bluuebunny/crossref_metadata_2025 huggingface dataset:
Affinity purification of the hepatic high-density lipoprotein receptor identifies two acidic glycoproteins and enables further characterization of their binding properties
Authors: Hidaka H , Fidge N H
Published: August 2015

Several high-density lipoprotein (HDL)-binding proteins, candidates for the putative HDL receptor, have recently been identified, including two membrane proteins: HB1 of 120 kDa and HB2 of 100 kDa, present in rat and human liver plasma membranes respectively. Further insights into their function however, have been hampered by poor recoveries of these hydrophobic peptides, and the present work was undertaken to improve yields and enable a more detailed investigation of their properties. A significant improvement has been achieved using two affinity chromatographic procedures, one exploiting the glycoprotein nature of the proteins and the other exploiting their ligand properties, which in combination resulted in considerable enrichment of HB1 and HB2. Thus DEAE-Sephacel fractionation (0.05-0.2 M-NaCl) of CHAPS-solubilized plasma membranes yielded active HDL-binding proteins which bound to concanavalin A-Sepharose or wheat-germ-lectin-Sepharose columns and retained their binding activity after eluting with methyl-alpha-D-mannoside or N-acetylglucosamine respectively. These glycoproteins were further purified by affinity chromatography using apo-HDL-Sepharose columns. Final purification required preparative SDS/PAGE. Investigation of the carbohydrate moieties of the proteins using glycosidases and two-dimensional gel electrophoresis revealed pI values ranging from 4.6 to 4.9 and from 4.5 to 4.7 for HB1 and HB2 respectively, which after treatment with neuraminidase shifted towards basic pH (5.4-5.7 and 5.3-5.5 respectively). The molecular masses were decreased to 115 kDa and 95 kDa respectively, demonstrating that sialic acid residues contributed significantly to the negative charge of the glycosylated peptides. Treatment with the enzyme peptide N-glycosidase F (N-glycanase) resulted in a decrease in molecular mass of HB1 and HB2 to 105 kDa and 80 kDa respectively, but endo-alpha-N-acetylgalactosaminidase (O-glycanase) treatment was not effective. Neither neuraminidase nor N-glycanase treatment destroyed activity, suggesting that sialic acids or N-linked oligosaccharides are not important determinants of HDL binding. Digestion of plasma membranes with trypsin or Pronase resulted in a loss of activity of both HB1 and HB2 that was not influenced by prior treatment with neuraminidase, suggesting that sialic acid residues play no protective role against proteolytic cleavage of HDL receptor proteins.
Phosphatidylcholine-dependent protein kinase C activation. Effects of cis-fatty acid and diacylglycerol on synergism, autophosphorylation and Ca2+-dependency
Authors: Chen S G , Kulju D , Halt S , Murakami K
Published: August 2015

A long-chain neutral phospholipid, dioleoylphosphatidylcholine, was found to support protein kinase C activation by cis-fatty acid and diacylglycerol (DAG). This effect of phosphatidylcholine (PC) is totally dependent on the presence of cis-fatty acid; PC greatly stimulates the cis-fatty acid-induced protein kinase C activity, but it does not activate protein kinase C at all, even in the presence of DAG, if cis-fatty acid is absent. DAG, however, plays a modulatory role in the presence of Ca2+; it further enhances the PC-potentiated cis-fatty acid activation of protein kinase C. Although the activities of all three protein kinase C subtypes tested (types I, II and III) are supported by this PC mechanism, type III is most sensitive to the DAG effect, and it is activated synergistically by cis-fatty acid and DAG. The potency of PC to support the synergistic activation of this subtype is equivalent to that of phosphatidylserine (PS). There are several differences, however, between PC- and PS-supported synergism observed in type III protein kinase C: (1) Ca(2+)-sensitivity is different; PC requires higher concentrations of Ca2+ (10-20 microM-Ca2+) than those required for PS (micromolar Ca2+); (2) PC/cis-fatty acid/DAG-induced autophosphorylation of protein kinase C subtypes (types I, II and III) is very weak, whereas PS/cis-fatty acid/DAG strongly stimulate autophosphorylation of these subtypes under the conditions at which both PC and PS systems fully activate the protein kinase C in terms of histone phosphorylation. These observations suggest that a neutral phospholipid such as PC may also participate in the activation and differential regulation of protein kinase C.
Processing of the phospholipid analogue phosphatidyl(N-sulphorhodamine B sulphonyl)ethanolamine by rat hepatocytes in vitro and in vivo
Authors: Verkade H J , Zaal K J M , Derksen J T P , Vonk R J , Hoekstra D , Kuipers F , Scherphof G L
Published: August 2015

