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atrial fibrillation ( af ) is the most common sustained arrhythmia in western countries , with an estimated 30 million patients affected by 2050 across united states and europe alone .
atrial fibrillation has a significant impact on morbidity mainly related to symptoms , heart failure , and thromboembolic events and is the most frequent arrhythmic cause of hospital admission in the usa [ 13 ] .
in addition , af is associated with excess mortality independently of thromboembolic complications . to date , the most effective treatment for af is radiofrequency catheter ablation , and pulmonary vein antrum isolation ( pvai ) is the mainstay of such an approach .
the major drawback of catheter ablation of af consists in its potential risk of periprocedural complications , with thromboembolic and hemorrhagic complications being among the most common and insidious ones [ 7 , 8 ] . despite the introduction of novel ablation technologies , such as open irrigation catheters , and the widespread use of systemic anticoagulation with heparin
, the risk of periprocedural thromboembolism remains not negligible , reaching up to 2.8% in large series [ 6 , 9 ] . with these premises ,
the development of novel strategies able to further decrease the risk of periprocedural thromboembolism without increasing the risk of bleeding is crucial .
oral anticoagulant therapy with warfarin is the current standard of care for the treatment of af , with a strikingly favorable balance between thromboembolic protection and bleeding complications when adequate monitoring of international normalized ratio ( inr ) is achieved . for years ,
discontinuation of warfarin before ablation associated with the periprocedural use of unfractioned and low molecular weight heparin and bridging with warfarin after ablation has been the most widespread anticoagulation protocol in patients undergoing catheter ablation of af [ 5 , 11 ] .
we firstly described the feasibility of af ablation without interruption of oral anticoagulation and recently reported that this approach is able to potentially abolish thromboembolic complications without increasing the risk of bleeding .
accordingly , af ablation without interruption of anticoagulation is nowadays the standard protocol in our center . in this paper , we summarize our approach to anticoagulation in patients undergoing af ablation .
patients eligible for catheter ablation of af are started on warfarin as outpatients , at least 2 months before the scheduled procedure .
all patients receive weekly inr monitoring during the 4 to 6 weeks preceding the procedure , with a target inr of 2 to 3 .
preprocedural transesophageal echocardiography ( tee ) is performed only in patients showing sub - therapeutic inr values in the month prior to the procedure .
patients who demonstrate inr values consistently above 2 in the month before the procedure are directly sent to ablation .
four venous accesses are obtained : two right and one left femoral venous accesses , and one right internal jugular vein access .
femoral venous accesses are usually obtained with the seldinger technique or with ultrasound guidance in difficult cases . the right internal jugular vein
is accessed with ultrasound guidance or under fluoroscopic guidance with a wire or a deflectable catheter advanced into this vein via the right femoral vein sheath .
a double transseptal puncture is performed with the assistance of intracardiac echocardiography ( ice ) [ 13 , 14 ] . before transseptal punctures ,
all patients receive a bolus of unfractionated heparin ( 10,000 units ) , followed by a noncontinuous infusion to maintain an activated coagulation time ( act ) > 300 seconds .
we found important the addition of unfractionated heparin to periprocedural therapeutic warfarin , since soft thrombus can still be observed on the transeptal sheath or left atrial catheters in patients with periprocedural therapeutic inr .
radiofrequency energy is delivered with an open irrigated ablation catheter with a maximum temperature of 42c , power up to 45 w , and flow rate of 30 cm/min .
intracardiac echocardiography is continuously used to monitor the electrode surface during ablation , to assist with catheter positioning and identify coagulum formation , and to monitor for complications including pericardial effusion .
an esophageal temperature probe is always inserted to assist with power titration during posterior wall ablation . at the end of the left atrial ablation , we partially reverse heparin anticoagulation with up to 40 mg of protamine guided by the act and remove the sheaths when the act is less than 250 seconds . with this approach
the majority of the major bleedings are related to cardiac tamponade , which can be effectively managed with emergent pericardiocentesis , together with heparin interruption and reversal with protamine , and warfarin reversal with fresh - frozen plasma or prothrombin complex concentrate .
if continued significant drainage or reaccumulation of the pericardial effusion occurs despite these measures , emergent open surgical exploration is considered .
notably , there is also no difference in rates of emergent surgical exploration between patients with periprocedural therapeutic inr and those who discontinue warfarin , although patients on therapeutic anticoagulation are more likely to have a larger amount of blood removed from their pericardium for stabilization and require more blood transfusion units .
all patients receive a single dose of aspirin ( 325 mg ) before leaving the electrophysiology laboratory and continue their warfarin dosage regimens to maintain a target inr of 2 to 3 .
after ablation the thromboembolic risk may be further increased by the procedure - related endothelial damage , which may activate the coagulation cascade and increase the risk of thromboembolism .
therefore , warfarin discontinuation may be associated with an increased thromboembolic risk also in the postprocedural period , since reachieving a therapeutic inr after the procedure may take several days , and the risk of left atrial thrombosis during af is strikingly time dependent .
all patients are strictly monitored for outcome and complications during overnight hospital stay , and on the following day prior to discharge using symptom assessment , serial neurological examinations , and puncture site checks .
all patients are instructed to call in case of any symptom development and to send weekly transtelephonic electrocardiogram transmissions for the first 5 months after ablation .
progress of recovery and symptoms are assessed as well by a dedicated nurse . in case of symptoms or suspected complications patients
are asked to seek medical attention at either a local emergency department or our emergency department or to follow up with their local physician .
moreover , all patients present for followup 3 to 4 months after ablation with the electrophysiologist who performed the procedure . with regard to the out - of - hospital long - term anticoagulation management , patients are referred to dedicated anticoagulation clinics with the aim of maintaining a stable therapeutic inr level .
we follow a standard , uniform , and validated protocol of long - term postprocedural anticoagulation management . briefly ,
oral anticoagulation is discontinued , regardless of the chads2 score , if patients do not experience any recurrence of atrial tachyarrhythmias , severe pulmonary vein stenosis ( pulmonary vein narrowing > 70% ) , and severe left atrial mechanical dysfunction , as assessed by transthoracic echocardiography .
patients with a chads2 score 1 experiencing early recurrence of af are maintained on warfarin for at least 6 months . in these patients
, warfarin is discontinued if there is no af recurrence in the last 3 months without antiarrhythmic drugs , and aspirin 81 to 325 mg is started . in case of new af recurrence after warfarin discontinuation in patients with a chads2 score 1 , oral anticoagulation
our approach to periprocedural anticoagulation has been extensively validated in previous work [ 12 , 13 , 17 ] . in particular , we found that a conventional anticoagulation approach , which included warfarin discontinuation with peri- and postprocedural bridging with unfractioned and low - molecular - weight heparin actually increases the risk of bleeding and thromboembolic complications , as compared to no periprocedural interruption of oral anticoagulation [ 12 , 13 , 17 ] .
our most recent report provides strong evidence that performing af ablation under therapeutic inr is a safe and effective approach to virtually abolish the risk of thromboembolic complications .
we reported a multicenter prospective comparison of three anticoagulation protocols in 9 centers performing the same ablation procedure .
a total of 6,454 patients were included in the study , of whom 2,488 underwent ablation with an 8-mm ablation catheter and preprocedural warfarin discontinuation ( group 1 ) , 1,348 underwent ablation with an open irrigated catheter and preprocedural warfarin discontinuation ( group 2 ) , and 2,618 underwent ablation with an open irrigated catheter without preprocedural warfarin discontinuation ( group 3 ) .
overall , periprocedural thromboembolic complications occurred in 39 ( 0.6% ) patients , with a rate of 1.1% in group 1 , and of 0.9% in group 2 .
these data support also the appropriateness of our approach to tee based on the intensity of anticoagulation in the month preceding the procedure , especially considering that 1,178 ( 45% ) group 3 patients had persistent af , and 498 ( 19% ) long - standing persistent af . at multivariable analysis , which adjusted for age , gender , coronary artery disease , type of af , heart failure , diabetes , hypertension , and prior stroke , the anticoagulation strategy of ablation with
a therapeutic inr was a strong independent predictor of lower periprocedural thromboembolic events ( odds ratio 0.54 , 95% confidence interval 0.32 to 0.89 , p = .017 ) .
with regard to bleedings , the pooled rate of major bleeding complications ( i.e. , bleeding requiring interventions including transfusions , hemopericardium , hemothorax , and retroperitoneal bleeding ) and pericardial effusion in patients who discontinued warfarin before the ablation procedure ( groups 1 and 2 ) was 1.1% , whereas in group 3 was 0.8% ( figure 1 ) .
if also minor bleedings were included , patients who discontinued warfarin before ablation procedure had a pooled rate of bleeding complications of 20.7% , while patients who were maintained on warfarin had a rate of 4.8% . translating such percentages into treatment effects , the net clinical benefit associated with af ablation without interruption of oral anticoagulation is overwhelming ( figure 2 ) , with an estimated 170 thromboembolic or hemorrhagic complications avoided every 1,000 patients ablated . in the most recent survey on af catheter ablation , cappato et al . reported thromboembolic and hemorrhagic complication rates in 16,039 patients undergoing af ablation between 2003 and 2006 in 521 centers distributed worldwide .
all these patients had oral anticoagulant discontinuation before the ablation procedure . accordingly , thromboembolic and bleeding complication rates in this large survey were fairly consistent with that reported in group 1 and 2 patients of our study [ 13 , 18 ] .
based on our findings it is estimable that , if periprocedural oral anticoagulation was not discontinued in all patients included in the survey , more than 2,700 thromboembolic or hemorrhagic complications would have been avoided worldwide from 2003 to 2006 .
therefore , there are cogent data supporting the benefit of af ablation without discontinuation of oral anticoagulation .
since the acute reversal of the anticoagulant effect of warfarin is possible only through infusion of fresh coagulation factors , we routinely type- and cross - match all patients , so that packed red blood cells and fresh frozen plasma are readily available for infusion in case of hemorragic complications . if the preprocedural inr is above 3.5 , we partially reverse the anticoagulant effect with one to two units of fresh frozen plasma . of interest ,
the strategy of ablation under therapeutic inr could also be more cost - effective compared to bridging therapy with enoxaparin , which is expensive and may be inconvenient for many patients .
radiofrequency catheter ablation of af without discontinuation of oral anticoagulation significantly reduces the risk of thromboembolic and minor bleeding complications .
maintenance of periprocedural therapeutic inr should be considered the anticoagulation strategy of choice among patients submitted to catheter ablation of af . | atrial fibrillation
( af ) can be cured by pulmonary vein antrum
isolation ( pvai ) in a substantial proportion of
patients .
the high efficacy of pvai is partially
undermined by a small but concrete
periprocedural risk of complications , such as
thromboembolic events and bleeding .
a correct
management of anticoagulation is essential to
prevent such complications .
performing pvai
without interruption of oral anticoagulation has
been demonstrated feasible by our group in
previous studies .
recently , we reported that
continuation of therapeutic warfarin during
radiofrequency catheter ablation consistently
reduces the risk of periprocedural
stroke / transient ischemic attack without
increasing the risk of hemorrhagic events . of
note , interrupting warfarin anticoagulation may
actually increase the risk of stroke even when
bridged with heparin .
the latter strategy is
also associated with an increased risk of minor
bleeding . with regard to major bleeding , we
found no significant difference between patients
with a therapeutic inr and those who were
bridged with heparin .
therefore , continuation of
therapeutic warfarin during ablation of af
appears to be the best anticoagulation strategy .
in this paper
we summarize our experience with
af ablation without interruption of
anticoagulation . |
breast cancer is the most frequently diagnosed cancer and the leading cause of cancer death in women worldwide , accounting for 23% ( 1.38 million ) of the total new cancer cases and 14% ( 458,400 ) of the total cancer deaths in 2008 . about half of the breast cancer cases and 60% of the deaths are estimated to occur in economically developing countries . in iran ,
also , patients with advanced stages of the disease are relatively younger ( about 10 years ) than their western counterparts [ 2 , 3 ] .
exposure to environmental pollutants such as metals including cadmium , chromium , nickel , and arsenic is classified in group 1 of the international agency for research on cancer categories of carcinogen ; it also reports lead as a suspected human carcinogen ( group 2a ) and also mercury as possibly carcinogenic to humans ( group 2b ) . in order to explain the role of metals in breast cancer incidence
, we should refer to the studies in the field of estrogenicity of metals that express estrogen - like activity in breast cancer cells and suggest several pathways to explain association of metals with human cancer [ 711 ] . on the other hand , cadmium , lead , and mercury as carcinogens belong to the group of selenium , the antagonistic elements that compete with selenium uptake as anticarcinogen .
the mechanism of se as an anticarcinogenic element is unknown , but several speculative hypotheses have been advanced .
se exerts its essential role in the formation of glutathione peroxidase , a selenoenzyme that protects body against oxidative injury and free radical damage so its suggested mechanism for cancer prevention includes effects upon programmed cell death , dna repair , carcinogen metabolism , and the immune system [ 1417 ] .
therefore , it seems that , according to the results of these researches , a considerable amount of literature has been published on the determination of metals in human breast cancer tissues which show various values in malignant and benign tissues in comparison with healthy tissues [ 1728 ] . in addition , all available studies have only focused on metal concentration totally in breast tissue , but the aim of this study is to determine cd , pb , hg , and se in various parts of breast tissue ( tegmen , tumor tissue , tumor adiposity , and tegmen adiposity ) in order to compare any significant differences which may exist between different parts of breast tissues .
also , this research may be the first report of metal concentration in iranian breast cancer tissues .
in order to determine cd , pb , hg , and se concentrations in different parts of malignant breast tissue , 14 removed samples by mastectomy surgery of women patients ( in age range 3050 ) were taken from imam hospitals located in uremia and separate tegmen , tumor , tumor adiposity , and tegmen adiposity of breast cancerous tissues were dried in freeze - dryer at 64c for 20 to 30 hours .
then , about 1 g of each part of separated breast tissue was put in the digestion tubes ( polytetrafluoro ethylene ) with 5 ml concentrated hno3 for 3 hours in 100c on hot block digester until the disappearance of brown fumes .
after cooling , 1 to 3 ml h2o2 30% for 1 hour on heater was added .
after cooling , it was filtered with whatman filter paper number 1 and then diluted with deionized water to final volume of 25 ml [ 26 , 30 ] .
pb and cd measurements were performed by using aas and with the graphite furnace technique model aa-670 and se analysis was done by icp - oes ( ultima 2ce ) .
the aas instrument was calibrated using aqueous standards of 10 , 30 , and 60 g / kg for pb and 0.5 , 2 , and 5 g / kg for cd in breast cancer tissues .
there was a good linear relation between absorbance and standard concentrations of pb and cd .
linearity was evaluated by calculating the r - square value , which was 0.998 for pb and 0.999 for cd .
the detection limit with aas was calculated as 3 times the sd of the blank sample divided by the slope of calibration curves ( 1.57 g / kg for pb and 0.18 g / kg for cd ) .
the detection limit of se with icp - oes was 0.2 g / kg .
about 0.020.04 g of dried samples was put on nickel boot for measuring hg by leco ama 254 advanced mercury analyzer ( usa ) . to evaluate the analytical potency of the proposed methodology , accuracy of total hg analysis
was checked by running three samples of standard reference materials ( srm ) , national institute of standards and technology ( nist ) , srm 1633b , srm 2709 , and srm 2711 in seven replications .
the detection limit of the used instrument was 1 g / kg of dry weight . to check for contamination ,
all of used glassware was acid - washed and one blank was analyzed after five samples .
table 1 presents mean , standard deviation , and range of pb , cd , hg , and se concentration in 4 parts of tegmen , tumor , tumor adiposity , and tegmen adiposity of 14 breast cancer tissues . as it can be seen in the table ,
the concentration data of pb in kolmogorov - smirnov test was not normal ( p < 0.05 ) , so by using the kruskal - wallis nonparametric test , no significant differences between pb values in parts of the breast tissues were shown ( p = 0.820 ) . to assay the differences between cd , se , and hg ,
one - way anova test was used and no significant differences were found between metals in the separated breast cancer tissues ( resp . ,
no significant difference between the concentrations of metals in four separate parts might be due to the very close blood relationship between tissues . a noticeable point in this research
is that the preparation of healthy tissues as control samples for comparing cancer tissues was not possible , so we compared our results with previous studies . despite the possible relation between cd and breast cancer exposure ,
its values in different parts of 14 breast cancer samples show mean concentration in tegmen ( 35.51 g / kg ) , tumor ( 45.04 g / kg ) , tumor adiposity ( 41.15 g / kg ) , and tegmen adiposity ( 32.95 g / kg ) .
table 2 compares cd , pb , and se concentration in breast tissues from results of available studies .
, so there were not any significant differences between cd in cancerous ( 20470 g / kg ) and healthy ( 31700 g / kg ) ones and the maximum concentration of cd among hitherto accessible studies has been reported .
the separation of the close parts of breast tissues in this study did not show any significant difference between them and , as it can be inferred from the table , the present results are in accordance with the results of strumylaite et al . that report the minimum cd values so far .
due to the multiple carcinogenic evidence of lead , its detection has been done in various tumors such as breast ones . according to table 2 , the considerable concentrations in tegmen ( 336.18 g / kg ) , tumor ( 327.50 g / kg ) , tegmen adiposity ( 396.52 g / kg ) , and tumor adiposity ( 365.73 g / kg ) are similar to the results of majewska et al . and kubala - kuku et al . .
also mercuric chloride has been widely considered as causative of tumors . among collected studies ,
rizk and sky - peck report the mean concentration of hg ( 770 g / kg dry weight ) in breast cancer tissues and , according to the results of this study , it is supposed that the accumulation of this metal in tegmen ( 29.10 g / kg ) , tumor ( 33.26 g / kg ) , tumor adiposity ( 28.04 g / kg ) , and tegmen adiposity ( 26.13 g / kg ) can have a role in carcinogenicity . as it can be seen in table 2 , according to the studies , the accumulation of se in breast cancer tissues has been determined .
the mean concentration of se is similar to the results of rizk and sky - peck , kue et al .
the noticeable issue is that metals are just one of the effective factors in carcinogenesis or anticarcinogenesis , so their clear mechanism would have been investigated .
this study just reports the concentration of some metals in breast cancer in women 's samples from selective hospital , as we know that different factors may affect the occurrence of cancer especially breast cancer in women all over the world and environmental pollutants such as metals from several sources could enter the human body and , by accumulation , increase , and intensification , they may cause the incidence of cancer .
this study showed that there were not any significant differences between metals concentration in different parts of breast cancer tissues .
this result might be because of close relation of separated parts of fatty breast organ .
in general , in puberty and presence of estrogen hormone , breast cells have been grown rapidly .
in normal situation , after sudden increased rate of estrogen and breast cell growth , hormone balance became in equilibrium and irregular cell proliferation was interrupted .
according to recent researches and hypotheses , it could be concluded that estrogen - like properties of metals could mostly influence hormonal responses by binding to estrogen receptors and disrupt endocrine system and finally increased proliferation of cells would be occurring .
we conclude , thus , that a high level of these heavy metals is accumulated in iranian cancerous breasts and their presence can be one of the reasons for breast cancer appearance . | breast cancer is the major cause of cancer morbidity and mortality between women in the world .
metals involved in environmental toxicology are closely related to tumor growth and cancer . on the other hand ,
some metals such as selenium have anticarcinogenic properties .
the aim of this study is to determine the concentration of cadmium , lead , mercury , and selenium in separated parts of tegmen , tumor , tumor adiposity , and tegmen adiposity of 14 breast cancer tissues which have been analyzed by graphite furnace atomic absorption ( aa-670 ) and icp - oes ( ultima 2ce ) .
