status scores by age and sex among 8,535 adults aged 50 years and over in northern rural viet nam a similar pattern was observed for qol . \n overall , the proportion of respondents with poor qol among men and women was 38.4 and 52.4% , respectively ( table 3 ) . \n distribution of whoqol scores by age and sex among 8,535 adults aged 50 years and over in northern rural viet nam \n fig . \n women were shown to have poorer health status than men . about 25.9% of men and 40% of women reported having both poor health status and poor qol . \n distribution ( % ) of study subjects by health status and quality of life , among 8,535 adults aged 50 years and over in northern rural viet nam . \n multivariate logistic regression analyses of the association between poor health status and poor qol , and socio - economic status are shown in table 4 . \n men were shown to be significantly less likely to have poor health status compared to women . \n people with lower educational levels had a significantly higher probability of having poor health status than those with higher educational levels . \n study subjects in the lowest wealth quintile were more likely to have poor health status than those belonging to the highest wealth quintile . \n there was no significant association between poor health status and marital status or the proportion of older people living in a household . \n factors associated with poor health status and poor quality of life among 8,535 adults aged 50 years and over in northern rural viet nam \n\t\t\t\t\t\t\t\t significant results ( 95% ci does not include 1 ) . \n similar to the pattern of health status , poor qol was shown to be significantly associated with women , older ages , lower educational levels and lower economic status . \n table 4 shows that the respondents whose families had more elderly people ( 75% people aged over 50 years and over in the same household ) were significantly less likely to have poor qol . \n women , older people , those with lower educational level , without any marital partnership , with fewer older people in the family and with lower economic status were more likely to have both poor health status and poor qol . \n of the total 8,874 people aged 50 and over living in the study setting at the time of the survey , there were 8,535 who participated in the study ( amounting to 96% ) . \n four percent of subjects were unable to participate as they were away ( 2.3% ) or were not healthy enough to take part in the survey ( 1.7% ) . \n the background characteristics of potential study subjects ( respondents and non - respondents ) are described in table 1 . \n there were no significant differences in socio - economic characteristics between the respondents and the non - respondents . \n table 2 presents the distribution of health status scores of the study population by age and sex . \n the overall mean health status score was 66.2 and median 65.0 . in both sexes , the average health status scores decreased with age . \n overall , the proportion of respondents with below - median health status among men and women was 39.1 and 58.3% , respectively . \n distribution of health status scores by age and sex among 8,535 adults aged 50 years and over in northern rural viet nam a similar pattern was observed for qol . \n overall , the proportion of respondents with poor qol among men and women was 38.4 and 52.4% , respectively ( table 3 ) . \n distribution of whoqol scores by age and sex among 8,535 adults aged 50 years and over in northern rural viet nam \n fig . \n women were shown to have poorer health status than men . about 25.9% of men and 40% of women reported having both poor health status and poor qol . \n distribution ( % ) of study subjects by health status and quality of life , among 8,535 adults aged 50 years and over in northern rural viet nam . \n multivariate logistic regression analyses of the association between poor health status and poor qol , and socio - economic status are shown in table 4 . \n men were shown to be significantly less likely to have poor health status compared to women . \n people with lower educational levels had a significantly higher probability of having poor health status than those with higher educational levels . \n study subjects in the lowest wealth quintile were more likely to have poor health status than those belonging to the highest wealth quintile . \n there was no significant association between poor health status and marital status or the proportion of older people living in a household . \n factors associated with poor health status and poor quality of life among 8,535 adults aged 50 years and over in northern rural viet nam \n\t\t\t\t\t\t\t\t significant results ( 95% ci does not include 1 ) . \n similar to the pattern of health status , poor qol was shown to be significantly associated with women , older ages , lower educational levels and lower economic status . \n table 4 shows that the respondents whose families had more elderly people ( 75% people aged over 50 years and over in the same household ) were significantly less likely to have poor qol . \n women , older people , those with lower educational level , without any marital partnership , with fewer older people in the family and with lower economic status were more likely to have both poor health status and poor qol . \n this article describes the pattern of health status and qol among older adults in a rural community of viet nam . \n it reveals socio - economic inequalities in health status and qol among older adults in the study setting . \n we found that a higher proportion of women reported both poor health status and poor qol compared to men . \n the findings are in line with recent studies on health status from other asian countries such as pakistan ( 13 ) , bangladesh ( 14 ) and singapore ( 15 ) . \n gender inequality in health has been well documented in the international literature ( 16 ) . \n the findings are also consistent with results from previous studies on qol that reported female disadvantages in both emotional and subjective well - being ( 1720 ) . \n one likely explanation could be that women are more likely to suffer from conditions that are debilitating but not fatal . \n the paradox is that women report poorer health but live longer , and this is true in almost every country in the world ( 21 ) . \n age was shown to be a factor significantly associated with poor health status and poor qol . \n this has been consistently shown in previous studies ( 1315 ) . in our setting , \n chronic diseases were shown to be more prevalent among women and older people ( 22 ) . \n we found that higher educational levels were significant positive predictors of both health status and qol among the study subjects . \n education is well known as an important factor for health , both among men and women , particularly in rural areas . \n education is assumed to have a positive effect on health status since persons with more education are assumed to be better informed about health matters , diet and disease prevention measures leading to better health conditions , consequently leading to higher qol . \n similarly , improvements in economic status are also likely to raise both health status and qol . \n in addition to providing means for purchasing health care , higher economic status can provide better nutrition , housing and recreational opportunities . \n the findings are consistent with previous studies in asia ( 1315 ) , in europe ( 23 ) and the americas ( 24 ) . \n this study also revealed a positive effect of having more older people living in the same family . \n this positive effect may be the result of mutually practised health beliefs and behaviours , shared physical environments and interpersonal relations between the older people in the same family . \n firstly , the cross - sectional nature of the data limited our ability to understand causal mechanisms that resulted in particular heath status and qol outcomes among the study population . secondly \n , low educational level and the presence of impaired cognition in older people might have led to inaccuracies in the self - reported data . \n our careful training and field supervision would have overcome this problem to some extent . in summary \n , this study provides cross - sectional evidence on patterns of health status and qol among older adults in rural viet nam . \n the findings reveal problems of inequality in health status and qol among older adults in the study setting by sex , age , education and economic status . given these findings , actions to enhance the health of disadvantaged people ( women , the elderly , less educated and lower economic status ) are needed in this setting . \n access the supplementary material to this article : indepth who - sage questionnaire ( including variants of vignettes ) , a data dictionary and a password - protected dataset ( see supplementary files under reading tools online ) . to obtain a password for the dataset , please send a request with sage data as its subject , detailing how you propose to use the data , to global.health@epiph.umu.se \n the authors have not received any funding or benefits from industry to conduct this study .", "role": "user" }, { "content": "backgroundto effectively and efficiently respond to the growing health needs of older people , it is critical to have an indepth understanding about their health status , quality of life ( qol ) and related factors . \n this paper , taking advantage of the indepth who - sage study on global ageing and adult health , aims to describe the pattern of health status and qol among older adults in a rural community of viet nam , and examine their associations with some socio - economic factors.methodsthe study was carried out in the bavi district , a rural community located 60 km west of hanoi , the capital , within the epidemiological field laboratory of bavi ( filabavi ) . \n face - to - face household interviews were conducted with people aged 50 years and over who lived in the filabavi area . \n the interviews were performed by trained surveyors from filabavi using a standard summary version sage questionnaire . \n both descriptive and analytical statistics were used to examine the patterns of health status and qol , and associations with socio - economic factors.resultshigher proportions of women reported both poor health status and poor qol compared to men . \n age was shown to be a factor significantly associated with poor health status and poor qol . \n higher educational level was a significant positive predictor of both health status and qol among the study subjects . \n higher economic status was also associated with both health status and qol . \n the respondents whose families included more older people were significantly less likely to have poor qol.conclusionthe findings reveal problems of inequality in health status and qol among older adults in the study setting by sex , age , education and socio - economic status \n . given the findings , actions targeted towards improving the health of disadvantaged people ( women , older people and lower education and economic status ) are needed in this setting .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: blue rubber bleb nevus syndrome ( brbns ) is a rare disorder characterized by venous malformations of the skin , gastrointestinal ( gi ) tract , and less commonly of other tissues in the body . \n we present the case of a 59-year - old man and a review of current literature to illuminate the diversity of ophthalmic symptoms , as well as potential treatment options for patients with blue rubber bleb nevus syndrome . \n the patient was initially diagnosed with a familial case of blue rubber bleb nevus syndrome as a child after developing multiple cutaneous lesions on his back , chest , face , and extremities . \n no major complications from the disease were apparent until july , 2012 ( apart from mild anemia ) , when he presented to his local hospital with loss of vision in his right eye , and chronic pain in the left eye . \n magnetic resonance angiography ( mra ) revealed a dural arteriovenous fistula ( davf ) in the left skull base ( figure 1 ( fig . \n 1 ) ) . a total of three endovascular embolization procedures were used to treat the patient s ophthalmic symptoms . \n partial occlusion of the davf after the first procedure failed to alleviate all of his optic symptoms , as the patient developed morning blurriness and diplopia . \n he appears to have slight pallor of the optic nerves , as well as imbalance and vertigo , but his overall health has greatly improved . \n approximately 200 reports of the syndrome have been documented to date . although some forms appear to have an autosomal dominant inheritance pattern linked to chromosome 9p , the majority of cases are sporadic . \n patients often present with small , compressible lesions of the skin and chronic anemia from gastrointestinal bleeding . \n diagnosis of the disease usually occurs from these common presentations as well as histological analysis . \n radical resection has had positive results in patients with gastrointestinal hemorrhage , and laser therapy has shown promise for cutaneous lesions , . \n we present here some of the few cases of ophthalmic symptoms associated with this disorder ( table 1 ( tab . \n 1 ) ) . past reports of orbital involvement of blue rubber bleb nevus syndrome have presented with a myriad of symptoms including : proptosis , enophthalmos , ptosis , vision loss , imbalance , occipital headaches , ecchymosis , and intraocular pressure . \n ophthalmic symptoms can be chronic or acute in nature , as with the case of a sudden intraorbital hemorrhage . \n cutaneous lesions were also present in most patients ( 10/11 cases ) , and gastrointestinal lesions in a subset of reports ( 5/11 cases ) \n . it can be difficult to differentiate brbns from mafucci s syndrome , klippel - trenaunay syndrome , diffuse neonatal angiomatosis , and rendu - osler - weber syndrome . based on our analysis \n , the common orbital symptoms of blue rubber bleb nevus syndrome may include : the presence of cutaneous venous malformations , chronic anemia , decreased visual acuity , and proptosis or enopthalmos of the eye . systemic therapy has not yet successfully treated optic cases of brbns , but has achieved varied outcomes in other forms of the disease . \n multiple reports have shown interferon- and corticosteroids to be unsuccessful in long - term treatment of multi focal , large gi lesions , . \n however , recent publications have suggested that sirolimus may be beneficial in the treatment of gastrointestinal bleeds and anemia , . \n therapy with sirolimus can cause many side effects including hypertension , immunosuppression , renal failure , anemia and hyperlipidemia . in these cases \n no significant adverse symptoms were recorded except for a slight elevation in cholesterol in one patient . \n sirolimus was used in combination with propranolol to reduce adverse effects , but cutaneous lesions and melena were noted 10 days after the physicians attempted to wean the patient off the medication . \n this agent may , therefore , be considered for short - term treatment of ophthalmic forms of brbns , but prolonged use of the drug may be needed to successfully reduce symptoms . \n unfortunately , long - term use of sirolimus will likely expose patients to the potential adverse side effects of the drug . \n surgical treatment of chronic ophthalmic manifestations has been successful in our case as well as the other patients in our literature review , . \n endovascular embolization has been used to alleviate symptoms in patients presenting with dural arteriovenous fistulas , and surgical excision with other venous malformations in the orbit . \n embolization procedures have reduced symptoms in all the cases presented , but multiple procedures were often required . \n understanding the critical symptoms and potential treatment options will optimize the management of patients with this extremely rare disease and hopefully lead to improved outcomes . \n the patient in the case report gave consent to the authors to write up this publication and gave permission to use the images . \n the patient in the case report gave consent to the authors to write up this publication and gave permission to use the images . \n ", "role": "user" }, { "content": "objective : blue rubber bleb nevus syndrome is a rare vascular disease most commonly associated with venous malformations of the skin and the gastrointestinal tract . \n few ophthalmic cases have been reported to date , and no clear treatment regimen exists . \n we describe the case of a 59-year - old man , along with a review of literature , to help in the future diagnosis and treatment of patients with the disease . \n methods : this paper is an observational case report and a review of medical literature on the syndrome from 1981 to present.results : our patient developed a dural arteriovenous fistula in his orbit after being diagnosed with a familial form of blue rubber bleb nevus syndrome . \n multiple endovascular embolization procedures eliminated all of his ocular symptoms . \n surgical procedures were also successful in other cases reviewed , and similar symptoms were seen across cases.conclusions : comparing our case with other ophthalamic reports in literature , surgical intervention appears to be a plausible long - term treatment for optic manifestations of blue rubber bleb nevus syndrome . \n systemic therapies , including sirolimus and corticosteroids , have had limited success in the long - term treatment of other forms of blue rubber bleb nevus syndrome , and therefore are not recommended in the treatment of ocular symptoms .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: marine sponges have \n emerged as excellent sources of structurally \n diverse natural products that possess activities \n against a number of important disease targets and as such serve as \n leads in medicinal chemistry programs . \n our laboratories have developed interests in the imidazole - containing \n alkaloids belonging to the oroidin or \n the leucetta families of natural \n products . \n these synthetic programs have largely centered on the discovery \n and development of new methods and strategies for the elaboration \n of simple imidazole derivatives rather than the de novo construction of the imidazole ring . in this manuscript \n , \n we describe the application of two such methods to the syntheses of \n three analogs of the imidazonaphthoquinone alkaloids kealiiquinone \n ( 1 ) and 2-deoxy-2-aminokealiiquinone \n ( 2 ) ( figure 1 ) . specifically the use of an intramolecular diels alder \n reaction to construct a polysubstituted dihydrobenzimidazole and oxidation of the imidazole c2-position via \n the intermediacy of imidazolium salts . \n kealiiquinone ( 1 ) was isolated from \n a leucetta \n sp . found off the islands of guam and saipan by the scheuer \n lab and characterized by a combination of spectroscopic methods and \n x - ray crystallography . \n more recently schmitz \n and co - workers reported the isolation and characterization of the \n 2-amino congener 2 from leucetta chagosensis collected in chuuk state ( part of the federated states of micronesia ) . in both cases , biological activity was not reported \n by the isolation groups , although the presence of the quinone would \n suggest that they might be good bio - michael acceptors . \n the biosynthetic \n details of the leucetta alkaloids are unknown , although \n it has been speculated that kealiiquinone ( 1 ) may be \n derived from the kealiinines ( 35 ) through oxidation of the c - ring . \n an \n alternative hypothesis has been posited involving the intermediacy \n and subsequent cyclization of the quinone - containing naamidine f ( 6 ) . \n prior to our efforts , only one report of a total synthesis of kealiiquinone \n has appeared from the ohta lab resulting in the synthesis of the 2-keto \n tautomer of the natural product \n . a second total \n synthesis of kealiiquinone ( 1 ) and the first of 2-deoxy-2-aminokealiiquinone \n ( 2 ) have been completed by our lab using a complementary \n bioinspired friedel crafts - like strategy to the approach described \n by ohta and co - workers . the synthetic version and a regioisomer \n were \n evaluated in a panel of cancer cell lines and shown to possess modest \n activity , but the activity profile suggested \n a possible unique mechanism of action . \n however , this facet does not \n appear to have been pursued further by the ohta group or anyone else . \n very recently , syntheses of the less oxidized kealiinines have been \n reported independently by two groups , including ours . \n the cytotoxicity of \n the kealiinines ( 35 ) , the kealiiquinones \n ( 1 and 2 ) , and several intermediates have \n been investigated and shown for the most part to possess modest activity \n ( ic50 = 2095 m in mtt growth assay with \n mcf7 breast cancer cell lines ) . \n we were intrigued by the opportunity \n offered by kealiiquinone ( 1 ) to demonstrate the utility \n of several aspects of imidazole chemistry that we have developed , as well as providing a platform to develop novel \n chemistry along the way . \n our experience with diels alder reactions \n of 4-vinylimidazoles suggested that an intramolecular variant of this \n chemistry would provide rapid and controlled access to polysubstituted \n benzimidazole ( 89 , figure 2 ) . in our hands \n we have found the 4-isomers to be superior in the cycloaddition \n chemistry and thus the choice of a benzyl protecting group in enyne 9 would permit the position - selective incorporation of the \n methyl substituent later in the synthetic sequence . whereas the cycloaddition \n chemistry would properly position substituents around the core framework , \n a method was needed for the installation of the 2,3-dimethoxyquinone \n moiety . while there were limited options for this transformation , \n chemistry reported by venuti , using a \n cyanide - catalyzed , benzoin - like condensation via \n elaboration at the c2-position of the imidazole via metalation \n and electrophilic quench would then deliver either of the two kealiiquinone \n natural products 1 and 2 from a common late - stage \n intermediate ( figure 2 ) . \n overview \n of the synthetic strategy toward kealiiquinone . as part of our exploratory efforts \n on the intramolecular diels alder and as a prelude to an approach \n to kealiiquinone we had prepared dihydrobenzimidazole 14 from n - benzyl protected enyne 13 in 83% yield . \n c \n and air , and while these conditions provided the required product 15 ( scheme 1 ) , it gave inconsistent \n yields and required extended reaction times for good conversions . \n subsequently , we found that the aromatization could be effected efficiently \n and reproducibly in the presence of mno2 , delivering 15 in 91% yield . \n since these preliminary experiments were successful , we \n decided \n to construct the full substrate which required the preparation of \n the anisyl substituted propiolic acid 12 . \n this was prepared \n through a sonogashira cross - coupling between p - iodoanisole \n and propargyl alcohol ( 85% ) . \n oxidation \n of the product with mno2 provided the aldehyde ( 85% ) and a subsequent pinnick oxidation produced \n the corresponding propiolic acid 12 ( 60% ) . \n the acid 12 was coupled with the benzyl protected vinylimidazole 10 through a dcc - mediated condensation delivering enyne 9 ( scheme 1 ) . \n subjection of the thus \n obtained enyne 9 to heating in toluene at reflux for \n 48 h resulted in the formation of the dihydrobenzimidazole 8 in excellent yield . \n oxidation with mno2 affords \n the aromatized benzimidazole 16 in good yield ( scheme 1 ) . \n the structure of 16 was confirmed \n through x - ray crystallography ( see si ) \n which nicely illustrated the relative location of the two aromatic \n substituents . \n of note is the slight pyramidalization \n of the imidazole n1 ( = 353.89 ) and the deviation \n from orthogonality around the c3c10 bond ( crystallographic \n numbering ) . \n presumably both structural effects minimize nonbonded \n interactions between the n - benzyl and anisyl moieties . \n at this stage , both the unsubstituted and p - methoxy \n systems 15 and 16 were taken through the \n same set of reactions to construct the imidazonaphthoquinone \n framework but in slightly different sequences as described in schemes 2 , 3 , and 4 . \n initially , it was decided \n to introduce the n - methyl group on the p - anisyl - containing substrate 16 , as this can be accomplished \n via the intermediacy of the imidazolium salt by hydrogenolysis of \n the benzyl group ( scheme 2 ) . \n conversion of 16 to the imidazolium salt 17 was accomplished \n uneventfully by treatment with methyl iodide , and subsequent catalytic \n hydrogenation of 17 delivered the corresponding debenzylated n - methyl benzimidazole derivative 18 . in our \n preliminary experiments we noted some variability in the yield of \n this reaction . \n ultimately however it was recognized that hydrolysis \n of the lactone occurred ; treatment of the crude hydrogenation product \n with aqueous acid recyclized the -hydroxy acid thus providing \n improved yields and reproducibility . \n this strategy for introduction \n of the methyl group has the advantage of avoiding potential selectivity \n issues in comparison to an approach involving direct methylation of \n the parent benzimidazole . \n reduction of \n the lactone to the corresponding diol 19 was accomplished \n effectively with dibal , setting the stage for the installation of \n the final ring . in order to accomplish this critical transformation , \n initial \n attempts to convert diol 19 to the phthaldehyde derivative 7 with common oxidants were thwarted by the formation of the \n lactone ; only swern conditions were partially successful . in this \n latter case \n there was some batch to batch inconsistency , the cause \n of which we were unable to track down . \n fortunately , this led us to \n investigate reagents that behaved mechanistically similarly to swern \n reagents ; among these , the best system we identified was reported \n by mukaiyama and co - workers involving sulfinimidoyl chlorides , e.g. , 20 in the presence of dbu . \n this \n reagent combination reproducibly resulted in the formation of the \n phthaldehyde 7 in yields between 65% and 75% . with the \n aldehyde in hand we were in position to evaluate venuti s method \n for its conversion into the corresponding dihydroxy benzoquinone 22 . \n gratifyingly , we found that \n the 2,3-dihydroxybenzoquinone 22 was \n produced reproducibly , albeit , in a modest yield of approximately \n 40% . at this stage , \n all that remained to be done to complete the total \n synthesis of kealiiquinone ( 1 ) was the conversion of \n the hydroxyl groups to methyl ethers and oxidation at c2 . \n the first \n task was accomplished by taking advantage of the fact that the oh \n groups are part of a vinylogous acid and as such reacted efficiently \n with tms - diazomethane , delivering dimethoxyquinone 23 . however , the introduction of functionality at c2 was significantly \n more difficult . \n we and others have routinely used metalation by deprotonation \n at c2 with a strong base ( buli or lda ) and followed that by trapping \n with a suitable electrophile : in this context either \n ( tmso)2 for kealiiquinone ( 1 ) or tsn3 for the amino congener 2 . \n multiple attempts to perform this \n chemistry with 23 as a substrate simply failed to produce \n the anticipated products ; indeed these reactions did not produce any \n products that we were able to characterize . experiments to trap out \n any metalated intermediates with d2o or methanol - d4 were similarly unsuccessful . \n although we have no experimental \n evidence to support our hypothesis , we suspect that electron transfer \n processes are intervening . as a means to circumvent these issues , \n we briefly attempted to introduce functionality at c2 earlier in the \n sequence by metallating with n - buli on lactones 16 or 18 ; however issues with solubility or deprotonation \n occurring on the n - benzyl or the lactone methylene \n compromised this strategy . \n alternative timings for introducing the \n c2-functionality were rejected as these options invariably increased \n the step count unacceptably . given \n this late stage failure \n to functionalize at c2 of the naphthimidazole , we began to contemplate \n our options for moving the synthesis forward , and it occurred to us \n that the imidazolium salts that were prepared as a means to selectively \n methylate the benzimidazoles may be easier to functionalize given \n the increased acidity of the proton at c2 in these systems . \n based on this hypothesis , we have developed \n an extremely mild method for introducing oxygen at c2 through exposure \n of the imidazolium ion to bleach or alternatively nitrogen can be \n introduced through exposure to n - chlorosulfonamides \n or n - chlorocarbamates ; this chemistry has been \n described in more detail elsewhere . with \n this method in hand we subjected the lactone imidazolium species 17 to bleach and found that it too underwent the oxidation \n to provide the 2-benzimidazolone 24 in good yield ( scheme 3 ) . \n an x - ray structure determination on the dibal - reduction \n product 25 unequivocally demonstrates the introduction \n of the 2-oxo moiety and served to confirm the relative location of \n the methyl and aryl groups . at this point \n we elaborated the benzimidazolone 25 in a largely analogous \n fashion to the corresponding phthaldehyde 26 . \n although \n the efficiency was lower , dialdehyde 26 was converted \n into the dihydroxyquinone 27 and subsequently the hydroxyl \n groups were converted into methoxy groups by treatment with tms - diazomethane \n ( 27 28 , scheme 3 ) . \n all that remained to complete the revised synthesis of \n kealiiquinone ( 1 ) was removal of the n - benzyl protecting group ; unfortunately this proved to be exceedingly \n difficult . while n - benzyl groups can be removed quite \n readily from amines under a variety of conditions , amides on the other \n hand tend to be much more challenging substrates . \n indeed it was found \n that standard reductive or oxidative conditions failed to remove the n - benzyl moiety . \n ultimately , \n we determined that heating the substrates in neat triflic acid resulted \n in debenzylation with reasonable efficiencies . \n unfortunately , in the case of the precursor to kealiiquinone , \n these reaction conditions led to the selective demethylation of the \n 7-methyl and thus the formation of 7-desmethylkealiiquinone \n ( 29 ) . in an effort to access the natural product , \n various \n attempts were made to remethylate the phenolic hydroxyl group . in \n our hands \n this resulted in overmethylation ( methylation of the imidazole \n nitrogen and the hydroxyl group ) or n - methylation \n under a variety of different conditions and with different methyl \n sources . \n an \n alternative strategy \n was explored concurrently with the one described above in which the \n introduction of the c2-substituent was delayed until the end of the \n synthesis . in part \n this was undertaken in order to not only evaluate \n the strategy in general but also establish whether the highly functionalized \n quinones were viable substrates in our oxidative chemistry . \n thus lactone 15 was reduced to the corresponding diol 30 and \n then converted to the phthaldehyde 31 upon treatment \n with 20 ( scheme 4 ) . \n conversion \n to the 2,3-dihydroxyquinone 32 was accomplished \n with glyoxal 21 , followed by methylation with tms - diazomethane . \n methylation of the imidazole nitrogen to prepare the imidazolium salt 34 was then carried out by exposure of 33 to \n methyl iodide in acetonitrile at reflux . \n initial oxidation experiments \n of 34 with bleach resulted in the formation of the imidazolone 35 , but the yields were rather low , ca . \n fortunately , we had established previously that other chloronium sources \n effect this chemistry , and so exposure of 34 to ncs and \n aqueous potassium carbonate delivered the imidazolone 35 in 60% yield . \n reaction of the imidazolone \n with tfoh at 55 c resulted in debenzylation and the formation \n of 4-desmethoxykealiiquinone ( 36 ) . with the imidazolium salt in hand \n , we also took the opportunity \n to evaluate the introduction of an imino group through exposure to \n an n - chlorocarbamate ( scheme 4 ) ; to date only simple benzimidazolium and imidazolium salts had \n been investigated in this chemistry . while \n the yield turned out to be quite modest , the reaction of 34 with n - chloro tert - butylcarbamate \n ( produced in situ from tert - butyl \n carbamate and t - buocl ) did produce the expected product 37 ( scheme 4 ) . \n a preliminary attempt \n to remove the bn- and boc - protecting groups by sequential treatment \n with tfoh and then tfa was not successful . one further strategy was evaluated to prepare the imidazolium \n salt \n near the end of the synthesis with a more easily removable n - substituent , but one that was capable of activating the \n oxidation ( scheme 5 ) . to test the feasibility \n of such an approach given that the n3-position is relatively hindered , \n lactone 18 was used as a model and converted to the corresponding \n dimethyl imidazolium salt 38 . \n gratifyingly , treatment \n of the salt with bleach resulted in c2-oxidation to deliver imidazolone 39 in good yield suggesting that late stage functionalization \n through application of our chemistry was achievable . frustratingly \n however , \n . specifically , attempted introduction of a mom \n group was unsuccessful , whereas the attempted incorporation of a sem \n group resulted in the net protonation \n of the imidazole and production of the imidazolium salt 41 . \n the significant downfield shift of \n the c2-proton absorbance in the h nmr spectrum of the \n product is characteristic of the formation of an imidazolium species . \n this was further confirmed when an x - ray crystal structure of this \n product was obtained , which clearly indicates the protonation of the \n nitrogen along with chloride as the counterion ( see supporting information ) . \n our assumption here is that the proximity \n of the aryl group to the nitrogen atom renders it sterically hindered \n and only small electrophiles can access it . \n attempts to oxidize the \n protonated imidazolium salt 41 with bleach to the corresponding \n imidazolone 42 were not successful . \n presumably , the basic \n nature of the reaction conditions simply results in deprotonation \n and inactivation of the imidazole . \n efficient \n routes have been developed to the heterocyclic framework \n of kealiiquinone through application of an intramolecular diels \n alder \n reaction of an enyne , c2-functionalization through a novel oxidation \n of imidazolium salts , and application