We have investigated the processing of the non-exchangeable fluorescent phospholipid analogue phosphatidyl(N-sulphorhodamine B sulphonyl)ethanolamine (N-Rh-PE) by rat liver cells. In the hepatocyte couplet system, N-Rh-PE was incorporated into the plasma membrane at 2 degrees C and readily internalized upon warming to 37 degrees C. Fluorescence was initially found to be concentrated in vesicles clustered throughout the cell, but subsequently it started to accumulate in pericanalicular vesicles, tentatively identified as lysosomes, and in the bile canalicular lumen. Analysis of cells and media by t.l.c. revealed the slow formation of at least two metabolites. After intravenous injection into bile-fistula rats of [9,10-3H-oleoyl]N-Rh-PE incorporated in small unilamellar liposomes, the initial rates of elimination from plasma of 3H and rhodamine label were virtually identical. However, biliary secretion of the 3H label (5.5% of dose at 2 h) was much slower than that of the rhodamine label (49.3% at 2 h). The rhodamine label in bile was chloroform-soluble, but not identical to the native molecule, and was resistant to phospholipase A2 and alkaline hydrolysis. To gain insight in the mechanism of the rapid bile secretion of this metabolite, we compared the processing of N-Rh-PE, its deacylated form [glycerophospho(N-sulphorhodamine B sulphonyl)ethanolamine; Gly-N-Rh] and the rhodamine label itself (sulphorhodamine B sulphonyl chloride; SRho). Intravenous injection of chloroform-soluble N-Rh-PE and of methanol/water-soluble Gly-N-Rh complexed with albumin both resulted in rapid bile secretion of chloroform-soluble fluorescent compounds (60.2% and 86.3% respectively at 2 h), which showed behaviour identical to that of the metabolite of liposomal N-Rh-PE on t.l.c. Methanol/water-soluble SRho was also rapidly secreted into bile (89.5% at 2 h) without being metabolized. Bile secretion of the chloroform-soluble metabolite of N-Rh-PE and of SRho was markedly impaired (-31% and -52% respectively) in GY Wistar rats, which express a genetic defect in the hepatobiliary transport of organic anions. Our data show that the rat hepatocyte is capable of modifying the structure of N-Rh-PE, a process which proceeds considerably faster in vivo than in vitro. The chloroform-soluble metabolite is subsequently rapidly removed via the bile. The canalicular organic anion transporting system, which is deficient in GY rats, appears to be involved in the excretion of this apolar product of hepatic metabolism.
A biochemical analysis of human periodontal tissue proteoglycans
Authors: Larjava H , Häkkinen L , Rahemtulla F
Published: August 2015

Proteoglycans synthesized by periodontal (gingival, periodontal ligament, dental follicle) fibroblasts were analysed by SDS/polyacrylamide and agarose gel electrophoresis after being labelled with radioactive sulphate. Medium, cell membrane and extracellular matrix fractions were analysed separately. Samples were treated with chondroitinase AC, chondroitinase ABC, heparitinase or a combination of chondroitinase ABC and heparitinase before electrophoretic separation of proteoglycans. Antibodies to versican and decorin were used to identify these molecules by Western immunoblots. For steady-state metabolic radiolabelling of fibroblasts, medium and cell membrane fractions contained about equal proportions of radiolabelled proteoglycans (about 43%), whereas less radioactivity (about 14%) was found in proteoglycans of the matrix fraction. Periodontal fibroblasts produce six major proteoglycans: versican, a high-molecular-mass chondroitin sulphate proteoglycan (CSPG); decorin, a dermatan sulphate proteoglycan (DSPG); a membrane-associated heparan sulphate proteoglycan (HSPG); two medium- or matrix-associated HSPGs; and a 91 kDa membrane-associated CSPG. Variation in decorin molecular size was observed in mass cultures of fibroblasts. Similar polydispersity in molecular size of decorin was seen in several clones established from one mass culture.
Demonstration of inhibitory guanine nucleotide regulatory protein (Gi) function in liver and hepatocyte membranes from streptozotocin-treated rats
Authors: Kirkham D M , Murphy G J , Young P
Published: August 2015

By using a defined plasma-membrane preparation, functional inhibition of adenylate cyclase activity by the inhibitory G-protein (Gi) was observed in liver and hepatocyte membranes from rats made diabetic by streptozotocin. These observations contrast with previous reports which have shown a defect in Gi in this diabetic animal model. These results suggest that Gi function is not impaired in the livers of streptozotocin-treated rats and that plasma-membrane preparation procedures should be clearly defined before ascribing Gi defects to a pathological state such as diabetes.
Exocytosis from permeabilized bovine adrenal chromaffin cells is differently modulated by guanosine 5′-[γ-thio]triphosphate and guanosine 5′-[β γ-imido]triphosphate. Evidence for the involvement of various guanine nucleotide-binding proteins
Authors: Ahnert-Hilger G , Wegenhorst U , Stecher B , Spicher K , Rosenthal W , Gratz M
Published: August 2015

1. In bovine adrenal chromaffin cells made permeable either to molecules less than or equal to 3 kDa with alphatoxin or to proteins less than or equal to 150 kDa with streptolysin O, the GTP analogues guanosine 5′-[beta gamma-imido]triphosphate (p[NH]ppG) and guanosine 5′-[gamma-thio]triphosphate (GTP[S]) differently modulated Ca(2+)-stimulated exocytosis. 2. In alphatoxin-permeabilized cells, p[NH]ppG up to 20 microM activated Ca(2+)-stimulated exocytosis. Higher concentrations had little or no effect. At a free Ca2+ concentration of 5 microM, 7 microM-p[NH]ppG stimulated exocytosis 6-fold. Increasing the free Ca2+ concentration reduced the effect of p[NH]ppG. Pretreatment of the cells with pertussis toxin prevented the activation of the Ca(2+)-stimulated exocytosis by p[NH]ppG. 3. In streptolysin O-permeabilized cells, p[NH]ppG did not activate, but rather inhibited Ca(2+)-dependent catecholamine release under all conditions studied. In the soluble cytoplasmic material that escaped during permeabilization with streptolysin O, different G-protein alpha-subunits were detected using an appropriate antibody. Around 15% of the cellular alpha-subunits were detected in the supernatant of permeabilized control cells. p[NH]ppG or GTP[S] stimulated the release of alpha-subunits 2-fold, causing a loss of about 30% of the cellular G-protein alpha-subunits under these conditions. Two of the alpha-subunits in the supernatant belonged to the G(o) type, as revealed by an antibody specific for G(o) alpha. 4. GTP[S], when present alone during stimulation with Ca2+, activated exocytosis in a similar manner to p[NH]ppG. Upon prolonged incubation, GTP[S], in contrast to p[NH]ppG, inhibited Ca(2+)-induced exocytosis from cells permeabilized by either of the pore-forming toxins. This effect was resistant to pertussin toxin. 5. The p[NH]ppG-induced activation of Ca(2+)-stimulated release from alphatoxin-permeabilized chromaffin cells may be attributed to one of the heterotrimeric G-proteins lost during permeabilization with streptolysin O. The inhibitory effect of GTP[S] on exocytosis is apparently not mediated by G-protein alpha-subunits, but by another GTP-dependent process still occurring after permeabilization with streptolysin O.
Glycaemia regulates the glucose transporter number in the plasma membrane of rat skeletal muscle
Authors: Dimitrakoudis D , Ramlal T , Rastogi S , Vranic M , Klip A
Published: August 2015