our results show that se and hg have maximum and minimum concentration , respectively .
statistical analysis reveals no significant differences between metal accumulations in different parts of cancer tissues ( p > 0.05 ) and this observation might be due to the close relation of separated parts of fatty breast organ .
thus , we could conclude that a high level of these heavy metals is accumulated in iranian cancerous breasts and their presence can be one of the reasons of cancer appearance . |
in the neonatal intensive care unit ( nicu ) population , hyponatremia is the most frequent encountered water and salt abnormality . with its broad differential diagnosis ,
the most frequent causes are renal salt loosing through an immature kidney and the use of drugs such as diuretics .
hypertension , however , is a relatively rare feature in children , especially in neonates , with an incidence in nicu 's ranging from 0.7% to 3.2% , and renal arterial thrombosis following umbilical arterial catheterization as the leading cause [ 2 , 3 ] .
a rare clinical presentation of unilateral renal arterial stenosis is the hyponatremic hypertensive syndrome ( hhs ) , characterized by activation of the renin angiotensin aldosterone ( raas ) system in the ischemic kidney , causing hypertension , and a counteracting effect on the other kidney , by different mechanisms leading to volume depletion and loss of electrolytes .
this syndrome is caused by unilateral renal ischemia , due to stenosis or occlusion of a ( branch of a ) renal artery , and also occurs in a variety of other underlying disorders .
so far , only a few reports of hhs in children are available , with polydipsia , polyuria , enuresis , weight loss , volume depletion , and various neurological and behavioural symptoms as presenting symptoms .
we present a case of hhs in a preterm infant , with an extremely low sodium concentration , and discuss the difficulties encountered in treatment and the irreversible neurological sequels due to this potentially life - threatening metabolic disturbance .
a preterm boy presented with extreme hyponatremia ( plasma sodium of 101 mmol / l ) at the 20th day after birth .
he was born from a nulliparous woman at a gestational age of 31 weeks and 4 days after an uncomplicated pregnancy , followed by spontaneous rupture of membranes and antenatal corticosteroid administration .
apgar scores were 9 and 10 at 1 and 5 minutes , and the birth weight was 2080 grams .
an umbilical arterial catheter was inserted directly after birth , for the purpose of blood pressure monitoring , and removed after 3 days .
furthermore an umbilical venous catheter and subsequently a peripheral central venous catheter were inserted for the purpose of parenteral feeding .
routine cerebral ultrasonography showed an image consistent with the gestational age and mild periventricular flaring .
he gained weight ( from 1900 grams at the 3rd day to 2100 grams at two weeks after birth ) . at the age of 3 weeks rejection to feeding ( until this moment consisting of 150 ml / kg / day breast milk with breast milk fortifier ) , weight loss ( to a minimum of 1960 grams ) , irritability , hyperthermia , and
cerebrospinal fluid analysis showed 219 leukocytes / mm with 12000 erythrocytes / mm , after a traumatic lumbar puncture , thus a meningitis could no be excluded and intravenous antibiotics were started .
intravenous fluids , with a total volume of 150 ml / kg / day , containing 8 mg / kg / min glucose and 5 mmol / kg / day sodium , were administered in the regional hospital for 2 days ( before return to the nicu ) .
the plasma sodium level had declined , from 140 mmol / l nine days before , to 101 mmol / l .
there were no sodium levels examined in the interval between , but the level at the onset of symptoms was established at 112
the boy returned to the nicu under suspicion of a syndrome of inappropriate antidiuretic hormone secretion ( siadh ) associated with the assumed meningitis , with initiated fluid restriction and sodium supplementation considered to be the appropriate therapy .
we saw a pale , irritated neonate with tachypnea , arterial hypertension ( 104/60 mmhg , mean 78 mmhg ) , opisthotonus , and abnormal synchronized extensions of arms and legs .
mmol / l , laboratory analysis revealed a mild hypokalemia , hypochloremia , and hypomagnesemia , with normal calcium and phosphate levels ( for detailed information on all important laboratory results , see table 1 ) .
plasma osmolarity was 219 mosmol / kg , and urea and creatinine levels were normal .
infection parameters were low , but liver enzymes and lactate were elevated ( in the blood drawn several shortly after the seizure ) .
furthermore an elevated plasma b - type natriuretic peptide ( bnp ) of 1228 pmol / l was found ( in children , there are no validated data on normal values available ) .
urinalysis showed no leucocytes , mild hematuria , low sodium and potassium , with proteinuria , glucosuria , and a urine osmolarity of 129 mosmol / kg .
the combination of hyponatremia and hypertension ( defined as a mean blood pressure of > 2 standard deviations for age and weight , in this case > 75 mmhg ) was suggestive of renal pathology .
abdominal doppler ultrasound showed a right renal arterial thrombosis , partially calcified , and an oedematous appearance of the left kidney .
it was suggested that the symptoms of this neonate resulted from an hhs secondary to a renal arterial thrombosis .
blood pressure levels further increased in the first hours to a maximum of 108/62 ( mean 96 ) mmhg .
we chose to carefully normalise the blood pressure with intravenous dihydralazine , causing the right kidney to become completely afunctional ( as demonstrated with 99 m technetium mag3 renography in combination with the findings on doppler ultrasound as mentioned before ) .
plasma sodium levels rose to , relatively fast within the first hours , above 120 mmol / l , and more slowly within the next 24 hours to normal .
the dihydralazine and sodium supplementation could be gradually stopped after a few days , after which a normal blood pressure and electrolyte levels were maintained without any additional therapy , also urinalysis returned to normal . within a few minutes after return to the nicu the boy developed convulsions ,
cerebral ultrasound and magnetic resonance imaging ( mri ) showed extensive white matter abnormalities and the presence of a sinus thrombosis of the superior sagittal , straight , and transverse sinus .
additional genetic screening revealed a mutation in the methylenetetra - hydrofolate reductase ( mthfr ) gene .
follow - up mri at one and two months of age showed extension of the white matter abnormalities , with secondary haemorrhage , vacuolisation , and cyst formation .
the gyration and myelinisation had increased , there were no signs of new ischemia , and all sinuses were recanalized .
the child showed abnormal neurological behaviour ( agitation , uncontrolled movements , and delayed motor development ) at three months of followup .
the parents of this child gave their informed consent to publication of this case report .
in this case report , we describe a 3-week - old preterm boy , with extreme hyponatremia , hypertension and a dramatic neurological outcome , as a result of hhs following umbilical arterial catheterization .
the typical combination of symptoms in hhs was first described in 1952 in adults , and the term hhs was established by brown et al . in 1965 .
the syndrome is not encountered frequently and in neonates it is even more rare , with renal arterial stenosis following umbilical arterial catheterization being one of the described causes [ 810 ] , accompanied by renal microthrombi in sepsis and an association with dexamethasone use .
the syndrome has been described more often in preterm than in term infants [ 811 ] and sometimes showed a lethal course [ 12 , 13 ] .
the high incidence of hyponatremia ( 30% ) reported in hypertensive neonates , suggests that hhs is probably more common than we think .
hhs is thought to be due to a complex interplay of different mechanisms , with unilateral renal hypoperfusion and a counteracting effect of the contralateral normal kidney as major hallmarks ( two - kidney - one - clip hypertension ) ( figure 1 ) .
the renal arterial thrombosis causes hypoperfusion of one kidney , which activates the raas system to cause hypertension .
the contralateral nonstenotic kidney reacts to this hypertension by excreting water and sodium ( pressure diuresis and natriuresis ) [ 12 , 15 ] .
the hypertension additionally stimulates the cardiac atrial natriuretic peptide ( anp ) and bnp to excrete more sodium and protein .
the resulting hypovolemia , probably together with an increased production of angiotensin ii , stimulates antidiuretic hormone ( adh ) , further aggravating the hyponatremia .
furthermore , aldosterone causes renal potassium loss , which in turn stimulates renin secretion , causing a vicious circle .
proteinuria , glucosuria , and hypercalciuria can also be present due to glomerular hyperfiltration in hypertension , increased renin activity , and probably even more extensive tubulointerstitial involvement .
in our case , the presence of an umbilical arterial catheter , and the finding of a mutation in the mthfr gene ( suggested to play role in homocysteine metabolism and enhancing atherosclerosis ) were thought to be contributing factors in the development of the renal arterial thrombosis . at the moment of presentation at the nicu
, our patient was thought to be already beyond the natriuretic phase of hhs ( probably the severe hyponatremia finally resulted in sodium retention , explaining the low urine sodium ) and most likely adh had been turned on in reaction to the volume depletion .
the initial sepsis - like presentation was retrospectively interpreted as the result of a combination of hypovolemia and central nervous system disturbances .
the prematurity was probably a major contributing factor leading to the severe outcome in this patient .
this could have led to more extreme hyponatremia because preterms have a relatively low sodium intake , reduced tubular sodium reabsorption , and decreased glomerular filtration rate , impairing free water excretion .
furthermore , recognizing the nonspecific clinical symptoms of hhs can be very difficult in a neonate .
the symptoms of hhs disappeared after normalising the blood pressure with a vasodilator agent ( dihydralazine ) , causing the ischemic kidney to become nonvital by totally abrogating the blood flow , destroying the juxtaglomerular cells , and hence stopping renin production .
there was a gradual decline in blood pressure in this patient , different from the potentially dangerous fall which can be seen with angiotensin - converting enzyme ( ace ) inhibitors . with severe volume depletion ,
cautious repletion is needed , which can probably also reduce arterial pressure by suppression of raas .
correction of longstanding hyponatremia should be managed carefully , to minimise the risk of developing cerebral shrinking .
final therapy of the underlying renal arterial stenosis was not necessary in this case , but can be achieved by balloon angioplasty , renal artery stenting , or uninephrectomy .
the remarkable irreversible neurological features in this case are most likely to be the consecutive effect of a hypertensive and hyponatremic encephalopathy , aggravated by a diminished cerebral circulation due to hypovolemia and a sinus thrombosis .
furthermore the convulsions could also have led to irreversible damage to the vulnerable preterm brain .
previous case reports in older children mainly mention reversible neurological symptoms , and even reversible findings on computer tomography or mri associated with hhs [ 19 , 2224 ] and only one infant dying from massive cerebral haemorrhage .
first , there is a lack of ( clinical and laboratory ) information about the period before the patient represented with the severe hyponatremia .
unfortunately , no detailed information on water balance or 24-hour urine volumes during the period in which the patient developed the hyponatremia was available .
as it is especially cumbersome to collect such data in newborns , this is very rarely done .
second , no data on plasma adh , rennin , and aldosterone are available to confirm our suggested diagnosis .
this case was meant to describe the complex pathophysiology of hhs , and the possible misleading clinical features in a neonate .
furthermore we want to underline the risk of severe irreversible neurological damage when there is a diagnostic delay .
we think that in evaluating a neonate with severe hyponatremia , hhs should be considered , especially if following umbilical arterial catheterization . | objective . to report the irreversible severe neurological symptoms following the hyponatremic hypertensive syndrome ( hhs ) in an infant after umbilical arterial catheterization
. design .
case report with review of the literature . setting . neonatal intensive care unit at a tertiary care children 's hospital . patient .
a three - week - old preterm infant . conclusions . in evaluating a neonate with hyponatremia and hypertension , hhs should be considered , especially in case of umbilical arterial catheterization . in case of diagnostic delay
, there is a risk of severe irreversible neurological damage . |
the first requirements of the no child left behind act ( nclb ) , which became law in 2002 , led every state to evaluate the adequate yearly progress ( ayp ) of students in math and language arts based on specific content standards in those subjects .
assessment requirements extended ( by statute , if not yet in practice ) to science in 2007 , where they encompass testing in three grade bands , 35 , 69 , and 1012 . in science , guidance for the production of content standards and curricula has been available through the national science education standards ( national research council [ nrc ] , 1996 ) and project 2061 benchmarks for science literacy ( american association for the advancement of science [ aaas ] , 1993 , 2009 ) , and through state and district administrative offices .
the standards governing ayp in all subjects were developed on a state - by - state basis because , until recently at least , there has been little support at the state level for common national standards .
most states have adopted common core standards for reading and math , although tremendous work remains to be done to implement these standards ( paulson , 2010 ; for current list , see http://www.corestandards.org/in-the-states ) .
the rationale behind standards and testing can be summarized by a familiar saying , invoked in the context of nclb by former u.s . secretary of education margaret spellings : what gets measured , gets done ( spellings , 2005 ) . in k12 education , this can be further translated as standards drive testing and testing drives curriculum ( popham , 2004 ) .
there is some evidence that standards can also drive achievement , although here the details of structure and implementation are more critical ( swanson , 2006 ) .
we make no argument in favor of or against standards - based instruction or nclb ; however , given that this is the reality of k12 public education in the united states at the current time , we do argue that understanding the quality of standards is useful and even necessary .
science will soon follow reading and math down the path of high - stakes outcomes and the drive to achieve ayp through curriculum , instruction , and testing
and so it is reasonable to examine science standards for quality and consistency across different states .
this is especially true in light of concerns that variations in defining proficiency allow states to manipulate standards to facilitate meeting ayp and to avoid sanctions ( kingsbury et al . , 2003 ; aspen institute , 2006 ) .
in fact , evidence from the national center for educational statistics ( nces ) demonstrates an incongruity between student proficiency at the state level and student performance on the gold standard of assessment , the national assessment of educational progress ( naep ) : there is a strong negative correlation between the proportions of students meeting the states proficiency standards and the naep score equivalents to those standards , suggesting that the observed heterogeneity in states reported percents proficient can be largely attributed to differences in the stringency of their standards ( nces , 2007 ) .
such findings suggest worrisome weaknesses in math and reading standards and reason for concern apropos science standards . to date , there have been few comprehensive evaluations of specific science content in state standards , and none by a professional scientific society , and their quality remains largely unknown ( gross et al . , 2005 ; hoffman and barstow , 2007 ) . in genetics ,
lists of core principles have been developed to guide learning for audiences ranging from undergraduate nonscience majors to health professionals ( hott et al .
, 2002 ; national coalition for health professional education in genetics [ nchpeg ] , 2004 ; smith et al . , 2008 ) ,
the american society of human genetics ( ashg ) has been interested in genetics education and k12 standards since it began documenting misconceptions that frequently appeared in essays submitted to its national dna day essay contest ( shaw et al . ,
2008 ) . in this paper , we present the results of an evaluation of the life sciences standards of every state to assess their adequacy with respect to genetics coverage .
as benchmarks , we developed and used a list of core genetics concepts that all students should understand by grade 12 as preparation for life in a world of healthcare and medicine that is increasingly informed by genetics .
this analysis of extant genetics standards hopefully can inform science education researchers and policy makers during the normal revision process for existing standards , which occurs at different intervals for different states , and during the development of common science standards .
a list of core concepts in genetics appropriate for students up to grade 12 was developed using an iterative process .
initial drafts of concepts were adapted by ashg staff from several previously published documents that included important or essential genetics content : development and evaluation of a genetics literacy assessment for undergraduates
( bowling et al . , 2008 ) ; the genetics concept assessment ( smith et al . , 2008 ) ; closing the gap ( dougherty , 2009 ) ; nchpeg core principles of genetics ( 2004 ) ; project 2061 benchmarks for science literacy ( aaas , 2009 ) ; national science education standards ( nrc , 1996 ) ; ap biology course description ( college board , 2010 ) ; and undergraduate nonmajor biology courses ( hott et al . , 2002 ) . using these documents as a baseline
, concepts were developed to adequately describe five major conceptual areas of genetics deemed essential to genetic literacy : nature of the genetic material , transmission genetics / patterns of inheritance , gene expression and regulation , genetic variation , and evolution . because our interest was the quality and comprehensiveness of genetics content in the state standards
, we did not develop benchmarks ( or analyze standards ) related to noncontent understandings or abilities , such as inquiry , the nature of science , or system standards .
advanced genetics topics , such as epigenetics , the regulatory roles of small rnas , chromatin remodeling , and others , also were not included on the grounds that : 1 ) they were too complex or detailed and basic genetic literacy did not require them of all students ; 2 ) many teachers would likely be unfamiliar with them ; and 3 ) most state standards would not be expected to include them .
initial drafts were reviewed , revised through 11 iterations , and ultimately approved by ashg 's information and education committee , a standing committee that advises ashg 's board of directors on issues related to genetics education .
its members have expertise in genetics content , science education , and teaching , including substantial expertise with high school curricula and pedagogy .
a total of 15 concepts were tested in a pilot analysis , and a final list of 19 concepts served as the benchmark concepts against which state science standards were compared ( table 1 ) .
table 1.scoring results for ashg 's genetics concepts averaged across all states by concept and by concept categorycore conceptaverage score ( all states)/adequacyconcept categoryaverage by category / adequacy1 .
dna is the genetic material for all species of living organisms.1.7/adequatenature of genetic material1.6/adequate2 .
alleles are passed from parent to offspring through the processes of replication , meiosis , and fertilization.1.4/inadequate5 . for traits primarily influenced by single genes , certain combinations of alleles
lead to predictable genotypic and phenotypic patterns of inheritance , illustrating mendel 's principles of segregation of alleles and independent assortment of genes.1.5/adequate6 .
polygenic ( or complex ) traits ( e.g. , height , blood glucose ) often show continuous variation within populations and are less predictable than single - gene traits.0.3/not addressed / absent7 .
this occurs through transcription of dna into rna and translation of mrna into protein.1.3/inadequategene expression and regulation0.9/inadequate9 .
different genes are turned on and off at specific times to form different types of cells and to influence the way different cells function.0.6/inadequate11 .
the functions of genes and their products can be affected by the environment and other genes at one or many steps involved in producing a trait.0.7/inadequate12 .
they can occur spontaneously during dna replication or they can be the result of damage by environmental factors.1.3/inadequategenetic variation1.1/inadequate13 .
mutations in dna , and sorting and recombination during meiosis , result in genetic variation.1.4/inadequate14 .
mutations may help , harm , or have little or no effect on an organism.1.2/inadequate16 .
some genetic diseases are inherited ( e.g. , tay - sachs ) , and others develop during life ( e.g. , cancer).0.8/inadequate17 . genetic variation and the phenotypic variation it leads to are the basis for evolution.1.5/adequateevolution1.5/adequate18 .
evolution by natural selection is a process by which inherited traits influence how likely an organism is to survive , reproduce , and pass those traits to its offspring.1.7/adequate19 .
the process of evolution occurs at a population level ( e.g. , not at the level of individual organisms ) , and takes place over generations ( e.g. , not within an individual organism 's lifespan).1.2/inadequatenumerical scores : 0 = not addressed / absent ; 1 = inadequately addressed ; 2 = adequate .
scoring results for ashg 's genetics concepts averaged across all states by concept and by concept category numerical scores : 0 = not addressed / absent ; 1 = inadequately addressed ; 2 = adequate .
the most current state standards as of summer 2009 were identified from the websites of the departments of education for all 50 states and the district of columbia .
standards related to genetics in any way ( i.e. , in all life or biological sciences categories ) from grades 912 were assembled in state - specific pdf files to aid reviewers .
where states used more than one section to group content , for example , introductory paragraphs or rubrics to supplement content listings , we tried to be as inclusive as possible .
in general , the only content excluded from consideration related to rare cases where content was explicitly identified as beyond the level at which all students should learn ( i.e. , exceeding the state 's view of essential content and , therefore , less likely to be taught or tested ) .
a simple online scoring guide was developed in - house to allow reviewers to open an appropriate pdf of the state 's standards , review them , and then use drop - down menus to record a score for each concept .
a box for written comments was also provided . in light of ongoing efforts to revise national standards documents , we chose not to analyze standards that appear likely to soon be replaced ( e.g. , the national science education standards or project 2061 benchmarks for science literacy ) .
a pilot test was conducted to evaluate the feasibility and reliability of our rating system .
we used a three - level grading system ( 0 = not addressed / absent , 1 = inadequate , 2 = adequate ) that was similar to methods used for evaluating earth science standards ( hoffman and barstow , 2007 ) .
two project staff members serving as pilot reviewers independently evaluated the standards of 10 randomly selected states against a draft set of ashg core genetics concepts that included 15 concepts ( n = 150 pairwise comparisons ) .
interrater reliability for averages across all concepts at a state - by - state level was calculated according to the methods of ebel ( 1951 ) , using the open - access calculator developed by solomon ( 2004 ) .
adequacy grades were assigned by converting state or concept averages according to : adequate = 1.52.0 ; inadequate = 0.61.4 ; not addressed / absent < 0.6 . to assemble the requisite experts to serve as evaluators of state standards , we recruited 167 members of ashg 's genetics education outreach network , a volunteer network of geneticists across the nation who work in k12 classrooms , help grade dna day essays , and/or provide career talks .
all have content expertise in genetics and varying levels of expertise with genetics education outreach . of those participating
, 77% held doctoral degrees ( mostly phd and md ) , 20% held master 's degrees , and just over 2% held bachelor 's degrees .
