Influence of stimulus intensity on the soleus H-reflex amplitude and modulation during locomotion

Diverging results have been reported regarding the modulation and amplitude of the soleus H-reflex measured during human walking and running. A possible explanation to this could be the use of too high stimulus strength in some studies while not in others. During activities like walking and running it is necessary to use a small M-wave to control the effective stimulus strength during all phases of the movement. This implies that the descending part of the H-reflex recruitment curve is being used, which may lead to an unwanted suppression of the H-reflex due to limitations imbedded within the H-reflex methodology itself.Accordingly, the purpose of the present study was to study the effect on the soleus H-reflex during walking and running using stimulus intensities normally considered too high (up to 45% Mmax).Using M-waves of 25–45% Mmax as opposed to 5–25% Mmax showed a significant suppression of the peak H-reflex during the stance phase of walking, while no changes were observed during running. No differences were observed regarding modulation pattern. So a possible use of too high stimulus intensity cannot explain the differences mentioned. The surprising result in running may be explained by the much higher voluntary muscle activity, which implies the existence of a V-wave influencing the H-reflex amplitude in positive direction.     

Characterization of human myotubes from type 2 diabetic and nondiabetic subjects using complementary quantitative mass spectrometric methods

Skeletal muscle is a key tissue site of insulin resistance in type 2 diabetes. Human myotubes are primary skeletal muscle cells displaying both morphological and biochemical characteristics of mature skeletal muscle and the diabetic phenotype is conserved in myotubes derived from subjects with type 2 diabetes. Several abnormalities have been identified in skeletal muscle from type 2 diabetic subjects, however, the exact molecular mechanisms leading to the diabetic phenotype has still not been found. Here we present a large-scale study in which we combine a quantitative proteomic discovery strategy using isobaric peptide tags for relative and absolute quantification (iTRAQ) and a label-free study with a targeted quantitative proteomic approach using selected reaction monitoring to identify, quantify, and validate changes in protein abundance among human myotubes obtained from nondiabetic lean, nondiabetic obese, and type 2 diabetic subjects, respectively. Using an optimized protein precipitation protocol, a total of 2832 unique proteins were identified and quantified using the iTRAQ strategy. Despite a clear diabetic phenotype in diabetic myotubes, the majority of the proteins identified in this study did not exhibit significant abundance changes across the patient groups. Proteins from all major pathways known to be important in type 2 diabetic subjects were well-characterized in this study. This included pathways like the trichloroacetic acid (TCA) cycle, lipid oxidation, oxidative phosphorylation, the glycolytic pathway, and glycogen metabolism from which all but two enzymes were found in the present study. None of these enzymes were found to be regulated at the level of protein expression or degradation supporting the hypothesis that these pathways are regulated at the level of post-translational modification. Twelve proteins were, however, differentially expressed among the three different groups. Thirty-six proteins were chosen for further analysis and validation using selected reaction monitoring based on the regulation identified in the iTRAQ discovery study. The abundance of adenosine deaminase was considerably down-regulated in diabetic myotubes and as the protein binds propyl dipeptidase (DPP-IV), we speculate whether the reduced binding of adenosine deaminase to DPP-IV may contribute to the diabetic phenotype in vivo by leading to a higher level of free DPP-IV to bind and inactivate the anti-diabetic hormones, glucagon-like peptide-1 and glucose-dependent insulintropic polypeptide.     

Structural basis for substrate recognition by Erwinia chrysanthemi GH30 glucuronoxylanase

Xylanase A from the phytopathogenic bacterium Erwinia chrysanthemi is classified as a glycoside hydrolase family 30 enzyme (previously in family 5) and is specialized for degradation of glucuronoxylan. The recombinant enzyme was crystallized with the aldotetraouronic acid β-D-xylopyranosyl-(1→4)-[4-O-methyl-α-D-glucuronosyl-(1→2)]-β-D-xylopyranosyl-(1→4)-D-xylose as a ligand. The crystal structure of the enzyme-ligand complex was solved at 1.39 ? resolution. The ligand xylotriose moiety occupies subsites -1, -2 and -3, whereas the methyl glucuronic acid residue attached to the middle xylopyranosyl residue of xylotriose is bound to the enzyme through hydrogen bonds to five amino acids and by the ionic interaction of the methyl glucuronic acid carboxylate with the positively charged guanidinium group of Arg293. The interaction of the enzyme with the methyl glucuronic acid residue appears to be indispensable for proper distortion of the xylan chain and its effective hydrolysis. Such a distortion does not occur with linear β-1,4-xylooligosaccharides, which are hydrolyzed by the enzyme at a negligible rate. DATABASE: Structural and experimental data are available in the Protein Data Bank database under accession number 2y24 [45].     