The number of glucose transporters was measured in isolated membranes from diabetic-rat skeletal muscle to determine the role of circulating blood glucose levels in the control of glucose uptake into skeletal muscle. Three experimental groups of animals were investigated in the post-absorptive state: normoglycaemic/normoinsulinaemic, hyperglycaemic/normoinsulinaemic and hyperglycaemic/normoinsulinaemic made normoglycaemic/normoinsulinaemic by phlorizin treatment. Hyperglycaemia caused a reversible decrease in total transporter number, as measured by cytochalasin B binding, in both plasma membranes and internal membranes of skeletal muscle. Changes in GLUT4 glucose transporter protein mirrored changes in cytochalasin B binding in plasma membranes. However, there was no recovery of GLUT4 levels in intracellular membranes with correction of glycaemia. GLUT4 mRNA levels decreased with hyperglycaemia and recovered only partially with correction of glycaemia. Conversely, GLUT1 glucose transporters were only detectable in the plasma membranes; the levels of this protein varied directly with glycaemia, i.e. in the opposite direction to GLUT4 glucose transporters. This study demonstrates that hyperglycaemia, in the absence of hypoinsulinaemia, is capable of down-regulating the glucose transport system in skeletal muscle, the major site of peripheral resistance to insulin-stimulated glucose transport in diabetes. Furthermore, correction of hyperglycaemia causes a complete restoration of the transport system in the basal state (determined by the transporter number in the plasma membrane), but possibly only an incomplete recovery of the transport system's ability to respond to insulin (since there is no recovery of GLUT4 levels in the intracellular membrane insulin-responsive transporter pool). Finally, the effect of hyperglycaemia is specific for glucose transporter isoforms, with GLUT1 and GLUT4 proteins varying respectively in parallel and opposite directions to levels of glycaemia.
The antigenicity of the carbohydrate moiety of an insect glycoprotein, honey-bee (Apis mellifera) venom phospholipase A2. The role of α1,3-fucosylation of the asparagine-bound N-acetylglucosamine
Authors: Prenner C , Mach L , Glössl J , März L
Published: August 2015

A rabbit polyclonal antiserum raised against honey-bee (Apis mellifera) venom phospholipase A2 (PLA2) contains antibodies that react exclusively with its glycosylated variants and cross-react with plant glycoproteins. The interaction of anti-(horseradish peroxidase) antiserum with PLA2 suggests the existence of a carbohydrate determinant common to both glycoproteins. E.l.i.s.a. binding and inhibition experiments, employing glycoproteins and glycopeptides of plant and animal origin with known N-glycan structures, in combination with chemical and enzymic deglycosylation, identified alpha 1,3-fucosylation of the asparagine-bound N-acetylglucosamine as the antigenic determinant. This fucose residue is present in the N-glycan of PLA2 and is frequently found in plant glycoproteins, whereas mammalian glycoproteins lack this modification.
Formation of salivary-mucosal pellicle: the role of transglutaminase
Authors: Bradway S D , Bergey E J , Scannapieco F A , Ramasubbu N , Zawacki S , Levine M J
Published: August 2015

The present investigation was carried out to identify salivary components of mucosal pellicles in vivo and explore further the mechanism of interaction between salivary molecules and buccal epithelial cells. By using specific antisera and immunoprotein blotting, high-(MG1) and low-(MG2) molecular-mass salivary mucins, amylase, salivary cystatins and proline-rich proteins were detected within mucosal pellicle in vivo. In addition, the data indicated that the mucins and proline-rich proteins could be cleaved into lower-molecular-mass products, whereas the proline-rich proteins could also be cross-linked into higher-molecular-mass complexes. The role of buccal epithelial cell transglutaminase in these interactions was further studied by utilizing purified iodinated amylase, neutral cystatin SN and acidic proline-rich proteins 1 and 3 (APRP1 and 3). After incubation with buccal epithelial cells in vitro 125I-labelled APRPs appeared to undergo a greater degree of cross-linking than 125I-labelled cystatin SN, as determined by SDS/PAGE/autoradiography. Amylase did not appear to be cross-linked at all. Recovery of 125I-labelled APRPs and 125I-labelled cystatin SN with epithelial cell envelopes after repeated extraction suggested that both molecules were cross-linked to envelope proteins, but that 125I-labelled APRPs were cross-linked to a greater degree than 125I-labelled cystatin SN. Cross-linking in buccal epithelial cell preparations was inhibited by an excess of methylamine hydrochloride, a transglutaminase substrate. In a further assessment of amylase, cystatin and APRPs as transglutaminase substrates, only APRP3 and a partially purified preparation of APRPs acted as an amine acceptor for the cross-linking of [14C]methylamine by purified transglutaminase, as determined by SDS/PAGE/fluorography. This reaction was completely inhibited by excess EDTA. The combined data from this study suggest that during mucosal pellicle formation multiple components of saliva adsorb to buccal epithelial cell surfaces, and that, within this group, selected components are enzymically cross-linked by an epithelial transglutaminase and/or proteolytically cleaved into smaller fragments.
Limited accumulation of cyclic AMP underlies a modest vasoactive-intestinal-peptide-mediated increase in cytosolic [Ca2+] transients in GH3 pituitary cells
Authors: Mollard P , Zhang Y , Rodman D , Cooper D M F
Published: August 2015

