Vitamin D Prevents Hypoxia/Reoxygenation-Induced Blood-Brain Barrier Disruption via Vitamin D Receptor-Mediated NF-kB Signaling Pathways

Maintaining blood-brain barrier integrity and minimizing neuronal injury are critical components of any therapeutic intervention following ischemic stroke. However, a low level of vitamin D hormone is a risk factor for many vascular diseases including stroke. The neuroprotective effects of 1,25(OH)2D3 (vitamin D) after ischemic stroke have been studied, but it is not known whether it prevents ischemic injury to brain endothelial cells, a key component of the neurovascular unit. We analyzed the effect of 1,25(OH)₂D₃ on brain endothelial cell barrier integrity and tight junction proteins after hypoxia/reoxygenation in a mouse brain endothelial cell culture model that closely mimics many of the features of the blood-brain barrier in vitro. Following hypoxic injury in bEnd.3 cells, 1,25(OH)₂D₃ treatment prevented the decrease in barrier function as measured by transendothelial electrical resistance and permeability of FITC-dextran (40 kDa), the decrease in the expression of the tight junction proteins zonula occludin-1, claudin-5, and occludin, the activation of NF—kB, and the increase in matrix metalloproteinase-9 expression. These responses were blocked when the interaction of 1,25(OH) )₂D₃ with the vitamin D receptor (VDR) was inhibited by pyridoxal 5’-phosphate treatment. Our findings show a direct, VDR-mediated, protective effect of 1,25(OH) )₂D₃ against ischemic injury-induced blood-brain barrier dysfunction in cerebral endothelial cells.     

Child Centred Approach to Climate Change and Health Adaptation through Schools in Bangladesh: A Cluster Randomised Intervention Trial

Background

Bangladesh is one of the most vulnerable countries to climate change. People are getting educated at different levels on how to deal with potential impacts. One such educational mode was the preparation of a school manual, for high school students on climate change and health protection endorsed by the National Curriculum and Textbook Board, which is based on a 2008 World Health Organization manual. The objective of this study was to test the effectiveness of the manual in increasing the knowledge level of the school children about climate change and health adaptation.

Methods

This cluster randomized intervention trial involved 60 schools throughout Bangladesh, with 3293 secondary school students participating. School upazilas (sub-districts) were randomised into intervention and control groups, and two schools from each upazila were randomly selected. All year seven students from both groups of schools sat for a pre-test of 30 short questions of binary response. A total of 1515 students from 30 intervention schools received the intervention through classroom training based on the school manual and 1778 students of the 30 control schools did not get the manual but a leaflet on climate change and health issues. Six months later, a post-intervention test of the same questionnaire used in the pre-test was performed at both intervention and control schools. The pre and post test scores were analysed along with the demographic data by using random effects model.

Results

None of the various school level and student level variables were significantly different between the control and intervention group. However, the intervention group had a 17.42% (95% CI: 14.45 to 20.38, P = <0.001) higher score in the post-test after adjusting for pre-test score and other covariates in a multi-level linear regression model.

Conclusions

These results suggest that school-based intervention for climate change and health adaptation is effective for increasing the knowledge level of school children on this topic.     

Bioelectricity Generation and Bioremediation of an Azo-Dye in a Microbial Fuel Cell Coupled Activated Sludge Process

Simultaneous bioelectricity generation and dye degradation was achieved in the present study by using a combined anaerobic-aerobic process. The anaerobic system was a typical single chambered microbial fuel cell (SMFC) which utilizes acid navy blue r (ANB) dye along with glucose as growth substrate to generate electricity. Four different concentrations of ANB (50, 100, 200 and 400 ppm) were tested in the SMFC and the degradation products were further treated in an activated sludge post treatment process. The dye decolorization followed pseudo first order kinetics while the negative values of the thermodynamic parameter ∆G (change in Gibbs free energy) shows that the reaction proceeds with a net decrease in the free energy of the system. The coulombic efficiency (CE) and power density (PD) attained peak values at 10.36% and 2,236 mW/m² respectively for 200 ppm of ANB. A further increase in ANB concentrations results in lowering of cell potential (and PD) values owing to microbial inhibition at higher concentrations of toxic substrates. Cyclic voltammetry studies revealed a perfect redox reaction was taking place in the SMFC. The pH, temperature and conductivity remain 7.5–8.0, 27(±2°C and 10.6–18.2 mS/cm throughout the operation. The biodegradation pathway was studied by the gas chromatography coupled with mass spectroscopy technique, suggested the preferential cleavage of the azo bond as the initial step resulting in to aromatic amines. Thus, a combined anaerobic-aerobic process using SMFC coupled with activated sludge process can be a viable option for effective degradation of complex dye substrates along with energy (bioelectricity) recovery.     