( ashg 's membership consists largely of academic basic researchers , clinical researchers , and genetic counselors . ) our rationale for using a large pool of experts external to the project team as reviewers was that the intent of a state 's standards apropos a specific concept should be clear . if a specific concept in the standards is not apparent to these experts , it likely will not be apparent to many curriculum developers and teachers .
using our criteria , such standards would be graded as either inadequate ( i.e. , present but not adequate ) or not addressed / absent . as hoffman and barstow ( 2007 )
have noted apropos standards , each state has its own development processes , generally involving a wide variety of people with different backgrounds , subject area expertise , teaching experience , special interests , and political agendas .
such variation is likely the case for those interpreting standards as well , thus arguing in favor of clarity .
in addition , by using a large number of external reviewers , we hoped to avoid the effects of systematic bias that may creep into analyses conducted by small , close - knit teams .
the evaluation and grading of standards is necessarily a subjective process , but by averaging across a number of independent judges , we believe a reasonably accurate assessment was attained .
just over half ( 94 ) of the evaluators reviewed two states , and the remainder ( 73 ) reviewed one state , with sufficient overlap to yield an average of 5.11 reviewers per state .
, we dropped the high and low scores from states having five or six reviewers .
thus , the analysis for all states was based on a minimum of three scores per concept ( four for 14 states ) .
averages by state across all concepts ( i.e. , per - state averages ) and averages by concept across all states ( i.e. , per - concept averages ) were calculated .
interrater reliabilities were evaluated by the methods of ebel and solomon , as described above . to establish a common reference frame for reviewers to use in their evaluation ( acknowledging both the vagaries of the state standards and likely differing interpretations ) ,
the reviewers were instructed by email as follows : please use your best judgment to match concepts with standards and do not assume too much .
an incomplete standard that evokes for you ( as an expert in genetics ) a natural elaboration encompassing the ashg core concept may not evoke that same connection for a non - expert high school teacher .
we need to evaluate what each standard actually says . at the same time , if the intent of a standard clearly matches the intent of an ashg core concept and differs only in wording , you should credit that standard appropriately .
a list of core concepts in genetics appropriate for students up to grade 12 was developed using an iterative process .
initial drafts of concepts were adapted by ashg staff from several previously published documents that included important or essential genetics content : development and evaluation of a genetics literacy assessment for undergraduates
( bowling et al . , 2008 ) ; the genetics concept assessment ( smith et al . , 2008 ) ; closing the gap ( dougherty , 2009 ) ; nchpeg core principles of genetics ( 2004 ) ; project 2061 benchmarks for science literacy ( aaas , 2009 ) ; national science education standards ( nrc , 1996 ) ; ap biology course description ( college board , 2010 ) ; and undergraduate nonmajor biology courses ( hott et al . , 2002 ) . using these documents as a baseline
, concepts were developed to adequately describe five major conceptual areas of genetics deemed essential to genetic literacy : nature of the genetic material , transmission genetics / patterns of inheritance , gene expression and regulation , genetic variation , and evolution . because our interest was the quality and comprehensiveness of genetics content in the state standards
, we did not develop benchmarks ( or analyze standards ) related to noncontent understandings or abilities , such as inquiry , the nature of science , or system standards .
advanced genetics topics , such as epigenetics , the regulatory roles of small rnas , chromatin remodeling , and others , also were not included on the grounds that : 1 ) they were too complex or detailed and basic genetic literacy did not require them of all students ; 2 ) many teachers would likely be unfamiliar with them ; and 3 ) most state standards would not be expected to include them .
initial drafts were reviewed , revised through 11 iterations , and ultimately approved by ashg 's information and education committee , a standing committee that advises ashg 's board of directors on issues related to genetics education .
its members have expertise in genetics content , science education , and teaching , including substantial expertise with high school curricula and pedagogy .
a total of 15 concepts were tested in a pilot analysis , and a final list of 19 concepts served as the benchmark concepts against which state science standards were compared ( table 1 ) .
table 1.scoring results for ashg 's genetics concepts averaged across all states by concept and by concept categorycore conceptaverage score ( all states)/adequacyconcept categoryaverage by category / adequacy1 .
dna is the genetic material for all species of living organisms.1.7/adequatenature of genetic material1.6/adequate2 .
alleles are passed from parent to offspring through the processes of replication , meiosis , and fertilization.1.4/inadequate5 . for traits primarily influenced by single genes , certain combinations of alleles
lead to predictable genotypic and phenotypic patterns of inheritance , illustrating mendel 's principles of segregation of alleles and independent assortment of genes.1.5/adequate6 .
polygenic ( or complex ) traits ( e.g. , height , blood glucose ) often show continuous variation within populations and are less predictable than single - gene traits.0.3/not addressed / absent7 .
this occurs through transcription of dna into rna and translation of mrna into protein.1.3/inadequategene expression and regulation0.9/inadequate9 .
different genes are turned on and off at specific times to form different types of cells and to influence the way different cells function.0.6/inadequate11 .
the functions of genes and their products can be affected by the environment and other genes at one or many steps involved in producing a trait.0.7/inadequate12 .
they can occur spontaneously during dna replication or they can be the result of damage by environmental factors.1.3/inadequategenetic variation1.1/inadequate13 .
mutations in dna , and sorting and recombination during meiosis , result in genetic variation.1.4/inadequate14 .
mutations may help , harm , or have little or no effect on an organism.1.2/inadequate16 .
some genetic diseases are inherited ( e.g. , tay - sachs ) , and others develop during life ( e.g. , cancer).0.8/inadequate17 . genetic variation and the phenotypic variation it leads to are the basis for evolution.1.5/adequateevolution1.5/adequate18 .
evolution by natural selection is a process by which inherited traits influence how likely an organism is to survive , reproduce , and pass those traits to its offspring.1.7/adequate19 .
the process of evolution occurs at a population level ( e.g. , not at the level of individual organisms ) , and takes place over generations ( e.g. , not within an individual organism 's lifespan).1.2/inadequatenumerical scores : 0 = not addressed / absent ; 1 = inadequately addressed ; 2 = adequate .
scoring results for ashg 's genetics concepts averaged across all states by concept and by concept category numerical scores : 0 = not addressed / absent ; 1 = inadequately addressed ; 2 = adequate .
the most current state standards as of summer 2009 were identified from the websites of the departments of education for all 50 states and the district of columbia .
standards related to genetics in any way ( i.e. , in all life or biological sciences categories ) from grades 912 were assembled in state - specific pdf files to aid reviewers .
where states used more than one section to group content , for example , introductory paragraphs or rubrics to supplement content listings , we tried to be as inclusive as possible .
in general , the only content excluded from consideration related to rare cases where content was explicitly identified as beyond the level at which all students should learn ( i.e. , exceeding the state 's view of essential content and , therefore , less likely to be taught or tested ) . a simple online scoring guide was developed in - house to allow reviewers to open an appropriate pdf of the state 's standards , review them , and then use drop - down menus to record a score for each concept .
a box for written comments was also provided . in light of ongoing efforts to revise national standards documents
, we chose not to analyze standards that appear likely to soon be replaced ( e.g. , the national science education standards or project 2061 benchmarks for science literacy ) .
a pilot test was conducted to evaluate the feasibility and reliability of our rating system .
we used a three - level grading system ( 0 = not addressed / absent , 1 = inadequate , 2 = adequate ) that was similar to methods used for evaluating earth science standards ( hoffman and barstow , 2007 ) .
two project staff members serving as pilot reviewers independently evaluated the standards of 10 randomly selected states against a draft set of ashg core genetics concepts that included 15 concepts ( n = 150 pairwise comparisons ) .
interrater reliability for averages across all concepts at a state - by - state level was calculated according to the methods of ebel ( 1951 ) , using the open - access calculator developed by solomon ( 2004 ) .
adequacy grades were assigned by converting state or concept averages according to : adequate = 1.52.0 ; inadequate = 0.61.4 ; not addressed / absent < 0.6 .
to assemble the requisite experts to serve as evaluators of state standards , we recruited 167 members of ashg 's genetics education outreach network , a volunteer network of geneticists across the nation who work in k12 classrooms , help grade dna day essays , and/or provide career talks .
all have content expertise in genetics and varying levels of expertise with genetics education outreach . of those participating
, 77% held doctoral degrees ( mostly phd and md ) , 20% held master 's degrees , and just over 2% held bachelor 's degrees .
( ashg 's membership consists largely of academic basic researchers , clinical researchers , and genetic counselors . ) our rationale for using a large pool of experts external to the project team as reviewers was that the intent of a state 's standards apropos a specific concept should be clear . if a specific concept in the standards is not apparent to these experts , it likely will not be apparent to many curriculum developers and teachers .
using our criteria , such standards would be graded as either inadequate ( i.e. , present but not adequate ) or not addressed / absent . as hoffman and barstow ( 2007 )
have noted apropos standards , each state has its own development processes , generally involving a wide variety of people with different backgrounds , subject area expertise , teaching experience , special interests , and political agendas .
such variation is likely the case for those interpreting standards as well , thus arguing in favor of clarity .
in addition , by using a large number of external reviewers , we hoped to avoid the effects of systematic bias that may creep into analyses conducted by small , close - knit teams .
the evaluation and grading of standards is necessarily a subjective process , but by averaging across a number of independent judges , we believe a reasonably accurate assessment was attained .
just over half ( 94 ) of the evaluators reviewed two states , and the remainder ( 73 ) reviewed one state , with sufficient overlap to yield an average of 5.11 reviewers per state .
, we dropped the high and low scores from states having five or six reviewers .
thus , the analysis for all states was based on a minimum of three scores per concept ( four for 14 states ) .
averages by state across all concepts ( i.e. , per - state averages ) and averages by concept across all states ( i.e. , per - concept averages ) were calculated .
interrater reliabilities were evaluated by the methods of ebel and solomon , as described above . to establish a common reference frame for reviewers to use in their evaluation ( acknowledging both the vagaries of the state standards and likely differing interpretations ) , the reviewers were instructed by email as follows : please use your best judgment to match concepts with standards and do not assume too much . an incomplete standard that evokes for you ( as an expert in genetics ) a natural elaboration encompassing the ashg core concept may not evoke that same connection for a non - expert high school teacher . we need to evaluate what each standard actually says . at the same time , if the intent of a standard clearly matches the intent of an ashg core concept and differs only in wording , you should credit that standard appropriately .
an important statistical measure for assessing the consistency of analysis ( or scoring ) by different reviewers is interrater reliability , which we calculated for the pilot and full evaluations .
values of 70% or higher are generally considered good to acceptable , and values of 80% or higher to be very good ( lombard et al . , 2010 ) .
interrater reliability in the pilot phase ( two reviewers ) , across 150 pairwise comparisons , was 63% at the per - concept level , and in only one instance ( 0.7% ) did the two reviewers differ by 2 grading steps ( i.e. , 0 , not addressed / absent , and 2 , adequate ) .
interrater reliability at the level of per - state averages ( i.e. , average of scores across all 15 concepts used in the pilot ) was 96% .
the difference in reliability at the two levels reflects the fact that states with overall weak ( or strong ) standards tend to have many individually weak ( or strong ) standards , and averaging smooths out differences that may be more apparent at the finer - grained level of individual concepts .
in addition , the very high degree of per - state reliability was not surprising , because the project staff who conducted the pilot analysis had spent considerable time assembling pdfs of the state standards and discussing scoring criteria and the meaning of agreement . also , the pilot reviewers noted that a number of concepts were difficult to align with state standards , because the benchmarking concepts themselves contained more than one distinct idea .
based on that finding , several benchmarking concepts containing more than one idea were divided into simpler , separate concepts , yielding a final draft with 19 concepts ( table 1 ) . at the conclusion of the full evaluation ,
the interrater reliability for the same 10 states that were used in the pilot was 79% .
the interrater reliability across all states analyzed ( i.e. , 50 states plus the district of columbia ) was 75% .
the per - concept reliability average was 71% , with four concepts having a reliability under 60% ( averaging 52% : concepts 1 , 11 , 18 , and 19 in table 1 ) .
however , there was virtually no correlation between interrater reliability ( by concept ) and concept score ( r = 0.16 ; unpublished data ) .
thus , although four concepts were more challenging for reviewers to evaluate in the standards ( i.e. , lower reliability ) , that difficulty did not translate into systematically higher or lower scores .
we first analyzed which genetics concepts were adequately addressed across the nation as a whole .
the 19 core concepts yielded average scores ( i.e. , across all states ) ranging from not addressed / absent ( 2 concepts ) to inadequate ( 12 concepts ) to adequate ( 5 concepts ) , and the five broader genetics categories encompassing those concepts had averages of adequate ( 2 ) and inadequate ( 3 ; see table 1 ) .
overall , average scores for many concepts were low ( e.g. , 7 out of 19 were < 1.0 ) , and the range of scores was broad both across and within categories , especially for transmission / patterns of inheritance , which comprised five concepts ranging from 0.31.5 .
no concept scored above 1.7 , and only five scored 1.5 or higher ( dna being common to all living organisms ; genes as units of information ; mendel 's laws of segregation and independent assortment ; genetic and phenotypic variation as the substrate for evolution ; and evolution by natural selection ) .
the five lowest - scoring concepts ranged from 0.30.8 ( continuous variation ; polygeny ; differential gene expression ; multifactorial causation ; and the connection between mutations and inherited vs. somatic genetic disease ) . the two lowest - scoring concepts ( 6 and 7 , at 0.3essentially not addressed ) are both related to complex traits .
when concepts were analyzed by category , the nature of the genetic material and evolution fared best , scoring adequate , while transmission / patterns of inheritance , gene expression and regulation , and genetic variation all were deemed inadequate .
average scores across all concepts were calculated by state and were normally distributed ( mean = 1.11 , median = 1.11 , and range = 0.511.72 ) . figure 1 shows the results of each state 's overall quality of genetics standards using a map depiction .
seven states ( i.e. , 14% ) had overall genetics standards deemed adequate by the reviewers ( delaware , illinois , kansas , michigan , north carolina , tennessee , and washington ) . among those states , michigan fared best , with an average of 1.72 , followed by delaware at 1.67 .
states judged to have inadequate standards and at the bottom of the ranking included alaska , colorado , idaho , louisiana , north dakota , nebraska , new jersey , oregon ( the only state to have an overall average of not addressed / absent with respect to genetics coverage ) , and wyoming , which all had average scores below 0.8 .
( for individual concept scores for each state , please see supplementary data at : www.ashg.org/education/pdf/stateconceptscores.pdf . )
figure 1.map of the united states summarizing the average quality of genetics standards on a state - by - state basis .
map of the united states summarizing the average quality of genetics standards on a state - by - state basis .
we also examined states for their comprehensiveness with respect to the benchmark genetics concepts that is , the number of individual genetics concepts ( out of the total 19 ) for which a state received scores ranking it as adequate ( independent of overall average ) .
figure 2 shows a map of the specific results by state and table 2 summarizes those results .
delaware and michigan also fared well by this measure , each having 15 concepts that scored as adequate . interestingly , both states scored inadequately on the same four concepts ( 6 , 7 , 10 , and 11 ) , which are the four lowest - scoring concepts overall ( see table 1 ) .
not surprisingly , states with high overall averages tended to have the largest number of concepts that adequately addressed the genetics benchmarks ( r = 0.80 ; unpublished data ) .
for example , kansas had a very high average of 1.6 , but addressed only 11 of 19 concepts adequately .
this is not far from the modal value of 10 adequate concepts across all states .
similarly , texas had a relatively high average score of 1.4 ( just below adequate ) , but had adequate treatment of only seven concepts . in both cases ,
the concepts that were addressed adequately tended to have consistently high ratings from all reviewers . across all states ,
the average number of benchmark concepts covered adequately was 7.4 , with an sd of 4.0 .
figure 2.map of the united states summarizing the comprehensiveness of genetics standards on a state - by - state basis .
the ashg benchmarks list included 19 concepts ( see table 1 ) ; colors indicate the total number of concepts rated by reviewers as adequate .
table 2.summary of comprehensiveness of genetics standards across all states and comparison with an independent evaluation of the overall quality of science standards.number of adequate genetics concepts ( out of 19)number of states ( including district of columbia)fordham scoring , science overallnumber of states15192a7111410b1271016c93616d7<37f15total5150columns 1 and 2 : see table 1 . columns 3 and 4 : data from the thomas b. fordham institute ( gross et al . , 2005 ) .
the fordham analysis used a letter grading system of a f and considered noncontent criteria , such as organization , seriousness , and inquiry . at the time of this study , iowa was not included because it did not have its own state standards .
map of the united states summarizing the comprehensiveness of genetics standards on a state - by - state basis .
the ashg benchmarks list included 19 concepts ( see table 1 ) ; colors indicate the total number of concepts rated by reviewers as adequate .
summary of comprehensiveness of genetics standards across all states and comparison with an independent evaluation of the overall quality of science standards .
columns 3 and 4 : data from the thomas b. fordham institute ( gross et al . , 2005 ) .
the fordham analysis used a letter grading system of a f and considered noncontent criteria , such as organization , seriousness , and inquiry . at the time of this study , iowa was not included because it did not have its own state standards .
three of the four lowest - scoring concepts deal with complex traits ( continuous variation , polygenic inheritance , and multifactorial causation [ concepts 6 , 7 , and 11 , respectively , in table 1 ] ) .
only kansas , north carolina , and tennessee were judged as having adequate coverage of complex trait genetics concepts ( unpublished data ) .
there was a modest but significant positive correlation between the overall quality of a state 's genetics standards and its treatment of complex traits ( r = 0.38 , p < 0.001 ; figure 3 ) .
figure 3.the relationship between the overall average quality of genetics standards to the average of the three concepts directly related to complex trait genetics ( concepts 6 , 7 , and 11 ; table 1 ) .
the relationship between the overall average quality of genetics standards to the average of the three concepts directly related to complex trait genetics ( concepts 6 , 7 , and 11 ; table 1 ) .
our analysis is based on the premise that the discipline of genetics / genomics ( and scientific disciplines , in general ) is structured around an intellectual core of fundamental concepts .
the dynamic nature of science has accustomed us to expect that our understanding of scientific details will change even as further research serves to strengthen the concepts at the core .
such is the case with genetics , where concepts such as the allelic nature of genes , basic processes of gene expression , mutations , genetic variation , and patterns of inheritance have been expanded and enhanced across more than a century .
other concepts , while less familiar , such as differential gene expression and the genetic basis of complex traits , have been recognized for decades as crucial , but only in the past 1020 yr have advances in technology enabled researchers to elucidate the genetic mechanisms by which they occur .