Broad antiviral activity of carbohydrate-binding agents against the four serotypes of dengue virus in monocyte-derived dendritic cells

Background

Dendritic cells (DC), present in the skin, are the first target cells of dengue virus (DENV). Dendritic cell-specific intercellular adhesion molecule 3-grabbing non-integrin (DC-SIGN) is present on DC and recognizes N-glycosylation sites on the E-glycoprotein of DENV. Thus, the DC-SIGN/E-glycoprotein interaction can be considered as an important target for inhibitors of viral replication. We evaluated various carbohydrate-binding agents (CBAs) against all four described serotypes of DENV replication in Raji/DC-SIGN⁺ cells and in monocyte-derived DC (MDDC).

Methodology/Principal Findings

A dose-dependent anti-DENV activity of the CBAs Hippeastrum hybrid (HHA), Galanthus nivalis (GNA) and Urtica dioica (UDA), but not actinohivin (AH) was observed against all four DENV serotypes as analyzed by flow cytometry making use of anti-DENV antibodies. Remarkably, the potency of the CBAs against DENV in MDDC cultures was significantly higher (up to 100-fold) than in Raji/DC-SIGN⁺ cells. Pradimicin-S (PRM-S), a small-size non-peptidic CBA, exerted antiviral activity in MDDC but not in Raji/DC-SIGN⁺ cells. The CBAs act at an early step of DENV infection as they bind to the viral envelope of DENV and subsequently prevent virus attachment. Only weak antiviral activity of the CBAs was detected when administered after the virus attachment step. The CBAs were also able to completely prevent the cellular activation and differentiation process of MDDC induced upon DENV infection.

Conclusions/Significance

The CBAs exerted broad spectrum antiviral activity against the four DENV serotypes, laboratory-adapted viruses and low passage clinical isolates, evaluated in Raji/DC-SIGN⁺ cells and in primary MDDC.     

The Highly Autoaggregative and Adhesive Phenotype of the Vaginal Lactobacillus plantarum Strain CMPG5300 Is Sortase Dependent

Lactobacilli are important for the maintenance of a healthy ecosystem in the human vagina. Various mechanisms are postulated but so far are poorly substantiated by molecular studies, such as mutant analysis. Bacterial autoaggregation is an interesting phenomenon that can promote adhesion to host cells and displacement of pathogens. In this study, we report on the identification of a human vaginal isolate, Lactobacillus plantarum strain CMPG5300, which shows high autoaggregative and adhesive capacity. To investigate the importance of sortase-dependent proteins (SDPs) in these phenotypes, a gene deletion mutant was constructed for srtA, the gene encoding the housekeeping sortase that covalently anchors these SDPs to the cell surface. This mutant lost the capacity to autoaggregate, showed a decrease in adhesion to vaginal epithelial cells, and lost biofilm-forming capacity under the conditions tested. These results indicate that the housekeeping sortase SrtA of CMPG5300 is a key determinant of the peculiar surface properties of this vaginal Lactobacillus strain.     

Randomised Trial of Planned Caesarean Section Prior to Versus after 39 Weeks: Unscheduled Deliveries and Facility Logistics - A Secondary Analysis

Objectives

To compare the impact of scheduling caesarean section prior to versus after 39 completed weeks of gestation on the occurrence of unscheduled caesarean section and rescheduling of the procedure.