DisPredict: A Predictor of Disordered Protein Using Optimized RBF Kernel

Intrinsically disordered proteins or, regions perform important biological functions through their dynamic conformations during binding. Thus accurate identification of these disordered regions have significant implications in proper annotation of function, induced fold prediction and drug design to combat critical diseases. We introduce DisPredict, a disorder predictor that employs a single support vector machine with RBF kernel and novel features for reliable characterization of protein structure. DisPredict yields effective performance. In addition to 10-fold cross validation, training and testing of DisPredict was conducted with independent test datasets. The results were consistent with both the training and test error minimal. The use of multiple data sources, makes the predictor generic. The datasets used in developing the model include disordered regions of various length which are categorized as short and long having different compositions, different types of disorder, ranging from fully to partially disordered regions as well as completely ordered regions. Through comparison with other state of the art approaches and case studies, DisPredict is found to be a useful tool with competitive performance. DisPredict is available at https://github.com/tamjidul/DisPredict_v1.0.     

Metabolite Profiling of Low-P Tolerant and Low-P Sensitive Maize Genotypes under Phosphorus Starvation and Restoration Conditions

Background

Maize (Zea mays L.) is one of the most widely cultivated crop plants. Unavoidable economic and environmental problems associated with the excessive use of phosphatic fertilizers demands its better management. The solution lies in improving the phosphorus (P) use efficiency to sustain productivity even at low P levels. Untargeted metabolomic profiling of contrasting genotypes provides a snap shot of whole metabolome which differs under specific conditions. This information provides an understanding of the mechanisms underlying tolerance to P stress and the approach for increasing P-use-efficiency.

Methodology/Principal Findings

A comparative metabolite-profiling approach based on gas chromatography-mass spectrometry (GC/MS) was applied to investigate the effect of P starvation and its restoration in low-P sensitive (HM-4) and low-P tolerant (PEHM-2) maize genotypes. A comparison of the metabolite profiles of contrasting genotypes in response to P-deficiency revealed distinct differences among low-P sensitive and tolerant genotypes. Another set of these genotypes were grown under P-restoration condition and sampled at different time intervals (3, 5 and 10 days) to investigate if the changes in metabolite profile under P-deficiency was restored. Significant variations in the metabolite pools of these genotypes were observed under P-deficiency which were genotype specific. Out of 180 distinct analytes, 91 were identified. Phosphorus-starvation resulted in accumulation of di- and trisaccharides and metabolites of ammonium metabolism, specifically in leaves, but decreased the levels of phosphate-containing metabolites and organic acids. A sharp increase in the concentrations of glutamine, asparagine, serine and glycine was observed in both shoots and roots under low-P condition.

Conclusion

The new insights generated on the maize metabolome in resposne to P-starvation and restoration would be useful towards improvement of the P-use efficiency in maize.     

High Throughput Sequencing Analysis of the Immunoglobulin Heavy Chain Gene from Flow-Sorted B Cell Sub-Populations Define the Dynamics of Follicular Lymphoma Clonal Evolution