The 4-chlorophenylthio analogue of cyclic AMP evoked profound and long-lasting changes in cytosolic [Ca2+] ([Ca2+]i) in pituitary-derived GH3 cells. However, vasoactive intestinal peptide (VIP), a hormone considered to act via cyclic AMP, was ineffective in modulating [Ca2+]i. The ability of VIP to modulate [Ca2+]i was enhanced by treatments that increased intracellular cyclic AMP. Much greater concentrations of intracellular cyclic nucleotides were achieved by the analogue than with VIP, under any condition. Thus cyclic AMP may play a prominent role in regulating [Ca2+]i in these cells, but the ability of hormones to stimulate its synthesis is limited, leading to a weak action on [Ca2+]i.
Effect of permethylation on the haemolytic activity of melittin
Authors: Ramalingam K , Bello J
Published: August 2015

The cytolytic activity of the bee venom toxin, melittin, is abolished on permethylation of the ammonium groups into quaternary trimethylammonium groups. The loss of activity in permethylated melittin may result partly from the absence of the hydrogen bonding potential and partly from steric effects involving the bulky trimethylammonium groups. Displacing the trimethylammonium groups away from the backbone to relieve steric effects (by acylating melittin with glycine or 5-aminopentanoic acid followed by permethylation) restored moderate activity at 5-fold increase in concentration.
Mammary development and milk secretion in transgenic mice expressing the sheep β-lactoglobulin gene
Authors: Wilde C J , Clark A J , Kerr M A , Knight C H , McClenaghan M , Simons J P
Published: August 2015

Mammary development and milk secretion were studied in transgenic mice which exhibited mammary tissue-specific expression of the sheep beta-lactoglobulin gene, and secreted significant quantities of the foreign protein in milk. Mammary development was unaffected by transgenesis. Tissue DNA content and the activities of several key enzyme markers of cell differentiation were similar in transgenic mice and non-transgenic controls. Milk yield, whether estimated by pup weight gain or measured by a 3H2O-dilution method, was unchanged by foreign gene expression. Gross milk composition, including milk protein concentration, was also similar in transgenic and non-transgenic animals, even though beta-lactoglobulin accounted for 29% of total milk protein. Therefore the foreign gene product was synthesized at the expense of endogenous milk proteins. However, transgenic mammary tissue in vitro exhibited a significantly higher rate of total protein synthesis than did control tissue. This suggested that a factor limiting milk protein synthesis or secretion in transgenic mice in vivo may have been removed by short-term explant culture of mammary tissue. The results emphasize that the use of transgenesis for manipulating milk composition may depend not only on high-level mammary-specific expression of the foreign gene, but also on the biosynthetic capacity of the mammary gland itself.
Novel linear and branched polyamines in the extremely thermophilic eubacteria Thermoleophilum, Bacillus and Hydrogenobacter
Authors: Hamana K , Niitsu M , Matsuzaki S , Samejima K , Igarashi Y , Kodama T
Published: August 2015

Novel tertiary branched tetra-amines, quaternary branched penta-amines, linear penta-amines and linear hexa-amines were distributed as the major polyamines in six obligately extremely thermophilic eubacteria belonging to Thermoleophilum, Bacillus or Hydrogenobacter. The major polyamine of Thermoleophilum album and Thermoleophilum minutum was identified as a quaternary branched penta-amine, 4,4-bis(3-aminopropyl)-1,8-diamino-4-azaoctane (NH2[CH2]3N+([CH2]3NH2)2[CH2]4NH2) by h.p.l.c., t.l.c. and g.c.-m.s. Hydrogenobacter thermophilus and Hydrogenobacter halophilus contained another quaternary branched penta-amine, 4,4-bis(3-aminopropyl)-1,7-diamino-4-azaheptane (NH2[CH2]3N([CH2]3NH2)2[CH2]3NH2) as the major polyamine, and tertiary branched tetra-amines (4-(3-aminopropyl)-1,7-diamino-4-azaheptane (NH2[CH2]3N([CH2]3NH2)[CH2]3NH2), 4-(3-aminopropyl)-1,8-diamino-4-azaoctane (NH2[CH2]3N([CH2]3NH2)[CH2]4NH2)) and 4,4-bis(3-aminopropyl)-1,8-diamino-4-azaoctane were confirmed as minor components. Bacillus schlegelii contained a branched tetra-amine, 4-(3-aminopropyl)-1,8-diamino-4-azaoctane, a branched penta-amine, 4,4-bis(3-aminopropyl)-1,8-diamino-4-azaoctane, a linear penta-amine, 1,16-diamino-4,8,13-triazahexadecane (NH2[CH2]3NH[CH2]3NH[CH2]4NH[CH2]3NH2) and linear hexa-amine(s), 1,20-diamino-4,8,12,17-tetra-azaeicosane (NH2[CH2]3NH[CH2]3NH[CH2]3NH[CH2]4NH[CH2]3NH2) and/or 1,20-diamino-4,8,13,17-tetra-azaeicosane (NH2[CH2]3NH[CH2]3NH[CH2]4NH[CH2]3NH[CH2]3NH2).
Purification and characterization of variants of acyl-CoA-binding protein in the bovine liver
Authors: Jensen M S , Højrup P , Rasmussen J T , Knudsen J
Published: August 2015