other groups ( several of which are referenced in methods ) have developed their own lists for their own purposes .
the ashg list drew heavily from those previously published lists , but our choices were guided by the specific goal of capturing the minimum number of concepts , at the appropriate level of specificity , that the authors and ashg 's information and education committee believe to be essential for genetic literacy and thus should be understood by all high school graduates . as with all such lists , our process necessitated choosing among concepts that should be included and concepts that should be omitted , and some readers may disagree with these choices .
we believe that ashg , a large and influential professional society in genetics , can make a contribution to the discussion of standards with its views on what constitutes a reasonably specific , comprehensive , and age - appropriate set of concepts in one of the major subdisciplines of biology .
consequently , our choices reflect a moderate level of specificity , and our benchmark list includes 19 concepts rather than , for example , 6 or 60 .
this acknowledges dramatic changes in genetics over the past two decades , such as the discipline 's increasing impact on medicine and direct - to - consumer genetic testing , and the fact that an understanding of additional concepts is needed to raise the standard of scientific literacy among the nation 's population .
the new concepts we propose can be taught using many different subconcepts and examples , which we do not prescribe .
overall , our analysis identifies substantial deficiencies in the treatment of genetics in state standards across the united states .
of ashg 's 19 benchmark concepts , 14 were treated inadequately ( averaged across all states , including the district of columbia ) , and 39 states had adequate coverage of fewer than 11 concepts ( table 1 and figure 2 ) .
two concepts ( continuous variation of complex traits and polygenic inheritance ) were virtually absent from state standards .
only evolution and the nature of the genetic material were adequately covered , while the broad categories of gene expression and regulation and genetic variation were not .
it is particularly troubling that the inadequate and not addressed / absent concepts represent ideas that are increasingly important as genetics / genomics matures and assumes practical importance in people 's lives . as dna sequencing and genotyping technologies
have become more powerful and less expensive ( at rates that vastly exceed moore 's law in computing ) , they have been applied more often in medical research .
it is now possible to investigate the genetic contributions to virtually any disease or normal trait , and our ability to apply knowledge of individual genetic variation to clinical treatment and outcomes is advancing .
for example , roughly 10% of drugs approved by the food and drug administration have labels with pharmacogenomics information , and genetic testing has already become common for certain conditions , such as breast cancer ( hamburg and collins , 2010 ) . as lanie et al .
have written , even if people grasp and understand these basic concepts [ mendelian genetic concepts ] , the impact of this knowledge will be limited . . .
it will not go far in helping the layperson understand the barrage of genetic information to which they are exposed through the media the vast majority of which deals with complex diseases and traits ( 2004 ) . without adequate standards dealing with mutations , gene regulation , and non - mendelian patterns of inheritance , such concepts may never make it into curricula and assessments .
the most - neglected concepts also demand higher orders of thinking than concepts that tended to be covered adequately .
for example , unlike the nature of the genetic material ( i.e. , dna ) , which deals largely with descriptive biology , gene expression ( concept 8) deals with the mechanisms of how genes operate and the functional consequences of those operations .
the latter involve understanding positive and negative feedback ( general ideas that also extend to engineering , computing , and economics ) and require thinking at levels of application , interpretation , and analysis . in parallel with being
poorly represented in state standards ( table 1 ) , assessment questions related to the concepts of gene expression ( concept 8) and differential gene expression ( concepts 9 and 10 ) elicited very few responses indicative of complete understan ding by grade 12 students on the 2000 naep ( 2006 ; table 3 ) .
table 3.students responses to genetics questions on the 2000 naep ( grade 12 ) indicate poor understanding of essential genetics conceptsnaep question ( related concept , this work , from table 1)% complete or essential answerswhat is a gene ?
( 2)21briefly explain how the information that a gene provides to a cell results in the production of a molecule such as hemoglobin .
( 8)1[e]xplain why kidney cells do not make hemoglobin , even though they contain the hemoglobin gene .
( 10)10give an example of an inherited disease that people might be able to treat by use of [ recombinant dna ] technology , and explain how you think the technology might be used to treat this disease .
( 16)5 students responses to genetics questions on the 2000 naep ( grade 12 ) indicate poor understanding of essential genetics concepts another specific area of weakness is the state standards treatment of mutations .
frequently , state standards do not distinguish between germline and somatic mutations and their connections to hereditary versus nonhereditary genetic disease ( concepts 14 and 16 , respectively ) .
failure to make this distinction may contribute to students conflating inherited disease with any disease influenced by genetics , a frequent student misconception ( shaw et al . , 2008 ) .
students poor understanding of these concepts is also reflected in their responses to a related question on the naep ( table 3 ) .
perhaps not surprisingly , dna as the genetic material ( concept 1 ) is well covered by most states , as it should be nearly 60 yr after the elucidation of dna 's structure ( average score of 1.7 , table 1 ) .
the organization of genetic information in the form of genes carried by chromosomes also fared well ( 1.5 ) .
evolution by natural selection ( concept 18 ) tied for best average score ( 1.7 ) , and was supported by strong coverage of genetic and phenotypic variation as the substrate for evolution ( concept 17 ; 1.5 ) .
these findings support earlier research indicating improvements in state standards coverage of evolution ( mead and mates , 2009 ) .
unfortunately , state standards are not doing as well framing the population - level and generational timescale at which evolution operates ( concept 19 ) .
learning goals related to single - gene inheritance patterns ( i.e. , mendelian inheritance , concept 5 ) are also adequately addressed nationwide in the state standards ( average score 1.5 ) , an encouraging but not unexpected finding .
instruction about genes , alleles , and mendelian segregation , including meiosis , is virtually ubiquitous in high school general biology , and these concepts continue to dominate instruction in equivalent undergraduate courses ( hott et al . , 2002 ) .
these concepts also lend themselves easily to problem - based learning , which engages higher - level critical thinking . unfortunately , there may be a downside to all this attention . in the context of the modern discipline 's broader view of genetics
, we may be skewing students understanding of the genotype phenotype connection ( dougherty , 2009 ) .
in essence , if we spend too much time on single - gene inheritance at the expense of polygenic inheritance and complex traits then we not only fail to convey modern genetics accurately , but we also risk giving students a false impression that most traits are inherited in the simple manner conveyed by the rare single - gene traits that are so often used as examples , such as cystic fibrosis and hemophilia .
in fact , patterns - of - inheritance misconceptions represent the second most problematic conceptual category in genetics identified by shaw et al .
not surprisingly , low scores for concepts related to complex traits and multifactorial causation ( concepts 6 , 7 , and 11 ) contributed substantially to low average scores for most states .
to get some sense of how our specific results compared with the quality of state science standards as judged more generally , we compared them with the results of an evaluation conducted by the thomas b. fordham institute ( table 2 , columns three and four ) .
differences in methodology prevent direct comparison ; however , weaknesses in state standards are apparent in both analyses .
first , states were not scored blindly , which may have allowed some unintentional reviewer bias , although our method of using multiple reviewers and dropping the high and low scores for states with five or six reviewers should have helped minimize such bias .
second , for a small number of states , it is possible that some concepts may have been addressed only at lower grade levels , in which case this analysis would have missed them .
the decision to focus exclusively on high school standards was based on a preliminary analysis that indicated that middle school standards were generally repeated in the high school standards for the states that included genetics concepts at both levels .
the repetition of genetics concepts in middle and high school standards , in the cases where we observed it , bolsters the argument by schmidt et al .
different states have different revision schedules ( and processes ) for their standards , and the performance of any given state with respect to genetics ( or any other content ) would be expected to change over time .
finally , our analysis considers only one variable ( i.e. , standards ) in the complex system that constitutes genetics education .
of course , the absence of concepts in a state 's standards does not mean that those concepts will not be taught .
knowledgeable teachers can address such concepts even in situations where high - stakes exams do not require them .
however , given the pressures of performing well on state exams and limited classroom time , teachers often prioritize the content specified by their states .
conversely , the fact that concepts are represented in a state 's standards does not guarantee that students will learn them ( schmidt et al . ,
for example , dna as the genetic material and the nature of genes were the best - covered concepts for any category on our benchmark list ( concepts 1 and 2 , table 1 ) ; however , only a minority of students gave complete or essential answers to a question dealing with these ideas on the 2000 naep .
those concepts have been taught since long before standards achieved their current prominence in education , and yet student understanding lags .
the fact that 44 states ( 86% ) had genetics standards judged to be inadequate and only 12 states ( 24% ) had 11 or more individual concepts judged to be adequate convincingly demonstrates ( in our view ) a need for improvement when states revise their life sciences standards .
ashg will encourage members of its genetics education outreach network to volunteer to assist with revisions in those states that allow such participation .
similar networks exist at other scientific professional societies and may offer a generalizable mechanism for involving scientists who are content experts , as well as knowledgeable about k12 education , in both the evaluation and improvement of content coverage in state standards .
standards are just one element of what should be an integrated and coherent teaching system , and changes to standards should be made only with full recognition of the effects those changes may have on other parts of the system .
crowded curricula , teacher professional development , and available instruction time , must all be taken into consideration . within genetics , new conceptual frameworks
for example , it may be possible to refocus instruction around complex traits , which have a greater capacity for carrying modern understandings of genetics ( as well as traditional ones ) than do the simpler , single - gene traits that are used so widely in high school classrooms now ( dougherty , 2009 ) . the nrc 's move to generate common science standards
a recent report supports the establishment of clear and common standards across states , noting that roughly one - half the teachers in all states agreed that the standards of their own states were not clear enough , and that 85% think having tougher academic standards would make at least a moderate impact on improving academic achievement
sound science instruction requires expertise in teaching ( content knowledge and pedagogy ) , exemplary curricula , strong assessment , and a supportive system ( e.g. , administration , professional development ) . in the current u.s .
system , standards are the foundation upon which curricula and instruction are built , and our work shows that the foundation , at least in genetics , is in need of repair .
we propose that the findings detailed in this paper are well positioned to guide the development of common core standards in science . | science education in the united states will increasingly be driven by testing and accountability requirements , such as those mandated by the no child left behind act , which rely heavily on learning outcomes , or
standards , that are currently developed on a state - by - state basis . those standards , in turn , drive curriculum and instruction .
given the importance of standards to teaching and learning , we investigated the quality of life sciences / biology standards with respect to genetics for all 50 states and the district of columbia , using core concepts developed by the american society of human genetics as normative benchmarks .
our results indicate that the states genetics standards , in general , are poor , with more than 85% of the states receiving overall scores of inadequate . in particular , the standards in virtually every state have failed to keep pace with changes in the discipline as it has become genomic in scope , omitting concepts related to genetic complexity , the importance of environment to phenotypic variation , differential gene expression , and the differences between inherited and somatic genetic disease .
clearer , more comprehensive genetics standards are likely to benefit genetics instruction and learning , help prepare future genetics researchers , and contribute to the genetic literacy of the u.s . citizenry . |
traction exerted by the posterior hyaloid has been implicated in the pathogenesis of several conditions such as proliferative vitreoretinopathy ( pvr ) , proliferative diabetic retinopathy ( pdr ) , penetrating eye trauma and macular holes .
therefore the surgical goal of any vitrectomy should be posterior hyaloid separation and removal of the vitreous as much as possible . despite the development of surgical techniques , at times
the surgeon may not be confident that the posterior hyaloid has actually been removed . in conditions characterized by breakdown of the blood retinal barrier such as pvr , uveitis , retinal vein occlusions and diabetic retinopathy , an intravenous injection of fluorescein sodium 1 to 2 days prior to the scheduled vitrectomy stains the vitreous green facilitating its identification.5 blood in the vitreous cavity coats the vitreous by adhering to its collagen fibrils making the normally transparent vitreous opaque and easier to visualize for removal . in eyes with no pre - existing vitreous hemorrhage , a small amount of autologous blood may be injected into the vitreous cavity to coat the vitreous.6 triamcinolone acetonide ( ta ) is a well - tolerated corticosteroid used for the treatment of diseases such as uveitis , diabetic macular edema ( dme ) and retinal vein occlusions .
once ta is injected into the vitreous cavity , its particles adhere to the vitreous gel facilitating visualization and identification ( figures 1 and 2).7 in addition , using ta may improve vitrectomy outcomes by reducing blood retinal barrier breakdown and preretinal fibrosis .
currently this is the most widely used technique to visualize the posterior hyaloid.4,6 a comparative study of fluorescein , icg , ta and tb concluded that the vitreous was best highlighted by ta.8 a recent study demonstrated that a 20% solution containing the natural dyes lutein and zeaxanthin , precipitates on the vitreous surface staining it orange.9
over the past two decades owing to refinements in instrumentation and surgical techniques , erm removal has been the typical indication for macular surgery .
the transparent erms pose a challenge even to experienced surgeons since incomplete removal of the erm is implicated in post - surgical recurrences10 which have been reported in up to 21% of cases.11 - 13 staining of the transparent erm allows identification of its entire extent and facilitates its visualization and complete removal .
staining of these erms facilitates surgery by improving visualization , characterization of the erm as diffuse or focal , and confirmation that the tissue requiring to be peeled is indeed an erm and not swollen nerve fiber layer.14 among the commercially available agents , tb is the dye of choice for erm peeling.4,15 this dye binds to degenerated cell elements and does not stain live cells or tissues with intact cell membranes since there is no uptake of the dye . as erms are mostly composed of dead glial cells ,
cataract surgeons have long used tb to stain the anterior capsule during phacoemulsification.16 erms stain prominently with 0.15% tb which is a relatively safe concentration.15 histopathological analysis of excised erm showed no retinal cells on the retinal side of the erm and no signs of apoptosis .
no rpe defects or signs of retinal toxicity have been reported in most studies as well.15,17,18 nevertheless , animal and in vitro studies show that dose dependent toxicity may appear with concentrations above 0.3%.15
the ilm is made of the basement membrane of mller cells representing an interface between the retina and the vitreous .
the significance of ilm peeling is in achieving closure of large and chronic idiopathic mhs.19 tangential traction from the ilm plays a key role in the pathogenesis of mh . in a post - mortem examination , the eye with successful mh closure
was characterized by an area of absent ilm surrounding the sealed mh.20 in contrast in another patient that had a re - opened mh , histopathological examination revealed an erm with ilm surrounding the open mh suggesting that traction from the ilm was partly responsible for recurrence.21 in other conditions such as dme and erm , ilm peeling has remained controversial .
surgical specimens often show fragments of the ilm interspersed among the erm.22 - 24 breaks in the ilm may provide access to the macular surface for cellular elements , thus ilm peeling may reduce the risk of recurrence following erm removal . even in cases with complete erm removal , recurrences may occur .
therfore by removing the ilm as well , the surgeon can be assured that the erm has been entirely removed .
thus ilm peeling has been proposed as a means to ensure complete removal of the erm and to deter recurrence.13,25 visual improvement in 65% to 90% of eyes following erm removal and a low recurrence rate of 5% with even fewer re - operations were reported and these results were achieved without any ilm peeling.26 - 28 sivalingam et al23 reported that long segments of ilm removal led to a worse visual outcome compared to less ilm removal .
in contrast , more recently bovey et al29 reported that the visual outcome of eyes in which the erm had ilm remnants were significantly more satisfactory than eyes without ilm remnants in erm .
since the ilm consists of footplates of mller cells , it is reasonable to assume that its removal may result in some type of injury .
electron microscopic evaluation of surgically removed ilm specimens revealed degenerated and necrotic mller cell processes.30 mild damage to the vitreoretinal interface following icg stained ilm removal has also been documented with electron microscopy.31 given that icg has been reported to cause retinal toxicity , it remains unclear whether this mild damage to the vitreoretinal interface is due to icg or mechanical peeling of the ilm.4 ilm peeling causes a selective delay in the recovery of focal macular erg b wave even 6 months after operation32 suggesting that some type of injury to the macular mller cells occurs after ilm peeling , since these cells are responsible for generating the b wave in the electroretinogram .
tadayoni et al 33 coined the term " dissociated optic nerve fiber layer " to describe a peculiar fundus appearance following erm stripping . in their series out of 100 eyes ,
43% manifested numerous arcuate striae slightly darker than the surrounding retina in the direction of optic nerve fibers appearing 1 to 3 months postoperatively .
mitamura et al34,35 used optical coherence tomography ( oct ) to further characterize these findings as dimples limited to the retinal nerve fiber layer .
more recently clark and colleagues36 described 3 to 5 dark striae radiating from the papilla to the macula in an arcuate fashion as early as 1 week to 1 month postoperatively on blue light autofluorescence and infrared reflectance imaging .
simultaneous oct through the striae revealed focal areas of swelling in the arcuate retinal nerve fiber layer .
these changes resolved spontaneously after a mean period of 2 months not resulting in worse post - operative visual acuity as compared to eyes without such changes.36 in some eyes , swelling of the arcuate retinal nerve fiber layer evolved into characteristic dimples of dissociated optic nerve fiber layer suggesting that both of these findings represent different time points on the same spectrum.36 it appears that both dissociated optic nerve fiber layer and the arcuate swelling of the nerve fiber layer do not affect visual function .
a number of factors including direct trauma by the surgical forceps as they grasp the ilm , subclinical trauma to the inner retina from mller cell damage , and mller cell degeneration have been speculated.36,37 more longitudinal studies are required to assess the clinical implications of these changes .
park et al13 showed that ilm peeling did not have a detrimental effect on visual acuity .
they compared the outcomes between 24 eyes that underwent pars plana vitrectomy with erm peeling but not ilm peeling , with 20 eyes that had undergone pars plana vitrectomy with both erm and ilm peeling .
they reported that visual outcomes were similar between the two groups but that recurrence rate was much higher in eyes without ilm peeling .
more recent studies have shown that ilm removal does not improve or worsen postoperative visual acuity.12,38 however , recurrence was reduced by ilm peeling.12 another purported advantage of ilm peeling is a higher likelihood of resolution of retinal striae.39 selective stains such as icg and bbg have a high afnity for basal membranes such as the ilm and a low afnity for collagen tissues such as erms .
when either dye is used to stain both layers before removing any tissue during surgery , the unstained erm is clearly depicted against the ilm , which is stained blue or green ( figure 4 ) .
some surgeons re - stain the macula to visualize the remaining ilm but others do not .
even with a single bbg staining , up to two thirds of eyes will have residual ilm remnants after erm.12,40,41 in a recent prospective study conducted by the pan american collaborative retina study group , there was little correlation between the surgeon s un - aided observation and the bbg stained observation of the ilm .
thus , if the surgeon believes in the significance of ilm peeling in erm operation , staining should be strongly encouraged.42 the first vital dye used for ilm staining was icg.43 since icg binds to the ilm , biomechanical stiffness of the ilm increases thus facilitating its peeling ( figure 5).44 however the initial enthusiasm for the use of icg has tempered following numerous reports of toxicity.4,45 a meta - analysis of chromovitrectomy with icg compared to peeling of the ilm without staining in mh surgery showed similar anatomic outcomes in both groups .
however the functional results in eyes in which the ilm was stained by icg were much worse.46 several strategies can be used to minimize potential icg toxicity .
the lowest concentration of icg which provides adequate visualization of the ilm should be used ; concentrations of 0.5% or less are relatively safe . to avoid leaving excess
icg in the eye is of high importance to limit the area of icg staining to the ilm that will be peeled . to protect the bare rpe of the mh floor , a small bubble of heavy perfluorocarbon liquid or viscoelastic
is placed over the mh . following partial fluid - air exchange , 1 to 2 drops of the icg solution
are placed over the macula for 30 to 60 seconds . then the icg is washed out and ilm peeling can be performed . an alternative to icg
is infracyanine green which , unlike icg , does not contain sodium iodide allowing it to be dissolved in 5% glucose and to generate an iso - osmotic solution . in vitro assays have shown that infracyanine green induced significantly less toxicity on rpe and retinal ganglion cell lines in comparison to icg.47 this toxicity was more time dependent rather than concentration dependent .