of an underutilized method for \n the formation of dihydroxy quinones from phthaldehydes . \n indeed , this \n work constitutes the first application of venuti s method for \n the de novo construction of a 2,3-dimethoxyquinone . \n unfortunately deprotection of an n - benzyl moiety \n from the penultimate intermediate resulted in concomitant demethylation \n delivering 7-desmethylkealiiquinone rather \n than the natural product . \n similar chemistry has been used to obtain \n related materials lacking the 4-methoxy group and the 2-oxo \n group which will enable the sar investigations of this unique family \n of natural products . \n nmr \n spectra were obtained at either 500 or 300 mhz ( for h \n nmr spectra ) or 125 or 75 mhz ( for c nmr ) in cdcl3 unless indicated otherwise . \n h nmr spectra were \n referenced to residual protiosolvent unless indicated otherwise ( chcl3 = 7.26 ppm ) . \n for spectra recorded in other solvents , \n residual meoh ( = 3.31 ppm ) or dmso ( = 2.50 ppm ) were \n used as internal references . \n c nmr spectra were recorded \n in cdcl3 ( unless otherwise indicated ) using residual chcl3 ( = 77.2 ppm ) as an internal reference . for spectra \n recorded in other solvents , residual meoh ( = 39.5 ppm ) and \n dmso ( = 49.0 ppm ) \n infrared \n ( ir ) spectra were obtained on neat samples ( atr spectroscopy ) or using \n either kbr pellets for solids or neat films on nacl plates for liquids \n ( transmission ) . \n analytical thin - layer chromatography ( tlc ) was performed \n on silica gel plates , 200 mesh with f254 indicator . \n visualization \n was accomplished by uv light ( 254 nm ) and/or a 10% ethanol solution \n of kmno4 . \n high resolution mass spectra ( hrms ) were recorded \n by electrospray ionization ( esi - tof ) or atmospheric - pressure chemical \n ionization ( apci - tof ) unless otherwise indicated . \n propiolic acid 12 ( 2.00 g , 11.4 mmol ) , alcohol 10 ( 2.94 g , 13.7 mmol ) , dmap ( 0.14 g , 1.2 mmol ) , and camphorsulfonic \n acid ( 0.16 g , 0.70 mmol ) were dissolved in ch2cl2 ( 50 ml ) and cooled to 78 c under n2 . \n dcc \n ( 3.50 g , 17.1 mmol ) dissolved in ch2cl2 ( 20 \n ml ) was added dropwise . \n the mixture was allowed to come to rt and \n stir for 2 h. the solids were filtered through celite and washed with \n ch2cl2 . the filtrate was reduced , and the resulting \n brown residue was purified by column chromatography ( 8:2 etoac / hexane ) \n to afford 9 as an off - white solid ( 2.53 g , 56% ) . \n mp : \n 6567 c ; h nmr : = 7.52 ( s , 1h ) , 7.49 \n ( d , j = 2.1 hz , 2h ) , 7.367.31 ( m , 3h ) , 7.157.13 \n ( m , 2h ) , 6.86 ( m , 3h ) , 6.58 ( d , j = 15.6 hz , 1h ) , \n 6.476.38 ( dt , j = 15.5 , j = 6.2 hz , 1h ) , 5.06 ( s , 2h ) , 4.82 ( d , j = 6.6 hz , \n 2h ) , 3.81 ( s , 3h ) ; c nmr : = 161.6 , 154.2 , 139.8 , \n 137.9 , 135.9 , 135.0 , 129.1 , 128.5 , 127.4 , 127.0 , 120.6 , 117.9 , 114.4 , \n 111.5 , 87.3 , 80.1 , 66.5 , 55.5 , 51.0 ; ir ( kbr , cm ) : 3105 , 2195 , 1899 , 1697 , 1602 , 1567 , 1541 ; hr - esims ( m / z ) : calcd for [ m + h ] c23h21n2o3 is 373.1547 , found 373.1564 . \n enyne 9 ( 2.54 g , 6.84 mmol ) was dissolved \n in toluene ( 200 ml ) and purged with n2 for 15 min . \n the \n solution was then heated at 130 c in a sealed tube for 48 h. \n the solvent was removed , and the resulting solids were washed with \n diethyl ether to afford 8 as an off - white solid ( 2.14 \n g , 84% ) . \n mp : 199201 c ; h nmr : = \n 7.44 ( s , 1h ) , 7.247.10 ( m , 5h ) , 6.85 ( dd , j = 5.7 , 1.8 hz , 2h ) , 6.64 ( dd , j = 5.1 , 1.5 hz , \n 2h ) , 4.68 ( dd , j = 16.5 , 7.5 hz , 2h ) , 4.43 ( d , j = 15.6 hz , 1h ) , 4.01 ( t , j = 8.7 hz , \n 1h ) , 3.83 ( s , 3h ) , 3.673.58 ( m , 1h ) , 3.06 ( dd , j = 15.0 , 8.7 hz , 1h ) , 2.73 ( dd , j = 17.1 , 15.9 hz , \n 1h ) ; c nmr : = 168.1 , 160.3 , 144.4 , 141.2 , 139.4 , \n 135.8 , 128.7 , 128.4 , 128.1 , 126.6 , 123.9 , 117.2 , 70.6 , 55.4 , 50.3 , \n 38.1 , 28.0 ; ir ( kbr , cm ) : 3465 , 2946 , 1737 , 1621 , \n 1512 , 1252 ; hr - esims ( m / z ) : calcd \n for [ m + h ] c23h21n2o3 373.1547 , found 373.1571 . to 14(11 ) ( 5.5 g , 16 mmol ) \n in ch2cl2 ( 250 ml ) was added mno2 ( 85% act . ) ( 15 g ) followed by heating at 40 c for \n 48 h. the solids were filtered through celite and washed with ch2cl2 . the filtrate was reduced to give the product \n as a white solid ( 5.0 g , 91% ) . \n mp : 185186 c ; h nmr : = 8.02 ( s , 1h ) , 7.85 ( s , 1h ) , 7.42 ( t , j = 7.5 hz , 1h ) , 7.30 ( t , j = 7.5 hz , 2h ) , 7.21 ( d , j = 7.5 hz , 1h ) , 7.187.13 ( m , 4h ) , 6.54 ( d , j = 7.5 hz , 2h ) , 5.37 ( s , 2h ) , 4.93 ( s , 2h ) ; c nmr : = 169.8 , 148.9 , 140.5 , 135.7 , 132.3 , 129.8 , 128.8 , \n 128.6 , 128.1 , 127.9 , 127.6 , 126.2 , 118.4 , 112.6 , 68.1 , 50.3 ; ir ( cm ) : 3005 , 2923 , 1752 , 1497 , 1363 , 1251 , 1130 , 1018 ; \n hr - esims ( m / z ) : calcd for [ m + na ] c22h16n2o2na \n 363.1104 , found 363.1114 . \n 8 ( 7.0 g , 19 mmol ) was dissolved \n in ch2cl2 ( 500 ml ) , and mno2 ( 18.0 \n g , 188 mmol ) was added . then the mixture was stirred at rt for 24 \n h. the solids were filtered and washed with ch2cl2 followed by meoh . \n the filtrate was concentrated to give 16 as an off - white solid ( 6.24 g , 89% ) . \n mp : 176178 c ; h nmr : = 8.01 ( s , 1h ) , 7.82 ( s , 1h ) , 7.217.16 \n ( m , 3h ) , 7.06 ( d , j = 8.9 hz , 2h ) , 6.83 ( d , j = 8.9 hz , 2h ) , 6.59 ( d , j = 7.5 hz , 2h ) , \n 5.35 ( s , 2h ) , 4.97 ( s , 2h ) , 3.84 ( s , 3h ) ; c nmr : \n = 170.0 , 159.8 , 149.4 , 148.9 , 140.5 , 135.8 , 132.7 , 131.0 , 128.7 , 128.0 , \n 127.6 , 126.2 , 124.2 , 118.5 , 113.3 , 112.4 , 68.0 , 55.3 , 50.3 ; ir ( kbr , \n cm ) : 2947 , 1739 , 1614 , 1500 , 1347 , 1253 ; hr - esims \n ( m / z ) : calcd for [ m + na ] c23h18n2o3na 393.1210 , \n found 393.1210 . \n 16 ( 0.37 g , 1.0 mmol ) \n was dissolved in toluene ( 20 ml ) , and mei ( 0.20 ml , 3.2 mmol ) was \n added . \n the mixture was heated at 120 c for 24 h during which \n additional aliquots of methyl iodide ( 0.5 equiv ) were added until \n no starting material could be detected by tlc . \n the solids were filtered \n and washed with etoac to afford 17 as a pale yellow solid \n ( 0.48 g , 93% ) . \n mp : 198200 c ; h nmr : \n = 10.50 ( s , 1h ) , 8.03 ( s , 1h ) , 7.257.12 ( m , 5h ) , 6.946.87 \n ( m , 4h ) , 5.43 ( s , 2h ) , 5.27 ( s , 2h ) , 4.32 ( s , 3h ) , 3.86 ( s , 3h ) ; c nmr : = 167.7 , 160.8 , 146.4 , 144.8 , 136.8 , 132.2 , \n 131.4 , 131.1 , 130.1 , 129.2 , 127.7 , 122.8 , 121.0 , 113.9 , 106.7 , 68.0 , \n 55.5 , 53.2 , 35.4 ; ir ( cm ) : 2939 , 1775 , 1612 , 1575 , \n 1501 , 1451 , 1378 , 1337 ; hr - esims ( m / z ) : calcd for [ m + h ] c24h21n2o3 385.1547 , found 385.1548 . to \n imidazolium salt 17 ( 0.44 g , 0.86 mmol ) was added etoh \n ( 50 ml ) and sat . \n nahco3 ( 5 ml ) , followed by 10% pd / c ( 0.10 \n g ) . \n the reaction mixture was heated at 80 c under h2 ( 1 atm ) for 24 h. the solids were filtered and washed with ch2cl2/meoh followed by water . \n the aqueous solution \n was then acidified to ph = 5 with 3 m hcl and stirred for 2 h. the \n resulting aqueous solution was extracted multiple times with ch2cl2 . \n the resulting solid was \n recrystallized from meoh to afford 18 as a white solid \n ( 0.19 g , 76% ) . \n mp : > 250 c ; h nmr ( dmso - d6 ) : = 8.32 ( s , 1h ) , 7.69 ( s , 1h ) , 7.55 ( d , j = 9.2 hz , 2h ) , 6.98 ( d , j = 9.2 hz , 2h ) , \n 5.39 ( s , \n 2h ) , 3.89 ( s , 3h ) , 3.80 ( s , 3h ) ; c nmr ( dmso - d6 ) : = 170.5 , 159.7 , 147.9 , 143.1 , 142.6 , 139.8 , 133.4 , 133.2 , \n 125.4 , 114.6 , 113.2 , 102.9 , 68.4 , 55.7 , 31.6 ; ir ( kbr , cm ) : 3044 , 2957 , 2835 , 1739 , 1603 , 1501 , 1453 ; hr - esims ( m / z ) : calcd for [ m + h ] c17h15n2o3 295.1077 , found 295.1092 . benzimidazole 18 ( 0.19 g , \n 0.65 mmol ) was dissolved in ch2cl2 ( 5 ml ) and \n cooled to 78 c under n2 . dibal - h ( 1 m in \n hexane ) ( 1.5 ml , 1.5 mmol ) was added dropwise , and the reaction mixture \n was allowed to warm to rt and stirred for 3 h. the reaction mixture \n was cooled to 0 c , and water was added slowly followed by meoh . \n the organic extract was separated , and the residual aqueous \n solution was extracted multiple times with ch2cl2 . \n the resulting white solid was triturated \n with et2o to afford 19 as a white solid ( 0.15 \n g , 79% ) . \n mp : 245247 c ; h nmr ( dmso - d6 ) : = 8.02 ( s , 1h ) , 7.58 ( s , 1h ) , 7.43 ( d , j = 8.6 hz , 2h ) , 6.98 ( d , j = 8.6 hz , 2h ) , \n 5.25 ( t , j = 5.5 hz , 1h ) , 4.83 ( d , j = 5.2 hz , 2h ) , 4.79 ( t , j = 4.8 hz , 1h ) , 4.43 ( d , j = 4.5 hz , 2h ) , 3.81 ( s , 3h ) , 3.79 ( s , 3h ) ; c nmr ( dmso - d6 ) : = 158.8 , 144.7 , 141.6 , 137.6 , \n 134.2 , 133.3 , 132.5 , 129.9 , 129.8 , 113.5 , 108.4 , 62.4 , 58.5 , 55.6 , \n 31.2 ; ir ( cm ) : 3074 , 2889 , 1608 , 1574 , 1340 , 1237 , \n 1174 , 1015 ; hr - esims ( m / z ) : calcd \n for [ m + h ] c17h19n2o3 299.1390 , found 299.1411 . \n diol 19 ( 0.10 g , 0.34 mmol ) \n was dissolved in ch2cl2 , and dbu ( 0.2 ml , 1.34 \n mmol ) was added . \n the solution was cooled to 78 c under \n n2 , and n - tert - butylbenzenesulfinimidoyl \n chloride ( 20 ) ( 0.22 g , 1.02 mmol ) dissolved in ch2cl2 ( 5 ml ) was added dropwise . \n the resulting residue \n was purified by column chromatography ( the silica gel was neutralized \n with 5% et3n prior to purification ) ( etoac to 2% meoh in \n etoac ) to afford 7 as an off - white solid ( 0.069 g , 70% ) . \n h nmr : = 10.61 ( s , 1h ) , 10.16 ( s , 1h ) , 8.09 ( s , 1h ) , \n 8.05 ( s , 1h ) , 7.49 ( d , 2h , j = 8.4 hz ) , 7.08 ( d , \n 2h , j = 8.4 hz ) , 3.97 ( s , 3h ) , 3.89 ( s , 3h ) ; c nmr : = 193.5 , 192.9 , 160.2 , 147.7 , 145.3 , 139.8 , \n 136.7 , 133.1 , 132.7 , 129.5 , 125.1 , 114.1 , 109.7 , 55.5 , 31.7 ; ir ( kbr , \n cm ) : 3105 , 2928 , 2881 , 1777 , 1751 , 1682 , 1663 , \n 1566 ; hr - esims ( m / z ) : calcd for \n [ m + h ] c17h15n2o3 295.1077 , found 295.1110 . \n to phthaldehyde 7 ( 74 mg , \n 0.27 mmol ) in etoh ( 1.5 ml ) was added 21 ( 50 mg , 0.81 \n mmol ) followed by the simultaneous addition of kcn ( 0.021 g , 0.33 \n mmol ) in h2o ( 0.5 ml ) and et3n ( 0.040 ml , 0.27 \n mmol ) . \n the reaction was stirred at rt for 15 min and then quenched \n with 10% hcl until a ph = 5 was attained . \n the aqueous \n layer was reduced , and the resulting solids were filtered and washed \n with acetone to give 22 as a red - orange solid ( 35 mg , \n 39% ) . \n h nmr ( dmso - d6 ) : = 9.73 ( brs , \n 2h ) , 8.34 ( s , 1h ) , 8.18 ( s , 1h ) , 7.16 ( d , j = 8.6 \n hz , 2h ) , 6.92 ( d , j = 8.6 hz , 2h ) , 3.93 ( s , 3h ) , \n 3.78 ( s , 3h ) ; c nmr ( dmso - d6 ) : = 181.8 , \n 181.3 , 158.9 , 148.8 , 146.8 , 141.9 , 139.4 , 137.1 , 135.8 , 131.1 , 129.9 , \n 127.5 , 121.5 , 113.4 , 109.6 , 55.6 , 31.9 ; ir ( cm ) : 2981 , 1642 , 1627 , 1608 , 1337 , 1243 , 1211 , 1021 ; hr - esims ( negative \n mode ) ( m / z ) : calcd for [ m h ] c19h13n2o5 349.0830 , found 349.0832 . to dihydroxyquinone 22 ( 23 \n mg , 0.066 mmol ) in thf ( 3 ml ) were added meoh ( 1 ml ) and tmschn2 ( 2.0 m in et2o ) ( 0.080 ml , 0.16 mmol ) with subsequent \n stirring at rt for 1 h. the reaction mixture was diluted with ch2cl2 and washed with sat . \n the resulting \n residue was purified by column chromatography ( 2:8 acetone / hexane \n to 1:1 acetone / hexane ) to give 23 as an orange residue \n ( 16 mg , 64% ) . \n h nmr : = 8.24 ( s , 1h ) , 8.00 ( s , \n 1h ) , 7.28 ( d , j = 8.6 hz , 2h ) , 7.02 ( d , j = 8.6 hz , 2h ) , 4.10 ( s , 3h ) , 4.01 ( s , 3h ) , 3.95 ( s , 3h ) , 3.86 ( s , \n 3h ) ; c nmr : = 182.3 , 182.0 , 159.1 , 148.9 , 147.4 , \n 146.5 , 136.8 , 130.6 , 130.2 , 128.8 , 128.3 , 122.8 , 114.5 , 113.8 , 108.8 , \n 61.4 , 61.3 , 55.3 , 31.7 ; ir ( cm ) : 2924 , 2852 , 1658 , \n 1618 , 1515 , 1456 , 1341 , 1307 , 1245 , 1215 , 1054 ; hr - apcims ( m / z ) : calcd for [ m + h ] c21h18n2o5 379.1288 , found \n 379.1296 . \n imidazolium salt 17 ( 1.0 g , \n 1.9 mmol ) in thf ( 40 ml ) was cooled to 0 c , and 5% naocl ( 30 \n ml ) was added dropwise . \n the reaction was diluted with etoac , and the aqueous \n layer was extracted with etoac ( 2 ) . \n the combined organic extracts \n were washed with h2o and then brine , dried ( anhyd . \n the resulting residue was \n purified by column chromatography ( 7:3 etoac / hexane ) to give 24 as an off - white solid ( 0.58 g , 75% ) . \n mp : 185187 \n c ; h nmr : = 7.127.09 ( m , 3h ) , 7.03 \n ( s , 1h ) , 6.94 ( d , j = 8.6 hz , 2h ) , 6.77 ( d , j = 8.9 hz , 2h ) , 6.586.56 ( m , 2h ) , 5.23 ( s , 2h ) , \n 4.79 ( s , 2h ) , 3.83 ( s , 3h ) , 3.57 ( s , 3h ) ; c nmr : \n = 169.8 , 159.7 , 155.6 , 141.8 , 136.4 , 136.2 , 131.3 , 128.2 , 127.8 , 127.2 , \n 126.0 , 124.0 , 123.6 , 117.1 , 113.1 , 99.5 , 67.7 , 55.3 , 45.8 , 27.8 ; ir \n ( cm ) : 1748 , 1705 , 1609 , 1517 , 1496 , 1352 , 1244 , \n 1028 ; hr - esims ( m / z ) : calcd for \n [ m + na ] c24h20n2o4na 423.1315 , found 423.1316 . \n imidazolone 24 ( 1.4 g , 3.5 mmol ) \n was dissolved \n in ch2cl2 ( 25 ml ) and cooled to 78 c \n under n2 . \n dibal - h ( 1 m in hexanes ) ( 9.8 ml , 9.8 mmol ) was \n added dropwise , then allowed to come to rt , and stirred for 4 h. the \n reaction mixture was cooled to 0 c , and water was added slowly \n followed by meoh . \n the resulting solids were washed with hexane and then et2o to give 25 as an off - white solid ( 1.17 g , 83% ) . \n mp : \n 205206 c ; h nmr : = 7.117.09 \n ( m , 3h ) , 7.03 ( s , 1h ) , 6.87 ( d , j = 8.6 hz , 2h ) , \n 6.70 ( d , j = 8.6 hz , 2h ) , 6.596.57 ( m , 2h ) , \n 4.81 ( s , 2h ) , 4.64 ( s , 2h ) , 4.38 ( s , 2h ) , 3.82 ( s , 3h ) , 3.46 ( s , 3h ) ; c nmr : = 159.4 , 155.5 , 137.1 , 134.0 , 132.5 , 131.5 , \n 130.1 , 128.1 , 127.2 , 126.9 , 125.8 , 125.6 , 113.4 , 108.5 , 65.1 , 59.5 , \n 55.4 , 45.4 , 27.4 ; ir ( cm ) : 3313 , 2930 , 1669 , 1611 , \n 1515 , 1398 , 1244 , 1181 ; hr - apcims ( m / z ) : calcd for [ m + h ] c24h25n2o4 405.1809 , found 405.1826 . \n diol 25 ( 0.22 g , 0.54 mmol ) was \n dissolved \n in ch2cl2 ( 10 ml ) and cooled to 78 c \n under n2 . \n dbu ( 0.32 ml , 2.2 mmol ) was added followed by 20 ( 0.38 g , 1.7 mmol ) in ch2cl2 ( 1 ml ) , \n and the reaction was stirred at 78 c for 30 min . sat . \n the organic layer was separated , and the aqueous layer was \n extracted with ch2cl2 ( 2 ) . \n the resulting residue was purified by column chromatography \n ( the silica gel was neutralized with et3n prior to purification ) \n ( 4:6 etoac / hexane ) to give 26 as an off - white solid ( 0.17 \n g , 77% ) . \n mp : 152155 c ; h nmr : = \n 10.40 ( s , 1h ) , 9.67 ( s , 1h ) , 7.68 ( s , 1h ) , 7.147.08 ( m , 3h ) , \n 6.92 ( d , j = 8.6 hz , 2h ) , 6.74 ( d , j = 8.6 hz , 2h ) , 6.54 ( d , j = 6.8 hz , 2h ) , 4.74 ( s , \n 2h ) , 3.84 ( s , 3h ) , 3.59 ( s , 3h ) ; c nmr : = 192.6 , \n 192.0 , 160.1 , 155.6 , 136.1 , 133.7 , 132.2 , 131.9 , 131.8 , 130.2 , 128.3 , \n 128.0 , 127.3 , 125.6 , 123.6 , 113.7 , 106.5 , 55.5 , 45.8 , 27.9 ; ir ( cm ) : 2934 , 1701 , 1675 , 1606 , 1513 , 1357 , 1243 , 1076 ; \n hr - esims ( m / z ) : calcd for [ m + na ] c24h20n2o4na \n 423.1315 , found 423.1309 . \n to phthaldehyde 26 ( 0.10 g , \n 0.25 mmol ) in thf ( 3 ml ) was added 21 ( 42 mg , 0.75 mmol ) \n followed by the addition of kcn ( 0.029 g , 0.44 mmol ) and k2co3 ( 0.17 g , 1.3 mmol ) in h2o ( 1 ml ) . \n the reaction \n was stirred at rt for 5 min , then quenched with 6 m hcl , and diluted \n with h2o ( 5 ml ) . \n the combined organic extracts were washed with h2o and then brine , dried ( anhyd . \n the resulting solids were filtered to give the dihydroxyquinone 27 as an orange solid ( 22 mg , 20% ) . \n mp : 204206 c ; h nmr ( dmso - d6 ) : = 9.65 ( s , 1h ) , 9.60 ( s,1h ) , \n 7.80 ( s , 1h ) , 7.137.11 ( m , 3h ) , 6.79 ( d , j = 8.6 hz , 2h ) , 6.63 ( d , j = 8.6 hz , 2h ) , 6.566.54 \n ( m , 2h ) , 4.44 ( s , 2h ) , 3.71 ( s , 3h ) , 3.50 ( s , 3h ) ; c \n nmr ( dmso - d6 ) : = 181.6 , 181.1 , 159.1 , 155.1 , 141.0 , \n 138.4 , 137.3 , 134.0 , 131.2 , 130.5 , 128.5 , 127.7 , 127.3 , 127.0 , 125.7 , \n 125.0 , 122.6 , 113.5 , 106.0 , 55.6 , 45.5 , 28.1 ; ir ( cm ) : 3240 , 1697 , 1654 , 1597 , 1514 , 1400 , 1283 , 1243 . \n hrms - esi ( negative \n mode ) ( m / z ) : calcd for c26h19n2o6 [ m h ] 455.1249 , found 455.1243 . \n the dihydroxyquinone 27 prepared \n above ( 22 mg , 0.048 mmol ) was dissolved in thf ( 3 ml ) , and meoh ( 0.5 \n ml ) was added followed by tmschn2 ( 2.0 m in et2o ) ( 0.07 ml , 0.15 mmol ) with subsequent stirring at rt for 30 min . \n the reaction was diluted with etoac and washed with 2 m na2co3 ( 2 ) and then brine . \n the \n resulting residue was purified by column chromatography ( 1:1 etoac / hexane ) \n to give 28 as an orange solid ( 15 mg , 65% ) . \n mp : 176178 \n c ; h nmr : = 7.82 ( s , 1h ) , 7.137.11 \n ( m , 3h ) , 6.79 ( d , j = 8.9 hz , 2h ) , 6.89 ( d , j = 8.9 hz , 2h ) , 6.586.56 ( m , 2h ) , 4.60 ( s , 2h ) , \n 4.03 ( s , 3h ) , 3.92 ( s , 3h ) , 3.81 ( s , 3h ) , 3.60 ( s , 3h ) ; c nmr : = 182.0 , 181.5 , 159.2 , 155.4 , 148.1 , 145.2 , 136.6 , \n 134.0 , 130.1 , 128.2 , 127.1 , 127.0 , 125.5 , 124.0 , 113.5 , 105.4 , 61.4 , \n 61.3 , 55.3 , 45.8 , 27.9 ; ir ( cm ) : 2950 , 1716 , 1651 , \n 1619 , 1601 , 1455 , 1241 , 1211 ; hr - esims ( m / z ) : calcd for [ m + na ] c28h24n2o6na 507.1527 , found 507.1533 . to 28 ( 30 mg , 0.062 mmol ) \n was added tfoh ( 2.5 ml ) , and the mixture was heated at 55 c \n in a vial for 4 h. the reaction was allowed to cool to rt and poured \n into etoac . \n the resulting \n residue was purified by column chromatography ( 8:2 etoac / hexane ) to \n give 29 as an orange solid ( 10 mg , 56% ) . \n mp : > 290 \n c ; h nmr ( dmso - d6 ) : = 10.95 ( s , \n 1h ) , 9.41 \n ( s , 1h ) , 7.63 ( s , 1h ) , 6.96 ( d , j = 8.6 hz , 2h ) , \n 6.75 ( d , j = 8.6 hz , 2h ) , 3.90 ( s , 3h ) , 3.82 ( s , \n 3h ) , 3.35 ( s , 3h ) ; c nmr ( dmso - d6 ) : \n = 181.9 , 181.7 , 157.2 , 155.3 , 148.4 , 145.6 , 134.4 , 133.1 , 130.3 , 127.0 , \n 126.4 , 124.5 , 123.2 , 115.8 , 104.9 , 61.2 ( 2c ) , 27.3 ; ir ( cm ) = 3200 , 2921 , 2851 , 1702 , 1656 , 1609 , 1515 , 1105 , 1059 , 1033 ; hr - esims \n ( m / z ) : calcd for [ m + na ] c20h16n2o6na 403.0901 , \n found 403.0911 . \n benzimidazole 15 ( 5.0 g , 15 \n mmol ) was dissolved in ch2cl2 ( 100 ml ) and cooled \n to 78 c under n2 . \n dibal - h ( 1 m in hexanes ) \n ( 41 ml , 41 mmol ) was added dropwise , and the mixture was allowed to \n come to rt and stirred overnight . \n the reaction mixture was then cooled \n to 0 c , and water was added slowly followed by meoh . \n the resulting solids \n were washed with et2o to give 30 as a white \n solid ( 3.2 g , 63% ) . \n mp : 191193 c ; h nmr : \n = 7.81 ( s , 1h ) , 7.79 ( s , 1h ) , 7.35 ( t , j = \n 8.0 hz , 1h ) , 7.23 ( d , j = 7.4 hz , 2h ) , 7.187.12 \n ( m , 3h ) , 7.08 ( d , j = 6.9 hz , 2h ) , 6.50 ( d , j = 6.9 hz , 2h ) , 4.91 ( s , 2h ) , 4.74 ( s , 2h ) , 4.49 ( s , 2h ) ; c nmr : = 146.0 , 143.4 , 136.5 , 135.9 , 135.1 , 134.1 , \n 131.5 , 130.4 , 128.6 , 128.1 , 128.0 , 127.7 , 127.7 , 126.0 , 121.2 , 65.4 , \n 59.6 , 49.6 ; ir ( cm ) : 3056 , 2856 , 1504 , 1441 , 1311 , \n 1175 , 1018 , 1000 ; hr - esims ( m / z ) : \n calcd for [ m + h ] c22h21n2o2 345.1598 , found 345.1604 . \n diol 30 ( 3.0 g , 8.7 mmol ) \n was dissolved in ch2cl2 ( 75 ml ) and cooled to \n 78 c under n2 . \n dbu ( 4.8 ml , 35 mmol ) was \n added followed by 20 ( 4.7 g , 22 mmol ) in ch2cl2 ( 3 ml ) , and the reaction was stirred at 78 \n c for 30 min . sat . \n the organic layer was separated , and the \n aqueous layer was extracted with ch2cl2 ( 2 ) . \n the resulting residue was purified by \n column chromatography ( the silica gel was neutralized with et3n prior to purification ) ( 6:4 etoac / hexane ) to give 31 as a white solid ( 2.0 g , 68% ) . \n mp : 167169 c ; h nmr : = 10.49 ( s , 1h ) , 9.88 ( s , 1h ) , 8.43 ( s,1h ) , \n 8.06 ( s , 1h ) , 7.44 ( t , j = 7.5 hz , 1h ) , 7.31 ( t , j = 8.0 hz , 2h ) , 7.23 ( d , j = 7.5 hz , 1h ) , \n 7.217.14 ( m , 4h ) , 6.51 ( d , j = 7.5 hz , 2h ) , \n 4.86 ( s , 2h ) ; c nmr : = 192.8 , 192.4 , 149.7 , 146.8 , \n 135.4 , 134.0 , 132.9 , 132.4 , 132.0 , 131.4 , 130.5 , 129.2 , 128.9 , 128.4 , \n 128.2 , 125.9 , 121.8 , 50.2 ; ir ( cm ) : 3070 , 2871 , \n 1756 , 1678 , 1595 , 1488 , 1455 , 1303 , 1213 ; hr - esims ( m / z ) : calcd for [ m + na ] c22h16n2o2na 363.1104 , found 363.1108 . \n to phthaldehyde 31 ( 150 mg , 0.44 mmol ) in thf was added 21 ( 0.037 g , 0.66 mmol ) followed by the simultaneous addition \n of kcn ( 29 mg , 0.44 mmol ) in h2o ( 1 ml ) and et3n ( 0.06 ml , 0.44 mmol ) . \n the reaction was stirred at rt for 15 min \n and then quenched with h2o ( 5 ml ) . \n 1 m hcl was added until \n ph = 5 was reached , and the aqueous layer was extracted with etoac \n ( 2 ) . \n an orange - red residue was \n recovered and recrystallized from meoh to give 32 as \n an orange - red solid ( 74 mg , 43% ) . \n mp : 280 c ( decomp . ) ; h nmr ( dmso - d6 ) : = 9.71 ( brs , 2h ) , 8.46 \n ( s , 1h ) , 8.30 ( s , 1h ) , 7.327.29 ( m , 1h ) , 7.197.16 \n ( m , 5h ) , 7.017.00 ( m , 2h ) , 6.45 ( d , j = 4.6 \n hz , 2h ) , 4.75 ( s , 2h ) ; c nmr ( dmso - d6 ) : \n = 181.6 , 181.1 , 151.2 , 146.6 , 141.8 , 139.6 , 137.6 , 136.5 , 135.5 , 129.3 , \n 129.1 , 128.8 , 128.2 , 127.8 , 127.8 , 127.1 , 125.9 , 122.9 , 118.7 , 49.3 ; \n ir ( cm ) : 2831 , 1662 , 1615 , 1595 , 1569 , 1343 , 1315 , \n 1206 , 1189 , 905 ; hr - apcims ( m / z ) : \n calcd for [ m + h ] c24h17n2o4 397.1183 , found 397.1196 . \n dihydroxyquinone 32 ( 84 mg , 0.21 \n mmol ) was suspended in thf ( 5 ml ) , and meoh \n ( 2.0 ml ) was added followed by tmschn2 ( 2.0 m in et2o ) ( 0.37 ml , 0.74 mmol ) with subsequent stirring at rt for \n 4 h. the reaction was diluted with etoac and washed with sat . \n the resulting residue was purified by column chromatography \n ( 1:1 etoac / hexane ) to give 33 as a yellow solid ( 45 mg , \n 54% ) . \n mp : 189191 c ; h nmr : = 8.65 \n ( s , 1h ) , 7.92 ( s , 1h ) , 7.37 ( t , j = 7.5 hz , 1h ) , \n 7.27 ( d , j = 8.0 hz , 2h ) , 7.237.18 ( m , 3h ) , \n 7.04 ( d , j = 7.5 hz , 2h ) , 6.57 ( d , j = 8.0 hz , 2h ) , 4.65 ( s , 2h ) , 4.07 ( s , 3h ) , 3.94 ( s , 3h ) ; c nmr : = 182.1 , 181.9 , 149.5 , 148.7 , 147.0 , 146.4 , 136.1 , \n 135.9 , 135.8 , 129.2 , 128.8 , 128.5 , 128.3 , 128.1 , 127.9 , 127.8 , 126.1 , \n 124.0 , 120.2 , 61.4 , 61.3 , 50.0 ; ir ( cm ) : 3009 , \n 2940 , 1649 , 1596 , 1441 , 1348 , 1291 , 1027 ; hr - esims ( m / z ) : calcd for [ m + h ] c26h21n2o4 425.1496 , found 425.1490 . \n to quinone 33 ( 45 mg , \n 0.11 mmol ) in ch3cn ( 8 ml ) was added mei ( 0.20 ml ) followed \n by heating at 70 c for 15 h. the reaction mixture was concentrated , \n and the resulting residue was triturated with et2o to give 34 as a red - orange solid ( 52 mg , 86% ) . \n mp : 184188 \n c ; h nmr : = 10.33 ( s , 1h ) , 8.49 ( s , 1h ) , \n 7.48 ( t , j = 7.5 hz , 1h ) , 7.39 ( t , j = 7.5 hz , 2h ) , 7.297.27 ( m , 3h ) , 7.18 ( d , j = 7.5 hz , 2h ) , 6.96 ( d , j = 7.5 hz , 2h ) , 4.93 ( s , \n 2h ) , 4.26 ( s,3h ) , 4.10 ( s , 3h ) , 3.98 ( s , 3h ) ; c nmr : \n = 180.2 , 180.0 , 149.3 , 147.7 , 146.4 , 134.9 , 132.9 , 132.9 , \n 132.3 , 132.2 , 131.1 , 129.4 , 129.3 , 128.9 , 128.6 , 128.0 , 127.0 , 111.8 , \n 61.6 ( 2c ) , 53.0 , 34.9 ; ir ( cm ) : 3031 , 2945 , 1718 , \n 1660 , 1609 , 1452 , 1314 , 1209 , 1052 , 916 ; hr - esims ( m / z ) : calcd for [ m ] c27h23n2o4 439.1652 , found 439.1634 . \n to imidazolium salt 34 ( 31 \n mg , 0.055 mmol ) in thf ( 1 ml ) was added 1 m k2co3 ( 0.27 ml ) and ncs ( 8 mg , 0.06 mmol ) , followed by stirring at rt \n for 30 min . the reaction was diluted with water ( 4 ml ) and extracted \n with etoac ( 2 5 ml ) . \n the resulting residue was purified \n by column chromatography ( 1:1 etoac / hexane ) to give 35 as a yellow solid ( 15 mg , 60% ) . \n mp : 160162 c ; h nmr : = 7.84 ( s , 1h ) , 7.33 ( t , j = 7.5 hz , 1h ) , 7.17 ( t , j = 7.5 hz , 2h ) , 7.137.09 \n ( m , 3h ) , 6.89 ( d , j = 6.9 hz , 2h ) , 6.53 ( d , j = 6.3 hz , 2h ) , 4.52 ( s , 2h ) , 4.03 ( s , 3h ) , 3.91 ( s , 3h ) , \n 3.60 ( s , 3h ) ; c nmr : = 181.9 , 181.4 , 155.4 , 148.1 , \n 145.3 , 136.5 , 135.2 , 134.0 , 131.4 , 129.0 , 128.2 , 128.1 , 127.9 , 127.5 , \n 127.1 , 125.6 , 125.5 , 123.7 , 105.5 , 61.4 , 61.3 , 45.7 , 27.9 ; ir ( cm ) : 2927 , 1709 , 1651 , 1620 , 1601 , 1452 , 1344 , 1209 , \n 1059 ; hr - esims ( m / z ) : calcd for \n [ m + na ] c27h22n2o5na 477.1421 , found 477.1438 . \n to imidazolone 35 ( 25 mg , \n 0.055 mmol ) was added tfoh ( 2 ml ) followed by heating at 55 c \n for 4 h. the reaction was quenched with water and neutralized with \n sat . \n the aqueous layer was extracted with etoac \n ( 2 ) , and the combined organic extracts were washed with brine , \n dried , and concentrated . \n the resulting residue was purified by column \n chromatography ( 6:4 etoac / hexane ) to give 36 as a yellow \n waxy solid ( 10 mg , 67% ) . \n h nmr : = 8.11 ( brs , 1h ) , \n 7.74 ( s , 1h ) , 7.497.43 ( m , 3h ) , 7.227.21 ( m , 2h ) , \n 4.07 ( s , 3h ) , 3.97 ( s , 3h ) , 3.47 ( s , 3h ) ; c nmr : \n = 181.9 , 181.5 , 154.2 , 147.6 , 146.0 , 135.3 , 134.1 , 131.6 , 129.1 , 128.3 , \n 128.1 , 127.8 , 124.3 , 123.0 , 105.5 , 61.5 , 61.4 , 27.4 ; ir ( cm ) : 2919 , 1708 , 1648 , 1623 , 1443 , 1332 , 1245 , 1194 , 1056 , 1026 ; hr - apcims \n ( m / z ) : calcd for [ m + h ] c20h17n2o5 365.1132 , \n found 365.1144 . to tert - butyl carbamate \n ( 19 mg , 0.16 mmol ) in ch2cl2 ( 2 ml ) was added t - buocl ( 0.02 ml , 0.17 mmol ) , followed by stirring at rt \n for 20 min . the reaction mixture was cooled to 0 c , and dbu \n ( 0.033 ml , 0.22 mmol ) was added followed by 34 in ch2cl2 ( 3 ml ) . \n the reaction was stirred at rt for \n 30 min and then diluted with water . \n the aqueous layer was extracted \n with ch2cl2 ( 2 ) , and the combined organic \n extracts were washed with brine , dried ( anhyd . \n the crude residue was purified by column \n chromatography ( 1:1 etoac / hexane ) to give 37 as a yellow - orange \n residue ( 10 mg , 30% ) . \n h nmr : = 7.98 ( s , 1h ) , 7.35 \n ( t , j = 7.5 hz , 1h ) , 7.18 ( t , j = \n 7.5 hz , 2h ) , 7.127.07 ( m , 3h ) , 6.84 ( d , j = 6.9 hz , 2h ) , 6.44 ( d , j = 6.9 hz , 2h ) , 4.70 ( s , \n 2h ) , 4.03 ( s , 3h ) , 3.90 ( s,3h ) , 3.68 ( s,3h ) , 1.46 ( s,9h ) ; c nmr : = 181.6 , 181.1 , 158.3 , 153.9 , 148.3 , 145.5 , 135.8 , \n 135.4 , 134.6 , 132.3 , 128.9 , 128.2 , 128.2 , 128.1 , 128.0 , 127.1 , 127.1 , \n 125.4 , 124.5 , 106.6 , 78.9 , 61.4 , 61.3 , 47.3 , 31.8 , 28.3 ; ir ( cm ) : 2929 , 1655 , 1581 , 1443 , 1308 , 1207 , 1147 , 1047 , \n 1026 ; hr - esims ( m / z ) : calcd for \n [ m + h ] c32h32n3o6 is 554.2286 , found 554.2290 . \n to benzimidazole 18 ( 25 mg , 0.085 mmol ) in toluene ( 10 ml ) was added mei ( 0.030 ml , \n 0.42 mmol ) , followed by heating to 140 c for 20 h. the solvent \n was reduced to give 38 ( 25 mg , 68% ) as an off - white solid . \n mp : 253256 c ; h nmr ( cd3od ) : \n = 8.13 ( s , 1h ) , 7.37 ( d , j = 8.6 hz , 2h ) , \n 7.08 ( d , j = 8.6 hz , 2h ) , 5.51 ( s , 2h ) , 4.18 ( s , \n 3h ) , 3.88 ( s , 3h ) , 3.54 ( s , 3h ) ; c nmr ( cd3od ) : = 168.9 , 160.9 , 145.2 , 136.7 , 131.1 , 130.8 , 130.5 , 121.8 , \n 113.3 , 106.3 , 68.4 , 54.6 , 36.2 , 32.9 ; ir ( cm ) : \n 2960 , 1772 , 1612 , 1577 , 1378 , 1241 , 1065 , 1018 , 823 ; hr - esims ( m / z ) : calcd for [ m ] c18h17n2o3 309.1234 , found 309.1229 . \n imidazolium salt 38 ( 10 mg , \n 0.023 mmol ) in thf was cooled to 0 c , and 5% naocl ( 0.3 ml ) \n was added followed by stirring for 10 min . \n the resulting solids were purified by ptlc ( etoac ) to give 39 as a white solid ( 6 mg , 73% ) . \n mp : 239240 c ; h nmr : = 7.26 ( d , j = 8.6 hz , 2h ) , 6.98 \n ( d , j = 8.6 hz , 2h ) , 6.98 ( s , 1h ) , 5.25 ( s , 2h ) , \n 3.87 ( s , 3h ) , 3.49 ( s , 3h ) , 2.92 ( s , 3h ) ; c nmr : \n = 170.0 , 159.9 , 155.4 , 141.5 , 135.8 , 131.3 , 128.9 , 124.1 , 123.4 , 116.8 , \n 113.3 , 99.3 , 67.9 , 55.3 , 30.3 , 27.7 ; ir ( cm ) : \n 2930 , 2838 , 1761 , 1700 , 1609 , 1520 , 1482 , 1357 , 1287 , 1248 , 1196 , \n 1179 , 1139 , 1078 ; hr - apcims ( m / z ) : calcd for [ m + h ] c18h17n2o4 325.1183 , found 325.1191 . \n to 18 ( 72 mg , 0.24 \n mmol ) in acetonitrile ( 10 ml ) was added semcl ( 50 mg , 0.30 mmol ) followed \n by stirring at 60 c for 1 h. the reaction mixture was cooled \n to 0 c , and the precipitated solids were filtered and washed \n with et2o to give 41 as a white solid ( 51 \n mg , 63% ) . \n mp : 255258 c ; h nmr ( cd3od ) : = 9.56 ( d , j = 4.6 hz , 1h ) , 8.06 ( s , \n 1h ) , 7.51 ( d , j = 8.6 hz , 2h ) , 7.12 ( d , j = 8.6 hz , 2h ) , 5.52 ( s , 2h ) , 4.19 ( s , 3h ) , 3.89 ( s , 3h ) ; c nmr ( cd3od ) : = 169.3 , 161.1 , 145.7 , 145.1 , 136.4 , \n 131.3 , 130.5 , 122.0 , 119.9 , 113.8 , 105.2 , 68.5 , 54.6 , 32.7 ; ir ( cm ) : 3155 , 2942 , 1728 , 1623 , 1506 , 1392 , 1243 , 1142 , \n 1110 , 1012 ; hr - esims ( m / z ) : calcd \n for [ m + na ] c17h14n2o3na 317.0897 , found 317.0903 .", "role": "user" }, { "content": "synthetic \n approaches to the imidazonaphthoquinone core \n of kealiiquinone and related leucetta - derived alkaloids \n are described . \n the polysubstituted benzimidazole framework can be \n constructed through intramolecular diels \n alder reactions of \n propiolate - derived enynes followed by oxidation . \n adjustment of the \n oxidation state of the thus formed lactone allows introduction of \n the 2,3-dihydroxybenzoquinone moiety through a presumed \n benzoin - like condensation between a phthaldehyde derivative and a \n masked glyoxal equivalent catalyzed by a cyanide ion . \n oxidation of \n the c2-position can be accomplished through application of an operationally \n simple treatment of an imidazolium salt with bleach , thus producing \n the corresponding 2-imidazolone . \n debenzylation of a late stage intermediate \n en route to kealiiquinone was compromised by concomitant o - demethylation upon treatment with triflic acid resulting in the \n formation of non - natural 7-desmethylkealiiquinone . \n other endgame strategies were evaluated ; however , these efforts did \n not lead to completion of a synthesis of kealiiquinone but did provide \n access to other closely related analogues .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: it is a condition in which urine culture reveals a significant growth of pathogens that is greater than 10 bacteria / ml , but without the patient showing symptoms of urinary tract infection ( uti ) . \n the apparent reduction in immunity of pregnant women appears to encourage the growth of both commensal and non - commensal microorganisms . \n the physiological increase in plasma volume during pregnancy decrease urine concentration and up to 70% pregnant women develop glucosurea , which encourages bacterial growth in the urine . \n pregnancy enhances the progression from asymptomatic to symptomatic bacteriuria which could lead to pyelonephritis and adverse obstetric outcomes such as prematurity , low birth weight and higher foetal mortality rates . \n the adverse effects of undiagnosed asymptomatic bacteriuria on mother and child have made researchers to suggest routine culture screening for all pregnant women attending antenatal clinic in order to prevent mother and child from any form of complication that may arise due to infection . \n however , in many hospitals in developing countries including nigeria , routine urine culture test is not carried out for antenatal patients probably due to cost implication and time factor for culture result ( usually 48 hours period ) instead many clinicians opt for the strip urinalysis method for assessing urine in pregnant women . \n the true picture of such urine specimen can not be fully assessed as the strip can not quantify the extent of infection in such a patient as well as provide antimicrobial therapy which is usually seen in the case of culture test . in many health centers in developing countries , the attention of clinicians and health care providers is usually on the presence of glucose and protein in urine specimens with less attention on possible asymptomatic infection . against this background , \n this work is aimed at determining asymptomatic bacteriuria among pregnant women attending antenatal clinic in a primary health centre in benin city , nigeria . \n study area : this study was carried out in benin city , edo state , nigeria . \n the state has approximately 4 million people and is bounded in the south by delta state , in the west by ondo state , in the north by kogi state and in the east by kogi and anambra states . \n iwogban primary health centre is a primary health institution that attends to the primary health needs of the people within and around the locality . \n it attends to cases such as antenatal clinic , malaria , diarrhea , postnatal immunization of infants and other minor ailments within the scope of available equipment and manpower . \n it has an average number of about 150 patients per day and it is situated on the outskirts of benin city . \n it is a rural setting with a fast growing population of new residents from the main city along side with the original inhabitants of the community . \n study population : this study was carried out at iwogban primary health centre , benin city , nigeria . \n a total of 1,228 pregnant women attending antenatal clinic at the centre were assessed for asymptomatic bacteriuria between january 2007 and december 2008 . \n exclusion criteria included signs and symptoms of uti , and antibiotic usage within one week . \n specimen collection and processing : clean - catch midstream urine was collected from each patient into a sterile universal container . \n samples were cultured on dried plates of blood agar and cysteine lactose electrolyte deficient agar ( cled ) , using a calibrated drop delivering 0.002ml of urine . \n colony counts yielding bacterial growth of 10/ml or more of pure isolates were regarded as significant for infection . \n similarly , 10ml of each patient urine was transferred into sterile centrifuge tubes and then centrifuged at 3000rpm for 10 - 15 minutes . \n the supernatant was discarded and the deposit examined microscopically at high magnification for pus cells , red blood cells , epithelial cells , casts , crystals , yeast - like cells and trichomonas vaginalis . \n the national committee for clinical laboratory standard ( nccls ) operating procedure ( 11 ) was followed . statistical analysis : statistical analysis was performed by the chi - square ( ) test . \n out of 1,228 pregnant women examined for asymptomatic bacteriuria , 556 were positive for significant bacteriuria , giving a prevalence of 45.3% . \n out of the 556 positive culture , 448 ( 80.6% ) yielded single bacterial isolates while 108 ( 19.4% ) yielded mixed bacterial isolates , giving a total of 664 . \n there was a significant difference in the prevalence of asymptomatic bacteriuria with respect to age ( p<0.001 ) ( table 1 ) . \n however , there was no significant difference with respect to trimester ( p=0.2006 ) ( table 2 ) . \n the prevalence of asymptomatic bacteriuria in pregnant women in this study was 45.3% . this is higher than the 23.9% from the study in sagamu , nigeria , \n it is lower than the 86.6% earlier reported in benin city , nigeria and 78.7% reported in abakaliki , nigeria . \n there was a significant difference in the prevalence of asymptomatic bacteriuria with respect to age group ( p < 0.0001 ) . \n age group 26 - 30 years had the highest percentage of infection ( 53.1% ) . \n this was closely followed by age group 36 - 40 ( 52.5% ) and 31 - 35 ( 52.1% ) respectively ( table 1 ) . \n advanced maternal age ( 35 years ) was reported as risk factor for asymptomatic bacteriuria in pregnancy . \n another reason could also be due to the fact that many women within this age bracket are likely to have had many children before the present pregnancy and it has been reported that multiparty is a risk factor for acquiring asymptomatic bacteriuria in pregnancy . \n there was no significant difference in the prevalence of asymptomatic bacteriuria with respect to trimester ( p= 0.2006 ) . \n the most prevalence organism observed in this study was escherichia coli ( 27.1% ) , followed by staphylococcus aureus ( 24.4% ) . \n this finding agrees with earlier reports[132123 ] who observed the same predominant trend in escherichia coli infection pattern . \n this could be due to the fact that urinary stasis is common in pregnancy and since most escherichia coli strains prefer that environment , they cause uti \n . another reason could be as a result of poor genital hygienic practices by pregnant women who may find it difficult to clean their anus properly after defecating or clean their genital after passing urine . \n this agrees with previous studies which observed an increasing trend in the prevalence of staphylococcus aureus infection among asymptomatic pregnant women . \n the other organisms isolated included klebsiella species , proteus species , citrobacter species , providencia species and pseudomonas aeruginosa . \n this study revealed that ciprofloxacin , ceftriaxone and augmentin were very effective against most of the urinary isolates . \n the upsurge in antibiotic resistant pattern seen in this study could be due to antibiotic abuse and self medication being practiced in many developing countries including nigeria . also low \n cost and availability of these drugs could be another contributing factor for antibiotic resistance in this locality . \n this is far more on a high side compared to the low prevalence in studies in ghana , and ethiopia . \n this is worrisome because uti in pregnancy may have serious consequences for both the mother and the child . \n routine urine culture test should be carried out on all antenatal women to detect asymptomatic bacteriuria .", "role": "user" }, { "content": "background : asymptomatic bacteriuria is the significant presence of bacteria in the urine of an individual without symptoms . in pregnancy , the apparent reduction in immunity of pregnant women tends to encourage the growth of pathogens.aim:this study was carried out to determine the prevalence of asymptomatic bacteriuria in pregnant women attending a primary health centre in benin city , nigeria.materials and methods : a total of 1,228 pregnant women were recruited for this study . \n all subjects were clinically identified to have no signs and symptoms of uti . \n clean catch midstream urine sample was collected from each patient into sterile universal container . \n the urine samples were examined microscopically and by cultural method . \n identification of isolates was by standard microbiological technique.result:a total of 556 ( 45.3% ) were positive for significant bacteriuria . \n there was a significant difference in the prevalence of asymptomatic bacteriuria with respect to age ( p < 0.0001 ) . \n trimester did not show any significant difference ( p = 0.2006 ) in the prevalence of asymptomatic bacteriuria . \n escherichia coli was the most predominant organism followed closely by staphylococcus aureus . \n ciprofloxacin , ceftriaxone and augmentin were found to be the most effective antibiotics against the urinary isolates.conclusion:asymptomatic bacteriuria is not uncommon among antenatal patients in the population studied . \n routine urine cultural test should be carried out on all antenatal patients in order to identify any unsuspecting infection . \n this measure will go a long way in reducing maternal and obstetric complications associated with pregnancy .