Four differently modified forms of acyl-CoA-binding protein (ACBP) were identified in ACBP purified from bovine liver. The majority of the purified ACBP was focused at pH 5.9 in isoelectric focusing and could be shown to be N-acetylated ACBP without any further modifications. Two minor peaks were focused at pH 5.25 and 4.85 respectively. Mass spectrometry and sequence determination showed that the pI 5.25 form was acetylated at Lys18 and that the pI 4.85 form was malonylated in the same position. Furthermore, it could be shown that non-enzymic glycosylation occurred during purification. The acetylated and malonylated variants of ACBP were only found in adult cattle.
Differential polypeptide expression in adipose tissue of lean and obese Zucker rats. Evidence of specifically repressed peptides in 7-day-old pre-obese rats
Authors: Laurent-Winter C , Dugail I , Quignard-Boulange A , Le Liepvre X , Lavau M
Published: August 2015

Using two-dimensional electrophoresis on total extracts of adipose tissue from young lean (Fa/fa) and obese (fa/fa) Zucker rats, we have investigated the existence of early events at the protein level, before obvious obesity. Our results indicate that the two genotypes do not differ at 3 days of age in terms of polypeptide pattern. By 7 days of age, two polypeptides are transiently repressed in the fatty genotype, leading us to suggest their potential involvement in the onset of obesity. However, most of the differences between the lean and obese rats are detected at 30 days of age, characterized by an increase in the accumulation of several peptides in the adipose tissue of obese rats, in good agreement with the multiple biochemical changes previously identified at this stage of the disease. These results present evidence of new peptides that may be of interest in the study of the obesity syndrome.
Alterations in the mucosal processing of iron in response to very-short-term dietary iron depletion and repletion
Authors: Topham R W , Eads C E , Butler B F
Published: August 2015

The transfer of control rats to a low-iron diet for only 24 h resulted in a 2-fold increase in iron uptake by brush-border membrane vesicles. Extension of the low-iron feeding period to 72 h or 2 weeks resulted in only small additional increases in iron uptake by vesicle preparations. In contrast, the transfer of iron-deficient rats to a control diet resulted in a progressive decrease in iron uptake by vesicles that reached a level equivalent to that of control rats in 2 weeks. 59Fe labelling of detergent extracts of these vesicle preparations provided evidence for the presence of an iron-binding protein composed of subunits of 52,000 Da. The changes in the 59Fe labelling of this protein component were consistent with the changes observed in iron uptake by intact brush-border membrane vesicles. The 59Fe-labelling profiles of mucosal ferritin and transferrin from a test dose also were changed substantially in response to very-short-term alterations in dietary iron. Even though changes in dietary iron rapidly altered iron uptake by brush-border membrane vesicles and the incorporation of 59Fe from the test dose into mucosal transferrin, changes in the incorporation of 59Fe into mucosal ferritin best reflected the actual changes in the transfer of iron from dose to plasma.
Cloning and expression of the thyrotropin-releasing hormone receptor from GH3 rat anterior pituitary cells
Authors: de la Peña P , Delgado L M , del Camino D , Barros F
Published: August 2015

Functional thyrotropin-releasing hormone (TRH) receptors have been expressed in Xenopus laevis oocytes following the microinjection of total and poly(A)+ RNA from GH3 rat anterior pituitary tumour cells. Under voltage-clamp conditions, application of the peptide induced a biphasic Ca(2+)-dependent chloride current. The amplitude of the initial, fast, component of the response was dependent on the concentration of the hormone and on the amount of mRNA injected. Size fractionation of poly(A)+ RNA on a continuous sucrose gradient and Northern blot analysis indicated that the receptor was encoded by an mRNA of approx. 3.5 kb. A 3.28 kbp cDNA encoding the TRH receptor has been cloned and sequenced. Full functionality of the predicted 412-amino-acid receptor protein was demonstrated by functional expression of cell surface receptors in Xenopus oocytes after both cytoplasmic injection of sense RNA transcribed in vitro from this cDNA and nuclear injection of the cDNA under the control of the Herpes simplex virus thymidine kinase promoter. The predicted protein contains seven putative membrane-spanning domains and shows significant sequence identify with some G-protein-coupled receptors. RNA blot analysis indicates that the mRNA for the TRH receptor is exclusively expressed in the pituitary gland. Expression studies performed with clones in which the 3′ region of the mRNA has been successively shortened indicate that the 3′ terminal region is not an important determinant for efficient functional expression in oocytes.
Interaction of bleomycin with a bent DNA fragment
Authors: Nightingale K P , Fox K R
Published: August 2015

The interaction of bleomycin with a kinetoplast DNA fragment has been examined using various footprinting techniques. This DNA adopts a bent structure and displays an unusually low gel mobility on account of its phased runs of adenines. The bleomycin-cobalt complex increases the mobility of this DNA fragment, in contrast with other DNAs which show a decreased rate of gel migration, suggesting that the antibiotic removes DNA bending, possibly via an unwinding mechanism. Removal of the bending is confirmed by hydroxy-radical footprinting which produces a more even ladder of bands in the presence of the ligand. Cleavage by bleomycin is at the sequence G-pyrimidine, though not all such sites are affected to the same extent and some cutting is found at GA and GG. DNase I footprinting confirms the antibiotic-binding sites but reveals that some strong cleavage sites do not yield footprints. Bleomycin renders adenines on the 3′ side of its cleavage sites (GT, GC and GA) hyper-reactive to diethyl pyrocarbonate.
Mutual coupling investigation between loop and two inverted‐L antennas operating below 1 GHz
Authors: Bahramzy P. , Svendsen S. , Pedersen G.F.
Published: August 2015