when the duration of exposure approached 30 minutes , early apoptotic changes were observed in both cell lines , with no significant difference in apoptotic rates at 2 different concentrations ( 0.025% and 0.05%).47 several clinical investigations have revealed that infracyanine green assisted ilm peeling during mh repair results in high anatomic and functional success rates.48 - 50 bbg also has high affinity for the ilm . in animal and in vitro studies
, bbg appears to be relatively safe at doses up to 0.25 mg / ml.4 bbg is not a fluorescent dye ( figure 6 ) .
contact with the rpe should be avoided since rpe atrophy has been documented following subretinal migration of bbg . in general , bbg appears to be a safer alternative to icg for ilm peeling .
furthermore , bbg can be beneficial as a neuroprotective agent.51 in vitro studies have shown that photoreceptors undergo apoptotic cell death when exposed to extracellular atp through the activation of p2rx7 purinergic receptors .
identification of all retinal breaks is of paramount importance for successful repair of a retinal detachment . on occasion
transretinal injection of tb into the subretinal space with a 41 gauge needle followed by heavy perfluorocarbon liquid injection into the vitreous cavity assists the identification of previously unclear retinal breaks.52
the research is persevering and numerous dyes including methyl violet , crystal violet , eosin y , sudan black b , methylene blue , toluidine blue , indigo carmine , fast green , light green , congo red , bromophenol blue , patent blue and evans blue have been investigated.53 , 54 an ideal dye should have excellent contrast and high biocompatibility.55 german investigators have designed and tested 3,3-di-(4-sulfobutyl)-1,1,1,1 tetramethyl - di-1h - benz[e]indocarbocyanine ( dss ) with good biocompatibility which can be a proper alternative for ilm staining.55 on the other hand , brazilian investigators argue that natural dyes may be safer alternatives than synthetic dyes.56 they have investigated several naturally produced dyes and reported that acai fruit ( euterpe oleracea ) , logwood ( haematoxylum campechianum ) , cochineal ( dactylopius coccus ) and old fustic ( maclura tinctoria ) extracts facilitated posterior hyaloid visualization similar to triamcinolone acetonide in cadaver eyes .
acai fruit ( euterpe oleracea ) , chlorophyll extract from alfalfa ( medicago sativa ) and cochineal ( dactylopius coccus ) extracts stained the ilm well in cadaver eyes.56 time will tell if any of these agents will end up in routine clinical use .
with the recent introduction of ocriplasmin , pharmacological vitreolysis is a reality.57 the greatest advantage of enzymatic vitreolysis is that a cleaner cleavage plane between the retina and other tissues of interest may be achieved by enzymatic means as compared to mechanical separation by vitrectomy . in the future it is likely that enzymatic vitreolysis will play an even more significant role in the management of disorders of the vitreoretinal interface .
transparent tissues such as the posterior hyaloid , erms and the ilm play an important role in several diseases of the posterior pole . surgical removal of these tissues is a principal objective which can be facilitated by staining with a variety of vital dyes . several dyes are currently used in routine clinical procedures , however the ideal staining agent has not yet been found and any dye which is injected intravitreally has the potential to become toxic . in general ,
the lowest concentration of dye that will stain the tissue of interest should be used .
the dye should also be placed for the shortest period of time to result in minimal color change as a faint stain is usually sufficient for peeling . keeping the light pipe as far away as possible from the retinal surface
ta highlights the vitreous most ideally , and tb and bbg best stain erms and the ilm , respectively . | adequate visualization and identification of the posterior hyaloid , epiretinal membranes and the internal limiting membrane are of paramount importance in modern vitreoretinal surgery .
chromovitrectomy is a term used for describing the vital dyes use in order to stain these transparent tissues and facilitate their manipulation during vitreous surgery .
this article reviews the indications , applications and characteristics of vital dyes in vitreoretinal surgery .
various dyes are currently being used in routine clinical procedures , however the ideal staining agent has not yet been found .
any dye which is injected intravitreally has the potential to become toxic .
triamcinolone acetonide is used to highlight the vitreous and is particularly beneficial in determining the attachment of the posterior hyaloid to the underlying retina .
trypan blue stains epiretinal membranes and facilitates their complete removal . both indocyanine green and brilliant blue g stain the internal limiting membrane properly
, however concerns over indocyanine green toxicity have made surgeons switch to brillliant blue g as a safer alternative . |
atherosclerosis is a pathological process that takes place in the major arteries and represents one of the causes of myocardial infarction ( mi ) , ischemic stroke , and peripheral artery disease .
atherosclerosis , a multifactorial condition determined by environmental as well as genetic factors , has a strong inflammatory component characterised by deregulation of gene expression in macrophages within atherosclerotic plaques , together with local overproduction of cytokines . among those , chemokines ( chemotactic cytokines )
have also been implicated in atherosclerosis on the basis of clinical studies and animal disease models .
the chemokine rantes , also known as cc chemokine ligand ( ccl5 ) , is involved in the activation and proliferation of t - lymphocytes and is considered as a major chemokine implicated in both the acute and chronic phases of inflammation .
tissue rantes expression is differentially regulated : in atherosclerosis , rantes mrna has been detected in the lymphocytes , macrophages , myofibroblasts , and endothelial cells of human arteries undergoing accelerated atherosclerosis but not in normal coronary arteries .
platelets , which are also implicated in the atherosclerotic process including myocardial infarction ( mi ) , are known to store rantes in intracellular granules in the basal state and to secrete it when stimulated with thrombin .
single nucleotide polymorphisms in the rantes gene promoter and intronic region have been described : c-28 g , g-403a ( relative to transcription start site ) , polymorphism in intron 1 ( in1.1 t / c ) , and in the 3untranslated regions [ 6 , 7 ] .
the case - control study investigating rantes snp g-403a in germans associated this snp with a susceptibility to coronary atherosclerosis ; in another study , in hungarians , however , this association was not significant . that rantes polymorphisms may play a role in cardiac pathology
was also suggested by a report enrolling patients with type 2 diabetes mellitus and end stage renal disease : the carriage of either rantes -403 g / a or intronic polymorphism ( in1.1 t / c ) was associated with all - cause mortality due mainly to cardiac events . concerning the effect of these snps on rantes expression , an increased gene expression was linked with the minor allele in the rantes gene at the nucleotide position -403 and -28 [ 11 , 12 ] .
however , in vitro functional data are more important for rantes intronic polymorphism t / c : in1.1c allele or haplotypes that include this variant showed a strong dominant association with rapid progression to aids .
furthermore , rantes gene polymorphisms were associated with the outcomes of hcv and hbv infections [ 13 , 14 ] . by analogy , atherosclerotic lesions ,
the underlying cause of coronary artery disease ( cad ) and mi , represent a series of highly specific cellular and molecular events of the inflammatory response . despite convincing evidence for a role of the rantes polymorphisms in different inflammatory diseases , their role in cad
requires further investigation . given the inflammatory component of coronary atherosclerosis and the lack of case - control investigations of more than one rantes polymorphism in mi
, this study examined the relationship between two rantes polymorphisms ( promoter g-403a , intron1.1 t / c ) with their haplotypes and myocardial infarction . in agreement with genetic epidemiology principles , our replication
study comprised two population cohorts ( czech , russian ) differing in ethnicity from those populations in which the rantes gene was investigated previously ( hungarian , german ) .
a total of 804 unrelated caucasian individuals were enrolled into this case - control study , of which 467 were mi cases and 337 healthy controls subjects .
two population groups were recruited : ( 1 ) czech ( olomouc ) with 223 mi patients and 140 control subjects and ( 2 ) russian ( novosibirsk ) with 244 cases and 197 controls from the monica project register .
mi events were defined on the basis of electrocardiographic ( ecg ) changes and biomarker ( troponin , alternatively creatine - kinase ) measurements .
local research ethics committees approved the studies and the subjects provided written informed consent for use of samples in these genetic studies and anonymous publication of the summary data .
genotype analysis for the rantes snps was performed by polymerase chain reaction with sequence - specific primers ( pcr - ssp ) .
the investigated snps included rantes promoter g-403a ( rs2107538 ) and rantes intron 1.1 t / c ( rs2280789 ) .
the sequences of specific primers were as follows : allele g , forward 5 atg gat gag gga aag gag g 3 , allele a , forward : 5 cat gga tga ggg aaa gga ga 3 , constant reverse : 5 ttc tct gct gac atc ctt ag 3 ; intron 1.1 t / c ; allele t , forward : 5 cct gat cag ttt ttc tgt ctt t 3 , allele c , forward : 5 cct gat cag ttt ttc tgt ctt c 3 , constant reverse : 5 tgt gag cta ctc atg act ac 3. the internal control primers and conditions for pcr were adopted from a phototyping methodology .
calculations were performed using the spss v.13.0 ( spss inc , chicago , il ) .
categorical data of the study population was reported as frequencies and percentage , the continuous data as means with standard deviation .
the distribution of the rantes genotypes and alleles were analyzed using the pearson 's contingency table -test .
a multiple variable logistic regression model was used to estimate the association between genotype and disease .
haplotype analysis was performed using an accelerated expectation - maximization algorithm ( software arlequin , version 3.000 ) .
first , we evaluated distribution of rantes polymorphisms in patients and control subjects ( for genotype and allele frequencies see table 2 ) . out of 467 and 337
individuals in each group , the genotype data for rantes intron 1 polymorphism was available in 465 patients and 330 controls as 2 patients and 7 control samples failed to amplify ; the overall failure rate was only 1.2% ( 9/804 ) . both the rantes -403 and rantes in1.1 genotypes were distributed as expected from the hardy - weinberg equilibrium ( p > .05 ) . in both the czech and russian populations
, there were no significant differences in the frequencies of genotypes between the mi cases and controls for the two rantes polymorphisms studied .
the distribution of rantes genotypes was similar also when both populations were evaluated together ( table 2 ) , except that the frequency of cc homozygous genotype in mi patients ( 2.8% ) was higher than observed in controls ( 0.9% ) , but this difference did not achieve a statistical significance .
further , we carried out standard allelic association analysis : minor allele frequencies for the two polymorphisms were calculated , and the odds ratio and 95% confidence interval for the rantes -403a and rantes in1.1c were 0.996 ( 95% ci 0.7401.340 ) and 1.054 ( 95% ci 0.7591.464 ) , respectively , ( p > .05 in both cases ) . also , when evaluating carrier status for -403a and in1.1c alleles , there were no significant differences between mi cases and controls ( p = .97 , df = 1 for aa+ag versus gg -403 genotypes ; and p = .75 , for cc+ct versus tt in1.1 genotypes ) .
next , the patients were divided into subsets according to gender and age at the time of the 1st myocardial infarction , and univariate analyses were performed .
there was no difference in genotype distribution when male and female cases were compared with controls of appropriate gender separately ( rantes g-403a : p = .863 for males and p
= .789 for females ; rantes in1.1 t / c : p = .148 for males and p = .744 for females ) .
no differences in genotype and alleles frequencies ( -403a and in1.1c ) were observed between the patients stratified according to the age of mi onset ( > 50 yr and < 50 yr ) ( table 3 ) .
also , in the logistic regression analysis , no evidence for association was found between the polymorphisms of the rantes gene and the risk for mi in our study population ( table 4 ) .
the combined genotypes ggtt and agtc were the most frequent ( case / control group 62.3%/64% and 18%/18.4% , resp . ) .
no statistically significant difference was observed in the rantes combined genotypes between the cases and controls ( = 4.53 , df = 7 , p = .717 ) .
however , we detected strong linkage disequilibrium between the two less common alleles for two rantes snps : allele a of the rantes promoter g-403a and c allele of the rantes intron 1 t / c ( d = 0.83 for patients with mi and d = .88 for control group ; p < 10 in both cases ) .
this case - control study aimed at exploring the association between coronary atherosclerosis manifested by myocardial infarction and two functional polymorphisms in the rantes ( ccl5 ) gene , the product of which is involved in atherosclerotic inflammation . in this two - population replication study ,
no significant differences in the distribution of rantes g-403a , and in1.1 t / c single nucleotide polymorphisms were observed between patients with mi and the control groups .
further , when the subjects were stratified according to their gender and age at the first mi episode , no significant relationships were revealed .
thus , our findings do not support the previous report of association between rantes promoter g-403a polymorphism and coronary artery disease .
importantly , our report represents the first investigation of two linked rantes polymorphisms and specifically analysis of their haplotypes in the slavonic population .
previous studies associating rantes snps with coronary atherosclerosis [ 8 , 9 ] concentrated on the rantes promoter polymorphism ( g-403a ) and did not explore the relationship between cardiovascular symptomatology and further polymorphisms in the rantes gene , including its intronic variant in1.1t / c .
only bger et al . , who investigated rantes promoter and intronic polymorphisms in type 2 diabetes / end stage renal disease , reported that patients carrying the rantes-403a or in1.1c allele had higher all - cause mortality risk , mainly due to cardiac events .
we , however , can not compare their data with our results , because boger 's study did not include a control population ( healthy controls ) and also estimated the effect of genetic predictors of survival and functional polymorphisms on mortality in another disease condition . a difference in exact disease phenotypes
may be another less prominent , but still relevant factor explaining the discrepancy between our negative data and the first case - control association study on rantes , which showed a trend indicating a higher risk for coronary artery disease for hungarian patients with the -403a allele .
the population size of their cohort and our slavonic population was identical ; however , the hungarian study enrolled patients with severe coronary artery disease ( cad ) not necessarily presenting with mi .
the luric ( the ludwigshafen risk and cardiovascular health ) study was the second to report the association for -403a allele .
we speculate that the deviation of distribution of g-403a genotypes from that expected from the hardy - weinberg equilibrium in controls of the luric study , may contribute to the discrepancy between luric and our findings concerning the relationship of the rantes -403a variant and cad .
though smaller in number of enrolled subjects , our czech and russian groups were strongly compliant with the hwe .
strong linkage disequilibrium was observed throughout the rantes gene , and in particular between the alleles at the four most polymorphic sites , -403 , -28 , in1.1 , and 3222 .
the in 1.1 t occurs in combinations with -403 g in 77% of genotyped european americans ( 5% had in1.1t/-403a haplotype ) , and allele in1.1c occurred with frequencies of 0.14 in european americans , 0.20 in african americans , and 0.34 in asians . in the present study , we revealed similar allele in1.1c frequencies in our investigated slavonic populations as in european americans , and confirmed strong linkage disequilibrium between two less common variants at rantes -403 and in 1.1 loci .
the functional effect of particular snps in the rantes gene and the level of its expression and/or secretion which may provide a mechanism explaining the observed association with cad , is still controversial [ 7 , 11 , 12 , 17 , 18 ] .
the -403a allele has been associated with increased transcription of the rantes gene , and further evidence of the functional importance of this variant is provided by a significant risk of atopic dermatitis , atopy , and asthma in adults but without studying the effect of the -28 or in1.1 loci .
other studies showed an upregulatory role for the -28 g allele but did not demonstrate upregulation for -403a [ 12 , 18 ] .
another study demonstrated that rantes in1.1c alelle was associated with reduced rantes expression , -28 g allele had a moderate role for expression and -403a allele was not regulatory .
discrepant data on correlations between rantes polymorphisms and level rantes expression for other common inflammatory and chronic infection diseases support the need for further investigation of the relationship between variability in rantes gene and its expression . in conclusion , in this two - population - based case - control study , no significant difference was detected in the distribution of two rantes gene polymorphisms between healthy controls and patients with myocardial infarction , indicating the lack of association between coronary atherosclerosis and rantes single nucleotide polymorphisms .
our results , therefore , do not support a major involvement of rantes g-403a and in1.1 t / c polymorphisms in genetic predisposition to myocardial infarction . | coronary artery inflammation is a critical process in the pathogenesis of myocardial infarction ( mi ) .
the chemokine ccl5/rantes ( regulated upon activation , normal t cells expressed and secreted ) is expressed in advanced atherosclerotic lesions .
functional polymorphisms of the rantes gene can , therefore , be involved in the pathogenesis of coronary artery disease .
we examined the association of polymorphisms in the rantes gene with myocardial infarction in slavonic populations of czech and russian origin .
a total of 467 post - mi patients and 337 control subjects were genotyped for rantes promoter g-403a ( rs2107538 ) and intron 1.1 t / c ( rs2280789 ) variants by pcr - ssp .
both rantes genotypes and allele frequencies did not differ between case and control groups .
haplotype - based analysis also failed to reveal an association between mi and investigated markers .
strong linkage disequilibrium was detected between particular rantes alleles .
the data do not support an association between rantes g-403a polymorphism and mi , as reported previously . |
the advent of antenatal ultrasound has allowed the diagnosis of conditions like large neck masses affecting the fetus which can compromise the airway after delivery . fetal magnetic resonance imaging ( mri )
has helped in better delineation and planning for perinatal intervention which will improve the survival of these babies .
ex - utero intra partum treatment ( exit ) is a procedure conducted for a fetus diagnosed with compromised airway . by this procedure
the airway can be secured while the fetus is on utero - placental circulation and hypoxia can be prevented .
there are many indications for exit but the commonest indication is for a fetus with compromised airway . the diagnosis of congenital high airway obstruction syndrome ( chaos ) , lesions in the mouth and neck compressing the trachea is a definite indication for exit . detailed planning and execution is essential for this procedure to be successful .
this procedure has been extensively described in literature and this is one of first few case reports from india .
we present our experience with exit procedure and the challenges and the optimal positioning of the mother , fetus and anesthesiologist which helped us in securing the airway .
a 29-year - old multi - gravida was referred to our hospital with an antenatal diagnosis of a large cystic hygroma diagnosed at 38 weeks of gestation .
fetal mri revealed a large multiseptated cystic mass ( measuring 12 8 cm ) involving both sides of the neck with compression over the trachea [ figure 1 ] . hence a difficulty in securing the airway was anticipated and exit procedure was planned .
a multispecialty team comprising of obstetricians , anesthesiologists , pediatric surgeons , neonatologists , and radiologists were involved in planning and executing the procedure . under general anesthesia
two teams of anesthesiologists were present , one for the mother and the other for the fetus .
apart from the routine monitoring , radial artery cannulation was done for invasive blood pressure monitoring .
patient was positioned in the supine position with left lateral tilt to prevent compression on the aorta .
the legs were slightly abducted in order to allow easy access to the fetus for intubation .
general anesthesia was commenced and maintained with higher concentration of isoflurane ( for complete uterine relaxation ) , oxygen , nitrous oxide , and muscle relaxants . before skin incision
, ultrasound was done to locate the placental edge which revealed two loops of umbilical cord around the fetal neck . after skin and uterine incision ,
fetal heart rate was monitored with an ultrasound probe and manually by umbilical cord pulsation .
ultrasound - guided aspiration of the cyst was done but since most of the cysts were microcystic , adequate decompression could not be obtained .
the neck was delivered up to the sternal notch [ figure 3 ] and then intubated .
intubation could be accomplished in 3 minutes and 47 seconds and airway was secured with a 3-mm i d oral endotracheal tube and then ventilated with 100% oxygen .
there was no fetal bradycardia and cord was clamped and handed over to the neonatologist for further resuscitation .
isoflurane was stopped and oxytocin 20 units in 500 ml ringer lactate solution was administered .
the neonate was taken up for definitive surgery after 4 hours of stabilization in the nicu .