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: recombinant pl 4-dehydrogenase ( pldh ) , pm - pyruvate aminotransferase ( ppat ) , and pn 4-oxidase ( pnox ) were prepared as described previously ( 1012 ) . \n recombinant e. coli alkaline phosphatase ( alp ) was prepared according to a standard protocol ( 13 ) . \n almond -glucosidase ( gls ) , pm , and pl ( 99.5% ) were purchased from sigma chemical co. ( st . louis , mo ) , pn and plp from nacalai tesque , inc . \n the pmp was a gift from daiichi fine chemicals co. ( takaoka , japan ) and png was prepared enzymatically from d - glucose and pn based on a transglucosidase activity of a bacterial -glucosidase ( 15 ) . \n food samples were bought at local markets and chosen at random in order to test and validate the method . \n the basic principle is to convert all vitamin b6 compounds specifically into a highly fluorescent vitamin b6-derivative 4-pla ( see fig . \n 1 ) . thus , seven reaction mixtures were used to determine seven vitamin b6 compounds . \n the filtration was done with a dismic 13 syringe filter ( pore size of 0.2 m , advantech , tokyo , japan ) . to make a control ( i.e. no - reaction test sample ) , hcl was added to the reaction mixture before addition of the food sample . for the determination of plp , the food sample was at first hydrolyzed with alp in the reaction 4 ( fig . \n 2 ) , and then the pl produced was determined as shown under the reaction for plp ( fig . \n 3 ) . both pmp and pnp were determined in the same way just like plp but using the additional enzymes and the substrate or the cofactor as shown under the reactions for pmp and pnp , respectively ( fig . \n first hydrolyzed with -glucosidase in the reaction mixture containing nah2po4 , which inhibited phosphatase activity in the almond -glucosidase preparation , and then pn was determined as shown under the reaction for png ( fig . \n 3 ) . the reaction mixtures used for determination of seven vitamin b6 compounds . illustrations of analytical procedures of seven reactions . \n see table 1 for quantification of the seven compounds from the hplc elution profiles of the seven reactions described above . \n the 4-pla was measured by a reversed - phase isocratic hplc method using a cosmosil 5c18ms - ii column ( 2504.6 mm ; nacalai tesque , kyoto , japan ) , and a mobile phase consisting of 20 mm potassium phosphate buffer ( ph 7.0 ) and 10% ( v / v ) methanol . \n the separation was made with a jasco hplc system ( jasco , tokyo , japan ) equipped with an as-2055 autosampler , a pu-2080 pump , and an fp-920 fluorescence detector . \n the fluorescence intensity of the eluted 4-pyridoxolactone was monitored at 430 nm after excitation at 360 nm . \n the pldh ( 10 ) , pnox ( 12 ) , ppat ( 11 ) , alp ( 16 ) , and -glucosidase ( 17 ) were assayed as described previously . \n one unit of enzyme was defined as the amount that catalyzed the formation of 1mol of product per min . \n the pldh , pnox , alp , and -glucosidase catalyze irreversible reactions ; ppat catalyzes a reversible reaction . however , when coupled with pldh the reversible reaction is forced to produce 100% conversion . \n for example , the enzymatic reactions shown in fig . 1 are 100% conversion of the substrates . \n the activities of the enzymes were assayed every time just before usage for enzymatic conversion of vitamin b6 compounds into 4-pla . \n chicken white meat and liver , egg yolk and white , dried anchovy , carrot , and garlic were analyzed . \n the edible wet part of the foods was used , with the exception of the dried anchovy , which was ground with a mini blender ( osaka chemical co. , osaka , japan ) . \n foods ( 0.11.0 g , dry or wet ) were homogenized in 5ml of 0.1 m hcl with a polytron homogenizer , and then the homogenate was incubated at 100c for 30min to liberate protein - bound plp . the suspension \n was cooled on ice , then 0.1 ml of 50% ( w / v ) trichloroacetic acid ( tca ) was added , and followed by incubation at 100c for 5min . after cooling on ice , ph of the mixture \n was adjusted to 7.5 by the addition of 0.5ml of 0.5 m tris - hcl ( ph 7.5 ) and 0.57ml of 1.0 m naoh . \n the mixture was centrifuged at 8,000g for 5min at 4c , and then the supernatant ( 550l ) was used for the enzymatic reactions . \n samples that were added the seven vitamin b6 compounds were similarly treated to assess recovery . \n spiking level was 2 pmol ( the amount in 100l applied to the hplc column ) . \n recombinant pl 4-dehydrogenase ( pldh ) , pm - pyruvate aminotransferase ( ppat ) , and pn 4-oxidase ( pnox ) were prepared as described previously ( 1012 ) . \n recombinant e. coli alkaline phosphatase ( alp ) was prepared according to a standard protocol ( 13 ) . \n almond -glucosidase ( gls ) , pm , and pl ( 99.5% ) were purchased from sigma chemical co. ( st . louis , mo ) , pn and plp from nacalai tesque , inc . \n the pmp was a gift from daiichi fine chemicals co. ( takaoka , japan ) and png was prepared enzymatically from d - glucose and pn based on a transglucosidase activity of a bacterial -glucosidase ( 15 ) . \n food samples were bought at local markets and chosen at random in order to test and validate the method . \n the basic principle is to convert all vitamin b6 compounds specifically into a highly fluorescent vitamin b6-derivative 4-pla ( see fig . \n the filtration was done with a dismic 13 syringe filter ( pore size of 0.2 m , advantech , tokyo , japan ) . to make a control ( i.e. no - reaction test sample ) , \n hcl was added to the reaction mixture before addition of the food sample . for the determination of plp , \n the food sample was at first hydrolyzed with alp in the reaction 4 ( fig . \n 2 ) , and then the pl produced was determined as shown under the reaction for plp ( fig . \n 3 ) . both pmp and pnp were determined in the same way just like plp but using the additional enzymes and the substrate or the cofactor as shown under the reactions for pmp and pnp , respectively ( fig . \n first hydrolyzed with -glucosidase in the reaction mixture containing nah2po4 , which inhibited phosphatase activity in the almond -glucosidase preparation , and then pn was determined as shown under the reaction for png ( fig . \n horizontal arrows mean the addition of the solutions to the reaction mixtures . see table 1 for quantification of the seven compounds from the hplc elution profiles of the seven reactions described above . \n the 4-pla was measured by a reversed - phase isocratic hplc method using a cosmosil 5c18ms - ii column ( 2504.6 mm ; nacalai tesque , kyoto , japan ) , and a mobile phase consisting of 20 mm potassium phosphate buffer ( ph 7.0 ) and 10% ( v / v ) methanol . \n the separation was made with a jasco hplc system ( jasco , tokyo , japan ) equipped with an as-2055 autosampler , a pu-2080 pump , and an fp-920 fluorescence detector . \n the fluorescence intensity of the eluted 4-pyridoxolactone was monitored at 430 nm after excitation at 360 nm . \n the pldh ( 10 ) , pnox ( 12 ) , ppat ( 11 ) , alp ( 16 ) , and -glucosidase ( 17 ) were assayed as described previously . \n one unit of enzyme was defined as the amount that catalyzed the formation of 1mol of product per min . \n the pldh , pnox , alp , and -glucosidase catalyze irreversible reactions ; ppat catalyzes a reversible reaction . \n however , when coupled with pldh the reversible reaction is forced to produce 100% conversion . \n for example , the enzymatic reactions shown in fig . 1 are 100% conversion of the substrates . \n the activities of the enzymes were assayed every time just before usage for enzymatic conversion of vitamin b6 compounds into 4-pla . \n chicken white meat and liver , egg yolk and white , dried anchovy , carrot , and garlic were analyzed . \n the edible wet part of the foods was used , with the exception of the dried anchovy , which was ground with a mini blender ( osaka chemical co. , osaka , japan ) . \n foods ( 0.11.0 g , dry or wet ) were homogenized in 5ml of 0.1 m hcl with a polytron homogenizer , and then the homogenate was incubated at 100c for 30min to liberate protein - bound plp . the suspension was cooled on ice , then 0.1 ml of 50% ( w / v ) trichloroacetic acid ( tca ) was added , and followed by incubation at 100c for 5min . \n after cooling on ice , ph of the mixture was adjusted to 7.5 by the addition of 0.5ml of 0.5 m tris - hcl ( ph 7.5 ) and 0.57ml of 1.0 m naoh . \n the mixture was centrifuged at 8,000g for 5min at 4c , and then the supernatant ( 550l ) was used for the enzymatic reactions . \n samples that were added the seven vitamin b6 compounds were similarly treated to assess recovery . \n spiking level was 2 pmol ( the amount in 100l applied to the hplc column ) . \n the chromatograms for the analysis of pl , pm , and pn in the chicken liver sample are shown in fig . \n 4 : they showed similar but definitely different patterns of the fluorescent peaks of 4-pla . \n the 4-pla was eluted at about 11 min as shown with an arrow , together with unknown fluorescent compounds in the sample . \n the control reaction showed a tiny peak of 4-pla ( pl , no - r in fig . \n 4a ) , its area increased when the liver sample was subjected to the enzymatic conversion with the reaction 1 in fig . \n 2 for converting pl into 4-pla ( pl , r ) in the sample . thus , the subtraction of fluorescence intensity of 4-pla ( pl , no - r ) from that of 4-pla ( pl , r ) gave the amount of 4-pla corresponding to the amount of pl in the sample . the chicken liver sample spiked with \n pl showed a higher peak ( pl+sd , r ) because the corresponding amount of 4-pla was additionally produced by the enzymatic conversion . \n in contrast , the chicken liver sample spiked with pl without the enzymatic conversion did not show this increase ; its peak height was the same as that of the control reaction mixture ( pl+sd , no - r ) . \n thus , pl in the chicken liver was satisfactory determined as shown in table 1 . \n elution patterns of the two control reaction mixtures and reaction mixture spiked or nonspiked with pm are shown in fig . \n two controls ( pm , no - r , and pm+sd , no - r ) showed the same peak height as those of ( pl , no - r and pl+sd , no - r ) , respectively . \n in contrast , the peak height of the reaction mixture ( pm , r ) was higher than that of the reaction mixture ( pl , r ) because the former peak was a total of pl and pm . \n standard pm ( 2 pmol ) was quantitatively converted into 4-pla and determined ( pm+sd , r ) . \n 4c . like pl and pm , pn was also quantitatively converted into 4-pla and determined ( pn+sd , r ) . \n the peak area of the reaction mixture ( pn , r ) was almost the same as that of the reaction mixture ( pl , r ) , showing that the chicken liver sample contained a very low amount of pn . \n hplc chromatograms for the analyses of free forms of vitamin b6 in the chicken liver sample . \n the food sample ( 5l ) was subjected to the enzymatic conversion ( the procedure for pl in fig . \n 3 ) and applied to hplc ( pl , r ) . the food sample , to which standard pl had been added , was subjected to the enzymatic conversion ( pl+sd , r ) . \n their controls were also done ( pl , no - r , and pl+sd , no - r ) . \n ( b ) pm was similarly determined as shown in the procedure for pm in fig . \n ( c ) pn was similarly determined as shown in the procedure for pn in fig . \n 5a , b , and c shows the chromatograms for the analyses of plp , pmp , and pnp . \n the standard plp , pmp , and pnp spiked could be accurately determined ( plp+sd , r ; pmp+sd , r ; and pnp+sd , r ) . \n the peak area of pmp , r was high because it contained 4-pla produced from pl , pm , plp , and pmp . \n thus , pmp content was calculated as shown in table 1 . although the chromatograms obtained in the png analysis is not shown , \n hplc chromatograms for the analyses of phosphoester forms of vitamin b6 in the chicken liver sample . \n arrows show 4-pla peaks . ( a ) plp was determined . the food sample ( 5l ) \n 3 ) and applied to hplc ( pl , r ) . the food sample , to which standard plp had been added , was subjected to the enzymatic conversion ( plp+sd , r ) . \n their controls were also done ( plp , no - r , and plp+sd , no - r ) . \n ( b ) pmp was similarly determined as shown in the procedure for pmp in fig . \n ( c ) pnp was similarly determined as shown in the procedure for pnp in fig . \n the results showed that the method could be applied for the determination of contents of individual vitamin b6 compound in the chicken liver sample . \n the contents of seven vitamin b6 compounds in the foods were successfully determined ( table 2 ) . \n the values are in nmol / g , except for the total content that also is calculated as pn , hcl . \n although the recovery varied a lot among foods , the recovery tests could be determined for all the foods , showing that the present method for preparation of the food samples was almost adequate for the presented method : the standard deviation of the recovery for chicken liver , egg yolk and white , and dried small anchovy are shown in ( table 2 ) . \n the recovery of plp in the egg yolk and the dried anchovy was only 14 and 29% , respectively . \n these results indicate that plp in these foods bind so strongly to the other food component(s ) that alp can not hydrolyze plp bound to them . \n thus , improvement of the sample preparation method is required , although it is better than hcl - hydrolysis ( 9 ) , in which recoveries of plp and pm in garlic could not be estimated . \n another explanation could be that the egg yolk and dried anchovy contain some compounds that inhibit alp . \n although it is not clear what happens , the bioavailability of plp in the egg yolk and anchovy may be low , because it is well known that alp activity correlates significantly with in vivo luminal disappearance of plp ( 19 ) . \n this method may not be applicable for the foods that give a too low recovery of the vitamin b6 compounds . \n in such cases , it is necessary to develop an improved preparation method for the food samples . \n contents of vitamin b6 compounds and png in chicken liver and white meat , egg yolk and white , dried small anchovy , carrot , and garlic ; recoveries of the standards are also shown nd means = not detectable . \n total contents ( as pn - hcl ) of vitamin b6 compounds and png , which have been determined by a microbiological method , are described in the japanese food table . \n they are 35.5nmol / g for chicken white meat ; 38.4 , chicken liver ; 15.4 , egg yolk ; 0 , egg white ; 6.5 , carrot ; 6.5 , garlic ; and 16.6 , dried small anchovy . \n the values in nmol / g were converted from values given as mg/100 g of an edible part in the japanese food table . \n the microbiological and the present methods gave similar values for the contents in the chicken white meat and liver . \n in contrast , higher values were obtained for the other samples with the present method ; in particular , the small dried anchovy had about 9-fold higher value . because its content is in nmol / g , dry weight , in contrast to the other samples , the value obtained by the present method seems to be rational , suggesting that vitamin b6 compounds in the dried small anchovy could not be measured accurately by the microbiological method . \n the preferred comparison would be to measure the same food samples by the microbiological method and the present method . \n the relative contents of png were 19 and 42% of the total content in garlic and carrot , respectively . \n the png contents in plant - derived food have been reported to vary from 0% ( cauliflower ) ( 3 ) to 92% ( orange juice ) ( 20 ) . \n the contents of pn , which is generally used as a supplement of vitamin b6 for fortified foods , were very low in the foods examined ; actually pn was not found in either chicken white meat or egg yolk . \n although analytical data for many other foods are required , the results suggest a need for studies to determine which of the vitamin b6 compounds should be used for fortification of the food . \n each of the foods analyzed showed a characteristic pattern of the seven vitamin b6 compounds . \n the dried anchovy contained high amounts of plp , pmp , and pnp ; the free forms were very low . \n traditionally , the dried small anchovy has been used for making stock soup in japan . however , recently , a new preparation for taking the whole body as a food has been developed , which is the sample used in this study . because pmp should be absorbed as pm in the intestine after the hydrolysis by the alp , which shows reactivity toward pmp as well as plp , pmp should have the same efficacy as pm in the anti - diabetic complications . \n thus , the chicken liver and the dried small anchovy may be candidates for foods functioning to prevent diabetic complications . \n the time needed for determination of seven vitamin b6 compounds in one food by a trained student was approximately 10 days . \n although the first analysis was finished within 2 days , the additional days were required to determine recoveries of the recovery test . \n thus , the development of a determination kit consisting of the seven reaction mixtures and the seven standard compounds would be necessary to facilitate easier routine analysis of the food samples . \n the chromatograms for the analysis of pl , pm , and pn in the chicken liver sample are shown in fig . \n 4 : they showed similar but definitely different patterns of the fluorescent peaks of 4-pla . \n the 4-pla was eluted at about 11 min as shown with an arrow , together with unknown fluorescent compounds in the sample . \n the control reaction showed a tiny peak of 4-pla ( pl , no - r in fig . \n 4a ) , its area increased when the liver sample was subjected to the enzymatic conversion with the reaction 1 in fig . \n 2 for converting pl into 4-pla ( pl , r ) in the sample . thus , the subtraction of fluorescence intensity of 4-pla ( pl , no - r ) from that of 4-pla ( pl , r ) gave the amount of 4-pla corresponding to the amount of pl in the sample . the chicken liver sample spiked with \n pl showed a higher peak ( pl+sd , r ) because the corresponding amount of 4-pla was additionally produced by the enzymatic conversion . \n in contrast , the chicken liver sample spiked with pl without the enzymatic conversion did not show this increase ; its peak height was the same as that of the control reaction mixture ( pl+sd , no - r ) . \n thus , pl in the chicken liver was satisfactory determined as shown in table 1 . \n elution patterns of the two control reaction mixtures and reaction mixture spiked or nonspiked with pm are shown in fig . \n two controls ( pm , no - r , and pm+sd , no - r ) showed the same peak height as those of ( pl , no - r and pl+sd , no - r ) , respectively . \n in contrast , the peak height of the reaction mixture ( pm , r ) was higher than that of the reaction mixture ( pl , r ) because the former peak was a total of pl and pm . \n standard pm ( 2 pmol ) was quantitatively converted into 4-pla and determined ( pm+sd , r ) . \n 4c . like pl and pm , pn was also quantitatively converted into 4-pla and determined ( pn+sd , r ) . \n the peak area of the reaction mixture ( pn , r ) was almost the same as that of the reaction mixture ( pl , r ) , showing that the chicken liver sample contained a very low amount of pn . \n hplc chromatograms for the analyses of free forms of vitamin b6 in the chicken liver sample . \n the food sample ( 5l ) was subjected to the enzymatic conversion ( the procedure for pl in fig . \n 3 ) and applied to hplc ( pl , r ) . the food sample , to which standard pl had been added , was subjected to the enzymatic conversion ( pl+sd , r ) . \n their controls were also done ( pl , no - r , and pl+sd , no - r ) . \n ( b ) pm was similarly determined as shown in the procedure for pm in fig . \n ( c ) pn was similarly determined as shown in the procedure for pn in fig . \n 5a , b , and c shows the chromatograms for the analyses of plp , pmp , and pnp . \n the standard plp , pmp , and pnp spiked could be accurately determined ( plp+sd , r ; pmp+sd , r ; and pnp+sd , r ) . \n the peak area of pmp , r was high because it contained 4-pla produced from pl , pm , plp , and pmp . \n thus , pmp content was calculated as shown in table 1 . although the chromatograms obtained in the png analysis is not shown , \n hplc chromatograms for the analyses of phosphoester forms of vitamin b6 in the chicken liver sample . \n arrows show 4-pla peaks . ( a ) plp was determined . the food sample ( 5l ) \n 3 ) and applied to hplc ( pl , r ) . the food sample , to which standard plp had been added , was subjected to the enzymatic conversion ( plp+sd , r ) . \n their controls were also done ( plp , no - r , and plp+sd , no - r ) . \n ( b ) pmp was similarly determined as shown in the procedure for pmp in fig . \n ( c ) pnp was similarly determined as shown in the procedure for pnp in fig . \n the results showed that the method could be applied for the determination of contents of individual vitamin b6 compound in the chicken liver sample . \n the contents of seven vitamin b6 compounds in the foods were successfully determined ( table 2 ) . \n the values are in nmol / g , except for the total content that also is calculated as pn , hcl . \n although the recovery varied a lot among foods , the recovery tests could be determined for all the foods , showing that the present method for preparation of the food samples was almost adequate for the presented method : the standard deviation of the recovery for chicken liver , egg yolk and white , and dried small anchovy are shown in ( table 2 ) . \n the recovery of plp in the egg yolk and the dried anchovy was only 14 and 29% , respectively . \n these results indicate that plp in these foods bind so strongly to the other food component(s ) that alp can not hydrolyze plp bound to them . \n thus , improvement of the sample preparation method is required , although it is better than hcl - hydrolysis ( 9 ) , in which recoveries of plp and pm in garlic could not be estimated . \n another explanation could be that the egg yolk and dried anchovy contain some compounds that inhibit alp . \n although it is not clear what happens , the bioavailability of plp in the egg yolk and anchovy may be low , because it is well known that alp activity correlates significantly with in vivo luminal disappearance of plp ( 19 ) . \n this method may not be applicable for the foods that give a too low recovery of the vitamin b6 compounds . \n in such cases , it is necessary to develop an improved preparation method for the food samples . \n contents of vitamin b6 compounds and png in chicken liver and white meat , egg yolk and white , dried small anchovy , carrot , and garlic ; recoveries of the standards are also shown nd means = not detectable . \n total contents ( as pn - hcl ) of vitamin b6 compounds and png , which have been determined by a microbiological method , are described in the japanese food table . \n they are 35.5nmol / g for chicken white meat ; 38.4 , chicken liver ; 15.4 , egg yolk ; 0 , egg white ; 6.5 , carrot ; 6.5 , garlic ; and 16.6 , dried small anchovy . \n the values in nmol / g were converted from values given as mg/100 g of an edible part in the japanese food table . \n the microbiological and the present methods gave similar values for the contents in the chicken white meat and liver . \n in contrast , higher values were obtained for the other samples with the present method ; in particular , the small dried anchovy had about 9-fold higher value . \n because its content is in nmol / g , dry weight , in contrast to the other samples , the value obtained by the present method seems to be rational , suggesting that vitamin b6 compounds in the dried small anchovy could not be measured accurately by the microbiological method . \n the preferred comparison would be to measure the same food samples by the microbiological method and the present method . \n the relative contents of png were 19 and 42% of the total content in garlic and carrot , respectively . \n the png contents in plant - derived food have been reported to vary from 0% ( cauliflower ) ( 3 ) to 92% ( orange juice ) ( 20 ) . \n the contents of pn , which is generally used as a supplement of vitamin b6 for fortified foods , were very low in the foods examined ; actually pn was not found in either chicken white meat or egg yolk . \n although analytical data for many other foods are required , the results suggest a need for studies to determine which of the vitamin b6 compounds should be used for fortification of the food . \n each of the foods analyzed showed a characteristic pattern of the seven vitamin b6 compounds . \n the dried anchovy contained high amounts of plp , pmp , and pnp ; the free forms were very low . \n both the plp and pl were high in the garlic . traditionally , the dried small anchovy has been used for making stock soup in japan . \n however , recently , a new preparation for taking the whole body as a food has been developed , which is the sample used in this study . \n because pmp should be absorbed as pm in the intestine after the hydrolysis by the alp , which shows reactivity toward pmp as well as plp , pmp should have the same efficacy as pm in the anti - diabetic complications . \n thus , the chicken liver and the dried small anchovy may be candidates for foods functioning to prevent diabetic complications . \n the time needed for determination of seven vitamin b6 compounds in one food by a trained student was approximately 10 days . \n although the first analysis was finished within 2 days , the additional days were required to determine recoveries of the recovery test . \n thus , the development of a determination kit consisting of the seven reaction mixtures and the seven standard compounds would be necessary to facilitate easier routine analysis of the food samples . \n the content of pn that is used for fortification of foods was very low in the foods examined . \n this work was partially supported by the ministry of education , culture , sports , science and technonogy- japan , grant - in - ais for exploratory research , 18650215 , 2007 , and by a kochi university president 's discretionary grant .", "role": "user" }, { "content": "backgroundthere is a need for a reliable and accurate method for quantification of each of the seven individual vitamin b6 compounds including pyridoxine--glucoside in foods.objectiveto determine pyridoxal ( pl ) , pyridoxamine ( pm ) , pyridoxine ( pn ) , pyridoxal 5-phosphate ( plp ) , pyridoxamine 5-phosphate ( pmp ) , pyridoxine 5-phosphate ( pnp ) , and pyridoxine--glucoside ( png ) in foods.designby specific enzymatic treatment , each of the seven vitamin b6 compounds was all converted into 4-pyridoxolactone , which is a highly fluorescent compound . in total , seven separate , enzymatic steps were performed for each sample . \n separation and quantification were performed with reversed - phase high performance liquid chromatography ( hplc ) coupled with fluorescence detection . for each sample type \n the result was corrected for the recovery based on spiked samples . \n the method was applied for analyses of chicken liver , chicken white meat , egg yolk , egg white , dried anchovy , carrots , and garlic.resultsthe recovery varied from 14 to 114% in chicken liver , chicken white meat , egg yolk , egg white , dried anchovy , carrot , and garlic . \n each food showed a characteristic distribution of the seven vitamin b6 compounds . \n the png was only found in low amounts ; that is , 1729nmol vitamin b6/g in the plant - derived foods , carrot and garlic . \n only egg white showed a lower content , 3nmol / g . \n overall the content in chicken liver , chicken white meat , and egg yolk had a total content of vitamin b6 between 42 and 51nmol / g . \n both pm and pmp were high in the chicken liver . \n in contrast , pl and plp were high in the chicken white meat . \n the main vitamin b6 in the egg yolk was plp . \n the dried anchovy contained high amounts of plp and pmp and a total content of 144nmol / g.conclusionsthe enzymatic - based hplc method was applied for the determination of seven vitamin b6 compounds in foods . \n their distribution in the foods varied significantly .