Methods

Secondary analysis from a multicentre randomised open-label trial including singleton pregnant women with a healthy foetus and a reliable due date. Women were allocated by computerized telephone randomisation to planned caesarean section at 38 weeks and three days or 39 weeks and three days. The outcomes were unscheduled deliveries with provided reasons, such as spontaneous labour onset or supervening complications, and any changes in the scheduled delivery date. Statistical analyses were according to intention-to-treat using Fisher’s exact test.

Results

From March 2009 to June 2011 1,274 women were included. Median difference in gestational age at delivery was six days. Compared to the 38 weeks group, the women in the 39 weeks group were more likely to have an unscheduled caesarean section (15.2% vs. 9.3%; RR 1.64, 95% CI 1.21; 2.22), to deliver between 6 pm and 8 am (10 % vs. 6%; RR 1.68, 95% CI 1.14; 2.47), or to have the procedure rescheduled (36.7% vs. 23%; RR 1.6, 95% CI 1.34;1.90).

Conclusions

Scheduling caesarean section after 39 weeks leads to a 60% increase in unscheduled caesarean sections and a 70% increase in delivery outside regular work hours as compared to scheduling of the procedure prior to 39 weeks.

Trial Registration

www.clinicaltrials.gov NCT00835003 http://www.clinicaltrials.gov/ct2/show/NCT00835003?term=NCT00835003&rank=1     

Evaluation of a Screening System for Obesogenic Compounds: Screening of Endocrine Disrupting Compounds and Evaluation of the PPAR Dependency of the Effect

Recently the environmental obesogen hypothesis has been formulated, proposing a role for endocrine disrupting compounds (EDCs) in the development of obesity. To evaluate this hypothesis, a screening system for obesogenic compounds is urgently needed. In this study, we suggest a standardised protocol for obesogen screening based on the 3T3-L1 cell line, a well-characterised adipogenesis model, and direct fluorescent measurement using Nile red lipid staining technique. In a first phase, we characterised the assay using the acknowledged obesogens rosiglitazone and tributyltin. Based on the obtained dose-response curves for these model compounds, a lipid accumulation threshold value was calculated to ensure the biological relevance and reliability of statistically significant effects. This threshold based method was combined with the well described strictly standardized mean difference (SSMD) method for classification of non-, weak- or strong obesogenic compounds. In the next step, a range of EDCs, used in personal and household care products (parabens, musks, phthalates and alkylphenol compounds), were tested to further evaluate the obesogenicity screening assay for its discriminative power and sensitivity. Additionally, the peroxisome proliferator activated receptor γ (PPARγ) dependency of the positive compounds was evaluated using PPARγ activation and antagonist experiments. Our results showed the adipogenic potential of all tested parabens, several musks and phthalate compounds and bisphenol A (BPA). PPARγ activation was associated with adipogenesis for parabens, phthalates and BPA, however not required for obesogenic effects induced by Tonalide, indicating the role of other obesogenic mechanisms for this compound.     

Ectomycorrhizal communities in a productive Tuber aestivum Vittad. orchard: composition, host influence and species replacement

Truffles (Tuber spp.) and other ectomycorrhizal species form species-rich assemblages in the wild as well as in cultivated ecosystems. We aimed to investigate the ectomycorrhizal communities of hazels and hornbeams that are growing in a 24-year-old Tuber aestivum orchard. We demonstrated that the ectomycorrhizal communities included numerous species and were phylogenetically diverse. Twenty-nine ectomycorrhizal taxa were identified. Tuber aestivum ectomycorrhizae were abundant (9.3%), only those of Tricholoma scalpturatum were more so (21.4%), and were detected in both plant symbionts with a variation in distribution and abundance between the two different hosts. The Thelephoraceae family was the most diverse, being represented by 12 taxa. The overall observed diversity represented 85% of the potential one as determined by a jackknife estimation of richness and was significantly higher in hazel than in hornbeam. The ectomycorrhizal communities of hornbeam trees were closely related phylogenetically, whereas no clear distribution pattern was observed for the communities in hazel. Uniform site characteristics indicated that ectomycorrhizal relationships were host mediated, but not host specific. Despite the fact that different plant species hosted diverse ectomycorrhizal communities and that the abundance of T. aestivum differed among sites, no difference was detected in the production of fruiting bodies.