fetal mri showing large multiseptated cystic mass compressing trachea fetal head and mass delivered intubation attempts failed fetus delivered up to sternal notch successful intubation
exit was initially described to remove the tracheal clip which was utilized to plug in fetuses with cdh ( congenital diaphragmatic hernia ) . by this procedure , the fetus is maintained on utero - placental circulation without hypoxia to the neonate .
later it was expanded to other procedures such as intubation for fetuses with large neck masses .
the exit procedure maximizes the chance of survival for theses fetuses with large neck masses .
martino et al . , had described respiratory distress in their five cases with antenatal detection of cervical teratoma .
two of them were planned for exit but lost one baby due to difficulty in intubation as the baby was delivered via naturalis due to preterm delivery .
our patient had a large neck mass compressing the trachea and successful intubation could be performed because of exit .
the philadelphia group has explained the procedure of exit in great detail and the procedure was performed as advised .
on reviewing the literature , mother has been positioned either in supine with left uterine displacement , modified lithotomy , or lithotomy .
murphy dj et al . had positioned the mother in the modified lithotomy position for better access during intubation of the fetus .
considering the technical difficulties for the obstetrician to deliver the fetal head in lithotomy position on one hand and easy accessibility it would offer for the intubation of the fetus on the other hand , we decided to position the mother supine with left tilt and lower limbs abducted for the anesthesiologist to stand in between the patient 's legs for better access to the fetal airway [ table 1 ] .
the ideal positioning of the fetus before intubation has been to deliver the head and neck upto thorax . in our patient ,
the fetal head with the neck mass was not delivered upto the thorax because of the two twists of the cord around the neck . as the positioning was sub - optimal
the head and neck up to the sternal notch facilitated the proper alignment of the oro - pharyngo - laryngeal axes and successful intubation .
it would be ideal to unwind the umbilical cord while the fetus is still in the relaxed uterus , but unwinding the tight loop could lead to undue traction on the cord compromising the fetal circulation .
optimal position of the mother , anesthesiologist , and fetus for intubation during exit the uterus should be relaxed in order to maintain placental circulation .
a uterine stapler minimizes blood loss and this is essential if a longer procedure is planned . due to non - availability of staplers and
as we had planned only for intubation we could perform hysterotomy without staplers and the blood loss was approximately 400 ml .
exit has a definite role in fetuses diagnosed with large neck masses where airway compromise is suspected .
optimal positioning of the mother , obstetrician , anesthesiologists , and fetus is of paramount importance for successful and safe exit , especially in fetus with cord around the neck .
thus , exit can be safely performed in indian set up , with adequate planning and execution . | ex - utero intrapartum treatment ( exit ) is performed for fetuses diagnosed with large neck masses .
a case report of a fetus diagnosed with a large cystic hygroma and cord around the neck who was delivered by exit is presented .
the airway challenges and optimal positioning is discussed . |
supplementary material is available for this article at 10.1007/s13659 - 012 - 0035-y and is accessible for authorized users .
| solenostemon scutellarioides is a commercially important ornamental plant . in present study
, it was attempted to establish a protocol for high frequency in vitro regeneration of s. scutellarioides .
caulogenesis was found to be significant in solid ms medium supplemented with 0.5 mg dm3 bap .
combination of ga3 ( 0.5 mg dm3 ) and bap ( 0.5 mg dm3 ) induced high frequency regeneration coupled with higher plant height .
the plantlets exposed to iba ( 1 mg dm3 ) exhibited significant root development in terms of first appearance of root and number of roots per shoot .
to improve its commercial acceptability in terms of plant architecture and foliage colouration , two weeks old in vitro grown plantlets were exposed to different culture conditions namely ms strength , sucrose concentration , ph and light .
a variable plant types with diverse ornamental traits were developed under different culture conditions .
electronic supplementary materialsupplementary material is available for this article at 10.1007/s13659 - 012 - 0035-y and is accessible for authorized users . |
tomato ( solanum lycopersicum ) is one of the most important vegetable crops cultivated worldwide .
tomato has a diploid ( 2n = 2x = 24 ) and relatively compact genome of approximately 950 mb .
genetic linkage maps of tomato have been created by crossing cultivated tomato ( s. lycopersicum ) with several wild relatives , s. pennellii , s. pimpinellifolium , s. cheesmaniae , s. neorickii , s. chmielewskii , s. habrochaites , and s. peruvianum .
introgression lines generated from a cross between s. lycopersicum and s. pennellii have contributed to the isolation of important loci and quantitative trait loci ( qtls ) related to fruit size by utilizing dna markers on the tomato - expen 2000 genetic map [ 49 ] .
such interspecies genetic mapping is effective because the divergent genomes provide many polymorphic dna markers .
in contrast , intraspecies mapping is less popular in tomato because of the low genetic diversity within cultivated tomatoes that has resulted from the domestication process and subsequent modern breeding .
recently , we developed snp , simple sequence repeat ( ssr ) , and intronic polymorphic markers using publicly available est information and bac - end sequences ( bess ) derived from
heinz 1706 , a standard line for tomato genomics [ 11 , 12 ] , and applied these markers to create linkage maps between micro - tom and either ailsa craig , a greenhouse tomato , or m82 , a processing tomato , by mapping 1,137 markers .
micro - tom , a dwarf cultivar , is regarded as a model cultivar for functional genomics of tomato because of several characteristics , including small size ( 20 cm plant height ) , short life cycle ( 3 months ) , existence of indoor cultivation protocols under normal fluorescent conditions , and high - efficiency transformation methods that have been developed for this line [ 1315 ] .
the dwarf phenotype of micro - tom is the result of mutations in at least two major recessive loci .
dwarf ( d ) encodes a cytochrome p450 protein , which functions in the brassinosteroid biosynthesis pathway .
another locus , miniature ( mnt ) , is suggested to associate with gibberellin ( ga ) signaling without affecting ga metabolism , but the causal gene has not been identified to date . in japan , micro - tom genomics resources have been extensively accumulated , mainly in the framework of the national bioresource project ( nbrp ) ( http://tomato.nbrp.jp/indexen.html ) .
large - scale ethyl methanesulfonate ( ems ) and gamma - ray - mutagenized populations have been created , and visible phenotype data have been accumulated [ 1820 ] . the availability of micro - tom genome sequence data will accelerate the mapping of mutant alleles .
. currently , about 90,000 bess are available at the sol genomics network ( sgn , http://solgenomics.net/ ) .
bac - end sequencing has been conducted for other crop species . in the rice indica cultivar
reference genome . as a result , 12,170 paired bess were mapped that covered 80% of the rice genome . recently
, bac - end sequencing has been performed in crop plants with higher genome complexity .
bess from a commercial sugarcane variety , an interspecific hybrid with complex ploidy , were generated to analyze microsynteny between sugarcane and sorghum . in wheat , which has a complex hexaploid genome , the short arm of chromosome 3a was flow sorted to make a bac library , and chromosome arm - specific bess were generated for dna marker development . in switchgrass , more than 50,000 ssrs were identified from 330,000 bess , and this enabled detailed analysis on the evolution of this species . a low level of genetic variation has been observed for cultivated peanuts .
polymorphic ssrs were accumulated from the bess and successfully used in the construction of a genetic map .
bac - end sequencing can be useful as a resource for performing comparative genomic studies through mapping of the sequences to a reference genome and by facilitating the development of polymorphic dna markers . in the present study , we generated 93,682 single - pass end sequences from a micro - tom bac library . to compare the structures between the reference tomato
heinz 1706 genome , mapping of unique ends was performed , and possible genome rearrangements and polymorphisms were identified .
micro - tom ( tomjpf00001 ) seeds were obtained from the nbrp ( mext , japan ) and sent to the clemson university genomics institute ( cugi ) for bac library construction .
the genomic dna was partially digested , and fragments were cloned into the hind iii site of pindigobac536 .
a total of 55,296 clones in escherichia coli dh10b cells were arrayed in 144 384-well plates . to analyze bess ,
the bac dnas were amplified using a templiphi large - construction kit ( ge healthcare , uk ) , and the end sequences were analyzed according to the sanger method , using a cycle sequencing kit ( big dye - terminator kit , applied biosystems , usa ) with a type 3730xl dna sequencer ( applied biosystems ) .
the resulting sequence reads were quality checked with phred [ 27 , 28 ] , allowing the identification and removal of low - quality ( qv < 20 ) sequences .
the 93,682 reads clearing the quality criteria were submitted to ddbj / genbank with accession numbers ft227487-ft321168 .
bes reads were subjected to similarity search using the blastn program [ 29 , 30 ] . to isolate unique sequences from repetitive ones ,
93,682 bess were searched against the repeat database in itag2.3 ( http://solgenomics.net/ ) using a cutoff e - value of less than 10 .
the remaining sequences were searched against the published version of the heinz 1706 genome ( sl2.40 ) , which was accessed from the sgn database ( http://solgenomics.net/ ) . from all of the blast alignments ,
bess were extracted according to the following criteria , suggested in a previous report : ( 1 ) sequence identity > 90% and alignment coverage > 50% ; ( 2 ) mapped positions of each pair of ends < 200 kb apart in the same chromosome ; ( 3 ) direction of each paired end is correct ; ( 4 ) blastn e < 10 ; ( 5 ) a minimum of one hit for one of the paired ends ; ( 6 ) no redundant chromosomal locations
. sequence polymorphisms ( snps and indels ) between micro - tom and heinz 1706 were predicted based on the blastn alignment . since we did not allow a gap exceeding 27 bases , only indels up to 26 bases in length were counted .
mapped data and snp / indel sites were made accessible through the database tomatomics at http://bioinf.mind.meiji.ac.jp / tomatomics/. bac clones are available upon request from nbrp tomato ( http://tomato.nbrp.jp/indexen.html ) .
the bac insert size distribution was deduced based on the mapping results . according to these results , 45.4% ( 6,396 out of 14,101 ) of the bacs ranged from 100 to 120 kb , with average and median sizes of 101.3 kb and 101.8 kb , respectively ( figure 1 ) . by multiplying by the number of clones ( 55,296 ) , this bac library covers
micro - tom bes mapping to the heinz 1706 genome was processed as indicated in figure 2 . by eliminating repetitive , redundant , and unmapped sequences
, 28,804 unique pair ends and 8,263 unique ends were selected .
paired - end sequences were mapped onto the reference tomato genome sequence , and 2,248 hypothetical bac contigs were constructed ( see details at tomatomics , http://bioinf.mind.meiji.ac.jp/tomatomics/ ) . the integrity of the hypothetical contigs was confirmed by linking to the dna markers on two genetic maps , amf2 and mmf2 ( see supplementary table 1 in supplementary material available online at doi:10.1155/2012/437026 ) .
the genome coverage of the hypothetical bac contigs was assessed by applying euchromatin / heterochromatin boundary information from the genetic map expen2000 .
the results indicated that the euchromatin coverage ranged between 45.1% and 71.1% ( average , 58.9% ) among the different chromosomes , while heterochromatin coverage ranged between 57.4% and 75.3% ( average , 67.3% ) .
the total physical length of the bac contigs was 495,833,423 bp , covering 65.3% of the total chromosomes ( table 1 ) . to assess the occurrence of genome rearrangements , micro - tom and the reference tomato
possible inversions , translocations , and insertions were considered . to eliminate an artificial effect ( e.g. , chimeric bac clones ) ,
only regions covered by more than two bac clones were selected . after removing regions that had cleared the criteria for extraction ( see section 2 ) but were either shown to be multicopy by manual evaluation of the blast results or displayed similarity to transposable elements , we obtained two cases of a possible rearrangement between micro - tom and heinz 1706 ( table 2 ) . on chromosome 2 ,
the size of this inversion could be 20220 kb depending on which end of the bac clone is inversed .
translocation and inversion were observed on chromosome 3 . for each of two bac clones ( mtbac041l05 and mtbac077o14 ) , one of the ends
was mapped to 6,601 kb of chromosome 3 , while the other end was mapped to 55,665 kb , more than 49 megabases apart .
snps and indels between micro - tom and heinz 1706 were identified . among the snps and indels
found , 171,792 were mapped on 12 chromosomes , and 2,635 were mapped on pseudomolecules with no chromosomal information ( sl2.40ch00 of the tomato whole - genome shotgun chromosomes ) ( table 3 and supplementary table 2 , see details at tomatomics ) . according to these results , among the mapped snps and indels , a total of 30,930 polymorphisms were found in the euchromatin ( 1 out of 3,565 bp ) , and 140,862 were found in the heterochromatin ( 1 out of 2,737 bp ) .
transversion - type snps were observed in 83,262 cases , while 60,631 were transition - type snps . among the 30,534 indels , single - base insertions ( on the sl2.40 version of the tomato whole - genome shotgun chromosomes ) were observed in 10,740 cases , and single - base deletions were seen in 17,064 cases .
the remainder were larger indels , ranging from 2 to 26 bp ( supplementary table 2 ) .
by selecting unique end sequences from 93,682 reads , 28,804 paired ends ( 14,402 pairs ) and 8,263 unpaired ends were obtained .
the majority of the nonselected sequences ( 43,598 ) were derived from repetitive regions . for the rest , 10,943 had redundant hits to the heinz 1706 genome , possibly including repetitive sequences that were not represented in the repeat database in itag2.3 ( http://solgenomics.net/ ) , 2,015 showed weak similarity , and 59 showed no similarity ( figure 2 ) .
considering that the genome has been previously estimated to be composed of 25% gene - rich euchromatin [ 31 , 32 ] , bes selection in this study ( 39.6% , ( 28,804 + 8,263)/93,682 ) ) could have eliminated repetitive regions to a moderate degree .
micro - tom was bred by crossing the home - gardening cultivars , florida basket and ohio 4013 - 3 .
the pedigree of ohio 4013 - 3 suggested that a wild relative species was used in the breeding history [ 18 , 33 ] .
the micro - tom genome is now being sequenced ( draft sequence data available at ddbj with the accession number dra000311 ) , and mapping of orphan bess to the de novo assembly of micro - tom genome data will help to clarify this question .
the total physical length of micro - tom bac contigs was 495,833,423 bp , which covers approximately 65.3% of the dna from all 12 chromosomes . in the kasalath rice bes analysis ,
chromosomal coverage in relation to the reference nipponbare pseudomolecule was about 80% , despite the lower number ( 78,427 ) of analyzed bess .
because we used the same criteria for repetitive sequence selection ( e < 10 ) , the discrepancy between the two studies might be due to the larger genome size of tomato ( 950 mb ) compared with rice ( 430 mb ) .
our micro - tom bac coverage is reasonable , taking into account the scale of the bac library used .
micro - tom has been considered as a model cultivar to promote functional genomics studies of tomato by taking advantage of its characteristics .
currently , many tools and platforms have been developed , and some of these are already available to the research community .
the present study characterized the overall polymorphisms found between micro - tom bess and the reference tomato heinz 1706 genome .
in addition , two possible genome rearrangement events , on chromosome 2 and chromosome 3 , were observed ( table 2 ) . in the case of translocation and inversion on chromosome 3 , a gene annotated as reverse transcriptase was found in the flanking region ( solyc03g104840.1 ) .
we speculate that this region was translocated by the activity of a retrotransposon , as it was in the case of sun .
enhanced expression of sun caused by a gene duplication event mediated by the retrotransposon rider led to an elongated fruit shape . in the future
, we plan to sequence the entire bac and expect that this will help us to characterize these events in more detail . in the case of the other rearrangement possibility , on chromosome 2
since these rearrangements took place in euchromatin , which is rich in genes , these regions could represent an interesting target to investigate their possible effects on phenotypic variation between micro - tom and the reference tomato .
we mapped the polymorphisms and depicted them , alongside maps showing covered regions and gaps , in figure 3 . on chromosomes 2 , 5 , and 11
, polymorphisms seemed to be concentrated in the heterochromatic regions ; however , this tendency was not clearly observed in the other chromosomes . for the other regions , the polymorphism discovery rate seemed to be somehow correlated with the bac coverage .
although our analysis indicated little possibility of large - scale genome rearrangement between micro - tom and heinz 1706 ( table 2 ) , this uneven polymorphism distribution suggests the existence of highly divergent chromosomal regions .
the gaps in the hypothetical micro - tom bac contigs could have resulted from low coverage of the bac library , but the occurrence of chromosomal segments specific to either micro - tom or
the ongoing micro - tom genome sequencing and de novo assembly of the micro - tom genome will clarify the genome structure in detail , enabling a more solid assessment of the differences between micro - tom and heinz 1706 .
we had previously developed snp markers among several cultivated tomatoes . by selecting snps through in silico analysis using public est information and
previously developed ssr markers , 1,137 markers were obtained and successfully mapped on linkage groups between micro - tom and either ailsa craig or m82 . in the present study , we identified 171,792 snps and indels and mapped them on 12 chromosomes .
the average density was 1 snp per 3,565 bp in euchromatin and 1 snp per 2,886 bp in the genome in general ( including both euchromatin and heterochromatin ) .
previously , large - scale micro - tom full - length cdna analysis and comparison of exon regions with those on the heinz 1706 genome revealed a mean sequence mismatch of 0.061% ( 1/1,640 bp ) .
we used the published version of the heinz 1706 genome sequence , which has higher coverage , giving rise to greater accuracy , although our selection may still contain sequence errors because bess are single - pass sequences .
the information provided in this study will be useful in the development of dna markers between micro - tom and cultivated tomatoes , which will facilitate a better understanding of the physiological and metabolic differences between them .
it would also be useful in the genetic mapping of micro - tom mutants through the generation of f2 segregating populations . | a total of 93,682 bac - end sequences ( bess ) were generated from a dwarf model tomato , cv .
micro - tom . after removing repetitive sequences ,
the bess were similarity searched against the reference tomato genome of a standard cultivar ,
heinz 1706 . by referring to the heinz 1706 physical map and by eliminating redundant or nonsignificant hits , 28,804 unique pair ends and 8,263 unique ends were selected to construct hypothetical bac contigs .
the total physical length of the bac contigs was 495 , 833 , 423 bp , covering 65.3% of the entire genome .
the average coverage of euchromatin and heterochromatin was 58.9% and 67.3% , respectively . from this analysis ,
two possible genome rearrangements were identified : one in chromosome 2 ( inversion ) and the other in chromosome 3 ( inversion and translocation ) . polymorphisms ( snps and indels ) between the two cultivars were identified from the blast alignments . as a result , 171,792 polymorphisms were mapped on 12 chromosomes . among these , 30,930 polymorphisms were found in euchromatin ( 1 per 3,565 bp ) and 140,862 were found in heterochromatin ( 1 per 2,737 bp ) .
the average polymorphism density in the genome was 1 polymorphism per 2,886 bp . to facilitate the use of these data in micro - tom research , the bac contig and polymorphism information are available in the tomatomics database . |
the innate immune system or the nonspecific immune system is the first and the oldest line of defense in organisms [ 1 , 2 ] .
it was the most dominant form of immunity before the evolution of the more sophisticated adaptive immunity .
it is comprised of various mechanisms which are responsible for rapid defense of the host organism against invasion by other factors in a nonspecific manner .
the innate immune system differs from the adaptive immune system in a way that while it is able to defend the body against pathogens , it is not able to impart long - lasting immunity to the host , unlike the latter . despite the evolution of the more complex and specific adaptive immunity
, innate immunity still continues to function as the primary line of defense for most organisms .
the antimicrobial action in the innate immunity is mediated by various antimicrobial proteins and peptides , which have been evolutionary conserved .
antimicrobial peptides are small peptides which demonstrate broad - spectrum antibiotic activity against various gram - positive and gram - negative bacteria , fungi , protozoa , and viruses [ 58 ] .
while the most common mechanism of action deployed by these peptides is perturbation of microbial cell membrane [ 911 ] , there are other mechanisms which are also prevalent [ 1216 ] . due to increasing resistance to antibiotics
use of antimicrobial peptides is one of the promising approaches which may lead to potential antimicrobial drugs [ 16 , 2024 ] .