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: esthetic restorative materials have been widely used in dentistry in both anterior and posterior restorations . \n currently , four types of direct esthetic restorations are commonly used in dentistry which includes - resin composites , poly - acid modified resin composites ( copolymer ) , glass - ionomers and resin - modified glass - ionomers . resin composites were introduced in the early 1970s and are widely used as direct esthetic restorations.1 however , composite restorations have several significant disadvantages despite their continuous improvements.2 despite the fact that composites are currently the most preferred material for restorations , the polymerization shrinkage of 2.6 - 7.1% prompting microleakage is one of the most often observed issues.3 - 14 microleakage is one of major drawbacks of composites which lead to the development of recurrent caries , post - operative sensitivity , enamel fracture , marginal staining , and eventual failure of restorations.4 studies have reported several ways to decrease this problem such as slowing down the composite polymerization rate , utilizing an incremental build - up technique or placement of low modulus in - between layer , and reducing the c factor ( the ratio of bonded to non - bonded restoration surfaces).5 silorane - based dental resin composite materials introduced to clinical dentistry with an intention to reduce the polymerization shrinkage . \n its matrix consists of siloxanes and oxiranes , which differ from the conventional methacrylate ( bis - gma ) composite matrix . \n manufactures claimed that the silorane - based composites produce low - polymerization shrinkage of < 1% volume by using archimedes method . \n silorane - based composites polymerize by open ring polymerization reaction minimizing polymerization shrinkage and polymerization stress . \n thus , it provided a high - performance bond to the tooth with significantly lower shrinkage than other methacrylate composites.6,7 moreover , oxirane and siloxane were hydrophobic in nature making silorane based composites chemically stable in aqueous solution with less water sorption . \n additionally , silorane - based dental resin composites showed less microleakage and cusp deflection compared to other bis - gma composite materials.8 - 11 by utilizing bis - gma based technology , nontraditional \n these contained sub - micrometer particles ( nano fillers ) to further improve the physical and optical properties of the resins . \n nanohybrid composites with bis - gma base has been recommended for both anterior and posterior restorations . \n further , it has been improved to overcome composite drawbacks of recurrent caries , post - operative sensitivity , enamel fracture , marginal staining , microleakage and eventual failure of restorations . to the best of our knowledge \n , there is a lack of reported studies on the comparative evaluation of microleakage of methacrylate based nanohybrid and microhybrid composite materials with that of silorane based microhybrid composite materials . \n hence , the present study is aimed to evaluate the microleakage of silorane based ( filtek p90 , using two - step self - etching prime and bond adhesive system ) composite material with that of methacrylate based ( filtek z250 using single bond universal total etching technique , and filtek z250 xt using single bond universal self - etching technique ) composite materials in class i restorations by using stereomicroscope . \n sixty extracted intact upper premolar teeth were collected , and removal of calculus and debris was carried out by hand scaling . \n teeth were kept in 5% sodium chloride solution at 37c for the duration of 1-month . \n teeth were prepared approximately 4 mm in length mesio - distally , 2 mm bucco - palatally in width and 3 mm in depth to receive standardized class i restoration by using prefabricated polyvinylsiloxane mold of 2 mm 4 mm . \n the corresponding depth of the cavity was obtained with a diamond bur marked 3 mm from the tip of the bur . marking on the bur was carried out by using carborundom disk ( b&d technologies co , nanyi street , qiaonan , panyu , guangzhou , 511400 china ) . \n all the class i cavities was prepared by using high - speed hand piece ( nsk , avtec dental , mount pleasant , sc , u.s.a . ) , under constant water irrigation . \n 90 cavosurface margins were prepared , and the dimensions of the cavities were checked with a boley gauge . \n later teeth were randomly divided into three ( n = 20 ) groups to receive restorative materials . \n paul , mn , usa ) with single universal bond ( 3 m espe , st . \n paul , mn , usa ) was used as per the manufacturer s instructions . a phosphoric acid etching gel 35% ( scotchbond universal etchant , 3 m espe , st . \n paul , mn , usa ) was applied to the prepared tooth structure for 15 s. rinsed with water and dried carefully without incorporating oil or water . \n then , the bonding agent applied with a scrubbing motion for 20 s and exposed to the gentle stream of air for 5 s , followed by light curing for 10 s. filtek z250 was placed in a cavity in two increments with each increment of 2 mm size as per the manufacturer s instructions and light cured for 40 s. filtek z250 xt resin composite shade a2 ( 3 m espe , st . \n paul , mn , usa ) with single bond universal self - etch ( 3 m espe , st . \n the bonding agent was applied with a scrubbing motion for 20 s and the gentle air stream was passed for 5 s before it could be light - cured for 10 s. composite filtek z250 xt was placed by using the same protocol as described in group a. low shrinkage resin composite filtek p90 shade a2 ( 3 m espe , st . \n paul , mn , usa ) with ls system adhesive primer and bond ( 3 m espe , st . \n the p90 primer was applied by using a micro brush for 15 s , mildly air - dried and finally light - cured for 10 s. the p90 bond was applied , and a gentle stream of air was passed and light - cured for 10 s. p90 was placed by using the same protocol as described in group a according to the manufacturer s instructions and light cured for 40 s. all the specimens were polished with a series of coarse , medium , fine and superfine polishing discs ( sof - lex ; 3m / espe , st . \n paul , mn , usa ) with an electric handpiece ( k10 ; kavo , leutkirch , germany ) at 15,000 rpm for 15 s. all composite increments were light - cured using elipar s10 led curing unit ( 3 m espe , seefeld , germany ) at a power density of 1000 mw / cm for 40 s in a soft start mode , whereas adhesive systems were light - cured using the same light - curing unit with power density for 10 s. the integrated radiometer was utilized to continuous monitoring of the light intensity . \n teeth were then subjected to thermocycling , according to the international organization for standardization standard 11405 for 500 cycles at 5c-55c with a 30-s dwell time . before the evaluation of the microleakage , the root apices of the teeth were completely sealed with sticky wax and two coats of nail varnish was applied over the crown root surfaces except for 1 mm around the restoration margins . \n teeth were then drenched in 2% methylene blue dye ( ph = 7 ) ( merck specialties private ltd , mumbai , india ) at 37c for 24 h,13 washed and dried . \n later on , teeth were embedded in the acrylic resin block sectioned longitudinally in bucco - lingual direction with a diamond saw mounted on a cutting machine ( isomet 1000 ; buehler 41 waukegan road lake bluff , illinois usa ) . \n stereomicroscopic ( motic microscopes , china ) examination was carried out to determine the extent of dye penetration at 40 magnification based on five - point scale . \n 0 = no dye penetration;1 = dye penetration limited to outer half of the axial wall;2 = dye penetration limited to inner half of the axial wall;3 = dye penetration reach the pulpal wall;4 = dye penetration beyond the pulpal wall \n . \n 0 = no dye penetration ; 1 = dye penetration limited to outer half of the axial wall ; 2 = dye penetration limited to inner half of the axial wall ; 3 = dye penetration reach the pulpal wall ; 4 = dye penetration beyond the pulpal wall . \n whitney test were executed at a 95% significance level to evaluate any significant differences among the composite materials . \n all the statistical analysis was performed using the spss 11.0 program ( spss inc . , \n paul , mn , usa ) with single universal bond ( 3 m espe , st . \n paul , mn , usa ) was used as per the manufacturer s instructions . a phosphoric acid etching gel 35% ( scotchbond universal etchant , 3 m espe , st . \n paul , mn , usa ) was applied to the prepared tooth structure for 15 s. rinsed with water and dried carefully without incorporating oil or water . \n then , the bonding agent applied with a scrubbing motion for 20 s and exposed to the gentle stream of air for 5 s , followed by light curing for 10 s. filtek z250 was placed in a cavity in two increments with each increment of 2 mm size as per the manufacturer s instructions and light cured for 40 s. \n paul , mn , usa ) with single bond universal self - etch ( 3 m espe , st . \n the bonding agent was applied with a scrubbing motion for 20 s and the gentle air stream was passed for 5 s before it could be light - cured for 10 s. composite filtek z250 xt was placed by using the same protocol as described in group a. \n paul , mn , usa ) with ls system adhesive primer and bond ( 3 m espe , st . \n the p90 primer was applied by using a micro brush for 15 s , mildly air - dried and finally light - cured for 10 s. the p90 bond was applied , and a gentle stream of air was passed and light - cured for 10 s. p90 was placed by using the same protocol as described in group a according to the manufacturer s instructions and light cured for 40 s. all the specimens were polished with a series of coarse , medium , fine and superfine polishing discs ( sof - lex ; 3m / espe , st . \n paul , mn , usa ) with an electric handpiece ( k10 ; kavo , leutkirch , germany ) at 15,000 rpm for 15 s. all composite increments were light - cured using elipar s10 led curing unit ( 3 m espe , seefeld , germany ) at a power density of 1000 mw / cm for 40 s in a soft start mode , whereas adhesive systems were light - cured using the same light - curing unit with power density for 10 s. the integrated radiometer was utilized to continuous monitoring of the light intensity . \n teeth were then subjected to thermocycling , according to the international organization for standardization standard 11405 for 500 cycles at 5c-55c with a 30-s dwell time . before the evaluation of the microleakage , \n the root apices of the teeth were completely sealed with sticky wax and two coats of nail varnish was applied over the crown root surfaces except for 1 mm around the restoration margins . \n teeth were then drenched in 2% methylene blue dye ( ph = 7 ) ( merck specialties private ltd , mumbai , india ) at 37c for 24 h,13 washed and dried . \n later on , teeth were embedded in the acrylic resin block sectioned longitudinally in bucco - lingual direction with a diamond saw mounted on a cutting machine ( isomet 1000 ; buehler 41 waukegan road lake bluff , illinois usa ) . \n stereomicroscopic ( motic microscopes , china ) examination was carried out to determine the extent of dye penetration at 40 magnification based on five - point scale . \n 0 = no dye penetration;1 = dye penetration limited to outer half of the axial wall;2 = dye penetration limited to inner half of the axial wall;3 = dye penetration reach the pulpal wall;4 = dye penetration beyond the pulpal wall . \n 0 = no dye penetration ; 1 = dye penetration limited to outer half of the axial wall ; 2 = dye penetration limited to inner half of the axial wall ; 3 = dye penetration \n whitney test were executed at a 95% significance level to evaluate any significant differences among the composite materials . \n all the statistical analysis was performed using the spss 11.0 program ( spss inc . , \n descriptive statistics of mean , standard deviations , minimum and maximum scores for dye penetration test as shown in table 1 . \n it is evident that silorane - based composite material filtek p90 showed a highest mean score of 2.45 0.89 , as compared to methacrylate based composite materials filtek z250 and filtek z250 xt . \n number of teeth showing dye penetration reaching the pulpal wall of the cavity was highest ( n = 14 ) with filtek p90 ( silorane ) composite material as compared to the other two composite materials ( graph 1 ) . dye penetration test . \n a highest mean rank of 33.65was observed with filtek p90 ( silorane ) , followed by 30.55 and 27.30 filtek z250 xt ( methacrylate ) and filtek z250 ( methacrylate ) , respectively . \n wallis test revealed no significant differences ( p > 0.05 ) ( table 2 ) . \n microleakage of composite restorations occurs because of stresses encountered along the tooth / restoration margin from polymerization shrinkage , temperature variation in the oral environment , and mechanical fatigue through repetitive masticatory loading.15 the most likely cause for this phenomenon is polymerization contraction , which is toward the stronger enamel composite joint and the light source.4 - 10 various methods to detect microleakage \n have been suggested including dye leakage method , the use of color producing microorganisms , radioactive isotopes , the air pressure method , neutron activation analysis , electrochemical studies , scanning electron microscopy , thermal and mechanical cycling , and chemical tracers.16 a newer silorane - based composite resin system was designed to overcome polymerization contraction issue.6,17 the present study investigated the microleakage of a new low - shrinkage resin composite and clinically successful methacrylate resin composites in class i restorations . \n these class i cavities was chosen due to the high c - factor that leads to greater polymerization stresses,18 as a result of confined contraction by number of bonded surfaces . \n we did not observe any significant differences in microleakage among the three composite resin systems . \n this could be due to the use of high intensity light in a continuous mode , self - etch hydrophilic adhesive systems , the increased c - factor of class i cavity and the relatively excess concentration of methylene blue dye used in this study . \n the results of this study showed that the novel low - shrink composite restorative material filtek p90 showed microleakage score very similar to z250 with total etch single universal bond and z250 xt with self - etch systems after thermocycling . \n assumption that microleakage of the cavities restored with the filtek p90 ( silorane ) compared to the other resin - based composite materials was due to the innate ring - opening polymerization reaction of the silorane monomers , which make - up the volumetric reduction as the molecules approximate each other . however , radical polymerization of the other resin based composites is obvious with a decrease in polymerization shrinkage stresses at the tooth / restoration border . \n therefore , methacrylate based composite resins better withstand thermocycling fatigue at the tooth / restoration interface . \n our study is in agreement with the study reported by ernst , who presented no differences in microleakage of teeth restored with silorane and methacrylate composite ( tetric ceram ) with adhesive ( clearfil se bond ) system.13 this finding was due to the fact that author used an all - in - one ( 7 generation ) experimental bond of silorane before produced by the company . \n whereas in the present study new bond produced with silorane composite system consisting of two - step and two components ( 6 generation ) was utilized . however , we disagree with study results of palin et al . \n , who have reported that the microleakage of silorane based composite material was lesser than that of methacrylate based composite after occlusal loading forces.19 - 21 \n within the limitation of this study , we conclude that all the restorative systems tested in this study exhibited microleakage \n . however , silorane based technology ( filtek p90 ) showed higher microleakage scores compared with the clinically successful methacrylate - based composite materials ( filtek z250 and filtek z250 xt ) .", "role": "user" }, { "content": "background : to evaluate the microleakage of silorane - based composite material ( filtek p90 ) with that of two homologous methacrylate - based composites materials ( filtek z250 and filtek z250 xt ) , by using two different bonding techniques.materials and methods : sixty extracted human maxillary first premolars prepared for standardized class i cavities ( 4 mm 2 mm 2 mm ) were randomly divided into three groups . \n group a ( n = 20 ) was filled with filtek z250 ( methacrylate ) using single bond universal total etching technique , group b ( n = 20 ) was filled with filtek z250 xt ( methacrylate ) using single bond universal self - etching technique and group c ( n = 20 ) restored with filtek p90 ( silorane ) with dedicated two - step self - etching prime and bond adhesive system ( p90 system adhesive ) . \n teeth were subjected to thermocycling regime ( 500 , 5 - 55c ) , and dye penetration by immersing in 2% methylene blue for 24 h. tooth sectioning was performed , and extent of the dye penetration was scored based on dye penetration scale to evaluate the microleakage . statistical analysis included descriptive statistics and inferential statistics of kruskal wallis test to compare the mean ranks between groups.results:there was no significant difference observed for microleakage among the three composite materials tested in the present study . \n however , the cavities restored with silorane ( filtek p90 ) based composite displayed higher microleakage than the filtek z250 , z250 xt.conclusion:all the restorative systems tested in this study exhibited microleakage , but the silorane technology showed more microleakage when compared to the methacrylate - based composite systems .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: they allow the distinction of focal nodular hyperplasia ( fnh ) from hca and the identification of 2 major hca subtypes which represent more than 80% of all hcas , namely , hnf1a - mutated hca ( h - hca , 3540% ) , and inflammatory hca ( ihca , 5055% ) ; 10% of ihcas being also -catenin mutated ( b - ihca ) . \n the remaining hcas are the -catenin - mutated hca ( b - hca , 10% ) and the unclassified hca ( uhca , which account for less than 10% ) . the immunohistochemical ( ihc ) classification of hca subtypes was derived from the above - mentioned molecular characterization and showed a good correlation with the molecular data . using specific ihc markers , such as liver fatty acid - binding protein ( lfapb ) , c reactive protein ( crp ) or serum amyloid a ( saa ) , glutamine synthetase ( gs ) , and -catenin , it is possible to identify all hca subgroups with good confidence . among these markers , \n gs is of major importance to identify patients at high risk of malignant transformation . indeed , abnormal gs staining is a strong argument to suggest -catenin activation . \n unfortunately , the rarity of these tumors in routine practice leads to the low availability of the specific ihc markers . to overcome this problem , \n the possibility to identify the 2 major hca subtypes using standard histological techniques has not been tested . \n the aim of this study was ( 1 ) to test routine histology in the diagnosis of hca subtypes and ( 2 ) to appreciate the contribution of gs combined with routine histology in the diagnosis of hca subtypes . \n all surgically removed hca cases were retrieved from our files from january 2000 to november 2011 . \n we excluded cases with specific etiologies such as glycogenosis , male hormone administration ( since they are very particular and rare conditions ) , cases of obvious hepatocellular carcinoma ( hcc ) possibly related to hca ( but without formal confirmation for a previous hca ) and cases with massive hemorrhage or necrosis ( but without sufficient nonnecrotic tissue available ) . for all patients ( 114 cases ) , \n the following data were available : clinical data : sex , age , body mass index ( bmi ) , oral contraception ( oc ) , and imaging data ( number and size of nodules ) , as well as liver enzymes ( ap , ggt ) and crp in blood , routine stainings on paraffin sections in tissue areas devoid or with minimal necrosis or hemorrhage , ihc stainings . \n since 2007 , ihc has been performed prospectively ; prior this date , it had been performed retrospectively in all cases , according to previous published studies ; the hca classification was based on ihc . \n briefly , taking into account our own experience previously published in pathology textbooks showing the very good correlation between molecular analysis and ihc for h - hca and ihca [ 5 , 6 ] : lfabp negativity in tumor ( t ) and positivity in nontumoral liver ( ntl ) was interpreted as h - hca ; saa or crp staining positive in t and negative in ntl was interpreted as ihca ( even though saa / crp detection could be more or less intense and homogeneous ) . \n once the diagnosis of h - hca was made , saa or crp staining was not mandatory , and once the diagnosis of ihca was made , lfabp staining was not mandatory in such cases . \n the expression of gs , a target gene of -catenin , was studied in all cases . \n gs staining was quoted as negative in the absence of abnormal expression ; and in this situation , -catenin staining was not mandatory . \n gs staining in t always differed from the normal distribution of gs which is limited to a few rows of centrilobular hepatocytes in the ntl . in t , gs could be either totally absent or restricted to the border between t and ntl and/or around some persistent veins within t. abnormal expression of gs characterized in the whole nodule by a strong positive staining diffuse or patchy is easy to interpret but is more difficult to interpret when the staining is faint or focal , limited to individual or groups of positive hepatocytes irregularly distributed within the tumor . in any case , \n abnormal gs staining was considered as a strong argument to suggest -catenin activation [ 3 , 4 ] . in these cases , \n aberrant -catenin nuclear labelling allowed the final diagnosis of b - hca or b - ihca . \n however , the absence of labelled nuclei on the section did not rule out this diagnosis because the number of labelled nuclei can be heterogeneously distributed , extremely limited , or even absent on the section . \n when all the specific ihc markers were negative , the hca was termed uhca . clinical data : sex , age , body mass index ( bmi ) , oral contraception ( oc ) , and imaging data ( number and size of nodules ) , as well as liver enzymes ( ap , ggt ) and crp in blood , routine stainings on paraffin sections in tissue areas devoid or with minimal necrosis or hemorrhage , ihc stainings . since 2007 , \n ihc has been performed prospectively ; prior this date , it had been performed retrospectively in all cases , according to previous published studies ; the hca classification was based on ihc . \n it corresponded to cases not included in the above series and concerning cases from our center ( prior 2000 ) or from cases sent for advice . for each case , 2 slides of h&e and gs ( including the tumoral and nontumoral part ) were available . \n group a included h - hca cases and ihca ( -catenin and non -catenin activated ) cases . \n group b included all other cases . for each group and subgroup , we analysed the clinical , imaging , and biological data . \n routine stainings ( h&e , trichrome ) as well as cd34 , keratin ( k)7 immunostainings of the 2 groups were reviewed ( t and ntl ) according to a flowchart ( see table 1 in supplementary material available online at http://dx.doi.org/10.1155/2013/417323 ) filled out by an experienced liver pathologist ( pbs ) . \n different pathological items were checked such as steatosis , sinusoidal dilatation , congestion , inflammation , thick arteries , ductular reaction . \n steatosis and sinusoidal dilatation were graded semiquantitatively as indicated in supplemental table 1 . in the ntl , \n routine slides from hca previously classified into subgroups using ihc were analyzed blindly by 2 observers ( pbs and cb ) , without knowing ihc results obtained previously . \n for each case the following diagnosis based on standard features was : h - hca , ihca , and b - ihca with 3 possibilities : yes , no , and possibly for each . for each case the pathological items had to be filled ( supplemental table 1 ) . if the features were obvious , the diagnosis was certain : h - hca , ihca ( group a ) or another type ( group b ) . \n the diagnosis of h - hca was uncertain when the result of the reading was h - hca possibly , \n abnormal gs staining in hca without characteristics of h - hca or ihca was a strong argument to suggest b - hca . \n in exceptional cases , we observed abnormal gs staining in h - hca . because of the rather straight criteria defined to categorize cases , disagreement among observers was extremely rare , and when it happens , slides were reviewed and a consensus reached . \n disparities between groups according to age , bmi score , size , and the number of nodules were analyzed with unpaired t - test . abnormal liver enzyme and crp levels in each group \n fisher 's exact test was used to compare different types of pathological abnormalities ( steatosis and sinusoidal dilatation ) . \n kappa statistic was used to measure the level of agreement between the standard histological including gs versus the complete set of immunohistochemical techniques to estimate proportions of hca subtypes . \n amongst the 114 cases analysed by standard morphology and ihc , there were 98 cases in group a : 45 h - hcas , 53 ihcas ( 9 were b - ihcas ) and 16 cases in group b : 4 b - hcas and 12 uhcas . \n the relevant clinical , biological , and pathological data are shown in figures 1 and 2 , table 1 and supplemental table 2 . \n this subgroup included 45 patients , all women ; 84.4% were under oc ; the median age was 40 . \n the median size of the largest nodule was 5.6 cm , and nodules were multiple in 62.2% . \n 75% of the patients had normal bmi ; crp and ap were rarely elevated , and 22.2% had mild elevation of ggt . \n interestingly enough , in 8 patients , micro / small h - hca ( < 1 cm ) were incidentally discovered on the surgical specimen removed for large tumor(s ) of different types including focal nodular hyperplasia ( 4 cases ) , 1 angiomyolipoma , 1 hcc ; in other 2 cases , there was a past history of melanoma , and the small nodules were suspected to be metastases . \n steatosis was the hallmark of h - hca , which usually exhibited a very characteristic aspect with lobulated contours ( figures 2(a ) and 3(a ) ) . \n steatosis was present in 95.6% of cases : severe ( > 60% ) , moderate ( 3060% ) , or mild ( 1030% ) in 25.5 , 44.2 , and 30.2% respectively ; it was totally absent in 2 cases only ( figure 3(b ) ) . \n most of the time , steatotic hepatocytes were intermingled with clear hepatocytes which were predominant in 2 cases , with mild / focal steatosis . \n sinusoidal dilatation was present in 24.4% and mild in the great majority of cases . in two cases \n micro - h - hca nodules were observed in surrounding liver in 53.3% of the cases . \n other types of nodules ( fnh and hemangioma ) were associated in 22.2% of cases . \n this group included 44 patients ( 39 women and 5 men ) ( figures 1(a)1(e ) ) . \n the median size of the largest nodule was 6 cm , and nodules were solitary in 63.6% . \n bmi scoring revealed a strong association between ihca and high bmi score ( figure 1(a ) , p < 0.05 ) . crp and ggt were raised in 68.2 and 75% , respectively ; ap , ggt , and crp were higher than in the other hca groups ( h - hca / b - hca / uhca ( p < 0.01 ) . \n the hallmark of ihca was the inflammation ( presence of inflammatory cells mainly around pseudoportal tracts , sometimes in foci dispersed inside the tumor ) and pseudoportal tracts ( with thick - walled arteries ) present in 97.7 and 90.9% , respectively , ( figure 2(b ) ) , as well as sinusoidal dilatation , which was major or moderate in 30 and 47.5% respectively but focal in 57.5% ( figures 2(c ) and 4(a ) ) . \n steatosis was present in 38.6% of cases but focal in 88.2% and moderate in 47% ( figures 2(a ) and 4(b ) ) . \n surprisingly , in 3 cases , micro h - hca ( < 5 mm , solitary or multiple in 2 or 1 cases , resp . ) confirmed by lack of lfabp expression was incidentally observed on the surgical specimen at distance of ihca . \n the non tumoral liver was steatotic in 27% , and mild sinusoidal dilatation was observed in 13.6% ; both steatosis and sinusoidal dilatation were more frequently observed in ihca group in comparison to others ( figure 2(c ) , p < 0.001 ) . \n comparison between the h - hca and ihca groups showed that multiple nodules were more frequently observed in the h - hca group than in the ihca group ( p < 0.05 , figure 2(d ) ) . \n it is important to note that inflammatory cells , pseudo - pt , or ductular reaction were rare , often not detected in other hca subgroups than ihca ( figure 2(a ) ) . \n however , due to the low number of patients in b - ihca group , no significance difference was observed when compared to h - hca group . in summary , considering standard histological features , steatosis ( frequency and area ) and sinusoidal dilatation ( frequency and area ) may represent the hallmark of h - hca and ihca , respectively , ( figure 2 , p < 0.05 and p < 0.01 , resp . ) . \n this group included 9 patients ( 3 men ) and represented 17% of all ihcas . \n the standard pathological data were identical to ihca ( table 1 , figure 4(c ) ) , and therefore b - ihca could not be identified as such using standard stainings ; however , abnormal gs staining helps for the right diagnosis ( figure 4(d ) ) . \n other subtypes . \n it included 16 patients , all women ( 4 b - hcas and 12 uhcas , median age 26 and 26.5 , resp . ) . \n b - hca subtype was more frequently detected in young patients ( age < 25 ; p < 0.01 , figure 1(d ) ) than in other hcas subtypes . \n areas with many thin dilated veins were observed in 3 cases . in all these 4 b - hca cases , \n uhcas were detected often during complications ( hemorrhage and necrosis ) associated with large tumors . \n no specific pathological features usually existed ( figure 5(a ) ) , but focal steatosis and sinusoidal dilatation were observed in 16.7 and 25% , respectively ( table 2 ) . in 2 cases , numerous thin dilated veins were focally observed . in 2 cases , a few pseudo glandular arrangements were noticed . in 6 cases , \n there were 3 false negative readings ( h - hca negative and ihca + /or possible / or negative . there were 2 cases with no formal diagnosis ( h - hca possible and ihca possible ) . in 3 cases \n correctly classified h - hca , gs was abnormal , with mild and patchy staining irregularly distributed within the tumor . among the 27 ihcas , 24 were correctly classified , as well as 4 out of the 10 b - ihcas . \n there were 3 false negative readings : 1 in the ihca and 2 in the b - ihca . \n there were 6 cases with no formal diagnosis ( 1 in ihca and 4 in the b - ihca series . among the 27 ihcas , \n the agreement between routine technics combined to gs compared to the all sets of ihc markers for the identification of the 2 major hca subtypes gave a kappa index of 0.89 , 0.91/0.54 ( group a ) , and 0.92 ( group b ) ( p < 0.01 ) for h - hca , ihca / b - ihca , and b - hca / uhca respectively , indicating a good agreement between them . \n classification of hca subtypes should allow the selection of patients at high risk of malignant transformation [ 710 ] . \n specific immunohistochemistry is regarded as a main tool to classify hca in clinical practice [ 2 , 11 , 12 ] and still remains the best method to make the differential diagnosis between fnh and hca [ 5 , 6 ] ; however , it may not be widely available due to the fact that hca is a rare and essentially benign disease . \n the present study demonstrates the value of routine histology to classify the 2 major hca subtypes : h - hca and ihca ( table 2 ) , which represents more than 80% of cases at least in europe , without the use of specific ihc . \n the diagnosis of h - hca was often easy on routine h&e due to the presence of fat , a sign often used on mri to identify this subtype [ 1316 ] . \n the major criterion to identify ihca was inflammation associated with pseudo - portal tracts containing thick - walled arteries , with frequent ductular reaction [ 5 , 6 ] . \n sinusoidal dilatation , a major criteria used by radiologist to identify ihca was less sensitive than inflammation to diagnose ihca . \n interestingly enough , it was not possible to identify b - ihca in the absence of gs staining . \n overall , it was possible to strongly suspect an h - hca and ihca / b - ihca on standard routine staining combined to gs in more than 90% of cases related to their hallmark features ( figures 3(a ) and 4(a ) ) and by default the group b - hca / uhca . \n interestingly enough , it was also possible to distinguish b - hca from uhca . moreover , this study outlines the limits of routine histology to identify with certainty h - hca and ihca ( including b - ihca ) . \n first , in h - hca , steatosis can be mild , focal , or rarely absent , and furthermore areas of sinusoidal dilatation can exist . \n on the other hand , in ihca and b - ihca , sinusoidal dilatation can be mild , focal , or absent ; inflammation limited to few portal tracts - like and steatosis generally focal could be severe . \n in addition , in these 2 main subgroups , as well as in all cases of hca , necrotic areas may lead to remodeling and more or less misleading features . in ihca \n particularly , fibrotic bands associated with inflammation and ductular reaction may mimic fnh on routine histology . \n additional ihc is mandatory to assess the right diagnosis . the fact that in the control set , we missed 6 out of 10 b - ihcas based on ihca criteria raises the question of the similarity between the 2 entities . \n the number of b - ihca was , however , too small to be entirely sure to draw any firm conclusion . \n if routine histology represents a reasonable means to identify the 2 major subtypes for pathologists with no access to specific ihc , the lack of identification of b - ihca remains a serious limitation . \n interestingly enough , this study demonstrates the impossibility to identify b - ihca using standard histology . \n surprisingly , none of the b - ihca in our series ( test and validation ) had cytological or architectural abnormalities such as rosette formation as previously mentioned in b - hca . \n abnormal gs expression is a very useful marker to orientate towards -catenin - activated hca ( inflammatory or not ) , particularly when the staining is strong , diffuse , or patchy , even in the absence of aberrant -catenin nuclear staining . \n however , gs reading is not always easy as underlined above , particularly when gs staining is faint and focal ; in these cases , the distinction between a truly abnormal staining and , a few positive cells around veins in the nodule is hard to make . indeed , this was the case in rare h - hca . \n cases of -catenin activation could be observed in h - hca , but this seems extremely rare . \n therefore , the main issue remains the differential diagnosis between ihca and b - ihca . in these difficult cases , \n if there is no nuclear -catenin staining , the issue can be solved only by molecular biology . \n more generally speaking , in the absence of specific complete set of ihc stainings , routine histology combined with gs can be considered as a reasonable surrogate approach for the identification of the 2 major hca subgroups . \n in addition , clinical , biological , and radiological data can help to make the right diagnosis of hca subtype [ 17 , 18 ] . \n we have to remember that the very peculiar pattern of gs immunostaining in fnh is particularly helpful for their diagnosis mainly when different types of nodules are associated with the same liver . \n finally , in the present work , we acknowledged that the h - hca and ihca / b - ihca subtypes using routine histology were obtained on surgical specimens . in a recent article \n , we demonstrate that typical h - hca can be recognized on biopsy on h&e staining ; this is also possible for ihca , but the % of correct diagnosis is lower . therefore , to avoid the difficulty of interpretation and the higher risk of errors on biopsies , ihc is highly recommended . \n ihc not only offers the possibility to identify hca subtypes , but also to confidently make the differential diagnosis between fnh and hca . \n the diagnosis of hca may be difficult particularly for the pathologists are not familiar with these rare tumors . \n characteristic features may exist which can be recognized by general pathologists on routine histology , allowing a good estimation of the 2 major hca subtypes ( > 80% ) . \n therefore , a palliative diagnosis approach can be proposed , when the panel of the antibodies is not available . however , in absence of these features , ihc remains the gold standard to assess the diagnosis of hca and its subtype . \n gs is essential to identify -catenin - activated hca ( inflammatory or not ) that is particularly important to detect patient with higher risk of malignant transformation .", "role": "user" }, { "content": "immunohistochemistry is a valid method to classify hepatocellular adenoma ( hca ) . \n the aim was to test the performance of routine histology combined to glutamine synthetase ( gs ) staining to identify the 2 major hca subtypes : hnf1 inactivated ( h - hca ) and inflammatory hca ( ihca ) . \n 114 surgical cases , previously classified by immunohistochemistry , were analysed . \n group a comprised 45 h - hcas , 44 ihcas , and 9 -catenin - activated ihcas ( b - ihca ) , and group b , 16 b - hca and unclassified hca ( uhca ) . \n steatosis was the hallmark of h - hca . \n ihca and b - ihca were mainly characterized by inflammation , thick arteries , and sinusoidal dilatation ; b - ihca could not be differentiated from ihca by routine histology . \n group b was identified by default . \n a control set ( 91 cases ) was analyzed using routine and gs stainings ( without knowing immunohistochemical results ) . among the 45 h - hcas and 27 ihcas , \n 40 and 24 were correctly classified , respectively . among the 10 b - ihcas , \n 4 were identified as such using additional gs . \n eight of the 9 hcas that were neither h - hca nor ihca were correctly classified . conclusion . \n routine histology allows to diagnose > 85% of the 2 major hca subtypes . \n gs is essential to identify b - hca . \n this study demonstrates that a palliative diagnostic approach can be proposed , when the panel of specific antibodies is not available .