To overcome antenna design challenges posed by the increasing number of frequency bands, the use of separate transmit (Tx) and receive (Rx) narrow‐band antennas is proposed. In addition, a second Rx antenna is utilised for multiple input–multiple output (MIMO) application. While sufficiently low coupling between two antennas is important so as to gain the benifits of MIMO, this becomes even more challenging when considering coupling between three antennas. In this reported work, this is addressed by utilising a loop antenna as one of the three antennas, where the other two will be conventional inverted‐L antennas. The measurements show that low coupling is possible between all three antennas, while having low correlation between the two Rx antennas.
DEC‐cache: dynamically electing candidate cache for low power utilising hearing policy
Authors: Joo Hyunwook , Lee Yong Surk
Published: August 2015

On‐chip cache memory is one of the largest power consumers in modern microprocessors. A dynamic way prediction scheme utilising a hearing policy is proposed for a low‐power level‐one cache design that handles power limit issues. The high prediction accuracy of the dynamically electing candidate (DEC)‐cache helps to prevent large miss penalties. Owing to the high prediction accuracy, the experimental results show that the DEC‐cache structure improves the energy‐delay product by 26% compared with the existing buffered dual‐mode cache.
Matched‐filter properties of linear‐frequency‐modulation radar signal reflected from a phase‐switched screen
Authors: Xu Letao , Feng Dejun , Wang Xuesong
Published: August 2015

The phase‐switched screen (PSS) imposes phase modulation onto the radar‐reflected signal so that its energy is redistributed into sidebands that lie outside the receiver pass‐band. However, when illuminated by a wideband linear frequency modulation (LFM) radar, the modulated reflected signal may still occupy the radar pass‐band if the LFM signal bandwidth is greater than the modulating frequency. Previous research about the reflectivity performance of a PSS against LFM radar illumination focuses mainly on the frequency spectrum characteristics. In this study, an investigation has been carried out to determine the matched‐filter properties of an LFM radar signal modulated by a PSS. Theoretical analyses and simulation results have proven that even though the radar receiver bandwidth has been corrupted to aliasing by component of the reflected signal, the PSS is shown to be equally effective in shielding a target and additionally provide a multiple‐decoy capability.
Fabrication and mechanical properties of low‐loading graphene nanosheets encapsulated on the surface of graphene nanosheets/polypropylene composites
Authors: Hsu Ping‐Cheng , Tsai I‐Shou
Published: August 2015

In recent years, numerous scientists have used graphene for reinforcing polymer‐based composites because of its outstanding electrical, thermal and mechanical properties. Based on this present reported work, a simple manufacturing process is reported for the fabrication of low‐loading graphene nanosheets (GNSs) coated on the surface of a polypropylene matrix. According to a scanning electron microscope micrograph, the GNSs were dispersed uniformly on the surface of polypropylene pellets. The mechanical properties and thermal stability of the GNSs /polypropylene composite plates were improved substantially by the addition of GNSs. X‐ray diffraction was used to determine that the two‐theta values of the GNSs /polypropylene composite plates correspond to the weight fraction of the added GNSs. Fourier transform infrared spectroscopy revealed that the vibration peak of the polypropylene methyl group decreased gradually when the amount of GNSs increased.
Konzeption und Erfassung der gesundheitsbezogenen Transitionskompetenz von Jugendlichen mit chronischen Erkrankungen – Entwicklung und Prüfung eines Selbstbeurteilungsinstrumentes
Authors: Herrmann-Garitz C. , Muehlan H. , Bomba F. , Thyen U. , Schmidt S.
Published: August 2015

Hintergrund: Es hat sich gezeigt, dass gesundheitsbezogene Versorgungs- und Selbstmanagementkompetenzen bei Jugendlichen erforderlich sind, um den Übergang in die Erwachsenenversorgung zu ermöglichen. Ziel ist, ein Instrument zur Erfassung der gesundheitsbezogenen Transitionskompetenz (TK) zu entwickeln und psychometrisch zu testen.

Methodik: Die Instrumentenentwicklung wurde in mehreren Schritten mit konzeptuellen und psychometrischen Analysen durchgeführt. Basierend auf qualitativen Analysen von Fokusgruppen mit chronisch kranken Jugendlichen und Experteninterviews wurde ein Itempool zur Erfassung von TK entwickelt. Das Instrument wurde in einer Pilotstudie mit N=323 Jugendlichen mit chronischen Erkrankungen getestet und finalisiert.

Ergebnisse: Es konnten die drei Subskalen „Arbeitswelt“ (3 Items), „krankheitsbezogenes Wissen“ (3 Items) und „Versorgungskompetenz“ (4 Items) mit guten psychometrischen Werten identifiziert werden, die sich auch zu einer Globalskala zusammenfassen lassen.