it has been observed that peptides which are predominantly cationic and hydrophobic in nature show potent antimicrobial activity [ 5 , 2529 ] .
many of these peptides including indolicidin from bovine neutrophiles , tripticin from porcine neutrophil granules , puroindoline from wheat seeds , combi-1 , a synthesized antimicrobial peptide , and lys h and lys c from lysozyme have been extensively studied .
although these peptides adopt various conformations the alpha - helical conformation with polar and nonpolar groups on opposite sides of the helix tends to be the most abundant [ 3537 ] .
the antimicrobial property of these amphipathic alpha - helical peptides increases sequentially with increase in net charge .
lactoferrin is an iron - binding glycoprotein which is found in most of the exocrine secretions such as milk , tears , nasal secretions , saliva , urine , uterine secretions , and amniotic fluids [ 3941 ] as well as in secondary granules of neutrophils .
it exerts a wide antimicrobial activity against a number of bacterial , viral , and fungal pathogens in vitro [ 4348 ] .
lactoferrin exerts its antimicrobial action not just in the form of the intact molecule but the monoferric lobes and active peptides of lactoferrin also have a role in the host defense against microbial disease [ 4952 ] .
lactoferrin is a rich source of cationic and hydrophobic antimicrobial peptides , which may be used against microbes [ 53 , 54 ] .
these antibacterial peptides which are a part of the polypeptide chain of lactoferrin and are released upon the proteolysis of this molecule by various proteolytic enzymes can be developed into clinically useful lead molecules for antimicrobial therapeutics [ 55 , 56 ] .
although it has been shown that native lactoferrin exerts its antimicrobial action through sequestration of iron [ 5759 ] , it is still unclear how the antimicrobial lactoferrin peptides act against the microbes .
though a number of studies have implicated these peptides in the binding to the outer membrane proteins of various bacteria or binding to microbial proteases [ 6062 ] , the structure - function interrelationships of these peptides have not yet been established .
a number of functional peptides are produced from lactoferrin by the action of proteolytic enzymes .
it is expected that these enzymes are present in the gastrointestinal tract as well as the site of microbial infection , and hence , they may contribute in the natural function of lactoferrin in the human body .
this paper reports the comparison of these peptides with other antimicrobial peptides found in nature as well as cross - species comparison of the sequences of these antimicrobial peptides from the native sequences of lactoferrin with the intent to draw evolutionary inferences of their function .
although many antimicrobial peptides from lactoferrin have been isolated and characterized , only three of them have been studied in detail .
these are lf1 - 11 , lactoferrampin , and lactoferricin . the sequences of these peptides indicate that these peptides belong to the n - terminal half of lactoferrin ( figure 1 ) .
hydrophobicity , cationicity , and helical conformation of these antimicrobial peptides are the important characteristics that determine their antimicrobial potency [ 9 , 63 , 64 ] .
all these peptides have high pi values ( > 9 ) and is expected to interact with negatively charged elements .
all three have different sequences , structural elements , and modes of action ( table 1 ) .
an attempt to analyze and decipher the structural and functional characteristics of three peptides is made in this paper .
lf1 - 11 , as its name suggests , is the n - terminal peptide of lactoferrin , comprised of the first eleven residues of the molecule .
this peptide has been shown to be highly effective against five multidrug - resistant acinetobacter baumannii strains and methicillin - resistant staphylococcus aureus and various candida species [ 67 , 68 ] .
the potent antimicrobial effect of lf1 - 11 was attributed to the first two arginines at the n - terminus of human lactoferrin .
this conclusion was based on the fact that when the second or third arginines were replaced by alanine , the candidacidal activity of the lf1 - 11 was observed . additionally , while lf1 - 11 , lf2 - 11 , and lf3 - 11 showed comparable candidacidal activities , the same was found compromised in the case of lf4 - 11 .
the importance of the three arginines ( r2r4 ) for the potent antimicrobial activity of this peptide was established when synthetic peptides lacking the first three n - terminal residues were found to be less effective in the killing of bacteria .
also , mutant lactoferrin lacking the first five n - terminal residues displayed decreased binding to bacterial lipopolysaccharide . in yet another study ,
stallmann et al . studied the efficacy of local prophylactic treatment with human lf1 - 11 in a rabbit model of femur infection and observed that hlf1 - 11 effectively reduced the development of osteomyelitis in a rabbit model .
it was speculated that the mechanism of antimicrobial action of lf1 - 11 is mitochondrial damage , with the extracellular atp being essential but not sufficient for lf1 - 11 to exert its candidacidal activity . in later studies
, it was found that uptake of calcium by mitochondria is vital for killing of candida albicans by the lf1 - 11 . in another study , it was found that lf1 - 11 is responsible for directing the gm - csf - driven monocyte differentiation toward macrophages that produces both pro- and anti - inflammatory cytokines . it was speculated that the peptide could be used as agent to empower the innate immune response of the host for infections .
these results demonstrated the importance of the further development of lf1 - 11 as a promising drug against microbial infections in patients who may have compromised immune systems .
the cellular target for the immunomodulatory activity of lf1 - 11 was found to be myeloperoxidase , to which lf1 - 11 binds and inhibits after entering the monocytes .
a molecular modeling study by the same group demonstrated that lf1 - 11 bound at the active site of the enzyme .
the importance of the first two arginines and the cysteine at the tenth position was further substantiated by the fact that peptides which did not possess these necessary residues were not as effective in binding with myeloperoxidase .
the sequence comparison of lf1 - 11 among the six species ( table 2 ) shows that unlike human lf1 - 11 , which contains three arginines in the positions 25 ( r2r4 ) , the peptide from other species contains only one arginine ( r3 ) .
yet , it is noteworthy that the r4 has been replaced by lysine , which is also a basic residue in all the other species , thereby maintaining the highly cationic nature of the peptide throughout the species . also , in all the cases except human , arginine occurs at the seventh position also .
the most significant change is seen in the second residue which is proline in all cases except in human and camel .
notably , the hydrophobic residues , v6 , and w8 are conserved in all the cases .
lactoferrampin , comprised of residues 268284 in the n1 domain of lactoferrin , has been identified as an antimicrobial peptide and plays a key role in membrane - mediated activities of lactoferrin [ 76 , 77 ] .
it exhibits broad antimicrobial action against several gram - positive and gram - negative bacteria , notably , bacillus subtilis , escherichia coli , pseudomonas aeruginosa , and staphylococcus aureus , as well as candidacidal activity .
the antimicrobial action of this peptide was also found to be more potent than the native lactoferrin .
the structure of lactoferrampin revealed an amphipathic alpha - helix which begins with the n - terminus and ends at the 11th residue , followed by a c - terminus tail .
it is reported that the cleavage of this peptide at both the termini resulted in considerable decrease of the candidacidal activity .
the c - terminal residues of lactoferrampin are most critical for its antimicrobial action , possibly because the c - terminus consists of several residues with positive charges which are clustered together .
but truncation of c - terminal side did not alter the ability of this peptide to adopt helical conformations .
also , substitution of the basic residues at the c - terminus led to decrease in potency of this peptide [ 77 , 79 ] .
the n - terminal residues , truncated up to the sequence 270284 , are essential for maintaining the structure of this peptide in a helical conformation .
the helical conformation of this peptide was found to be critical for the potency against gram - positive bacteria as established when the bactericidal activities of two lactoferrampin peptides , lactoferrampin 265284 and lactoferrampin 268284 , were compared .
lactoferrampin 265284 , which consists of additional three residues , asp - leu - ile , showed a broader specificity since the asp - leu - ile sequence increases the tendency of this peptide to assume an alpha - helical conformation .
both the peptides possessed bactericidal activity against certain species of gram - positive and gram - negative bacteria .
compared to lactoferrampin 268284 , higher concentrations of lactoferrampin 265284 were required to kill the gram - negative bacteria , e. coli and p. aeruginosa .
the killing activity expressed as lc50 value ( the concentration that produced 50% reduction in viable counts of the microorganisms ) was found to be about 5.8 mol / l for lactoferrampin 268284 which is about 4 times higher than lactoferrampin 265284 .
the mode of action of this peptide on bacteria is by bacterial membrane binding and membrane disruption .
it is established that lactoferrampin is internalized within few minutes with the bacterial membrane permeabilization followed by cellular damage [ 81 , 82 ] .
distinct vesicle - like structures by the lactoferrampin peptide were also observed by freeze - fracture transmission electron microscopy in the membrane of c. albicans . it is speculated that this peptide exerted detergent - like activity , disturbing the hydrophobic interphase of the lipid bilayer .
several studies have revealed that the determinants for antimicrobial action are the orientation and structure of bovine lactoferrampin in bacterial membranes [ 78 , 8385 ] .
the solution structure of bovine lactoferrampin suggests that it adopts an amphipathic alpha - helical conformation across the first 11 residues of the peptide but remains comparatively random at the c - terminus [ 78 , 85 ] .
the interaction between the n - terminal tryptophan residue and model membranes of varying composition was evaluated suggesting that w1 is inserted into the membrane at the lipid / water interface .
along with this , the orientation of the phenyl side chain of f11 found to be in same direction as the indole ring of w1 also suggested that the amphipathic n - terminal helix anchors the peptide to membrane with these two residues that facilitates peptide folding [ 78 , 86 ] .
the same group has suggested that the hydrophobic patch in between the two residues as well as leu , ile , and ala side chains are responsible for interaction between the peptide and the hydrophobic core of a phospholipid bilayer .
in addition , the helix capping residues asp - leu - ile in the n - terminus of the peptide has been found to mediate the depth of membrane insertion by enhancing the affinity for negatively charged vesicles .
bovine lactoferrampin had been shown to have greater affinity to acidic phospholipids than that to neutral phospholipids .
haney et al . have speculated a two - step model of antimicrobial action by this peptide where the c - terminus positive charge cluster helps in the primary attraction of lactoferrampin to the membrane followed by the helix formation at the n - terminus that interacts to the surface of the bacterial lipid bilayer .
the sequence comparison of lactoferrampin from six different species shows uniform preponderance of cationic amino acid residues among hydrophobic residues ( table 3 ) .
the hydrophobic domain contains w1 in all the species that is involved in membrane insertion .
bovine lactoferrampin 268284 has a net positive charge of 5 + at neutral ph with hydrophobic domain .
the hydrophobic moment ( ) of the peptide which is a measure of lipophilicity was found to be 5.42 .
in contrast , the human lactoferrampin has a net charge of 2 + resulting in reduced antimicrobial activity .
however , by increasing the net positive charge near the c - terminal end of human lactoferrampin , a significant increase in its antibacterial and candidacidal activity was obtained .
the basic amino acid residues crucial for the antimicrobial action were found to be conserved among all the six species .
lactoferricin is a multifunctional , 25-residue peptide that is generated upon cleavage of native lactoferrin by pepsin and represents amino acid residues number 1741 in lactoferrin .
the lactoferricin peptide is different from the other peptides described so far as it contains a disulfide bond between residues cys 20 and cys 37 in human lactoferrin and cys 19 and cys 36 in bovine lactoferrin .
the peptide has an abundance of basic amino acids like lysine and arginine as well as hydrophobic residues like tryptophan and phenylalanine .
the first report on lactoferricin in 1992 described this peptide to be more potent as an antibacterial agent in comparison with the intact lactoferrin and it was demonstrated to cause a rapid loss of colony - forming capacity in most of its targets .
however , some strains like pseudomonas fluorescens , enterococcus faecalis , and bifidobacterium bifidum strains were found to be resistant to lactoferricin .
the antibacterial activity of this peptide was attributed to its action of releasing lipopolysaccharide from bacterial strains and , hence , disruption of cytoplasmic membrane permeability after cell binding [ 8991 ] .
apart from having a broad antibacterial spectrum , lactoferricin was found to be highly potent against candida albicans [ 89 , 92 , 93 ] .
recently , it has also been shown to have antiviral [ 94 , 95 ] and antiprotozoal activities .
it also displayed other activities like inhibition of tumor metastasis and induction of apoptosis in human leukemic cells .
the mechanism of action of lactoferricin was attributed to 11-amino - acid amphipathic alpha - helical region which is positioned on the outer surface of the n - lobe of lactoferrin .
the proline at the 26th position ( p26 ) was found to be essential for the antibacterial activity , and it was speculated to be responsible for disruption of the helical region , and hence the helicity of the peptide was predicted to be an essential aspect of the antibacterial action of this peptide .
lactoferricin was found to be produced in the human stomach , indicating that this peptide is definitely generated in vivo for host defense .
the comparison of the antimicrobial activities of lactoferricin from human , bovine , murine , and caprine showed that bovine lactoferricin was the most potent .
the minimal inhibitory concentration ( mic ) of lactoferricin b differs according to their source .
a comparison of the mic values of lactoferricin shows that bovine lactoferricin is the most potent .
the mic of bovine lactoferricin against certain e. coli strains has been found to be around 30 g / ml while that derived from human is more than 100 g / ml . the efficiency of antibacterial activity of bovine lactoferricin is due to the presence of high amount of net positive charge ( + 8 ) and hydrophobic residues ( primarily w6 , w8 , and m10 ) [ 51 , 88 ] .
the action of this peptide is also dependent on the ph [ 89 , 103 ] .
it was shown that only six central residues ( 49 ) among the twenty five residues of the peptide are required for its antimicrobial activity .
it may be noted that the tetrapeptide krds is present only in human lactoferrin while it shows variations in the sequence of others .
it has been reported that krds inhibits platelet aggregation [ 105 , 106 ] .
the bovine lactoferricin has been demonstrated to interact with the negatively charged elements in the membrane of susceptible bacteria and disrupt the cell membrane .
a synthetic peptide derived from human lactoferricin has been found to be effective in depolarizing the bacterial cytoplasmic membrane with a loss of ph gradient .
the permeabilizing effect of bovine lactoferricin causes membrane disruption resulting in inhibition of macromolecular biosynthesis and ultimately cell death .
the mode of action is however different in gram - positive and gram - negative bacteria . in gram - negative bacteria
antimicrobial peptides act on lipopolysaccharides and in gram - positive bacteria they act on lipoteichoic and teichoic acids .
in addition to antimicrobial properties , lactoferricin derived from human and bovine origin has also been found to be effective in inhibiting the classical complement pathway .
this implicates a role of these peptides in suppression of inflammatory effects caused by bacteria .
the sequence analysis of the lactoferricin from various species indicates that unlike bovine lactoferricin there is only one tryptophan at position 6 ( table 4 ) .
a further exception to this is equine which has no tryptophan residue in either of the positions .
this shows that the two tryptophans in lactoferricin are important for its optimal activity against microbes .
the solution structures of lactoferricin have been determined from bovine and human sources .
the bovine lactoferricin adopted a distorted antiparallel beta sheet , in complete contrast with its conformation in the intact lactoferrin , as observed in the structures obtained by x - ray crystallography .
however , the solution structure of human lactoferricin was closer to its structure in native lactoferrin since the amphipathic helix was preserved from gln14 to lys29 . however , the beta - sheet character was not observed in the solution structure of human lactoferricin either .
lipopolysaccharide ( lps ) , the outer membrane component of gram - negative bacteria , is one of the major causes of endotoxin - induced production of inflammatory cytokines and septic shock .
lactoferrin has been shown to neutralize the effect of lps - induced toxicity by binding to lps [ 115 , 116 ] .
the cationic peptide derived from lactoferrin which is responsible for this interaction and release of lps is first identified to be lactoferricin .
the residues from 28 to 34 of lactoferrin corresponding to the region in human lactoferricin have been identified to have a high affinity for binding to lps .
the initial binding of the peptide with e. coli has been found to be due to interaction with bacterial lps .
further studies have shown that bovine lactoferricin can arrest the lps - induced cytokine release by suppressing the il-6 response in human monocytic cells stimulated by lps .
a synthetic peptide corresponding to the antibacterial region of human lactoferricin was also found to facilitate depolarization of the bacterial cytoplasmic membrane , loss of the ph gradient , and a bactericidal effect in e. coli .
modelling studies using synthetic peptides derived from human and bovine lactoferricin have shown that these cationic peptides with their positively charged residues first interact with lps carrying negative charges .
this is followed by hydrophobic interactions between the tryptophan residues of the peptides and the lipid a molecule of lps to promote structural disorganization .
similarly , a synthetic peptide corresponding to 11 residues of human lactoferricin near its n - terminus has been found to bind to lps and neutralize the lps - induced adverse effects in vitro and in monocytes [ 121 , 122 ] . in yet another study using nmr , it has been observed that this peptide folds into a
conformation formed by its hydrophobic core and the two clusters of hydrophilic residues of the peptide targets the two phosphate moieties of lipid a in lps .
the antimicrobial peptides found in nature are classified into four groups according to a combination of their sequence homologies , functional similarities , and common three - dimensional structures .
the four groups include group 1 , which consists of linear , cationic , and amphipathic - helical peptides , for example , cecropins , magainins , bombinins , and temporins ; group 2 , which consists of -strands connected by intramolecular disulfide bridges , for example , human -defensin-2 , tachyplesins , and protegrins ; group 3 , which consists of linear peptides with an extended structure , characterized by overrepresentation of one or more amino acids , for example , tritrpticin and indolicidin ; and group 4 , which consists of peptides containing a looped structure , for example , bactenecin , brevinins , and esculentin . in the light of the above classification , human lf1 - 11 ( grrrsvqwcav )
consists of a highly variable loop region and a short -strand and is arginine rich , and hence can be classified in group 4 .
however , the same can not be said about lf1 - 11 from other species , since their conformation may be similar to the human lf1 - 11 in the structure , but they are not rich in arginines .
the arginine - rich fragment of this peptide is similar to other cationic arginine - rich peptides found in nature which have cell - penetrating activity and hence can traverse the plasma membrane of eukaryotic cells .
a significant example of arginine - rich peptide that has cell - penetrating property is arginine - rich hiv tat peptide ( grkkrrqrrrppq ) . on the other hand
, lactoferrampin belongs to group 1 which consists of linear , cationic , and amphipathic - helical peptides .
the alpha - helical amphipathic character of lactoferrampin has been compared with other group i peptides like magainins , bombinins , cecropin a , and temporins and are depicted by the helical wheel representation of the peptides in which the charged and polar residues are found aligned along one side and most of the amino acids with nonpolar side chains occupy the opposite side of the helical cylinder ( figure 3 ) .
the spatial segregation of the hydrophobic and hydrophilic residues designates the amphipathic nature of the peptides [ 127 , 128 ] .
these peptides upon interaction with target membranes fold into an amphipathic -helix with one face of the helix predominantly containing the hydrophobic amino acids and the opposite face the charged amino acids .
the presence of a prominent hydrophobic face is observed in the helical wheel representations of magainin 2 , bombinin , and temporin ( figures 3(a ) , 3(b ) , and 3(c ) ) whereas a pronounced cationic domain is present on the hydrophilic surface of the helical wheel diagram of bovine lactoferrampin like that in cecropin a ( figures 3(d ) and 3(e ) ) .
the positively charged domain is more distinct in bovine than human lactoferrampin ( data not shown ) .
the analysis suggests that there is very little similarity in the amino acid sequence within the group ; however there is a distinct trend in the distribution of different types of residue , that is , hydrophobic and charged , polar , and so forth , within the secondary structure of the helix .
sequence similarities between lactoferricin and dermaseptin and magainins suggest that lactoferricin may act as an amphipathic alpha - helix .
the active hexapeptide fragment within bovine lactoferricin peptide showed distinct similarities with lf1 - 11 and other amphipathic tryptophan and arginine - rich antimicrobial peptides found in other sources apart from lactoferrin ( table 5 ) .
bovine lactoferricin contains two tryptophan residues at positions 6 and 8 and two arginines at positions 4 and 5 .
these two amino acids have chemical properties which are one of the critical components of antimicrobial peptides .
enzymatic digestion of lactoferrin results in the generation of antimicrobial peptides which display antimicrobial properties , in some cases , with greater potency than the native lactoferrin , possibly for the protection of neonates against the invading pathogens .
these peptides , all from the n - lobe of lactoferrin , show a remarkable similarity to cationic antimicrobial peptides found in other invertebrate and vertebrate species .
though there may be minor variations in the sequence and the conformational features among these lactoferrin peptides from various species , the basic framework tends to be similar and conserved .
this indicates that these peptides play a significant role in the antimicrobial function of this protein .
the antimicrobial effect of cationic peptides of different origins is due to cytoplasmic membrane disruption of the target cell as well as immunomodulation .
the difference in their functional properties is due to the difference in their amino acid composition inspite of sharing amphipathic and cationic characteristics .
the presence of all these antimicrobial peptides in a single domain of lactoferrin suggests that the protein acts on the membrane interface and disturbs the membrane integrity resulting in its antimicrobial activity .
since lactoferrin is found in the milk and is ingested throughout the life of all neonates and most adults , it may be an excellent agent for administration to humans . in the future , | lactoferrin is a multifunctional , iron - binding glycoprotein which displays a wide array of modes of action to execute its primary antimicrobial function .
it contains various antimicrobial peptides which are released upon its hydrolysis by proteases .
these peptides display a similarity with the antimicrobial cationic peptides found in nature . in the current scenario of increasing resistance to antibiotics ,
there is a need for the discovery of novel antimicrobial drugs . in this context , the structural and functional perspectives on some of the antimicrobial peptides found in n - lobe of lactoferrin have been reviewed .
this paper provides the comparison of lactoferrin peptides with other antimicrobial peptides found in nature as well as interspecies comparison of the structural properties of these peptides within the native lactoferrin . |
aspirin - exacerbated respiratory disease ( aerd ) is a syndrome characterized by chronic hyperplastic rhinosinusitis , nasal polyposis , asthma , and aspirin sensitivity , as described in 1922 by szczeklik et al .