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: colorectal cancer is the third most common malignancy worldwide and the fourth most common in the united states , with estimated 146,970 new cases diagnosed in 2009 . in the united states , \n approximately 49,920 cancer - related deaths were attributed to a colorectal malignancy in 2009 , making it second only to mortality from lung cancer . despite the high incidence of colorectal carcinoma that is diagnosed yearly \n , the majority of lesions are resected with curative intent ( 70%80% ) and colorectal cancer - related deaths account for 20%30% of those diagnosed and treated surgically , making it a highly curable malignancy if identified in its early - stages . \n approximately , 55% of colorectal cancers diagnosed on screening are found to present with stage i or ii disease [ 2 , 3 ] . patients with stage i or stage ii colon cancer have a greater than 60% 5-year survival , and for patients with stage i or stage ii rectal cancer , there is a greater than 50% 5-year survival [ 4 , 5 ] . \n because of this , colorectal screening exams in asymptomatic patients have become the recommended standard of care for average risk patients at the age of 50 and at the age of 40 or younger for those in the moderate and high risk groups . \n approximately , 70% of colorectal lesions occur distal to the splenic flexure , and looking at colon cancer alone , approximately 25% are found in the sigmoid , 10% at the rectosigmoid junction , and 4%6% located in the descending colon . \n anatomically , the left colon has a smaller diameter than the right , and as a result , left - sided carcinomas can cause varying degrees of intraluminal occlusion and patients more frequently present with obstructive symptoms . \n large bowel obstructions present a challenging clinical scenario for the physician in the diagnosis , operative management , and the timing of colonic surveillance . \n patients presenting with advanced lesions causing partial or high - grade large bowel obstruction commonly have a distant history or no history of previous colonic surveillance . \n this fact is of clinical importance because the diagnosis of a primary colorectal malignancy is accompanied by an overall incidence of synchronous colorectal cancers between 2% and 10% and a frequency of synchronous adenomatous polyps ranging from 15% to 50% [ 713 ] . \n interestingly , when compared by stage , no statistically significant difference has been found in the overall and disease - free survival between synchronous and primary cancers [ 14 , 15 ] . \n high - grade or completely obstructing lesions may require emergent surgery , and preoperative colonoscopic surveillance by traditional colonoscopy , ct colonography , or barium enema is typically not recommended due to the lack of accuracy . \n colonoscopy has a high rate of unsuccessful passage of the colonoscope as well as the risk of colonoscopic perforation at the site of obstruction or the more proximal cecum secondary to barotrauma . \n this particular scenario leaves the patient without adequate visualization and evaluation of the proximal colon prior to their operative resection ( figures 1 and 2 ) . \n the issue of when and how to best monitor these patients is still debated and evolving , with the recent literature advocating intraoperative , on - table colonic lavage and colonoscopy . \n the current literature and management options for this unique clinical scenario are presented ( table 1 ) . \n colonoscopic screening and surveillance has been proven to be a diagnostic and therapeutic modality that can result in an early diagnosis of colorectal cancer at a curable stage as well as identify and remove premalignant polypoid lesions . \n quality control measures require cecal intubation during colonoscopy to classify the study as satisfactory and complete , thus decreasing the likelihood of failure to detect malignant or premalignant lesions . \n an incomplete colonoscopy is defined as failure of cecal intubation ( or ileocolonic anastomosis ) with nonvisualization of anatomic features ( when present ) , such as the ileocecal valve , appendiceal orifice , or terminal ileum . using a population - based cohort , neerincx and colleagues investigated the rates and reasons for incomplete colonoscopy , as well as the subsequent work - up conducted to evaluate the nonvisualized colon , and the incidence of premalignant and cancerous lesions missed on incomplete colonoscopy . \n several factors were cited to explain the occurrence , most commonly looping scope / excessively long colon ( dolichocolon ) ( 20.4% ) , patient discomfort ( 15.3% ) , obstructing tumor ( 13.9% ) , and insufficient bowel preparation ( 12.1% ) , with stenosis ( 3.7% ) and severe inflammation ( 3.5% ) being less common . during an 18-month follow - up period after incomplete index colonoscopy , \n patients with incomplete colonoscopies due to stenosis ( 78.9% ) , severe inflammation ( 77.8% ) , and obstructing tumor ( 74.6% ) were most likely to receive a secondary examination during the follow - up period . \n barium enema was found to be the most common secondary investigation ( 47.5% ) , despite its low diagnostic yield and miss rate for colorectal cancer detection , which has been reported as high as 22% . \n other modalities for secondary colon examination included repeat colonoscopy ( 20.5% ) , computed tomography colonography ( 4.3% ) , abdominal computed tomography ( 20.9% ) , and surgical exploration ( 6.8% ) . in individuals who underwent follow - up colonic evaluation after incomplete endoscopy , \n colorectal cancer was diagnosed in 3.5% of patients , with advanced adenoma found in 0.8% of patients , therefore noting overall that advanced neoplasia went undetected in 4.3% of patients due to incomplete colonoscopy . as described by neerincx et al . \n , obstructing colorectal tumors may lead to incomplete colonoscopy due to inability to pass the colonoscope beyond the lesion . \n morrin and colleagues advocated for the use of virtual computed tomographic ( ct ) colonography to image and detect synchronous lesions in the colon proximal to an obstructing colorectal lesion . developed in the 1990s , \n virtual ct colonography uses ionizing radiation to create endoluminal radiologic images in multiple planes of an air - insufflated , cleansed colon , while simultaneously imaging extracolonic regions of the body . using this modality , \n lesions greater than 1 cm can be detected with similar accuracy to conventional colonoscopy and with detection rates superior to barium enema . in the morrin et al . \n study , virtual ct colonography correctly staged 81% of colorectal cancers . in patients with incomplete colonoscopy , 97%100% of all colonic segments were adequately visualized on virtual ct colonography , serving as an accurate preoperative imaging option that could be used to investigate for synchronous proximal lesions , thus assisting with operative planning [ 1820 ] . in a study reported by wong et al . , air - inflated magnetic resonance colonoscopy ( mrc ) was as accurate for identification of colonic lesions as colonoscopy . \n as described earlier , while colonoscopy , barium enema , and ct colonography are widely accepted as the principle screening tools for the detection of colorectal cancer , they may be contraindicated and therefore not frequently performed during the hospitalization of patients in need of emergent resection for large bowel obstruction due to carcinoma . in the acute setting of complete or near - total colonic obstruction , where many patients will require an emergent operation , preoperative evaluation of the colon \n palpation of the colon and intraoperative colonic lavage and on - table colonoscopy are the only current diagnostic and therapeutic modalities available in the immediate perioperative time period . \n although preoperative ct imaging may identify a proximal synchronous lesion(s ) along with a high - grade , distal symptomatic obstruction , therapeutic colonoscopic intervention would be delayed until months postoperatively unless intraoperative techniques are used . \n throughout history , intent - to - cure surgical resection has remained the treatment of choice for localized regional colorectal cancer . \n surgical resection of obstructing carcinomas of the colon is associated with significant morbidity and mortality . however \n , surgical management of completely obstructive left - sided colon cancer remains a topic of discussion , particularly as operative techniques and technological advancements have evolved over time . of major concern \n is the theory that failure to detect a synchronous lesion proximal to the site of obstruction may lead to inadequate operative procedures at the time of initial surgical intervention , thus leaving unaddressed pathology behind and necessitating subsequent therapeutic interventions , including possible repeat surgical resection . \n historically , early studies favored performing a subtotal colectomy in the setting of left colon obstruction secondary to tumor [ 23 , 24 ] . at that time \n , subtotal colectomy allowed removal of synchronous cancers and prevented possible development of metachronous lesions in the colon proximal to an obstructing lesion that had not been properly evaluated endoscopically in the preoperative period . \n however , depending on the length of remaining colon and rectum , frequent bowel movements and even fecal incontinence have been reported in patients who underwent subtotal colectomy . over time , the concept of a one - stage resection with intraoperative colonic lavage and primary anastomosis evolved as a potential surgical alternative to subtotal colectomy . on - table \n colonic irrigation was first described by muir in 1968 . before that time , when a segmental resection was performed on an unprepared colon , standard practice was to create a colostomy ( hartmann 's procedure ) at the initial laparotomy . in the uncleansed colon , \n primary anastomosis was deferred at the time of initial surgery due to the anticipated risk of forceful stress associated with passing a solid load across a newly created surgical anastomosis . \n acknowledging the known advantages of operating on a prepped colon , muir investigated feasible techniques to allow a primary anastomosis to be performed when operating on a colon with a high fecal load preoperatively . since proper preoperative preparation of the bowel can be difficult or impossible to achieve when an obstructing colorectal lesion is present , muir proposed washing out the obstructed colon at the time of operation . \n the standard dissection and mobilization are performed as usual , the bowel is clamped and divided distal to the lesion , and the mobilized intestinal segment containing the lesion was inserted into a sterile polythene bag . \n proximal to the lesion , a tube was inserted , and irrigation solution was instilled to lavage the colon . \n the irrigation tubing was then removed , the lesion resected , and the operation completed with a primary anastomosis on the clean and empty colon . modifying muir 's lavage technique , dudley et al . proposed another method of on - table colonic irrigation in 1980 . \n the region of the lesion was dissected , and the bowel was divided distal to the lesion . the proximal bowel was mobilized and a sterile , rigid plastic tube was inserted through the cut end of this portion of bowel . \n a small enterotomy in the terminal ileum was made , through which a foley catheter was inserted and passed across the ileocecal valve . \n the balloon was inflated , and the catheter was secured into place with a purse - string suture . \n irrigation solution was instilled antegrade through the foley catheter , and feculent waste was drained through the rigid plastic tube distally . \n after the colon was determined to be clean , the specimen was resected , and primary anastomosis of the bowel was completed . \n the foley catheter was either removed and the enterotomy closed after resection of the lesion , or the catheter was brought out through a stab wound in the skin to create a tube cecostomy if clinically indicated . \n dudley also noted that by leaving the tube cecostomy in place , a contrast study could be completed in an antegrade fashion if assessment of the anastomosis is necessary in the postoperative period . \n 's technique for on - table antegrade colonic lavage , with only slight modification of the catheter location ( through the appendix , terminal ileum , or cecum ) to deliver irrigation and decrease fecal load proximal to the obstructing lesion in preparation for a primary anastomosis . \n however , colonic lavage via this technique can be cumbersome and time consuming to perform and is associated with possibility of contamination of spilled fecal content into the operative field . acknowledging these concerns , park et al . \n proposed a one - stage resection and primary anastomosis with intraoperative antegrade irrigation in patients with obstructing left - sided colon cancer . \n the proximal end of the irrigation device was connected with the dilated colon proximal to the obstructing lesion . \n the distal end of the device contained two ports : one for drainage of colonic contents and one for insertion of the irrigation catheter . \n after successful colonic irrigation , an on - table colonoscopy was subsequently performed by inserting a colonoscope through the irrigation port . \n intraoperative endoscopic detection of a synchronous lesion prompted the surgeon to increase the extent of resection in 17% of patients when biopsy - proven malignancy was determined on frozen section . \n the device employed in this study was one of the first that facilitated a one - stage procedure in patients with obstructing left - sided colon cancer , as well as enabled a simultaneous on - table colonoscopy to be performed both quickly and easily . determined to decrease incidence of contamination during on - table colonic lavage , buyukgebiz described a technique utilizing a camera sleeve to control irrigation drainage and spillage from the colon in patients with colonic obstruction due to tumor who underwent one - stage colon resection with primary anastomosis . \n the colon was divided proximal to the obstructing mass then introduced it into a nylon sleeve traditionally used to sterilely cover intraoperative camera equipment . \n the sleeve was sutured into place in a telescopic fashion to contain the fecal contents during colonic irrigation and washout via an antegrade catheter inserted through the appendix into the cecum . \n the in - sleeve method eliminated the need for irrigation drainage catheters , which have diameters smaller than that of the colonic lumen , making them prone to becoming clogged with feculent washout material . \n additionally , by washing out the colon , an intraoperative colonoscopy could be effectively performed by introducing the colonoscope through the sleeve and into the transected colon . \n this technique was found to be a safe and quick means to decrease fecal load in obstructed colons , thus decreasing risk of contamination and allowing proper colon preparation for segmental resection with primary anastomosis . \n in addition , it provided successful means to perform intraoperative colonoscopy and evaluate the proximal colon on - table for synchronous lesions [ 19 , 29 ] . intraoperative colonoscopy can be utilized to evaluate the colon that could not be properly endoscopically assessed during the preoperative period . while the entire colon can be manually examined to assess for synchronous lesions at the time of laparotomy , heald and bussey reported that up to 69% of synchronous cancer lesions could not be detected by palpation of the colon . \n given the high failure rate and lack of sensitivity of operative palpation , intraoperative colonoscopy may serve as a useful adjunct . \n however , some concerns have been raised regarding on - table colonoscopy ; in order to be properly performed , colonoscopy requires distension of the bowel with intraluminal air insufflation that may subsequently distort visualization and compromise the planned procedure , especially during laparoscopic surgery . \n if the bowel is over distended during insufflation , the colon may be at risk for mucosal , serosal , or mesenteric tears , regional hypoperfusion of tissue , and potential ischemia . \n carbon - dioxide may be used for colonoscopic insufflation and offers the advantages of rapid absorption and alleviation of bowel distension when compared to atmospheric , nitrogen - rich air . \n nakajima et al . utilized co2-insufflated colonoscopy during laparoscopic colonic resections and found this modality to be feasible , safe , and practical , as it minimized bowel distension without negatively affecting operative exposure or the subsequent procedure . \n additionally , it allowed the entire colon to be assessed endoscopically , rather than manually palpated , further facilitating the planned resection to be performed in a minimally invasive fashion . \n most recently , sasaki et al . investigated whether intraoperative colonic irrigation and on - table colonoscopy may be useful for more accurate diagnosis of colorectal cancer before colectomy in patients with completely obstructive left - sided cancer . during a one - stage procedure , intraoperative colonic irrigation was completed using a y - shaped irrigation catheter consisting of a working port and a drainage tube . \n after irrigation was completed , an intraoperative colonoscope was inserted through this device , and the bowel proximal to the obstructing tumor was endoscopically examined for presence of synchronous neoplastic lesions . in this study , \n synchronous adenomatous polyps were detected in 26.8% of patients receiving intraoperative colonic irrigation and colonoscopy , 4.0% of which were determined to be carcinoma on pathologic evaluation . in this study , they reported mean operative time to be 28 minutes longer when on - table colonic irrigation and intraoperative colonoscopy were included in the procedure ( 271 minutes versus 243 minutes ) . with intraoperative detection of these synchronous lesions , \n surgical intervention could be performed during the same procedure , thus deferring the need for a second laparotomy . \n found their protocol to be safe with no mortality , low morbidity , and without significant differences in complication rates . \n several studies have also shown that operative planning may need to be altered if a synchronous lesion is present . \n conducted a prospective study in an effort to evaluate the need to augment the extent of surgical resection due to detection of a synchronous colorectal lesion . in this series , \n synchronous lesions were present in 45.6% of patients , with 24% of these lesions influencing the degree of surgical resection . \n overall , in 11% of cases , the presence of a synchronous lesion influenced operative management and dictated a more extensive operation than would have been performed with only treating the primary lesion . \n kim and park advocated the use of intraoperative colonoscopy to evaluate for synchronous lesions , reporting that 37.2% of on - table endoscopies performed in their study detected synchronous lesions , leading to additional surgical procedures completed at the time of initial operation in 13.7% of cases in order to remove these lesions . in this notion \n , the overall goal of operative management should therefore be to perform a procedure with intent to cure , assess for synchronous pathology , and address all pathology present at time of the initial surgical intervention to avoid additional operations . \n while surgical resection is the primary treatment for localized colorectal cancer , development of local , regional , and/or distant recurrence has been reported to occur in 30% to 50% of patients . \n greater than 90% of these recurrences have occurred within a 5-year interval following surgery . when detected \n early , surgical intervention is the treatment of choice for resectable recurrences and for new primary tumors . \n recurrence after resection of colorectal cancer is most common in the liver ( 33% ) followed by lung ( 22% ) , local recurrence ( 15% in colon and 35% in rectum ) , and regional lymph nodes ( 14% ) . \n less common but still noteworthy is the development of second primary or metachronous lesion in 3% of patients . though the importance of postoperative surveillance for detection of these lesions is widely acknowledged , no consensus has been established regarding timing and modalities for follow - up protocols in colorectal cancer patients . in 2009 , \n scheer and auer performed a literature review exploring current guidelines from various institutions , including the american society of clinical oncology , the national comprehensive cancer network , and the european society of medical oncology , among others . in their meta - analysis \n , it became apparent that intensive surveillance after curative resection of colorectal cancer improved overall survival , allowing early reoperation to address asymptomatic recurrences . \n however , different approaches were noted in follow - up protocols , with large variation in frequency of postoperative office visits , serum cea level monitoring , colonoscopy , and radiologic studies of the chest and abdomen . \n office visits are important for surveillance monitoring , allowing the physician to discuss results of investigations , reinforce behavioral modifications , and provide counseling regarding disease process . \n advocates of cea monitoring will trend levels postoperatively , as levels will become elevated in approximately 75% of patients with colorectal recurrence . depending on the threshold level for abnormal value , sensitivity and specificity for detecting recurrence \n most studies have agreed that cea monitoring is most sensitive for hepatic and retroperitoneal lesions and least sensitive for local recurrence and pulmonary lesions . \n therefore , cea is used most commonly as a trended value , triggering further work - up if levels are noted to rise . \n after resection of colorectal cancer , follow - up colonoscopy is recommended to screen recurrence at the anastomosis and to assess metachronous lesions . \n mulder et al . reported a significantly higher incidence of metachronous colorectal cancer in patients who had a prior resection for colorectal cancer when compared to an age- and sex - matched population . \n of note , these patients had a 1.4 incidence ratio of metachronous lesions within a 3-year interval after initial colorectal cancer diagnosis and resection . \n the presence of a synchronous lesion at time of initial colorectal cancer resection was noted to be the only significant risk factor for development of metachronous lesion , yielding a relative risk of 13.9 ( 95% ci 4.741.0 ) . \n therefore , short - term interval ( ideally less than 3 years ) surveillance should be recommended for follow - up colonoscopy , possibly sooner for those patients diagnosed with synchronous tumors . \n couch and coworkers conducted a retrospective study investigating postoperative colonoscopic surveillance , noting that complete preoperative screening did not translate to a lower incidence of neoplasia detected on first postoperative colonoscopy . \n additionally , new neoplastic lesions and recurrences amenable to resection were detected within two years from time of operation . \n advocate a suggested time interval of no more than two years between initial surgery for colorectal cancer and first postoperative surveillance colonoscopy . in their meta - analysis , scheer and \n auer stated that most randomized trials evaluating surveillance for colorectal cancer recurrence after resection of primary lesion reported a median observation period of 5 years or less . aside from colonoscopy , several imaging modalities are available to assess the colon , as well as extracolonic anatomy . \n chest x - ray or thoracic ct can be used if pulmonary lesions are suspected . abdominal ct and/or ultrasound \n american society of clinical oncology current guidelines recommend yearly ct scan of the abdomen for the first 3 years following surgery . \n a newer imaging modality , ct colonography , may play a role in surveillance after curative resection of colorectal cancer due to its ability to assess for both colonic and extracolonic lesions . \n kim and colleagues reported an 81.8% per - patient and 80.8% per - lesion sensitivity for ct colonography when detecting advanced neoplasia , with similar sensitivities of 80% and 78.5% , respectively , for evaluation all adenomatous lesions . \n ct colonography was noted to have a specificity of 93.1% , with negative predictive values of 100% for adenocarcinoma , 99.1% for advanced neoplasia , and 97% for all adenomatous lesions . \n more recently , 18fdg - pet has been investigated as a potential diagnostic imaging modality , specifically in patients with an elevated cea and normal colonoscopy . \n while this option had a high diagnostic yield for detection of lesions , it is not currently considered a cost - effective method for routine postoperative surveillance . \n as discussed earlier , the need for postoperative colonoscopic surveillance after resection is paramount due to the elevated incidence of synchronous and metachronous lesions in those patients with a history of colorectal malignancy . \n typically , in patients who have undergone preoperative colonoscopic evaluation , colonoscopy is performed within one year after colorectal resection . in the patient without a recent preoperative evaluation ( within 5 years ) , who presents with an acute colonic obstruction that necessitates emergency surgical intervention without intraoperative colonic lavage and colonoscopy \n , it would be recommended to survey that patient in less than one year after resection , preferably 36 months postoperatively . in this specific scenario , \n if a biopsy - proven malignancy or sessile polyp not amenable to endoscopic resection is found on postoperative colonoscopy , the patient will need to be subjected to two abdominal operations for colorectal resection within a 6-month period . given the age , functional status , and burden of comorbidity of each patient \n , this predicament can be a challenging one , but one that is potentially avoidable . \n at the current state of technology , multiple modalities exist to visualize the colon in the asymptomatic patient , as well as during operative resection and the perioperative period . despite this fact , \n a large portion of the population remains unscreened for colorectal lesions , and high - grade obstruction or other emergent conditions from colonic carcinoma can occur without previous detection . on - table colonic lavage and colonoscopy during the time of resection \n is a technique that has been utilized for many years but recently is becoming more refined . \n the goal of recent research is to develop a safe , efficient , high - yield procedure that adds little to overall operating room time and postoperative morbidity while give the surgeon the necessary information to make a proper oncologic decision about the extent of resection to provide our patients the best chances for successful outcome . \n it is a technique and a technology that requires further use and study but with continued evolution and improvement looks to be a viable option in our intraoperative armamentarium .", "role": "user" }, { "content": "it is not always possible to evaluate patients that present acutely with carcinoma of the colon and rectum for synchronous lesions . \n patients that require emergent surgery necessitate urgent and efficient operation . \n patients with lower gastrointestinal bleeding , perforation , or obstruction represent a challenging subset of patients with colorectal cancer . an organized approach to these patients in the effort not to overlook \n a synchronous carcinoma is important . \n the present paper provides an evidenced - based approach to this special situation .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: condylar injuries are often a matter of discussion and controversy among maxillofacial topics ( 1 ) . \n these traumas constitute 1116% of all facial fractures ( 25 ) and 3040% of mandibular fractures ( 37 ) . \n this area is superiorly confined to the line passing through the sigmoid notch and anteriorly to the line obliquely connecting the sigmoid notch to the masseter tuberosity ( 8) . this area has a great clinical value due to important components such as the facial nerve and temporomandibular joint ( tmj ) . \n both of these are prone to functional disability due to either the fracture itself , or the surgical intervention . \n different stages of dislocation , displacement , comminution and fracture line are dependent on the force magnitude , point of application and transmission , and the patient s mandibular position and occlusion , during the trauma ( 8) . \n one of the reasons that cause condylar fractures to be of paramount importance in maxillofacial surgery , is that many ensuing complications are connected to a condylar fracture , whether it is treated or not . even though early clinical results are seen to be good , complications such as pain , restriction of movement , muscle spasms , mandibular deviation , malocclusion , pathologic alterations of the tmj , osteonecrosis , facial asymmetry , and ankylosis may be evident later ( 913 ) . \n other complications related to the condylar fracture are ranged from tympanic bone fracture , fracture of mandibular fossa of temporal bone with or without dislocation of the condylar segment into the middle cranial fossa , injury to the cranial nerves , vascular damage and bleeding , growth inhibition and arteriovenous fistula ( 14 , 15 ) . \n as there are controversies around management of condylar fractures , different treatment modalities are suggested in literature ( 16 ) . \n the aim of this study was to review the different treatment strategies of these fractures in adults : closed reduction with maxillomandibular fixation ( mmf ) open reduction with internal fixation ( orif ) endoscopic - assisted reduction with internal fixation ( erif ) \n closed reduction was historically considered the standard treatment for subcondylar fractures ( 17 ) . the assumption that a closed reduction will result in fewer complications , though the esthetics and functionality will be just the same as an orif treatment , induced this method as the common procedure . \n the noninvasive nature of this method will consequently result in a very little to no facial nerve damage or scaring ( 18 ) . despite this knowledge , and much debating , \n authors are still unable to agree on an absolute treatment plan for management of condylar fractures . \n some publications , state no marked difference between the results of open or closed treatments ( 16 , 1921 ) . closed treatment however , is considered undesirable in some other studies ( 17 , 22 , 23 ) . \n unfortunately no clinical trial large enough is available on this matter to answer these questions . \n it is mentioned in different articles that duration of mmf has a range of two to six weeks ( 8) . the shortest time for mmf is chosen by surgeons , to avoid possible ankylosis of the temporomandibular joint ( 24 ) . \n the precise reason for the ankylosis is yet to be explained but in theory , the bleeding in the joint , hematoma and the following fibrosis and bone generation due to a trauma , could cause the undesired ankylosis ( 25 , 26 ) . \n it would thus be wise to assume that tmj ankylosis is a result of a direct trauma to the joint capsule or condylar head therefore the risk of ankylosis is minor in subcondylar fractures : hence , longer periods of mmf should be considered for a better union in fractures distant from the joint capsule ( 8) . upon conclusion of the gathered information , \n the authors suggest 46 weeks of mmf in the subcondylar fractures in which the tmj capsule is not involved . \n this protocol is also applicable to bilateral fractures in which the fractured segments are not displaced . \n however in bilateral fractures , segments are more prevalently prone to displacement , thus an orif approach is recommended . since the introduction of internal fixation appliances , open reduction of condylar fractures is becoming more popular . \n few studies emphasize on the indications of this treatment procedure ( 27 ) , but as a general rule , the need for anatomic reduction in complex fractures is one reason to employ the open reduction and internal fixation approach . nowadays , many of the upcoming articles are discussing the better results of the surgical treatment of the condylar fractures , for better occlusion , bone morphology and tmj function ( 20 , 2831 ) . \n others however argue that a surgical access to the mandibular condyle , would increase the risk of trauma to the facial nerve and leave unpleasant scaring ( 27 , 3234 ) . \n hypertrophic scaring in 7.5% of cases are said to be quite tangible and significant ( 27 ) . \n however the superior anatomic reduction and better functional results , induce orif as the treatment of choice in patients with complex conditions \n . another advantage would be the immediate capability of the patient to move the injured area , reducing the mmf required time and thus lowering the risk of ankylosis ( 8) . in cases