Schlussfolgerungen: Die Skalen bieten eine ökonomische Möglichkeit, die notwendigen Kompetenzen im Vorfeld und im Verlauf der Transition standardisiert zu erfassen. Dies ermöglicht auch die Erfassung der Wirksamkeit von transitionsbezogenen Maßnahmen (z. B. Schulungen).
Attracting Lagrangian coherent structures on Riemannian manifolds
Authors: Karrasch Daniel
Published: August 2015

It is a wide-spread convention to identify repelling Lagrangian Coherent Structures (LCSs) with ridges of the forward finite-time Lyapunov exponent (FTLE) field and to identify attracting LCSs with ridges of the backward FTLE. However, we show that, in two-dimensional incompressible flows, also attracting LCSs appear as ridges of the forward FTLE field. This raises the issue of the characterization of attracting LCSs using a forward finite-time Lyapunov analysis. To this end, we extend recent results regarding the relationship between forward and backward maximal and minimal FTLEs, to both the whole finite-time Lyapunov spectrum and to stretch directions. This is accomplished by considering the singular value decomposition (SVD) of the linearized flow map. By virtue of geometrical insights from the SVD, we provide characterizations of attracting LCSs in forward time for two geometric approaches to hyperbolic LCSs. We apply these results to the attracting FTLE ridge of the incompressible saddle flow.
Using evaluation to improve medical student rural experience
Authors: Moffatt Jennifer J. , Wyatt Janine E.
Published: August 2015

Objective The aim of this evaluation was to see whether interventions implemented to improve the Rural Medicine Rotation made this a more effective rural medical education experience. Multiple interventions targeting the student experience, lecturers and preceptors were implemented. Methods A quasi-experimental design using pre- and post-measures was used. The participants were all University of Queensland, School of Medicine, Rural Medicine Rotation students who completed the 2009 and 2010 rural medicine rotation evaluations. There were 769 students, with an 84% response rate in 2009 and an 80% response rate in 2010. In addition, all the 25 program preceptors who were visited in 2009 and the 34 who were visited in 2010 participated in the study. Results The implementation of interventions resulted in significant improvement in three outcome measures, namely teaching effectiveness, provision of an environment supportive of learning in a rural/remote setting and opportunities for professional growth. Two of the three other outcome measures – ensuring a safe clinical placement and opportunities for procedural skills experience and development – were very positively evaluated in both 2009 and 2010. Conclusions The interventions contributed to a more effective rural medical education experience, providing students with the opportunity to develop skills and knowledge relevant for rural medicine and to gain an understanding of the context in which rural medicine is practiced. What is known about the topic? Many Australian medical schools offer students rural-based educational opportunities based on the premise that placing medical students in a rural setting may ultimately lead to them choosing careers in rural medicine. However, there is a paucity of evidence on the factors that are considered necessary for medical students to gain a positive rural experience of short conscripted rural placements. What does the paper add? This paper identifies successful interventions to the rotation and placements that provide a positive experience of the rural clinical placement for students. These interventions occurred within an ongoing evaluation program embedded in the rotation. What are the implications for practitioners? Through ongoing evaluation, interventions can be selected and implemented that succeed in contributing to students having a positive rural clinical placement experience. This paper demonstrates how an embedded continuous improvement program serves to provide direction for ongoing modifications.
Treatment and outcomes of polymerase chain reaction-confirmed early syphilis
Authors: Read Phillip J. , Guy Rebecca , Jeoffreys Neisha , Baker David , Shields Matthew , Donovan Basil
Published: August 2015

Background
Syphilis is resurgent among gay and bisexual men (GBM) despite effective treatment and widely available diagnostic serology. The polymerase chain reaction assay for Treponema pallidum (TP-PCR) is available, but little is known about the clinical features and outcomes for patients testing positive by TP-PCR. Methods: Clinical data were collected from four medical practices for patients recording a positive TP-PCR result between 2004 and 2011. Demographic, serological, treatment and reinfection details were obtained. Results were stratified by HIV status and whether treatment conformed to international guidelines. Results: 220 patients were positive for TP-PCR, of whom 92% were GBM. Seventeen (8.1%) were positive by TP-PCR before seroconversion. Almost one-third (32.1%) received treatment beyond that recommended in guidelines, and this was associated with HIV status (40.3% HIV positive vs 22.4% HIV negative, P < 0.01). All but one patient with adequate follow up achieved serological cure. There was no significant difference in time to serological cure between the groups receiving standard therapy or enhanced therapy (95 vs 108 days; P = 0.67) or between HIV positive and negative patients (93 vs 104 days, P = 0.06). Nineteen patients were reinfected during follow up. Conclusion: TP-PCR aids early diagnosis of syphilis and may be reactive before conventional serological tests. Treatment outcomes for TP-PCR-positive early infection were excellent, but a significant proportion of patients received non-standard therapy. Expanded use of syphilis PCR testing in at-risk populations is recommended since early identification and treatment is likely to be important in controlling the current epidemic in GBM.
Engagement in group sex among geosocial networking mobile application-using men who have sex with men
Authors: Phillips Gregory , Grov Christian , Mustanski Brian
Published: August 2015

Background: Men who have sex with men (MSM) remain the group most affected by the HIV epidemic in the United States. At least one-quarter of MSM report engagement in group sex events (GSEs), which can pose a risk for HIV transmission and acquisition. In this study, identification of event-level correlates of sexual and drug use behaviours at GSEs was sought to better inform prevention activities. Methods: For this study, participants were recruited via banner and pop-up advertisements placed on a geosocial networking mobile phone application that MSM use to meet men. Results: Of the 1997 individuals who completed the study screener, 36.0% reported participating in at least one GSE in the prior year. In multivariable logistic regression analysis, attendance at a GSE in the past year was significantly associated with older age, full/part time employment and being HIV positive. Of the men who attended a GSE, more than half reported condomless anal sex (CAS) with at least one of their partners (insertive: 57.7%; receptive: 56.3%). MSM who indicated drug use had significantly higher odds of having insertive CAS [odds ratio (OR) = 2.45; 95% confidence interval (CI): 1.37–4.39] and receptive CAS (OR = 3.60; 95% CI: 1.96–6.63) at their last GSE. Conclusions: The high prevalence of HIV-positive MSM engaging in group sex, coupled with their greater odds of CAS, poses a significant risk for HIV/STI transmission within the group sex setting. More research is needed to determine patterns of condom use at these events and whether seroadaptive behaviours are driving CAS.
Curvature estimates for minimal submanifolds of higher codimension and small G-rank
Authors: Jost J. , Xin Y. L. , Yang Ling
Published: August 2015