[ 1 , 2 ] . the prevalence of aerd is variable ; stevenson and szczeklik reported in 2006 that aerd occurs in 3% of adult patients with asthma in the united states , with the onset of symptoms during the third decade of life , and that it is more common in women than in men , with approximately 70% versus 30% in europe and 57% versus 43% in the usa .
the mechanisms underlying aspirin intolerance are not fully defined , with current research focusing on cyclooxygenase 1 ( cox-1 ) inhibition by aspirin and other nsaids diverting arachidonic acid metabolism from cox pathways to the lipoxygenase ( lo ) pathway .
this leads to increased synthesis of the cysteinyl - leukotrienes ( lt ) , ltc4 , ltd4 , and lte4 , resulting in bronchoconstriction , mucus hypersecretion , and possibly the development of polyps and urticaria .
the biological plausibility of this hypothesis fact has led to the search for polymorphisms in genes responsible for lt synthesis , to explore associations between these polymorphisms and local tissue levels of the proteins .
other factors such as polymorphisms in the genes for proinflammatory cytokines including tnf , il1b , il6 , and il8 are involved in chronic inflammatory and autoimmune diseases .
interleukin 1 ( il-1 ) is a cytokine associated with inflammatory responses and found in two forms , il-1 ( produced by the il1a gene ) and il-1 ( il1b ) , with both genes located on chromosome 2 .
il-1 is expressed in nasal polyps , nasal epithelium , macrophages , activated t lymphocytes , and monocytes ; its expression is regulated by adhesion molecules , and others inflammatory cytokines .
il1b polymorphisms have been associated with inflammatory bowel disease and gastric cancer among other diseases .
recently , genetic polymorphisms in proinflammatory cytokines such as il-1 have been recognized as key players in the pathogenesis of asthma .
similarly , il-8 has been implicated in the asthmatic inflammatory process , and genetic variation in this cytokine has been associated with both the susceptibility and the severity of this disease . in the present study
, we have investigated the frequencies of polymorphisms in the genes encoding these two cytokines in aerd patients .
patients with aspirin - exacerbated respiratory disease ( aerd ) ( n = 78 ) and aspirin - tolerant asthma ( ata ) ( n = 135 ) were recruited from the allergy and otolaryngology departments at the instituto nacional de enfermedades respiratorias ( iner ) . healthy control subjects ( hcs )
all participants underwent simple spirometry , inhaled methacholine challenge , and nasal challenge with lysine - aspirin ( l - asa ) according to international guidelines to determine the degree of bronchial hyperresponsiveness and confirm aerd diagnosis .
the aerd group had positive l - asa and methacholine challenges , the asthmatic group had positive methacholine challenge , but negative l - asa challenge , and the healthy control subjects were volunteers with negative l - asa and methacholine challenges .
a positive l - asa challenge was defined as a decrease of at least 40% in total nasal airflow after l - asa application compared with baseline measures ; methacholine challenge was considered positive with a decrease of at least 20% in forced expiratory volume in one second ( fev1 ) compared with baseline fev1 after the administration of different concentrations of methacholine ( beginning with 0.03 mg / ml , increasing gradually the concentration twice every 2 minutes until the concentration of 32 mg / ml ) ; in case it does not have it , it is considered negative .
the study was approved by the science and bioethics committees of iner , and all participants gave their informed consent [ 8 , 9 ] .
patients and healthy control subjects had ancestry of at least two generations born in mexico and were thus considered to be mexican mestizo in descent [ 10 , 11 ] .
peripheral blood was drawn by venipuncture , and genomic dna was obtained using the commercial bdtract dna isolation kit ( maxim biotech , san francisco , calif , usa ) .
the dna was quantified by absorption of ultraviolet light at 260 nm wavelength using an actgene spectrophotometer ( actgene , inc .
we selected two polymorphisms in two genes related to chronic inflammation : rs16944 in il1b and rs4073 in il8 .
allelic discrimination of snps rs16944 ( il1b ) and rs4073 ( il8 ) was performed by real - time pcr ( rt - pcr ) on a 7300 real time pcr system ( applied biosystems , calif , usa ) using taqman commercial probes ( applied biosystems , usa ) for each of the polymorphisms mentioned above and followed the cycling program : preread 50c , 1 minute ; absolute quantitation : 50c , 2 minutes , 1 cycle ; 95c , 10 minutes , 1 cycle ; 95c , 15 seconds , 60c 1 min , 40 cycles ; postread 50c , 1 minute .
the results were assessed taking into account the allelic discrimination and absolute quantitation in all samples ; additionally , we included four contamination controls per plate ( nontemplate controls ) .
the fluorescence signal detectors used were vic which was assigned to the b allele and fam assigned to the a allele for both snps .
statistical analysis was performed between groups of cases ( aerd and ata ) versus the healthy control subjects by test with 3 2 tables , using spss ( v. 15.0 ) software for windows , to identify the difference between the allele and genotype frequencies of each polymorphism evaluated .
in addition , odds ratios and 95% confidence intervals were calculated with epi - info ( v. 6.04 ) software .
we performed a genetic association study of the il8 ( rs4073 ) and il1b ( rs16944 ) gene polymorphisms in the three groups .
genetic data for the snps included in this study are shown in table 1 ; minor allele frequency ( maf ) of the polymorphisms tested in healthy control subjects had a similar distribution to that reported in international databases ( table 1 ) .
gene frequencies for each genotype within the three subject groups are shown in table 3 .
the frequency of genotypes aa , at , and tt of the il8 rs4073 snp was not statistically significant and different between the three studied groups .
analysis of gg and ga genotypes of the il1b ( rs16944 ) snp for aerd and ata patients versus the healthy control subjects showed nonstatistically significant associations .
interestingly , the aa genotype showed increased frequency in the aerd patients when compared to the ata group ( gf = 0.19 versus 0.07 ) ; this association was statistically significant ( p = 0.018 , or 2.98 , and ci 1.177.82 ) ( tables 3 and 4 ) and was not found when aerd or ata groups were compared to healthy control subjects .
there is no difference in aa versus ag + gg using contrast of healthy control subjects versus aerd patients ( data not shown ) .
the airways of aspirin - sensitive patients are characterized by chronic inflammation with cell infiltration even when they are not exposed to aspirin or other nsaids .
in addition to alterations in the metabolism of arachidonic acid , several proinflammatory cytokines have been associated with aerd .
the role of il-1 , however , has not been investigated previously . here , we report that aerd patients show an increased frequency of the il1b-511 polymorphism ( rs16944 aa genotype ) compared to aspirin - tolerant asthmatics .
interleukin-1 has been reported to be involved in the genesis of both asthma [ 13 , 14 ] and chronic rhinosinusitis with nasal polyposis in a turkish population .
most studies have attempted to establish the association of polymorphisms in the il1b promoter gene , mainly at positions-511 g / a ( rs16944 ) and -31 c / t .
for example , park et al . , in 2004 , did not find any association between these polymorphisms and either asthma or atopy in a korean population . in 2007 , erbek et al .
described a susceptibility for developing nasal polyps associated with the il1b-511 polymorphism ( rs16944 ) , but mfuna endam et al .
, in 2010 , failed to reproduce this finding in canadian patients with chronic rhinosinusitis . in 2003 , allen et al .
did not find any association with il1 gene polymorphisms in asthmatic families ( n = 244 ) , but they reported an association with the dna microsatellite d2s308 in these asthma families ( p = 0.00001 ) . in parallel ,
karajalein et al . evaluated 245 patients with asthma and nasal polyposis and did not find any association between the polymorphism il1b-511 c / t and nasal polyps .
evidence for the role of il-1 in pulmonary immune responses has been gathered in murine models of allergic asthma using il-1r1-deficient [ il-1r1 ( / ) ] mice ; changes observed in these mice include significant reduction of pulmonary eosinophilic inflammation , diminished goblet cell hyperplasia , and reduction of cell recruitment to the lungs , as compared to control balb / c mice .
however , there are no studies linking gene promoter polymorphisms and levels of expression of the cytokine in the lung microenvironment . in the present study
, we have demonstrated that the frequency of the il1b-511 polymorphism ( rs16944 aa genotype ) is three - fold higher in aerd ( 19.2% ) than in ata patients ( 7.4% ) , suggesting that patients carrying this polymorphism may exhibit genetic susceptibility to develop aerd .
findings on the biological functionality of the rs16944 polymorphism have not been consistent across studies .
the aa genotype has been associated with higher gastric mucosal levels of il-1 in bacterial infections , while mononuclear cells from subjects with the gg genotype showed an increased release of il-1 after stimulation with lipopolysaccharide .
recent studies suggest that the functional role of rs16944 may depend on il1b promoter region haplotypes including rs16944 [ 2326 ] .
although the findings are inconsistent , these previous studies suggest that rs16944 could affect the expression levels of il-1. our report is the first demonstration of the involvement of il1b polymorphism in aerd .
the sample size is relatively small , particularly in the aerd group , which may limit the statistical power , so it would be desirable to replicate our findings in an independent population .
future studies must investigate whether this cytokine is released in the airways of aerd patients and whether its levels relate to genetic polymorphisms in the il1b gene .
interleukin-8 has been implicated in asthma and found in high concentrations in bronchoalveolar lavage fluid of patients with acute asthma exacerbations .
in fact , polymorphic il8 alleles ( 251 t and 781c ) have been associated with asthma in a european population [ 7 , 27 ] , but not in asthmatics of korean origin ; the differences could be explained by the different ethnic populations studied . in contrast , korean asthmatics were found to show four nonsynonymous amino acid substitutions in the il-8 receptor a ( il8ra ) and an association of one synonymous variation in il8rb . in the present study ,
we did not find a significant difference in the rs4073 a / t between the aerd group and ata patients . | aspirin exacerbated respiratory disease ( aerd ) is characterized by chronic hyperplastic rhinosinusitis , nasal polyposis , asthma , and aspirin sensitivity .
the mechanisms which produce these manifestations of intolerance are not fully defined , current research focuses on cyclooxygenase 1 ( cox-1 ) inhibition , metabolism of arachidonic acid , and the cox pathway to the lipoxygenase ( lo ) route , inducing increased synthesis of leukotrienes ( lt ) .
the biological plausibility of this model has led to the search for polymorphisms in genes responsible for proinflammatory cytokines synthesis , such as il1b and il8 .
we performed a genetic association study between il8 - 251 ( rs4073 ) and il1b-511 ( rs16944 ) polymorphisms in aerd , aspirin - tolerant asthma ( ata ) , and healthy control subjects . using allelic discrimination by real - time pcr
, we found statistically nonsignificant associations between aerd , ata , and healthy control subjects for the gg and ga genotypes of il1b ( rs16944 ) .
interestingly , the aa genotype showed an increased frequency in the aerd patients versus the ata group ( gf = 0.19 versus 0.07 , p = 0.018 , or 2.98 , and 95% ci 1.177.82 ) .
this is the first observation that il1b polymorphisms are involved in aerd .
thus , future studies must investigate whether interleukin-1 is released in the airways of aerd patients and whether it relates to genetic polymorphisms in the il1b gene . |
traditionally , students learned in the classroom through interaction with the teachers , which required his physical presence and confidence in asking questions to clear his doubts
. many students may not take part in the discussion openly due to peer pressure and fear of asking a question that may appear silly and unimportant .
if the absorbing capacity of the student differs from the normal , he may find it difficult to follow the lectures , which are usually done quite fast within the time frame allotted for that particular lecture and the teacher has to disseminate the information in a cramped style .
the use of various aids like transparencies or powerpoint slides has not really helped in learning as they do not stimulate the processing of information delivered by the student . during contemporary medical education ,
portable and enduring and might enhance the recall of information leading to improved test performance .
note - taking by students is generally seen as an integral and essential part of the process of learning from lectures .
few studies have reported that the lecture handouts could improve understanding of the topic and students test performance . to further add on , many lecturers are now routinely publishing lecture handouts on the web for students to access .
this method of dissemination of information may provide many advantages over conventional paper - based distribution as it provides links to other online materials .
the technological advancement has made all the other methods redundant in the sense that students can learn at their own pace , contact the lecturer anytime , they want to post him questions and get an answer , explanation or clarification from the lecturer . at the same time
, the learning process is made easier by providing the students all the information in presentation slides .
handouts could be a valuable aid to guide the students to study in an organized way and is easy for them to cope with their exam stress .
writing in the blank spaces in handout during a lecture allows the students to be alert and active , and they learn by writing .
the present study was undertaken to find out the efficacy of the skeleton handouts against full lecture handout during ophthalmology theory lectures .
this was a pre- and post - test based experimental study involving 6 semester medical students ( n = 108 ) . in our syllabus ,
the ophthalmology theory lectures are covered during the 6 semester of medical training while the clinical posting in ophthalmology takes place during the 7 and 8 semester .
the topics of nearly equal difficulty level were identified with lecture gap of around 1-month . during the first intervention
, the full lecture handouts were distributed to each member of the class 4 days prior to concerned lecture . on the day of lecture ,
a pretest in the form of multiple choice questions was conducted for the whole class , and the post - test with the same questions was conducted soon after the lecture .
the same type of intervention was done for skeleton handouts as well after a gap of 1-month .
one hundred and eight students participated during the intervention with full handout whereas one hundred and four students participated in the skeleton handout intervention .
closed question regarding preference of students to either type of handouts was asked after both the interventions and the percentage of preference was noted .
data were processed using microsoft excel and analyzed using spss version 12 ibm , malaysia .
dependent sample t - test was used to compare the before and after test score in intervention groups , that is , skeleton handouts and full handouts .
anova with repeated measures was used to compare the change score from pretest between skeletal handouts and full handouts .
all the 108 students of the class participated in the intervention through full lecture handout in comparison to one hundred and four students participating in the skeleton handout intervention .
table 1 reveals that the posttest score ( 4.85 0.363 in skeleton handout and 2.61 0.771 in full lecture handout ) were significantly increased in comparison to respective pretest scores ( 1.85 1.275 in skeleton handouts and 1.92 1.09 in full lecture handouts ) [ table 2 ] . paired comparison of pretest and posttest scores in skeletal handout and full handout comparison of change score from pre - test between skeletal handout and full handout ( n=104 ;
those who participated in both full handout and skeleton handout interventions ) anova with repeated measures was used for the comparison of change score from pre - test between intervention groups .
skeleton handout has a mean change score from pretest 3.0 with sd 1.246 , which is higher than full handout having mean change score from pretest 0.70 with sd 1.238 .
there was a significant difference of mean change score between a skeletal handout and full handout ( p < 0.001 ) . in this study ,
the majority of the students preferring full handouts expressed that it was easy to revise at the last minute , and it could save a lot of work in the preparation and assimilation of the details about the subject taken
. however , most of the students among those who preferred skeleton handouts expressed that it encouraged them to stay focused during the class and all of them concurred that they could add points in the skeleton handouts that were being elaborated in the class .
they also said that they could get some more points and added on to the skeleton handouts by referring to information beyond textbooks .
handouts are a useful supplement that can support further learning in noncontact hours and also give a framework on which the students can learn .
they clarify expectations during the lectures and can encourage focused note - making by leaving specific areas for students to add notes in their own words or connect to other areas of learning .
provision of full slide presentation has decreased the students attentiveness as they already have the notes on what the lecturer is going to teach in the class with the belief that they can always refer to it at their leisure . however
, the skeleton handouts will provide an opportunity for the students to ponder over the subject and increase the understanding of the subject by the scaffold given .
the students who were given skeletal handouts out - performed when compared with students who were not given . in our study , the mean post - test value was significantly higher when the students were given skeleton handouts , which were similar to the one was done by russell et al .
they have concluded that learning by medical students was improved when they recorded notes in class .
however , the majority of the students in our study , preferred to have full handout ; probably for their easy referral during the examination .
when students are given full handouts , they often substitute this for class attendance and are less likely to attend the lecture , which sometimes is disheartening to the lecturer . perhaps the compromise that best assists students is the provision of partial or guided notes .
skeleton outlines or handouts can provide a scaffold for accurate student note - taking while still requiring their attention , active engagement , and attendance . guided rather than complete notes provided prior to class time can also allow students to prepare and review material for concepts in need of further explanation with the added benefits of increasing student engagement with the content and improving their accuracy of note - taking .
it has been stated in one of the literatures that the students who generated their own outlines or study questions learned more from a lecture than did students who used materials provided by the lecturer .
the limitation of our study was that we could not conduct a randomized control study , but we plan to conduct a randomized control study with long - term outcome measure in the next batch of students .
although the majority of the students wished to have full handouts of the ophthalmology lectures , they could perform better after going through skeleton handouts .
| background : although lecture handouts are commonly given to students during theory lectures , students perception , as well as their performance , can vary depending on the type of handouts they receive for information processing.methodology:this is a quasi - experimental study involving 6th semester medical students .
the study was conducted during theory lectures on ophthalmology .
the two types of notes given to the students were comprehensive handout and a skeleton handout , which included some lecture notes but required substantial annotation by the students .
pre - test and post - test in the form of multiple choice questions were conducted before and after the lecture session , respectively.results:there was a significant difference of mean score of pre- and post - test between skeletal handout ( pre = 1.85 1.275 , post = 4.85 0.363 ) and full handout ( pre = 1.92 1.09 post = 2.61 0.771 ) with p < 0.001
. however , the students responses to questionnaires indicated a strong preference for much detailed handouts as essential to preparation for examinations.conclusion:the student can improve their performance during examination while working on skeletal handouts during theory lectures in spite of showing a preference for complete handouts . |
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