of major displacement and loss of functional occlusion , when a closed reduction is not possible , orif is indicated and the result would overcome the presumptive complications ( 29 ) . to access the fractured area , different approaches are available and surgeons would decide upon a suitable approach , based on experience and personal taste . \n some of these approaches are periauricular , submandibular , retromandibular antroparotid and retromandibular transparotid . the retromandibular antroparotid with a two centimeter incision under the ear lobule is utilized in this center \n . post - surgical scaring is less evident in this approach and no entry to the parotid gland occurs , preventing complications such as salivary fistula ( 35 ) . \n erif is said to be embracing the benefits of both the closed and the open techniques . \n the resulting scar is very minor , no facial nerve damage is expected and a functional occlusion is ensured with anatomic reduction . an excellent visibility is achieved through the endoscope . \n this method is also swifter in comparison to other extraoral techniques ( 36 ) . according to literature \n , it is said that by using the endoscope , a variety of methods are possible . \n ( 3739 ) , although an intraoral approach is most commonly used by the surgeons utilizing an endoscope ( 40 , 41 ) . \n this approach is also used in this center , and a small incision on the skin suffices the entry of the trocar . \n the majority of the subcondylar fractures can be managed by this method , however an orif procedure would be the treatment of choice in cases of significant displacements . despite the preponderance and imperfections of the mentioned methods , the goal is achieving the premorbid and normal function of the patient with minimal damage . pointing out \n the best procedure is certainly not easy assay and many factors influence the matter , so the aim of this review was to summarize the available methods , and their advantages and disadvantages , in order to help make the decision easier . \n due to different techniques and materials and more importantly the position and kind of the subcondylar fracture , choosing the right treatment protocol is a challenge every time . despite \n the many clinical researches available on management of condylar fractures , it still remains a matter of debate ( 36 ) . \n the modern day surgical techniques such as the endoscope - assisted technology , reducing the morbidity rate , again raises the question of choosing either the open or closed treatment technique ( 42 ) . \n the closed technique is continues to be the most commonly used for medium to large displacements ( 43 ) . \n retrospective and prospective studies in literature comparing open and closed treatments , report that the surgical procedure has no superiority to the closed technique as in function , range of motion , occlusion , contour , sensory or motional function retrieval . \n the most common complaint of patients after six months was chronic pain after the closed reduction and a noticeable scar after the surgical treatment ( 22 , 44 , 45 ) . \n m. gupta et al . concluded that after both treatment protocols , the patients presented similar occlusion , retention of the reduced part s fixation , trismus index , mandibular movements and chewing sufficiency . \n the notable differences were the patient s discomfort during mandibular movements , chewing , swelling , neurogenic complications and parotid fistula on occasion after surgery ( 1 ) . \n however several radiologic studies demonstrate a better anatomical reduction after the orif treatment ( 19 , 4549 ) . the study of conservative treatment of 466 condylar fractures carried out by zachariades et al . , concluded that the closed technique is the treatment of choice for most of the subcondylar fractures , unless when there is nt a recoverable occlusion , severe displacement , loss of height of ramus or an edentate patient where the open reduction is indicated ( 50 ) . \n ( 48 ) mentioned that the two indicators for surgery are the condylar angle of 1045 and the loss of more than two millimeters of the ramus height . \n these studies also concluded that despite the surgical technique , anatomic reduction , mouth opening , lateral and forward movements of the mandible and the lower rate of pain and discomfort of the patients were considered beneficial in the orif group . \n the study by lee jw et al . , and al - moraissi et al . \n . however there are some studies about complications of open surgeries which indicate that permanent deformity and dysfunction after surgery is very rare ( 27 ) , also the mandibular and condylar mobility is better ( 29 , 30 ) and opening incisor pathway is much more normal ( 31 ) and also due to not decreasing the ramus height , less asymmetry is visible ( 28 ) . \n kokmueller et al . ( 36 ) in 2012 carried out a study to assimilate the endoscope - assisted transoral treatment and closed reduction of subcondylar fractures . both closed reduction and \n patients undergoing surgery had more complaints in the short run , but they were presented with fewer symptoms in long term follow ups . \n patients with endoscopic treatment had better occlusion parameters both in early and late follow up sessions . \n it is mentioned several times in literature that the surgical complications of patients are far less , when treated transorally by the endoscopic - assisted method ( 19 , 23 , 53 , 54 ) . \n authors using the endoscopic method to treat subcondylar fractures believe that a single intraoral incision would be sufficient to reduce a laterally displaced condyle . \n even for lateral dislocations of the subcondylar area , an intraoral approach is suggested , due to the unperceptive scar , no matter how challenging it may be ( 40 , 41 ) . \n it is wise to emphasize on the short duration of this procedure also ( 41 ) , butfor the surgeons who are newly utilizing this method , there is probably much more time to perform the anatomic reduction and do the surgery than the conventional extraoral methods . \n but according to the learning curve that is considered for endoscopic treatment , this time will soon be decreased ( 55 ) . for more complicated malocclusions and bigger dislocations \n closed reduction was historically considered the standard treatment for subcondylar fractures ( 17 ) . the assumption that a closed reduction will result in fewer complications , though the esthetics and functionality will be just the same as an orif treatment , induced this method as the common procedure . \n the noninvasive nature of this method will consequently result in a very little to no facial nerve damage or scaring ( 18 ) . despite this knowledge , and much debating , \n authors are still unable to agree on an absolute treatment plan for management of condylar fractures . \n some publications , state no marked difference between the results of open or closed treatments ( 16 , 1921 ) . closed treatment however , is considered undesirable in some other studies ( 17 , 22 , 23 ) . \n unfortunately no clinical trial large enough is available on this matter to answer these questions . \n it is mentioned in different articles that duration of mmf has a range of two to six weeks ( 8) . the shortest time for mmf is chosen by surgeons , to avoid possible ankylosis of the temporomandibular joint ( 24 ) . \n the precise reason for the ankylosis is yet to be explained but in theory , the bleeding in the joint , hematoma and the following fibrosis and bone generation due to a trauma , could cause the undesired ankylosis ( 25 , 26 ) . \n it would thus be wise to assume that tmj ankylosis is a result of a direct trauma to the joint capsule or condylar head therefore the risk of ankylosis is minor in subcondylar fractures : hence , longer periods of mmf should be considered for a better union in fractures distant from the joint capsule ( 8) . upon conclusion of the gathered information , \n the authors suggest 46 weeks of mmf in the subcondylar fractures in which the tmj capsule is not involved . \n this protocol is also applicable to bilateral fractures in which the fractured segments are not displaced . \n however in bilateral fractures , segments are more prevalently prone to displacement , thus an orif approach is recommended . \n since the introduction of internal fixation appliances , open reduction of condylar fractures is becoming more popular . \n few studies emphasize on the indications of this treatment procedure ( 27 ) , but as a general rule , the need for anatomic reduction in complex fractures is one reason to employ the open reduction and internal fixation approach . nowadays , many of the upcoming articles are discussing the better results of the surgical treatment of the condylar fractures , for better occlusion , bone morphology and tmj function ( 20 , 2831 ) . \n others however argue that a surgical access to the mandibular condyle , would increase the risk of trauma to the facial nerve and leave unpleasant scaring ( 27 , 3234 ) . \n hypertrophic scaring in 7.5% of cases are said to be quite tangible and significant ( 27 ) . \n however the superior anatomic reduction and better functional results , induce orif as the treatment of choice in patients with complex conditions \n . another advantage would be the immediate capability of the patient to move the injured area , reducing the mmf required time and thus lowering the risk of ankylosis ( 8) . in cases of major displacement and loss of functional occlusion , when a closed reduction is not possible , orif is indicated and the result would overcome the presumptive complications ( 29 ) . to access the fractured area , different approaches are available and surgeons would decide upon a suitable approach , based on experience and personal taste . \n some of these approaches are periauricular , submandibular , retromandibular antroparotid and retromandibular transparotid . the retromandibular antroparotid with a two centimeter incision under the ear lobule is utilized in this center . \n post - surgical scaring is less evident in this approach and no entry to the parotid gland occurs , preventing complications such as salivary fistula ( 35 ) . \n erif is said to be embracing the benefits of both the closed and the open techniques . \n the resulting scar is very minor , no facial nerve damage is expected and a functional occlusion is ensured with anatomic reduction . \n this method is also swifter in comparison to other extraoral techniques ( 36 ) . according to literature \n , it is said that by using the endoscope , a variety of methods are possible . \n ( 3739 ) , although an intraoral approach is most commonly used by the surgeons utilizing an endoscope ( 40 , 41 ) . \n this approach is also used in this center , and a small incision on the skin suffices the entry of the trocar . \n the majority of the subcondylar fractures can be managed by this method , however an orif procedure would be the treatment of choice in cases of significant displacements . despite the preponderance and imperfections of the mentioned methods , the goal is achieving the premorbid and normal function of the patient with minimal damage . pointing out the best procedure is certainly not easy assay and many factors influence the matter , so the aim of this review was to summarize the available methods , and their advantages and disadvantages , in order to help make the decision easier . \n due to different techniques and materials and more importantly the position and kind of the subcondylar fracture , choosing the right treatment protocol is a challenge every time . despite \n the many clinical researches available on management of condylar fractures , it still remains a matter of debate ( 36 ) . \n the modern day surgical techniques such as the endoscope - assisted technology , reducing the morbidity rate , again raises the question of choosing either the open or closed treatment technique ( 42 ) . \n the closed technique is continues to be the most commonly used for medium to large displacements ( 43 ) . \n retrospective and prospective studies in literature comparing open and closed treatments , report that the surgical procedure has no superiority to the closed technique as in function , range of motion , occlusion , contour , sensory or motional function retrieval . \n the most common complaint of patients after six months was chronic pain after the closed reduction and a noticeable scar after the surgical treatment ( 22 , 44 , 45 ) . \n m. gupta et al . concluded that after both treatment protocols , the patients presented similar occlusion , retention of the reduced part s fixation , trismus index , mandibular movements and chewing sufficiency . \n the notable differences were the patient s discomfort during mandibular movements , chewing , swelling , neurogenic complications and parotid fistula on occasion after surgery ( 1 ) . \n however several radiologic studies demonstrate a better anatomical reduction after the orif treatment ( 19 , 4549 ) . the study of conservative treatment of 466 condylar fractures carried out by zachariades et al . , concluded that the closed technique is the treatment of choice for most of the subcondylar fractures , unless when there is nt a recoverable occlusion , severe displacement , loss of height of ramus or an edentate patient where the open reduction is indicated ( 50 ) . \n ( 48 ) mentioned that the two indicators for surgery are the condylar angle of 1045 and the loss of more than two millimeters of the ramus height . \n these studies also concluded that despite the surgical technique , anatomic reduction , mouth opening , lateral and forward movements of the mandible and the lower rate of pain and discomfort of the patients were considered beneficial in the orif group . \n the study by lee jw et al . , and al - moraissi et al . \n however there are some studies about complications of open surgeries which indicate that permanent deformity and dysfunction after surgery is very rare ( 27 ) , also the mandibular and condylar mobility is better ( 29 , 30 ) and opening incisor pathway is much more normal ( 31 ) and also due to not decreasing the ramus height , less asymmetry is visible ( 28 ) . \n kokmueller et al . ( 36 ) in 2012 carried out a study to assimilate the endoscope - assisted transoral treatment and closed reduction of subcondylar fractures . both closed reduction and \n patients undergoing surgery had more complaints in the short run , but they were presented with fewer symptoms in long term follow ups . \n patients with endoscopic treatment had better occlusion parameters both in early and late follow up sessions . \n it is mentioned several times in literature that the surgical complications of patients are far less , when treated transorally by the endoscopic - assisted method ( 19 , 23 , 53 , 54 ) . \n authors using the endoscopic method to treat subcondylar fractures believe that a single intraoral incision would be sufficient to reduce a laterally displaced condyle . even for lateral dislocations of the subcondylar area , \n an intraoral approach is suggested , due to the unperceptive scar , no matter how challenging it may be ( 40 , 41 ) . \n it is wise to emphasize on the short duration of this procedure also ( 41 ) , butfor the surgeons who are newly utilizing this method , there is probably much more time to perform the anatomic reduction and do the surgery than the conventional extraoral methods . but according to the learning curve that is considered for endoscopic treatment , this time will soon be decreased ( 55 ) . for more complicated malocclusions and bigger dislocations \n overall it is safe to declare that endoscopic surgery is certainly a good replacement for approaches through the skin , for subcondylar fractures . avoiding the complications related to the traditional open technique such as the facial nerve damage , scarring and those related to the close technique such as the lengthened mmf , non - anatomical reduction and difficulties associated with mandibular movements , have deemed the endoscopic method a favorite among surgeons . with enhanced experience of the surgeons toward the endoscopic technique the controversies over the right treatment choice would slowly subside .", "role": "user" }, { "content": "condylar injuries are often subjected to discussion and controversy in maxillofacial surgery as they constitute many of the facial fractures . \n the condylar area has a great clinical value due to its important components . \n vital components in this area are susceptible to functional disability due to either the fracture itself or the subsequent surgical intervention . \n each of the strategies for the management of these fractures has its advantages and disadvantages . \n as there are controversies around management of condylar fractures , different treatment modalities are suggested in literature , so this paper reviews different treatment strategies of the following types of fractures in adults : 1-closed reduction with maxillomandibular fixation , 2-open reduction with internal fixation , 3-endoscopic - assisted reduction with internal fixation . in conclusion , \n we declare that the endoscopic surgery is certainly a good replacement for approaches through the skin , for subcondylar fractures , but still more randomized clinical trials are needed to be carried out on this issue .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: bread wheat ( triticum aestivum l. ) is a traditional crop in spain , which is the fifth wheat producer of the european union , devoting 14 million hectares to the species in 2010 ( eurostat 2011 ) . \n however , during the 20th century investments in wheat breeding in spain were irregular and genotype replacement relied mostly on the introduction of foreign germplasm . \n wheat breeding activities in spain started in the 1920s , mainly with the classification of local landraces and bulk selection within them ( royo & briceo - flix 2011 ) . \n however , the early breeding programmes were interrupted by the spanish civil war ( 193639 ) , which severely damaged the incipient efforts of cereal researchers to produce improved wheat genotypes . during the two decades after the war \n , breeding activities were devoted to further improving traditional wheat genotypes through selection and hybridization , and foreign improved cultivars , mainly from italy and france , were successfully introduced . \n some of the cultivars released during the 1950s ( such as the italian cultivars mara and impeto and the french cultivar estrella ) carried the rht8c dwarfing gene and probably the photoperiod - insensitive gene ppd1 ( worland et al . \n 1998 ) , which has been reported to affect plant height , cycle length and crop productivity ( borojevic & borojevic 2005 ) . \n the advent of cimmyt semi - dwarf germplasm in the late 1960s had a great impact on spanish agriculture . \n it was rapidly adopted in southern spain , but the french germplasm continued to be grown in northern spain ( royo & briceo - flix 2011 ) . the major dwarfing genes rht1b and rht1d were introduced with cimmyt germplasm , and have been reported to result in yield increases of c. 15% in near - optimal conditions ( worland & snape 2001 ) . a study conducted in durum wheat \n , whose history is parallel to that of bread wheat , indicated that new sources of alleles were used in the crosses that led to the cultivars released in spain since 1990 ( martos et al . \n understanding the crucial characteristics of the crop associated with the genetic gains achieved through breeding is an essential step for improving the current knowledge of yield - limiting factors and the design of future breeding strategies . \n historical series of genotypes have been deployed and used in several countries to assess the genetic gains achieved during a period of time through selection and breeding ( austin et al . \n . however , information about gains in yield and related traits in spain is scarce . \n the number of grains / m and , more precisely , the number of grains / spike , have been reported to be the main traits contributing to yield rises under a wide range of environmental conditions ( perry & d'antuono 1989 ; donmez et al . \n 2007 ) , whereas changes in grain weight have proven to be null ( austin et al . \n 1989 ; brancourt - hulmel et al . \n 2003 ) or even negative ( siddique et al . \n 1989 ; royo et al . \n increases in the number of spikes / m of bread and durum wheat during the 20th century have also been detected , mainly under mediterranean conditions ( perry & d'antuono 1989 ; canevara et al . \n 2007 ) , and they have been attributed to increases in the number of spikes / plant ( royo et al . \n 2007 ) . from the point of view of biomass production and allocation , past genetic gains in bread wheat yield have been associated widely with increases in harvest index ( hi ) and decreases in plant height ( canevara et al . \n the introduction of dwarfing genes reduced the size of the vegetative organs , thus enhancing the availability of assimilates to the reproductive organs of the plant , expressed as a higher number of grains / spike ( lvaro et al . \n ( 1980 ) proposed 062 as a theoretical upper limit for hi , but hi rises beyond 050 are difficult to attain ( fischer & edmeades 2010 ) . \n however , in some countries current levels are close to this upper limit and further yield improvements must therefore be based on increasing biomass while maintaining hi ( pfeiffer et al . \n the uneven history of bread wheat introduction in spain during the last century may have led to the boosting of different plant attributes in different phases , as has been reported in other countries where the paths followed to achieve yield gains during the last century differed across periods ( austin et al . \n 1994 ) . through a study of the main periods of release of bread wheat genotypes in spain , \n the objective of the present work was to quantify yield breeding progress during the 20th century and to identify the key traits associated with it . \n a collection of 28 bread wheat genotypes representative of those most widely grown in spain during the 20th century was assembled . \n the genotypes were grouped into four breeding periods : ( i ) spanish landraces grown before 1940 ; ( ii ) initial cultivars derived from crosses , including those improved and grown from the mid-1940s until the green revolution ( termed old - bred in a previously published paper , sanchez - garcia et al . \n 2012 ) ; ( iii ) intermediate cultivars introduced in spain from 1970 and mostly derived from semi - dwarf germplasm of cimmyt and french cultivars ; and ( iv ) modern cultivars , released during the last decade of the 20th century ( table 1 ) . \n table 1.description of the genotypes used in the study and the breeding periods to which they belonggenotypepedigreeoriginyear of release in spainexperiments*local landracesaragon 03selection of the landrace catalan de monte.spain<1940multi-location ( ml ) and lleida ( ll)barbillalandracespain<1940ml and llcandeallandracespain<1940ml and llchamorrolandracespain<1940mlnegrillolandracespain<1940mlinitialmaraautonomia / aquilaitaly1947llimpetofrassineto-405/villa - gloriitaly1950ml and llestrellamon - desir / ardito//mouton - a - epi - rouge / k-3/3/mouton - epi - rougefrance1952ml and llpan 247l-4/mentanaspain1955ml and llintermediatesiete cerrospenjamo-62/gabo-55cimmyt1969mlyecoraciano-67(sib)//sonora-64/klein - rendidor/3/siete - cerros-66cimmyt1972llcajemeciano-67(sib)//sonora-64/klein - rendidor/3/siete - cerros-66cimmyt1972ml and llanzalerma - rojo-64//norin-10/brevor/3/3*andes - enanocimmyt / usa1974ml and llastralfortunato / yga/3/florence / aurore//g-4france1974mlmariuscadet//thatcher / vilmorin-27/3/ariana / funduleafrance1980ml and llrinconadaunknownspain1981ml and llalcalunknownspain1984mlcartayakuz / buho//kal / bbcimmyt1984mlsoissonsiena / hn-35france1990ml and llmodernbancalunknownspain1991mlpinznunknownspain1992mlgazulunknownspain1992ml and lletechovpm / talent//fidel/3/declicfrance1995mlbabuiunknownspain1997mlberdununknownfrance1998mlisengrainapollo / soissonsfrance1998ml and llcraklinunknownfrance1999mlcalifa surunknownspain2001ml and ll*ml = genotypes included in eight multi - location experiments in 2006 and 2007 ; ll = genotypes included in experiments conducted in lleida in 2008 and 2009 . \n description of the genotypes used in the study and the breeding periods to which they belong ml = genotypes included in eight multi - location experiments in 2006 and 2007 ; ll = genotypes included in experiments conducted in lleida in 2008 and 2009 . \n twenty - six genotypes of the collection were tested in multi - location experiments harvested in 2006 and 2007 , but sown in november or december of the previous year ( table 1 ) . \n experiments were conducted at four sites representative of the main bread wheat growing areas in spain : burgos , in the north , with cold winters and mild summers ; lleida in the north - east , where terminal stress was moderate and the plots were irrigated ; albacete in the south - east , with cold winters and very hot and dry summers ; and cordoba in the south - west , with warm winters and very hot summers ( table 2 ) . \n table 2.experimental sites , description of the environmental conditions and management practicessitemulti - location experiments ( ml)lleida experiments ( ll)burgos ( nw)lleida ( ne)albacete ( se)cordoba ( sw)lleida ( ne)coordinates425n , 339w4140n , 020e3903n , 0205w3751n , 0448w4140n , 020ealtitude ( m asl)86420070093200soil texturesandy loamfine loamclay loamclay loamfine loamsoil classification*gleyic cambisolcalcic cambisolcalcic cambisoleutric fluvisolcalcic cambisolharvest years200620072006200720062007200620072008200820092009water regimerainfedrainfedirrigatedirrigatedrainfedrainfedrainfedrainfedrainfedirrigatedrainfedirrigatedexperiment codeb06b07l06l07a06a07c06c07l08rl08il09rl09ienvironmental conditions from : sowing to headingsowing to anthesisthermal time ( gdd ; c)940986117013999651183134912391342129911441014et0 ( mm)282263347247303344259228273259243239tmin ( c)050421351010624831302327tmax ( c)121119128138138141170166141142134129water input ( rainfall + irrigation , mm)212172345282106165274174102252183234environmental conditions from : heading to maturityanthesis to maturityet0 ( mm)240240262345476454394437170155113160tmin ( c)9277125133124117146132115113118127tmax ( c)265225281274282294301273231230270273water input ( rainfall + irrigation , mm)67145801948421331621331338654weed controlbromoxinil + ioxinil + mcpplinuron + trifluralin and bromoxinil + ioxinil + mcppmcpp and mcpaclodinafop - propargyl and tribenuron - methyllinuron + trifluralin and bromoxinil + ioxinil + mcppn applied ( kg n / ha)seed bed2424800121245450000top dressing8585173784040129129100903040p applied ( kg p2o5/ha)4545150110363611511512812068113k applied ( kg k2o / ha)4545150183121200213200113188sowing datedec 12dec 1nov 19nov 15dec 20dec 6dec 22dec 12nov 20nov 19nov 24nov \n 27harvest datejul 4jul 23jun 23jul 12jul 17jul 27jun 30jul 12jul 10jul 14jul 13jul 1mean yield ( kg / ha)290953624472649214732771339748454207541654347215gdd = growing degree days.tmin and tmax are average minimum and maximum daily temperatures , respectively.et0 is the reference evapotranspiration computed by the penman \n fao methodology ( allen et al . 1998).*soil - type classification according to fao ( 1998 ) . \n experimental sites , description of the environmental conditions and management practices gdd = growing degree days . \n soil - type classification according to fao ( 1998 ) . for a more detailed study , a subset of 16 genotypes representative of the entire collection ( table 1 ) was selected and further evaluated in four field experiments in lleida during two additional crop seasons ( 2008 and 2009 ) under two contrasting water regimes : irrigated and rainfed . \n irrigated experiments received supplementary water inputs of 150 mm in 2008 and 96 mm in 2009 ( see details in table 2 ) . \n sowing density was 350 seeds / m in the multi - location experiments conducted in 2006 and 2007 and 450 seeds / m in the experiments conducted in lleida in 2008 and 2009 . \n all plots were mechanically harvested at ripening and yield was expressed as 120 mg / g of grain moisture . \n ( number of spikes / m , number of grains / spike and mean grain weight ( mgw ) ) and the number of plants / m and stems / plant in lleida experiments , were determined from the whole plants pulled up from a 1 m long sample taken from a central row of each plot before harvesting . in the experiments conducted in lleida in 2008 and 2009 , \n plant height was measured in 10 main tillers from the tillering node to the top of the spike excluding the awns , number of spikelets / spike , number of grains / spikelet and spike length were assessed for main stems and tillers separately . \n spikes and grains were weighed after being oven - dried at 70 c for 48 h. the rest of the sample was used to determine the total number of stems / plant and the proportion of them bearing spikes ( fertile tillering ) and , after drying , aboveground biomass and grain weight , which were used to calculate hi . \n combined anova were performed for all the studied traits . in the model used for the anova of multi - location experiments ( 2006 and 2007 ) , \n the genotype effect was partitioned into breeding period considered as a fixed effect and genotype within breeding period considered as a random effect . \n absolute genetic gain ( agg ) and relative genetic gain ( rgg ) during the 20th century were computed as the slope of the linear regression between the absolute or relative value of the trait and the year of genotype release . \n relative values were computed for each genotype as a percentage with regard to the average value of all genotypes . \n stepwise regression analysis was conducted with the rgg for yield in each experiment as the dependent variable and the climatic features depicted in table 2 as the independent variables . \n regression models ( linear or bilinear when appropriate , y = bx+a(x < />c)+bc ( x < />c ) , with the slope of the second segment equal to zero ) were also fitted to the relationships between variables and those between variables and the year of release of the genotypes , using the tablecurve 2d v2.03 software ( jandel 1991 ) . \n the combined anova of yield data from multi - location experiments revealed a significant effect of most factors ( data not shown ) . \n the mean yield of the experiments ranged from 1473 kg / ha in albacete in 2006 ( a06 ) to 6492 kg / ha in lleida in 2007 ( l07 , table 2 ) . when the whole set of experiments was considered together , \n yield was estimated to increase significantly at a rate of 351 kg / ha / yr , which represents an rgg of 088%/yr ( table 3 , fig . \n 1 ) , whereas the regression equation fitted to the relationship between year of release and yield for intermediate and modern genotypes was not significant ( fig . \n 1 ) . given the statistical significance of the site year genotype ( breeding period ) interaction , genetic gains in yield and yield components were computed for each experiment . \n the results indicated great variability between experiments , with agg for yield ranging from 77 kg / ha / yr in a06 to 665 kg / ha / yr in c07 , and rgg ranging from 037 in b07 to 159%/yr in c06 ( table 3 ) . \n the coefficient of determination of the relationship between the mean yield of each experiment and the estimated agg was r = 037 ( p = 0106 ) , and that of the relationship between the mean yield of each experiment and the rgg was r = 002 ( p = 0738 ) . the average daily minimum temperature from sowing to heading was the only variable entered in the stepwise regression model built to explain variations in yield rgg using climatic features as independent variables ( fig . \n 2 ) . \n table 3.agg and rgg in yield and yield components of bread wheat determined in eight multi - location experiments ( d.f . : \n 25 ) , with standard deviation ( s.d . ) of yield and yield components agg and rgg values across environmentsexperiment code*yieldspikes / mgrains / spikemgwagg ( kg / ha / yr)rgg ( % /yr)rpagg ( spikes / m / yr)rgg ( % /yr)rpagg ( grains / spike / yr)rgg ( % /yr)rpagg ( 10 g / grain / yr)rgg ( % /yr)rpb0625086048<0001227047040<0001022087058<00010112040044<0001b07200370320003nsns00007760110370200020nsns0040320l0642093070<00012140480340002018041037<000100810220230013l0754082070<0001nsns0090151019054038<0001nsns0110089a0680520290004nsns005028401103801500470110037040<0001a071304501500480170460210018nsns0030394c0654159069<00011890380240011027078062<0001nsns0120074c0767137080<0001nsns0080158036108079<0001nsns0050268sd21804350193005500830266001730096overall351088082<00011360300280006020060055<000100460140120079*experiment codes : a , albacete ; b , burgos ; c , cordoba ; l , lleida ; 06 , 2006 ; 07 , 2007 . \n fig . \n 1.aggs in grain yield of 26 bread wheat genotypes released in spain during the 20th century . \n each point was obtained by averaging data of eight environments and three blocks per environment . \n breeding periods are identified with triangles for local landraces , diamonds for initial cultivars , squares for intermediate cultivars and circles for modern cultivars . \n 2.relationship between the rgg for yield and the average daily minimum temperatures from sowing to heading . \n each point corresponds to one of eight experiments identified as follows : squares , burgos ; diamonds , albacete ; triangles , lleida and circles , cordoba . \n agg and rgg in yield and yield components of bread wheat determined in eight multi - location experiments ( d.f . : \n 25 ) , with standard deviation ( s.d . ) of yield and yield components agg and rgg values across environments experiment codes : a , albacete ; b , burgos ; c , cordoba ; l , lleida ; 06 , 2006 ; 07 , 2007 . aggs in grain yield of 26 bread wheat genotypes released in spain during the 20th century . \n each point was obtained by averaging data of eight environments and three blocks per environment . \n breeding periods are identified with triangles for local landraces , diamonds for initial cultivars , squares for intermediate cultivars and circles for modern cultivars . \n relationship between the rgg for yield and the average daily minimum temperatures from sowing to heading . \n each point corresponds to one of eight experiments identified as follows : squares , burgos ; diamonds , albacete ; triangles , lleida and circles , cordoba . \n the number of grains / spike and , to a lesser extent , the number of spikes / m were the yield components most affected by breeding activities during the 20th century , as they increased at a relative rate of 060%/yr and 030%/yr , respectively , while no significant changes were observed for mgw when data were averaged across experiments ( table 3 ) . \n the number of spikes / m showed significant increases in b06 , l06 , and c06 , while no significant changes were detected in the remaining experiments . similarly to the findings for yield , increases in the number of grains / spike were significant in all the experiments , with rgg ranging from 037%/yr in b07 to 108%/yr in c07 . \n changes in mgw were only significant in three experiments , with rgg ranging from 022 to 040%/yr ( table 3 ) . \n although the genetic gains for yield and yield components depended on the experiment , differences between experiments were of a quantitative nature because all significant rates of change were positive for spikes / m and grains / spike , but negative for mgw . \n none of the climatic features considered contributed significantly to explaining genetic changes in yield components . \n the results showed that differences between breeding periods were significant for yield , grains / spike and mgw , but not for number of spikes / m ( data not shown ) . \n the breeding period was the factor that explained the largest proportion of variability for grains / spike ( data not shown ) . \n grain yield during the 20th century increased by 88% , but the largest rises occurred before the end of the intermediate period , since no significant differences in yield and yield components appeared between intermediate and modern cultivars ( table 4 ) . \n the initial cultivars out - yielded the local landraces by 30% , while an additional yield increase of 37% was attained by the intermediate cultivars . \n yield gains were due to increases of 20% in the number of spikes / m and of 65% in the number of grains / spike , but a reduction of 10% in mgw . \n the improved yield of initial cultivars in comparison with the landraces was due to their greater number of grains / spike ( which increased by 58% ) , despite the fact that the number of spikes / m was similar and mgw was 16% lower ( table 4 ) . \n changes in yield components between initial and intermediate cultivars ranged from 45% for grains / spike to 88% for mgw , but none of them were significant . \n table 4.mean values of yield and yield components for 26 bread wheat genotypes released in different periods in spain and grown in eight multi - location experiments . \n the standard error of the differences ( s.e.d . ) of the means of yield and yield components are presentedbreeding periodd.f.yield ( kg / ha)spikes / m \n * grains / spikemgw ( 10 g)landraces42457401223367initial23198 ( 30)432 ( 77)352 ( 58)307 ( 16)intermediate84397 ( 37)459 ( 62)368 ( 45)334 ( 88)modern84628 ( 52)483 ( 52)367 ( 03)330 ( 12)s.e.d.274395245192*the number of spikes / m could not be determined in a07 . \n mean values of yield and yield components for 26 bread wheat genotypes released in different periods in spain and grown in eight multi - location experiments . \n the standard error of the differences ( s.e.d . ) of the means of yield and yield components are presented the number of spikes / m could not be determined in a07 . \n data from four experiments conducted in lleida during 2 years under two water regimes ( l08r , l08i , l09r and l09i ) with a subset of 16 representative genotypes were used to assess the changes caused by breeding on biomass production and allocation . \n the results indicate that yield gains were due to increases in hi , with aboveground biomass being unchanged ( fig . \n hi increased from 024 for the landrace aragon 03 to 048 for the newest cultivar , califa sur . \n however , improvements in hi were not linear across the century , as a plateau in hi was reached after 1961 ( fig . \n although no significant relationship was found between yield or year of release and biomass when the 16 genotypes were considered together , positive and significant relationships between biomass and yield and between biomass and year of genotype release appeared when landraces were removed from the analyses ( broken lines in fig . \n crop biomass increased significantly and linearly from initial cultivars to modern ones , but the great variability within periods for this trait prevented differences between breeding periods from being significant in the anova ( data not shown ) . \n 3.relationship between hi , aboveground biomass , fertile tillering , number of stems / plant at maturity and plant height and both yield and year of release of 16 bread wheat genotypes widely grown in spain during the 20th century . \n breeding periods are identified with triangles for local landraces , diamonds for initial cultivars , squares for intermediate cultivars and circles for modern cultivars . \n relationship between hi , aboveground biomass , fertile tillering , number of stems / plant at maturity and plant height and both yield and year of release of 16 bread wheat genotypes widely grown in spain during the 20th century . \n breeding periods are identified with triangles for local landraces , diamonds for initial cultivars , squares for intermediate cultivars and circles for modern cultivars . \n the average number of plants / m at maturity was 380 in 2008 and 416 in 2009 , without significant differences between genotypes ( data not shown ) . \n the introduction of initial cultivars reduced the number of stems / plant at maturity ( fig . \n 3f ) , but caused a rise in the proportion of stems bearing spikes ( fig . 