We obtain new curvature estimates and Bernstein type results for a minimal

n
−

n-

submanifold in

R

n
+
m

,

m
≥
2

\mathbb {R}^{n+m},\, m\ge 2

, under the condition that the rank of its Gauss map is at most 2. In particular, this applies to minimal surfaces in Euclidean spaces of arbitrary codimension.
Lipid partitioning at the nuclear envelope controls membrane biogenesis
Authors: Barbosa Antonio Daniel , Sembongi Hiroshi , Su Wen-Min , Abreu Susana , Reggiori Fulvio , Carman George M. , Siniossoglou Symeon
Published: August 2015

Partitioning of lipid precursors between membranes and storage is crucial for cell growth, and its disruption underlies pathologies such as cancer, obesity, and type 2 diabetes. However, the mechanisms and signals that regulate this process are largely unknown. In yeast, lipid precursors are mainly used for phospholipid synthesis in nutrient-rich conditions in order to sustain rapid proliferation but are redirected to triacylglycerol (TAG) stored in lipid droplets during starvation. Here we investigate how cells reprogram lipid metabolism in the endoplasmic reticulum. We show that the conserved phosphatidate (PA) phosphatase Pah1, which generates diacylglycerol from PA, targets a nuclear membrane subdomain that is in contact with growing lipid droplets and mediates TAG synthesis. We find that cytosol acidification activates the master regulator of Pah1, the Nem1-Spo7 complex, thus linking Pah1 activity to cellular metabolic status. In the absence of TAG storage capacity, Pah1 still binds the nuclear membrane, but lipid precursors are redirected toward phospholipids, resulting in nuclear deformation and a proliferation of endoplasmic reticulum membrane. We propose that, in response to growth signals, activation of Pah1 at the nuclear envelope acts as a switch to control the balance between membrane biogenesis and lipid storage.
Development of the Parkland-UT Southwestern Colonoscopy Reporting System (CoRS) for evidence-based colon cancer surveillance recommendations
Authors: Skinner Celette Sugg , Gupta Samir , Halm Ethan A , Wright Shaun , McCallister Katharine , Bishop Wendy , Santini Noel , Mayorga Christian , Agrawal Deepak , Moran Brett , Sanders Joanne M , Singal Amit G
Published: August 2015

Objective Through colonoscopy, polyps can be identified and removed to reduce colorectal cancer incidence and mortality. Appropriate use of surveillance colonoscopy, post polypectomy, is a focus of healthcare reform.

Materials and Methods The authors developed and implemented the first electronic medical record–based colonoscopy reporting system (CoRS) that matches endoscopic findings with guideline-consistent surveillance recommendations and generates tailored results and recommendation letters for patients and providers.

Results In its first year, CoRS was used in 98.6% of indicated cases. Via a survey, colonoscopists agreed/strongly agreed it is easy to use (83%), provides guideline-based recommendations (89%), improves quality of Spanish letters (94%), they would recommend it for other institutions (78%), and it made their work easier (61%), and led to improved practice (56%).

Discussion CoRS’ widespread adoption and acceptance likely resulted from stakeholder engagement throughout the development and implementation process.

Conclusion CoRS is well-accepted by clinicians and provides guideline-based recommendations and results communications to patients and providers.
Descriptive analysis of medication errors reported to the Egyptian national online reporting system during six months
Authors: Shehata Zahraa Hassan Abdelrahman , Sabri Nagwa Ali , Elmelegy Ahmed Abdelsalam
Published: August 2015

Objectives This study analyzes reports to the Egyptian medication error (ME) reporting system from June to December 2014.

Methods Fifty hospital pharmacists received training on ME reporting using the national reporting system. All received reports were reviewed and analyzed. The pieces of data analyzed were patient age, gender, clinical setting, stage, type, medication(s), outcome, cause(s), and recommendation(s).

Results Over the course of 6 months, 12 000 valid reports were gathered and included in this analysis. The majority (66%) came from inpatient settings, while 23% came from intensive care units, and 11% came from outpatient departments. Prescribing errors were the most common type of MEs (54%), followed by monitoring (25%) and administration errors (16%). The most frequent error was incorrect dose (20%) followed by drug interactions, incorrect drug, and incorrect frequency. Most reports were potential (25%), prevented (11%), or harmless (51%) errors; only 13% of reported errors lead to patient harm. The top three medication classes involved in reported MEs were antibiotics, drugs acting on the central nervous system, and drugs acting on the cardiovascular system. Causes of MEs were mostly lack of knowledge, environmental factors, lack of drug information sources, and incomplete prescribing. Recommendations for addressing MEs were mainly staff training, local ME reporting, and improving work environment.

Discussion There are common problems among different healthcare systems, so that sharing experiences on the national level is essential to enable learning from MEs. Internationally, there is a great need for standardizing ME terminology, to facilitate knowledge transfer. Underreporting, inaccurate reporting, and a lack of reporter diversity are some limitations of this study.

Egypt now has a national database of MEs that allows researchers and decision makers to assess the problem, identify its root causes, and develop preventive strategies.