3 g , h ) , which thereafter remained unchanged . \n initial cultivars were also a first step in the reduction of plant height , which decreased steadily until the end of the century ( fig . \n the increase in fertile tillering was associated with increases in hi of up to 032 ( fig . \n 4a ) , and with a decrease in the total number of stems / plant that started with the introduction of the first improved cultivars ( fig . \n , increases in hi beyond 036 were associated with a reduction in plant height that started with the release of initial cultivars ( fig . \n 4.relationships between ( a ) hi and fertile tillering , ( b ) hi and plant height , and ( c ) fertile tillering and number of stems / m , of 16 bread wheat genotypes widely grown in spain during the 20th century . \n breeding periods are identified with triangles for local landraces , diamonds for initial cultivars , squares for intermediate cultivars and circles for modern cultivars . \n equations and r values are only presented for significant relationships at p<005 ( d.f . : \n 15 ) . relationships between ( a ) hi and fertile tillering , ( b ) hi and plant height , and ( c ) fertile tillering and number of stems / m , of 16 bread wheat genotypes widely grown in spain during the 20th century . each point represents the mean value of one genotype across four experiments . \n breeding periods are identified with triangles for local landraces , diamonds for initial cultivars , squares for intermediate cultivars and circles for modern cultivars . \n equations and r values are only presented for significant relationships at p<005 ( d.f . : \n the contribution of the main and secondary spikes to yield improvement during the 20th century was investigated in the experiments conducted in lleida in 2008 and 2009 with 16 genotypes . \n the weight at ripening of main and secondary spikes increased significantly during the last century , with rgg of 057%/yr and 075%/yr , respectively ( fig . \n these increases were mainly based on a boost in the number of grains / spike , which accounted for rgg of 058%/yr and 070%/yr in main and secondary spikes , respectively ( fig . \n the largest gain in spike weight occurred with the introduction of initial cultivars , which caused an increase of 31% in main spike weight and an increase of 77% in the number of grains / spike . \n the rise in the weight of secondary spikes was evenly distributed during the century . \n 5.relationships between main spike ( m , closed symbols ) and secondary spike ( s , open symbols ) components and year of release of 16 bread wheat genotypes widely grown in spain during the 20th century . \n breeding periods are identified with triangles for local landraces , diamonds for initial cultivars , squares for intermediate cultivars and circles for modern cultivars ( d.f . : \n relationships between main spike ( m , closed symbols ) and secondary spike ( s , open symbols ) components and year of release of 16 bread wheat genotypes widely grown in spain during the 20th century . \n breeding periods are identified with triangles for local landraces , diamonds for initial cultivars , squares for intermediate cultivars and circles for modern cultivars ( d.f . : \n the increases in the number of grains / spike were due to improvements in both the number of spikelets / spike , which increased at a rate of 021%/yr and 029%/yr on main stem and tillers , respectively ( fig . \n 5d ) , and the number of grains / spikelet , which increased at a rate of 042%/yr in both types of spike ( fig . \n 5e ) . genetic gains in the number of grains of main spikes contributed by initial cultivars were related to the improvement of both the number of spikelets / spike , which increased by 29% , and the number of grains / spikelet , which rose significantly , by c. 37% . \n the slopes of the linear models fitted for each spike component were compared between main and secondary stems , but no significant differences between them were found ( data not shown ) . \n the evaluation of historical series of genotypes is a common approach used in several countries to quantify the achievements of past breeding efforts ( austin et al . \n 1989 ; \n the present study examined a set of 26 bread wheat genotypes widely grown in spain in different periods during the 20th century , in order to estimate the genetic gain in yield and to understand its causes through the identification of crop traits associated with yield improvements . \n the agg for yield was 351 kg / ha / yr , which equated to an rgg of 088%/yr . \n these results are comparable or slightly higher than those reported by previous studies conducted with bread wheat in other countries and with durum wheat in spain ( royo et al . \n rgg for yield has been reported to be of 051%/yr in the uk from 1900 to 1985 ( austin et al . \n 1989 ) , 066%/yr in france from 1962 to 1988 ( brancourt - hulmel et al . \n 2003 ) , 068%/yr in the united states ( cox et al . \n 1988 ) and 070%/yr in siberia ( morgounov et al . \n 2010 ) . \n the fact that some of the genotypes most widely grown in spain during the last century were derived from cimmyt germplasm may explain the coincidence between the yield rgg estimated in the present study and the 088%/yr reported by cimmyt for the period 196288 ( sayre 1996 ) . when rgg for yield was calculated for each of the eight environments in which field experiments were conducted , the range of variation was greater than 400% , suggesting a very high dependence of the estimated genetic gains on the environmental conditions . \n previous studies have attributed these dissimilarities to the different yield potential of the environments ( brancourt - hulmel et al . \n 2003 ) , to factors affecting old and modern genotypes in an opposite way , such as the incidence of host - specific pathogens ( cox et al . \n 1988 ) , and to the environmental characteristics that induce genotype environment interactions ( gei ) . \n gei is a main constraint in assessing genetic gains , especially in countries with great environmental variations and large yield fluctuations between years , as is typical in mediterranean environments . \n the bias created by the assessment of yield gains in only one environment was overcome in the present study by conducting experiments under a wide range of environmental conditions involving the main wheat - growing areas of spain . in the present study , \n the differences between the mean yield of the eight environments explained 037 of the variations in yield agg . \n although this value was not significant , probably due to the limited number of environments involved in the present study , it accounted for a considerable proportion of the observed variability , thus showing a tendency of the mean yield of the environment to partially explain differences in the estimated agg in yield . \n in contrast , variations in yield rgg across environments were largely explained by the average minimum daily temperatures from sowing to heading ( 081 of the total rgg variance ) . \n the lowest rgg were recorded in the environments with the lowest minimum temperatures before heading independently of the mean yield of the environment , indicating that the yield differences between landraces and improved cultivars increased as the minimum daily temperatures before heading increased . in view of the results of a previous study conducted with the same set of genotypes ( sanchez - garcia et al . \n 2012 ) , this result may be interpreted in adaptation terms as : ( i ) the specific adaptation of spanish landraces to cold environments , ( ii ) the wider adaptation of modern cultivars in comparison with local landraces and ( iii ) the good performance of modern spring genotypes in warm spanish environments . \n average minimum daily temperature has been reported to be a major factor for wheat yield formation under mediterranean conditions ( siddique et al . \n 1990 ; le houerou 2004 ) and is among the main variables determining gei on yield in spain ( voltas et al . \n 2005 ; sanchez - garcia et al . \n the importance of temperature before heading in adaptation of wheat to mediterranean conditions has also been demonstrated in durum wheat ( royo et al . \n 2010 ) . \n of the three main yield components , the number of grains / spike was the one most related to wheat breeding achievements , as it explained up to 067 of yield increases in spain during the last century , the remaining 033 being explained by the number of spikes / m , while no consistent changes were observed in mgw . \n the relationship between the number of grains / spike and genetic gains in yield is universally accepted in bread and durum wheat , as it has been identified in many studies involving a wide range of environmental conditions ( waddington et al . \n 1986 ; austin et al . \n 1989 ; perry & d'antuono 1989 ; canevara et al . \n / spike were significant in all the environments , while increases in number of spikes / m were not consistent in all of them . \n this could be associated with the larger gei component of the variance recorded for number of spikes / m ( 028 of total variation ) in comparison with yield ( 014 ) and number of grains / spike ( 018 ) . \n the non - significant differences between breeding periods in the number of spikes / m could also be related to the data missing in one of the experiments . \n moreover , increases in the number of spikes / m were only significant in three environments characterized by dry and warm grain - filling periods , which probably caused a reduction in the tiller survival in the landraces , the latest - heading genotypes ( sanchez - garcia et al . \n 2012 ) , which increased the differences in spikes / m between landraces and improved cultivars . \n the great environmental effects on the number of spikes / m found in the present study are consistent with the variability of results of previous studies evaluating genetic changes in spikes / m over time , which range from a steady amount ( waddington et al . \n 1995 ) to large increases ( austin et al . \n 1989 ; donmez et al . \n 2001 ; shearman et al . \n 2005 ) , with slight improvements also being reported from experiments conducted under mediterranean conditions ( perry & d'antuono 1989 ; canevara et al . \n the classification of genotypes into historical breeding periods made it possible to identify two main episodes contributing to genetic improvements in yield in spain during the last century . \n the first one , which caused a yield increase of 30% , was the transition from traditional landraces to the first cultivars released from the earliest spanish breeding programmes and the improved cultivars introduced in spain from foreign countries , mainly italy and france during the 1950s . \n the increase estimated for this period matches with the improvement that occurred in 1956 in the wheat yield of spanish farms ( calderini & slafer 1998 ) . \n the reduction in plant height during this period was accompanied by an increase in the proportion of tillers bearing spikes and a huge rise in hi . \n the yield rise derived from the introduction of initial cultivars was partially caused by an improvement of c. 58% in the number of grains / spike despite a reduction in grain weight of c. 16% in comparison with local landraces . \n three of the four initial cultivars included in the present study ( estrella , originally named etoile - de - choisy , impeto and mara ) carry the rht8c dwarfing gene and probably the photoperiod - insensitive gene ppd1 ( worland et al . \n 1998 ) , which have been reported to reduce plant height , enhance spike fertility , and consequently improve grain yield ( borojevic & borojevic 2005 ) . \n the present results indicate that the great increase in the grain number of initial cultivars was sufficient to compensate for the reduction in grain weight , thus resulting in higher yields . \n these results evidence the success of the work conducted by wheat breeding pioneers , such as nazareno strampelli in italy ( salvi et al . 2012 ) and josep pan in spain . \n a second relevant breeding episode occurred with the introduction , in the late 1960s , of cultivars derived from cimmyt semi - dwarf germplasm , such as the widely grown cvars siete cerros , yecora , cajeme , anza and cartaya , and of european cultivars of winter growth - habit , such as the french cvars marius and soissons . \n this new germplasm further reduced plant height and increased the number of tillers / plant while maintaining their rate of fertility . \n this transition brought a yield increase of 37% in comparison with the initial cultivars , but it was due to slight increases in all three main yield components rather than significant rises in any of them . \n the wide adaptation of foreign cultivars released during this period ( sanchez - garcia et al . \n 2012 ) may partially explain their great success in terms of both productivity and adoption by farmers . \n the evidence of a non - significant yield increase during the period 19702000 suggests that a plateau was reached for yield gains in spain during the last 30 years of the century , agreeing with the results reported for other countries where slight or non - significant yield increases were observed ( sener et al . \n one of the physiological approaches used to understand yield formation consists of considering yield as a function of the biomass per unit area at maturity and the hi or proportion of the aboveground biomass partitioned to grain ( yield = aboveground biomass hi ) . \n the introduction of initial cultivars during the 1950s did not significantly modify the crop biomass , but caused a 60% increase in hi , which rose from 025 to 040 . \n the huge improvement in hi in this first stage could be associated with three factors : ( i ) a reduction in plant height of c. 10% , very probably due to the presence in initial cultivars of rht8c and ppd1 genes , ( ii ) a 28% decrease in the number of stems / plant and ( iii ) an increase from 075 to 093 in the proportion of stems bearing spikes . \n tillering reduction has been described as an advantageous trait under terminal drought stress , as it diminishes soil water use prior to anthesis ( richards 1988 ; duggan et al . \n a decrease in plant height and the number of stems / plant and an increase in the proportion of them bearing spikes probably reduced competition between the growing spikes and the growing stems , enhancing the partitioning of assimilates to the spike ( fischer & stockman 1986 ) and thus resulting in an increase in the weight of main spikes . \n improvements in the number of spikelets / spike and in grain setting were not accompanied by an enlargement of the spikes , so spike compactness augmented . \n increases in spike fertility and decreases in grain weight have been reported to be pleiotropic effects of the rht8c dwarfing gene and the photoperiod insensitive gene ppd1 ( canevara et al . \n the efforts made by local breeders during the early decades of the 20th century to increase grain setting probably improved the adaptation of wheat to mediterranean environments , where abiotic stresses during grain filling are the main cause of floret abortion ( lvaro et al . \n 2008b ) . the introduction of intermediate cultivars caused a slight , not significant , increase in aboveground biomass , but caused a second , smaller rise in hi , which reached values of c. 045 that were not further improved by the cultivars released during the 1990s . \n rises in hi during this phase could be explained by a further reduction of 17% in plant height and increases of c. 55% in the number of tillers / plant , while their rate of fertility was maintained . \n given the limited scope for raising hi beyond 050 ( fischer & edmeades 2010 ) , further yield improvements must rely on biomass production , as was reported for the cultivars released in the uk between 1983 and 1995 ( shearman et al . \n however , this does not seem to be the case in spain , where neither the yield changes nor the 40% increase in biomass during the last decade of the century were significant . \n the great differences induced by the environment in the rates of genetic gain found in the present study may have important implications in the current scenario of global climate change . \n the 18 c rise in average global temperature predicted for 2050 ( meehl et al . \n 2007 ) will force farmers to grow genotypes which are more tolerant , among other factors , to high temperatures ( ceccarelli et al . \n the large effect of the climatic variables , particularly temperature , over the estimated genetic gains of the present study emphasizes the role played by adaptation mechanisms over past breeding achievements , and the need to continue their improvement to face future environmental changes . \n the highest genetic yield gains observed in environments with greater minimum temperatures before heading suggest that the genotypes released recently in spain are better adapted than the old ones to high temperatures . \n therefore , the adaptation trends related to breeding progress in spain during the last century appear to be in the right direction to meet the expected temperature increases .", "role": "user" }, { "content": "summarya collection of 26 wheat genotypes widely grown in spain during the 20th century was evaluated in eight contrasting environments in order to quantify breeding achievements in yield and associated traits . from 1930 to 2000 , yield increased at a rate of 351 kg / ha / yr or 088%/yr , but estimations of relative genetic gain ( rgg ) were environment - dependent . \n rgg estimated for yield were positively associated with the average minimum daily temperatures from sowing to heading in the testing environments ( r2 = 081 ; p < 001 ) . \n the number of grains / spike and the number of spikes / m2 increased at a rate of 060%/yr and 030%/yr , respectively , while grain weight remained unchanged . \n the present study detected two main episodes of yield improvement during the century . \n the first one coincided with the introduction , during the 1950s , of the first improved cultivars derived from intra - specific crosses , which increased the yield of landraces by 30% due to an increase of c. 58% in the number of grains / spike , accompanied by a 16% reduction in grain weight . these initial cultivars ( termed old - bred in a previous study by sanchez - garcia et al . 2012 ) exhibited a higher harvest index ( hi ) , increased from 025 to 040 , but maintained the same aboveground biomass at maturity as the landraces ( despite reducing both plant height and the number of tillers / plant ) due to increases in the proportion of tillers bearing spikes . \n the second yield gain occurred after the introduction , in the early 1970s , of semi - dwarf germplasm from cimmyt ( international maize and wheat improvement centre ) and some french cultivars . \n this new germplasm further reduced plant height , improved hi up to 045 and increased the number of tillers / plant while maintaining their rate of fertility , thus resulting in a yield gain of c. 37% . \n the cultivars released during the last decade of the century did not contribute to significant yield improvements .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: cancer is the third leading cause of death worldwide , and lung cancer is the second most commonly occurring non - cutaneous cancer with the highest mortality rate among all cancers . \n there are 1.38 million new cases of lung cancer every year , along with 1.23 million deaths due to lung cancer.1 based on the histological types of the cells , lung cancer is categorized into two major types , small cell lung cancer ( sclc ) and non - small cell lung cancer ( nsclc ) . \n sclc occurs almost exclusively in heavy smokers , and makes up about 14% of all lung cancers . \n nsclc is a general term for several different types of lung cancer , including squamous cell carcinoma , in which the malignant cells appear larger in size in comparison to sclc . \n collectively , nsclc subtypes contribute to about 86% of all lung cancers.2 the average survival time for patients with sclc is about 10 months . \n the survival time for patients with nsclc varies depending on the stage of the disease . \n patients with an advanced stage of nsclc also have an average survival time of 10 months.3 the 5-year survival rate for sclc is only 6% , and for nsclc , the rate is merely 16% , making the average rate 14% , one of the lowest rates of all cancers.3,4 similar to that of many other cancers , the molecular basis of lung cancer is genetic mutation . \n when mutation occurs , the expression levels of related genes and their products are altered . \n genes in a cell interact with each other in pathways that regulate cell growth , division , dna repair , and apoptosis.5,6 unduly expressed or under expressed genes dysregulate bio - chemical pathways including metabolic pathways and signaling pathways . \n , one needs to understand the molecular mechanisms of carcinogenesis , cancer progression and metastasis and to understand how the cancer related genes interact in networks and pathways . \n the development of microarray technology has made it possible to describe the cellular and molecular states of cells quantitatively and to study the expression levels of many genes simultaneously.710 microarray gene expression profiling of cancer tissues and other disease tissues has been performed by many researchers.816 several groups of researchers conducted gene expression study exploring mrna gene expression differences in normal cells and different types of lung cancer cells including several non - small cell lung cancer cells.1416 in this study , we utilized three squamous cell lung cancer microarray datasets1416 generated by previous researchers to identify dysregulated genes and their associated signaling pathways and to assess the involvement of hedgehog pathway in squamous cell lung cancer . \n the first dataset was generated by a group of researchers to identify a list of genes that can be used to classify different types of lung cancer.14 the dataset includes the microarray gene expression profiles of 127 adenocarcinoma specimens , 21 squamous cell lung carcinomas , and 20 pulmonary cardinoids , as well as 17 normal cell samples . \n the second dataset was created by another group of researchers who worked more specifically with squamous cell lung carcinomas . \n their work paired normal lung tissue and squamous cell lung cancer tissue from 5 patients , and analyzed their results to find which genes were expressed differently in squamous cell cancer compared to normal cells.15 the relations and interactions between the altered genes were also investigated . \n the third dataset was produced by a group aimed at identifying highly dysregulated genes with biological significance in tumor progression.16 the dataset has 46 tumor samples , including 13 adenocarcinomas and 32 squamous cell lung carcinomas , and 45 paired nontumor control samples . in the study , \n gene expression levels were investigated as a function of tumor type , stage and differentiation grade . \n the raw data ( cel files ) were provided for the first dataset , and affymetrix s microarray suite 5 ( mas5 ) implemented in genespring software version 11.0 ( silicon genetics , redwood city , ca ) was used to normalize the expression levels of genes for the dataset . \n the median of each sample was transformed to the median of all samples for both datasets . \n we then carried out statistical analysis to identify the differentially expressed genes between squamous cell lung cancer cells and normal cells . \n for the first dataset , two sample welch t - test17 was deployed to test the null hypothesis that the mean expression level of a gene from cancer cells is the same as the mean expression level of the gene from normal cells . for the second dataset , \n , we assumed that samples were independently selected random samples from the two populations : squamous cell lung carcinomas tissue and normal lung tissue and the population distributions are approximately normal ; this is equivalent to assume that most of the time the fluctuation of gene expression levels within cancer tissues or within normal lung tissues is not that far from the average levels and the fluctuation is symmetric to the average levels.1416 multiple testing corrections based on benjamini and hochberg false discovery rate ( fdr ) were conducted.18,19 the two - tailed p - value 0.05 was used as the cutoff for significance to obtain a set of genes that were significantly altered in squamous cell lung carcinomas . \n for the third dataset , the list of differentially expressed genes provided in the supplementary table to the original paper was used . \n we then found the intersection of the three sets to obtain the genes that are commonly dysregulated in all three datasets . \n the set of commonly altered genes was analyzed using the software ipa ( ingenuity systems , http://www.ingenuity.com ) . \n top functional groups were identified using fisher s exact test20 with p - value 0.05 as the cutoff for significance and marked with the ratio of the number of dysregulated genes that are belong to the functional group to the total number of genes in ipa knowledge database that are annotated to the group . \n significance of association between the differentially expressed genes and the canonical pathways recorded in ipa knowledge database was evaluated with the same measures , p - value from fisher s exact test and the concentration ratio . \n in addition , we examined the expression levels of genes that are associated with hedgehog ( hh ) signaling pathway . \n hh signaling plays important roles in a variety of normal cellular processes including embryogenesis , patterning of organ structures , tissue repair , cellular proliferation and survival.21,22 the analysis of the connection between the altered genes with hh pathway was performed using ipa and fisher s exact test . \n sorting all the p - values for the genes , we can determine which genes have the highest difference in expression levels in the samples , and therefore are most likely to have a difference in expression levels in the population . \n there are 1951 genes that were significantly down regulated in the first dataset , 532 in the second dataset and 1685 genes in third dataset while 1909 , 474 , 1404 are up regulated respectively in the first , second and third datasets . \n 395 differentially expressed genes are common to all the three dataset , including 148 up - regulated and 247 down - regulated . these genes are presented in supplemental tables 1 and 2 . \n the top 30 up / down regulated genes are shown in tables 1 and 2 and the fold change of their expression levels are also presented in figure 1 . since the experiments conducted for the three datasets were carried out in different settings using different microarray chips , in order to compare the fold change of each gene across different experiments , fold change values were normalized for each dataset . \n the biological functions associated with the 395 commonly dysregulated genes were obtained using ipa , as they are most likely to be connected to the mechanism of cancer growth . \n the results show that the genes are associated with many molecular and cellular functions as well as different types of diseases including cancer and genetic disorder . \n the top molecular and cellular functions are presented in table 3 . to examine dysfunctional signaling pathways \n , we utilized all altered genes ( without fdr correction ) that are common to all three datasets . \n fdr correction was not conducted for identifying genes since the focus was on individual pathways . \n there are 1122 differentially expressed genes with 506 commonly up - regulated and 616 down - regulated . \n the core analysis using ipa suggests that over 100 canonical signaling pathways are implicated in squamous cell lung cancer ( supplemental table 3 ) . \n the top altered pathways include cell cycle : g2/m dna damage checkpoint regulation , rar activation , wnt/-catenin signaling , mtor signaling , hif1 signaling , as well as egf signaling . \n the top 40 signaling pathways implicated in squamous cell lung carcinoma are shown in figure 2 . the hedgehog signaling pathway has been implicated in the development of different human cancers including prostate cancer , colon cancer and basal - cell carcinoma . \n gli1 and gli2 are the two key members of gli family and together they have 223 downstream target molecules collected in the ipa knowledge database . \n 15 of them were considerably differentially expressed in squamous cell lung cancer cells with fdr corrected p - values less than 0.05 in all three datasets ( supplemental tables 1 and 2 ) . \n eight of them , cdc45 , cdk1 , csta , hdac1 , jag1 , jup , krt15 , krt17 , were up - regulated ; 7 of the genes , cxcl2 , fhl1 , foxa2 , papss2 , rhob , sap18 , syne1 , were down \n . the fisher s exact test on the 37 genes yields a p - value of 0.0484 ( < 0.05 ) which indicates the strong association of hedgehog signaling with non - small cell lung cancer . \n genes in a cell interact with each other through a sequence of orchestrated events in pathways . \n aberrant cell signaling along the pathways breaks the organized events , promotes proliferation , inhibits apoptosis , induces angiogenesis , and facilitates the cancer to spread . \n our study combined three microarray datasets to explore more stable biomarkers or drug targets for squamous cell lung cancer and the pathways implicated . \n the results indicate that the expression levels of many genes were altered and multiple pathways were dysfunctional in squamous cell lung carcinoma ( supplemental tables 1 and 2 , fig . \n 1 ) . the altered genes encode cytokines , enzymes , growth factors , kinases , transporters transmembrane/ nuclear receptors and transcription regulators . \n the molecules were located in multiple cellular locations in extracellular space , cytoplasm , nucleus , and plasma membrane . \n as we can see from table 1 , the top up- regulated genes include members of the keratin family , keratin 6a , keratin 15 , keratin 16 , and keratin 17 . \n keratins are intermediate filament forming proteins that provide mechanical support and perform a diversity of other functions in epithelial cells.2325 all four of them are epithelial keratin genes . \n they are associated with molecular functions such as structural constituent of cytoskeleton and protein binding and participate in cytoskeleton organization , cell differentiation , regulation of cell proliferation , epidermis development , intermediate filament cytoskeleton organization and ectoderm development . \n the proteins encoded by keratin 6a are type ii cytokeratins ( neutral - basic ) . \n these cytokeratins organize in pairs of heterotypic keratin chains , which are co - expressed during differentiation of simple and stratified epithelial tissues . \n keratin 16 and 17 are paired with keratin 6 while keratin 15 is paired with keratin 5 in squamous epithelial cells . \n it has been reported that keratins can be used as diagnostic markers in human carcinomas.25 a recent review on keratin summarized the evidence for active keratin involvement in cancer cell invasion and metastasis , as well as in treatment responsiveness.25 however , their involvement in squamous cell lung carcinoma requires further investigation . \n the protein encoded by this cdk1 is a member of the ser / thr protein kinase family . \n it is a catalytic subunit of the highly conserved protein kinase complex known as m - phase promoting factor , which is essential for g1/s and g2/m phase transitions of eukaryotic cell cycle . \n the kinase activity of this protein is controlled by cyclin accumulation and destruction through the cell cycle . \n the phosphorylation of this protein also plays important regulatory roles in cell cycle control.2628 cdk1 is involved in many signaling pathways including apoptosis signaling ; atm signaling ; cell cycle : g2/m dna damage checkpoint regulation ; cyclins and cell cycle regulation ; role of chk proteins in cell cycle checkpoint control ; and sonic hedgehog signaling . in addition , several other cell - division cycle proteins including cdc20 and cdc45 were up - regulated . \n several lim domain proteins , fhl1 , limch , and lmo3 , are in this group . \n the lim domain is a double - zinc finger motif found in a diversity of proteins . \n it is reported that the primary role of lim domains is in protein - protein interaction , through the formation of dimers with identical or different lim domains or by binding distinct proteins . \n lim - only proteins are implicated in the control of cell proliferation.29,30 fhl1 that encodes a member of the four - and - a - half- lim - only protein family was significantly down regulated . \n expression of these family members occurs in a cell and tissue specific mode and the proteins are involved in many cellular processes . \n it was suggested that the inhibitory effects of fhl1 on lung cancer cell growth were associated with both the g1 and the g2/m cell cycle arrest associated with a marked inhibition of cyclin a , cyclin b1 and cyclin d as well as the induction of the cyclin dependent kinase inhibitors p21 and p27.30 conversely , several members of cyclin family ( cyclin a2 , cyclin b1 , cyclin b2 and cyclin dependent kinase i ) were found up - regulated in lung cancer in this study ( supplemental table 1 ) while cyclin dependent kinase inhibitors cdkn1a and cdkn1b were consistently down - regulated in all three datasets ( although the down - regulation levels were not above the statistical significant threshold ) . \n down - regulated rhob is a member of the rho gtp - binding protein family . \n the rho family plays a crucial role in the dynamic regulation of the actin cytoskeleton . \n studies suggested that the proteins of the family also have a diversity of functions in membrane pathways . \n the rhob was shown to localize to vesicles of the endocytic compartment , suggesting a potential function in regulation of endocytic traffic . \n the expression of active rhob causes a delay in the intracellular trafficking of the epidermal growth factor ( egf ) receptor through regulating prk1.3133 the signaling pathway investigation in this study reveals that many of the cancer related pathways reported in the literatures are also responsible for squamous cell lung carcinoma . \n they include cell cycle regulation pathway ( p = 1.8e-7 ) , p53 tumor - suppressor pathway ( p = 4.2e-5 ) , il-8 signaling ( p = 1.9e-4 ) , wnt--catenin pathway ( p = 0.0038 ) , mtor signaling ( p = 0.0042 ) , and egf signaling ( p = 0.010 ) ( fig . \n cell cycle : g2/m dna damage checkpoint regulation pathway is on the top of the altered pathways . \n it prevents cells from entering mitosis when dna is damaged , providing an opportunity for repair and stopping the proliferation of damaged cells and helping to maintain genomic stability.26 14 out of 49 molecules on the pathway were significantly altered in lung cancer including the cyclin - dependent kinase 1 ( cdk1 ) which is essential to drive cells into mitosis ( supplemental tables 1 and 2 ) . \n p53 activation is induced by a number of stress signals , including dna damage , oxidative stress and activated oncogenes . \n 17 out of 49 molecules on the pathway were significantly altered , including up - regulated serpinb5 and tp63 . \n serpin peptidase inhibitor , clade b ( ovalbumin ) , member 5 ( serpinb5 ) , also known as maspin , is a tumor suppressor gene . \n it was reported to block the growth , invasion , and metastatic properties of mammary tumors . \n its expression was down- regulated in breast tumors , prostate cancer and uterine cervix cancer.3436 it was reported that serpinb5 over - expression increased the rate of apoptosis of both preneoplastic and carcinomatous mammary epithelial cells and reduced tumor growth through a combination of reduced angiogenesis and increased apoptosis.32 however , the role of serpinb5 in the inhibition of tumor growth is rather complex . \n in contrast to its reduced expression levels in breast cancer and other types of cancer , serpinb5 s over - expression in pancreatic carcinoma was also reported.37 a recent study shows how it works in regulating cell growth when tightly coupled with the thrombin receptor protease activated receptor-1 ( par-1).38 \n our study indicates that serpinb5 is significantly over - expressed in squamous cell lung carcinoma in all three datasets ( table 1 ) . \n the tumor protein tp63 is another up - regulated gene ( table 1 ) that encodes a member of the p53 family of transcription factors . \n its over - expression has already been documented in lung carcinoma.39 egfr encodes a receptor for members of the epidermal growth factor family . \n binding of the protein to a ligand induces receptor dimerization and tyrosine autophosphorylation and leads to inappropriate activation of the anti - apoptotic ras signaling cascade , eventually leading to uncontrolled cell proliferation . both egfr and the signal transducer and activator stat1 were up - regulated in the three datasets ( based on the p - values without fdr correction ) . \n egf receptor signaling has long been studied in searching for the treatment of lung cancer40,41 and egfr inhibitors against lung cancer have been explored.4245 in principle , it should be feasible to regulate the activity of signaling pathways with noncytotoxic agents . \n gefitinib and erlotinib are two agents used in the treatment of non - small cell lung carcinoma . \n these drugs are epidermal growth factor receptor ( egfr ) tyrosine kinase inhibitor , which interrupt signaling through egfr in target cells.44,45 they improve response rate and survival , especially in the subpopulation of patients who carry certain mutations in egfr gene . \n however , clinical trials show that egfr tyrosine kinase inhibitors , like many other drugs , have not been successful in curing cancer.4448 another important cancer related pathway is wnt/ -catenin signaling pathway . \n wnt signaling pathway is a network of proteins best known for their roles in embryogenesis and cancer , but also involved in normal physiological processes in adult animals.46,47 wnt proteins are secreted morphogens that are required for basic developmental processes , such as cell - fate specification , progenitor - cell proliferation and the control of asymmetric cell division , in many different species and organs.49,50 wnt ligand binds to its receptor and stabilizes cytoplasmic -catenin through inhibition of the -catenin degradation complex . \n -catenin is then free to enter the nucleus and activate wnt - regulated genes through its interaction with tcf family transcription factors and associated recruitment of coactivators . \n although conflicting roles of wnt-5 a in cancer have been reported,51 our data shows it is significantly up - regulated and the wnt inhibitor wif1 is down - regulated . \n the p - values without fdr correction are consistently below the threshold in all three datasets . \n mtor pathway has been extensively explored in some other cancers , including renal cell carcinoma , and large clinical trials have reported promising progression - free survival advantage with mtor inhibitor drugs in this disease . \n our report suggests that mtor signaling may be involved in squamous cell lung carcinoma as well , and exploring this pathway in more depth in squamous cell carcinoma of the lung may be needed.52,53 many hurdles remain in the race to find a better treatment to cancer . \n recently a cellular screen for antagonists of gli - mediated transcription revealed that a small molecule gant61 was able to selectively inhibit the gli - mediated gene transactivation , which is the last step of the hh pathway.54 hh signaling is tightly coupled with wnt signaling.47 it plays important roles in a variety of normal cellular processes including embryogenesis , patterning of organ structures , tissue repair , cellular proliferation and survival.21,22 the signaling pathway is activated when hedgehog signaling ligands , such as sonic hedgehog , bind to the membrane receptor patched ( ptch ) , and thus stop the suppression of the signaling molecule smoothened ( smo ) caused by ptch . \n smo then activates the cytoplasmic protein sufu , which releases the gli family of transcription factors , gli1 and gli2 . \n once the gli molecules are released , they translocate to the nucleus , where they bind to target genes to start transcription.21,22 the signaling process is shown in figure 3 . \n hh signaling pathway is an important regulator of the embryonic development and becomes reactivated in cancer . \n it has been activated in several human cancers including prostate cancer , colon , and small cell lung cancer although the pathway seems to be inactive in most normal adult tissues.5558 the inhibition of this pathway could offer a selective way of controlling just the tumor cells . in our data , gli2 was up - regulated although the significantly altered expression level of gli1 was not observed . \n on the other hand , our investigation into the group of gli1 and gli2 downstream molecules using ipa revealed that significant number of genes was altered in the group ( table 5 ) . \n the p - value 0.0484 from the fisher s exact test is below the commonly used threshold 0.05 ; indicating hh signaling pathway is involved in squamous cell lung carcinoma . \n sorting all the p - values for the genes , we can determine which genes have the highest difference in expression levels in the samples , and therefore are most likely to have a difference in expression levels in the population . \n there are 1951 genes that were significantly down regulated in the first dataset , 532 in the second dataset and 1685 genes in third dataset while 1909 , 474 , 1404 are up regulated respectively in the first , second and third datasets . \n 395 differentially expressed genes are common to all the three dataset , including 148 up - regulated and 247 down - regulated . these genes are presented in supplemental tables 1 and 2 . \n the top 30 up / down regulated genes are shown in tables 1 and 2 and the fold change of their expression levels are also presented in figure 1 . since the experiments conducted for the three datasets were carried out in different settings using different microarray chips , in order to compare the fold change of each gene across different experiments , fold change values were normalized for each dataset . \n the biological functions associated with the 395 commonly dysregulated genes were obtained using ipa , as they are most likely to be connected to the mechanism of cancer growth . \n the results show that the genes are associated with many molecular and cellular functions as well as different types of diseases including cancer and genetic disorder . \n to examine dysfunctional signaling pathways , we utilized all altered genes ( without fdr correction ) that are common to all three datasets . \n fdr correction was not conducted for identifying genes since the focus was on individual pathways . \n there are 1122 differentially expressed genes with 506 commonly up - regulated and 616 down - regulated . \n the core analysis using ipa suggests that over 100 canonical signaling pathways are implicated in squamous cell lung cancer ( supplemental table 3 ) . \n the top altered pathways include cell cycle : g2/m dna damage checkpoint regulation , rar activation , wnt/-catenin signaling , mtor signaling , hif1 signaling , as well as egf signaling . \n the top 40 signaling pathways implicated in squamous cell lung carcinoma are shown in figure 2 . \n the hedgehog signaling pathway has been implicated in the development of different human cancers including prostate cancer , colon cancer and basal - cell carcinoma . \n gli1 and gli2 are the two key members of gli family and together they have 223 downstream target molecules collected in the ipa knowledge database . \n 15 of them were considerably differentially expressed in squamous cell lung cancer cells with fdr corrected p - values less than 0.05 in all three datasets ( supplemental tables 1 and 2 ) . \n eight of them , cdc45 , cdk1 , csta , hdac1 , jag1 , jup , krt15 , krt17 , were up - regulated ; 7 of the genes , cxcl2 , fhl1 , foxa2 , papss2 , rhob , sap18 , syne1 , were down . the fisher s exact test on the 37 genes yields a p - value of 0.0484 ( < 0.05 ) which indicates the strong association of hedgehog signaling with non - small cell lung cancer . \n genes in a cell interact with each other through a sequence of orchestrated events in pathways . \n aberrant cell signaling along the pathways breaks the organized events , promotes proliferation , inhibits apoptosis , induces angiogenesis , and facilitates the cancer to spread . \n our study combined three microarray datasets to explore more stable biomarkers or drug targets for squamous cell lung cancer and the pathways implicated . \n the results indicate that the expression levels of many genes were altered and multiple pathways were dysfunctional in squamous cell lung carcinoma ( supplemental tables 1 and 2 , fig . \n 1 ) . the altered genes encode cytokines , enzymes , growth factors , kinases , transporters transmembrane/ nuclear receptors and transcription regulators . \n the molecules were located in multiple cellular locations in extracellular space , cytoplasm , nucleus , and plasma membrane . as we can see from table 1 , \n the top up- regulated genes include members of the keratin family , keratin 6a , keratin 15 , keratin 16 , and keratin 17 . \n keratins are intermediate filament forming proteins that provide mechanical support and perform a diversity of other functions in epithelial cells.2325 all four of them are epithelial keratin genes . \n they are associated with molecular functions such as structural constituent of cytoskeleton and protein binding and participate in cytoskeleton organization , cell differentiation , regulation of cell proliferation , epidermis development , intermediate filament cytoskeleton organization and ectoderm development . \n the proteins encoded by keratin 6a are type ii cytokeratins ( neutral - basic ) . \n these cytokeratins organize in pairs of heterotypic keratin chains , which are co - expressed during differentiation of simple and stratified epithelial tissues . \n keratin 16 and 17 are paired with keratin 6 while keratin 15 is paired with keratin 5 in squamous epithelial cells . \n it has been reported that keratins can be used as diagnostic markers in human carcinomas.25 a recent review on keratin summarized the evidence for active keratin involvement in cancer cell invasion and metastasis , as well as in treatment responsiveness.25 however , their involvement in squamous cell lung carcinoma requires further investigation . \n the protein encoded by this cdk1 is a member of the ser / thr protein kinase family . \n it is a catalytic subunit of the highly conserved protein kinase complex known as m - phase promoting factor , which is essential for g1/s and g2/m phase transitions of eukaryotic cell cycle . \n the kinase activity of this protein is controlled by cyclin accumulation and destruction through the cell cycle . \n the phosphorylation of this protein also plays important regulatory roles in cell cycle control.2628 cdk1 is involved in many signaling pathways including apoptosis signaling ; atm signaling ; cell cycle : g2/m dna damage checkpoint regulation ; cyclins and cell cycle regulation ; role of chk proteins in cell cycle checkpoint control ; and sonic hedgehog signaling . in addition , several other cell - division cycle proteins including cdc20 and cdc45 were up - regulated . \n several lim domain proteins , fhl1 , limch , and lmo3 , are in this group . \n the lim domain is a double - zinc finger motif found in a diversity of proteins . \n it is reported that the primary role of lim domains is in protein - protein interaction , through the formation of dimers with identical or different lim domains or by binding distinct proteins . \n lim - only proteins are implicated in the control of cell proliferation.29,30 fhl1 that encodes a member of the four - and - a - half- lim - only protein family was significantly down regulated . \n expression of these family members occurs in a cell and tissue specific mode and the proteins are involved in many cellular processes . \n it was suggested that the inhibitory effects of fhl1 on lung cancer cell growth were associated with both the g1 and the g2/m cell cycle arrest associated with a marked inhibition of cyclin a , cyclin b1 and cyclin d as well as the induction of the cyclin dependent kinase inhibitors p21 and p27.30 conversely , several members of cyclin family ( cyclin a2 , cyclin b1 , cyclin b2 and cyclin dependent kinase i ) were found up - regulated in lung cancer in this study ( supplemental table 1 ) while cyclin dependent kinase inhibitors cdkn1a and cdkn1b were consistently down - regulated in all three datasets ( although the down - regulation levels were not above the statistical significant threshold ) . \n down - regulated rhob is a member of the rho gtp - binding protein family . \n the rho family plays a crucial role in the dynamic regulation of the actin cytoskeleton . \n studies suggested that the proteins of the family also have a diversity of functions in membrane pathways . \n the rhob was shown to localize to vesicles of the endocytic compartment , suggesting a potential function in regulation of endocytic traffic . \n the expression of active rhob causes a delay in the intracellular trafficking of the epidermal growth factor ( egf ) receptor through regulating prk1.3133 the signaling pathway investigation in this study reveals that many of the cancer related pathways reported in the literatures are also responsible for squamous cell lung carcinoma . \n they include cell cycle regulation pathway ( p = 1.8e-7 ) , p53 tumor - suppressor pathway ( p = 4.2e-5 ) , il-8 signaling ( p = 1.9e-4 ) , wnt--catenin pathway ( p = 0.0038 ) , mtor signaling ( p = 0.0042 ) , and egf signaling ( p = 0.010 ) ( fig . \n cell cycle : g2/m dna damage checkpoint regulation pathway is on the top of the altered pathways . \n it prevents cells from entering mitosis when dna is damaged , providing an opportunity for repair and stopping the proliferation of damaged cells and helping to maintain genomic stability.26 14 out of 49 molecules on the pathway were significantly altered in lung cancer including the cyclin - dependent kinase 1 ( cdk1 ) which is essential to drive cells into mitosis ( supplemental tables 1 and 2 ) . \n p53 activation is induced by a number of stress signals , including dna damage , oxidative stress and activated oncogenes . \n 17 out of 49 molecules on the pathway were significantly altered , including up - regulated serpinb5 and tp63 . \n serpin peptidase inhibitor , clade b ( ovalbumin ) , member 5 ( serpinb5 ) , also known as maspin , is a tumor suppressor gene . \n it was reported to block the growth , invasion , and metastatic properties of mammary tumors . \n its expression was down- regulated in breast tumors , prostate cancer and uterine cervix cancer.3436 it was reported that serpinb5 over - expression increased the rate of apoptosis of both preneoplastic and carcinomatous mammary epithelial cells and reduced tumor growth through a combination of reduced angiogenesis and increased apoptosis.32 however , the role of serpinb5 in the inhibition of tumor growth is rather complex . \n in contrast to its reduced expression levels in breast cancer and other types of cancer , serpinb5 s over - expression in pancreatic carcinoma was also reported.37 a recent study shows how it works in regulating cell growth when tightly coupled with the thrombin receptor protease activated receptor-1 ( par-1).38 our study indicates that serpinb5 is significantly over - expressed in squamous cell lung carcinoma in all three datasets ( table 1 ) . \n the tumor protein tp63 is another up - regulated gene ( table 1 ) that encodes a member of the p53 family of transcription factors . \n its over - expression has already been documented in lung carcinoma.39 egfr encodes a receptor for members of the epidermal growth factor family . \n binding of the protein to a ligand induces receptor dimerization and tyrosine autophosphorylation and leads to inappropriate activation of the anti - apoptotic ras signaling cascade , eventually leading to uncontrolled cell proliferation . \n both egfr and the signal transducer and activator stat1 were up - regulated in the three datasets ( based on the p - values without fdr correction ) . \n egf receptor signaling has long been studied in searching for the treatment of lung cancer40,41 and egfr inhibitors against lung cancer have been explored.4245 in principle , it should be feasible to regulate the activity of signaling pathways with noncytotoxic agents . \n gefitinib and erlotinib are two agents used in the treatment of non - small cell lung carcinoma . \n these drugs are epidermal growth factor receptor ( egfr ) tyrosine kinase inhibitor , which interrupt signaling through egfr in target cells.44,45 they improve response rate and survival , especially in the subpopulation of patients who carry certain mutations in egfr gene . \n however , clinical trials show that egfr tyrosine kinase inhibitors , like many other drugs , have not been successful in curing cancer.4448 another important cancer related pathway is wnt/ -catenin signaling pathway . \n wnt signaling pathway is a network of proteins best known for their roles in embryogenesis and cancer , but also involved in normal physiological processes in adult animals.46,47 wnt proteins are secreted morphogens that are required for basic developmental processes , such as cell - fate specification , progenitor - cell proliferation and the control of asymmetric cell division , in many different species and organs.49,50 wnt ligand binds to its receptor and stabilizes cytoplasmic -catenin through inhibition of the -catenin degradation complex . \n -catenin is then free to enter the nucleus and activate wnt - regulated genes through its interaction with tcf family transcription factors and associated recruitment of coactivators . \n although conflicting roles of wnt-5 a in cancer have been reported,51 our data shows it is significantly up - regulated and the wnt inhibitor wif1 is down - regulated . \n the p - values without fdr correction are consistently below the threshold in all three datasets . \n mtor pathway has been extensively explored in some other cancers , including renal cell carcinoma , and large clinical trials have reported promising progression - free survival advantage with mtor inhibitor drugs in this disease . \n our report suggests that mtor signaling may be involved in squamous cell lung carcinoma as well , and exploring this pathway in more depth in squamous cell carcinoma of the lung may be needed.52,53 many hurdles remain in the race to find a better treatment to cancer . \n recently a cellular screen for antagonists of gli - mediated transcription revealed that a small molecule gant61 was able to selectively inhibit the gli - mediated gene transactivation , which is the last step of the hh pathway.54 hh signaling is tightly coupled with wnt signaling.47 it plays important roles in a variety of normal cellular processes including embryogenesis , patterning of organ structures , tissue repair , cellular proliferation and survival.21,22 the signaling pathway is activated when hedgehog signaling ligands , such as sonic hedgehog , bind to the membrane receptor patched ( ptch ) , and thus stop the suppression of the signaling molecule smoothened ( smo ) caused by ptch . \n smo then activates the cytoplasmic protein sufu , which releases the gli family of transcription factors , gli1 and gli2 . \n once the gli molecules are released , they translocate to the nucleus , where they bind to target genes to start transcription.21,22 the signaling process is shown in figure 3 . \n hh signaling pathway is an important regulator of the embryonic development and becomes reactivated in cancer . \n it has been activated in several human cancers including prostate cancer , colon , and small cell lung cancer although the pathway seems to be inactive in most normal adult tissues.5558 the inhibition of this pathway could offer a selective way of controlling just the tumor cells . in our data , \n gli2 was up - regulated although the significantly altered expression level of gli1 was not observed . \n on the other hand , our investigation into the group of gli1 and gli2 downstream molecules using ipa revealed that significant number of genes was altered in the group ( table 5 ) . \n the p - value 0.0484 from the fisher s exact test is below the commonly used threshold 0.05 ; indicating hh signaling pathway is involved in squamous cell lung carcinoma . \n by combining three microarray datasets , we created a more comprehensive and complete gene expression analysis of squamous cell lung cancer . \n the list of altered genes is more reliable than the ones obtained from a single setting of experiments , therefore providing a more dependable set of lung cancer biomarkers/ drug targets . \n the results of the analysis indicate numerous genes were altered and multiple pathways were dysfunctional in squamous cell lung carcinoma . \n as discussed above , many of the altered genes have been implicated in different types of carcinoma although some of the altered genes are organ - specific . \n hh signaling is implicated in squamous cell lung cancer , opening the door for exploring new cancer therapeutic treatment using gli antagonist gant 61 . while active in many different types of cancer , hh pathway \n is believed to be mostly inactive in normal adult tissues , making it a good target for cancer treatments . \n consequently , the treatment based on gant 61 is likely to cause few side effects . \n ", "role": "user" }, { "content": "lung cancer is the second most commonly occurring non - cutaneous cancer in the united states with the highest mortality rate among both men and women . in this study \n , we utilized three lung cancer microarray datasets generated by previous researchers to identify differentially expressed genes , altered signaling pathways , and assess the involvement of hedgehog ( hh ) pathway . \n the three datasets contain the expression levels of tens of thousands genes in normal lung tissues and squamous cell lung carcinoma . \n the datasets were combined and analyzed . \n the dysregulated genes and altered signaling pathways were identified using statistical methods . \n we then performed fisher s exact test on the significance of the association of hh pathway downstream genes and squamous cell lung carcinoma.395 genes were found commonly differentially expressed in squamous cell lung carcinoma . \n the genes encoding fibrous structural protein keratins and cell cycle dependent genes encoding cyclin - dependent kinases were significantly up - regulated while the ones encoding lim domains were down . over 100 signaling pathways were implicated in squamous cell lung carcinoma , including cell cycle regulation pathway , p53 tumor - suppressor pathway , il-8 signaling , wnt--catenin pathway , mtor signaling and egf signaling . \n in addition , 37 out of 223 downstream molecules of hh pathway were altered . \n the p - value from the fisher s exact test indicates that hh signaling is implicated in squamous cell lung carcinoma.numerous genes were altered and multiple pathways were dysfunctional in squamous cell lung carcinoma . \n many of the altered genes have been implicated in different types of carcinoma while some are organ - specific . \n hh signaling is implicated in squamous cell lung cancer , opening the door for exploring new cancer therapeutic treatment using gli antagonist gant 61 .", "role": "assistant" } ]

[ { "content": "You are a medical writer. Summarize the following article: progeria is a rare premature aging syndrome characterized by retarded physical development , abnormal facies , skeletal abnormalities and early onset of scleroderma . \n it was first described in 1886 by jonathan hutchinson and was also described independently in 1897 by hastings gilford . \n affected children are normal at birth and growth retardation is noticed from first year of life . by the second year \n , skin becomes thin and shiny in some areas but lax and wrinkled in other areas . \n two siblings , a 14-year - old male child and a 13-year - old female child , born to third - degree consanguineous parents , presented with abnormal facies and prematurely aged appearance . \n history from the mother revealed that the antenatal period was uneventful during both the pregnancies . \n both the children were delivered at term by cesarean section and their birth weight was more than 2.75 kg . \n mile stones were attained normally till 9 months of age after which growth retardation was noticed in both the children . \n height of the male and female child was 91 and 90 cm , respectively , while their weight was 11 and 10 kg , respectively . they had bird like facies with patent anterior fontanalle , frontal bossing , prominent eyes , protruding ears , beaked nose and micrognathia [ figure 1 ] . \n eyebrows were absent in the male and sparse in the female . they had high pitched voice and their intelligence was normal . \n beaked nose , protruding ears and micrognathism in both children crowded and irregularly erupted teeth dermatological examination revealed shiny hyperpigmented skin over forhead and thin , and wrinkled skin over hands and feet . \n there were hypopigmented macules interspersed between hyperpigmented macules in the chest and abdomen [ figure 3 ] . \n thin and wrinkled skin over the hands with dystrophic nails their blood count , hemoglobin percentage , blood sugar , renal and liver parameters and lipid profile were within normal limits . \n total thyroxine ( tt4 ) , thyroid stimulating hormone ( tsh ) , growth hormone ( gh ) levels were normal and there was no evidence of primary endocrinological dysfunction . \n usg abdomen revealed a mild hepatomegaly in the male child and no significant findings in the female child . \n echo showed mild anterior mitral leaflet thickening and trivial aortic regurgitation in the male child and mitral valve prolapse with trivial mitral regurgitation in the female child but their left ventricular function was normal . \n male child complained of visual disturbances and on ophthalmologic examination , he was found to have hypermetropia and pseudopapillitis . \n the female child had a swelling in the gums and was diagnosed as peripheral ossifying fibroma in the dental opd . \n ct and x - ray skull showed normal facial and skull bones with open anterior fontanalle . \n a diagnosis of progeria was made based on the clinical findings of short stature with bird like facies , hair and nail changes and normal intelligence . \n short stature , bird like facies and normal intelligence are distinctive features of progeria . in our case , the skin changes and normal intelligence excluded the other disorders with short stature and abnormal facies like bird headed dwarfism . the early age at onset excluded pangeria . \n absence of disproportionately large extremities , ocular changes and photosensitivity helped to exclude cockayne syndrome . \n it is usually caused by a new ( sporadic ) mutation during the early division of the cells in the child . \n it is usually genetically dominant ; therefore , parents who are healthy will normally not pass it on to their children . \n they typically live about 13 years , although many have been known to live into their late teens and early twenties and rarely some individuals may reach their forties . clinically , healthy parents with affected siblings in the family favor autosomal recessive pattern of inheritance , as in our case . \n there are very few case reports[68 ] of progeria in siblings with an autosomal recessive pattern of inheritance , which have been documented in india . \n we add on to this rare case list by reporting progeria in two children of the same family , suggesting an autosomal recessive pattern of inheritance .", "role": "user" }, { "content": "progeria , also known as hutchinson - gilford syndrome , is an extremely rare , severe genetic condition wherein symptoms resembling aspects of aging are manifested at an early age . \n it is an autosomal dominant disorder . \n it is not seen in siblings of affected children although there are very few case reports of progeria affecting more than one child in a family . \n here we are presenting two siblings , a 14-year - old male and a 13-year - old female with features of progeria , suggesting a possible autosomal recessive inheritance .", "role": "assistant" } ]