Understanding the dynamics of evolution of Follicular Lymphoma (FL) clones during disease progression is important for monitoring and targeting this tumor effectively. Genetic profiling of serial FL biopsies and examples of FL transmission following bone marrow transplant suggest that this disease may evolve by divergent evolution from a common ancestor cell. However where this ancestor cell resides and how it evolves is still unclear. The analysis of the pattern of somatic hypermutation of the immunoglobulin gene (Ig) is traditionally used for tracking the physiological clonal evolution of B cells within the germinal center and allows to discriminate those cells that have just entered the germinal center and display features of ancestor cells from those B cells that keep re-circulating across different lymphoid organs. Here we investigated the pattern of somatic hypermutation of the heavy chain of the immunoglobulin gene (IgH-VH) in 4 flow-sorted B cells subpopulations belonging to different stages of differentiation, from sequential lymph node biopsies of cases displaying diverse patterns of evolution, using the GS-FLX Titanium sequencing platform. We observed an unexpectedly high level of clonality, with hundreds of distinct tumor subclones in the different subpopulations from the same sample, the majority detected at a frequency <10⁻². By using a lineage trees analysis we observed in all our FL and t-FL cases that the oligoclonal FL population was trapped in a narrow intermediate stage of maturation that maintains the capacity to undergo SHM, but was unable to further differentiate. The presence of such a complex architecture highlights challenges currently encountered in finding a cure for this disease.     

An optimized and validated RP-HPLC/UV detection method for simultaneous determination of all-trans-Retinol (Vitamin A) and α-Tocopherol (Vitamin E) in human serum: Comparison of different particulate reversed-phase HPLC columns

A novel, simple and fast reversed-phase HPLC/UV method was developed, optimized for various chromatographic conditions, and validated according to international guidelines for simultaneous determination of all-trans-retinol and α-tocopherol in human serum using retinyl acetate as internal standard in the concentration of 0.5 μg/ml. A liquid-phase extraction was applied to the 250 μl of serum with n-hexane–dichloromethane mixture (70:30, v/v), in two steps, using ethanol–methanol mixture (95:5, v/v) for protein precipitation and BHT (butylated hydroxy toluene) as stabilizer for sample preparation. Both analytes were analyzed on Kromasil 100 C₁₈ column (150 mm × 4.6 mm, 5 μm), Brownlee analytical (Perkin Elmer) C₁₈ column (150 mm × 4.6 mm, 5 μm), and Supelco (Supelcosil) LC-18 column (150 mm × 3 mm, 3 μm), protected by a Perkin Elmer C₁₈ (30 mm × 4.6 mm, 10 μm; Norwalk, USA) pre-column guard cartridge, at 292 nm wavelength, using methanol–water (99:1, v/v), in isocratic mode as mobile phase applied at flow rate of 1.5 ml/min and 1 ml/min for both 5 μm and 3 μm columns, respectively. Complete separation of all the analytes was achieved in 3 and 6 min on 3 μm and 5 μm columns, respectively by injecting 20 μl of sample into the HPLC system by autosampler, keeping column oven temperature at 25 °C. Different particulate reversed-phase chromatographic columns were evaluated in order to select the best column in terms of sensitivity, selectivity, resolution and short run time of both the analytes and it was concluded that 3 μm columns are better to be used in clinical set up as well as in laboratories for the separation of these analytes in a shorter time as compared with 5 μm columns. The method was validated and applied for the analysis of all-trans-retinol and α-tocopherol in the serum of human volunteers.     

Induction of phytochelatins and antioxidant defence system in <Emphasis Type="Italic">Brassica juncea</Emphasis> and <Emphasis Type="Italic">Vigna radiata</Emphasis> in response to chromium treatments

Growth performance, chromium (Cr) accumulation potential and induction of antioxidative defence system and phytochelatins (PCs) were studied in hydroponically grown Brassica juncea (Indian mustard) and Vigna radiata (mungbean) at various levels of Cr treatments (0, 50, 100, 200 μM Cr). B. juncea accumulated twofolds and threefolds higher Cr in root and shoot, respectively than in V. radiata. Compared to B. juncea, V. radiata was found to be particularly sensitive to Cr as observed by the severity and development of Cr toxicity symptoms and decreased growth. Induction of PC and enzymes of antioxidant defence system were monitored as plant’s primary and secondary metal detoxifying responses, respectively. There was induction of PC and enzymes of antioxidant defence system in both the plants. PCs were induced significantly in roots and shoot of both the plants at all the levels of Cr treatments. Significantly higher activities of superoxide dismutase (SOD), ascorbate peroxidase (APX), catalase (CAT) and glutathione reductase (GR) were observed in shoot of B. juncea than V. radiata at all the levels of Cr treatments. Induction of PCs along with antioxidant defence system in response to Cr stress suggests the cumulative role of PCs and antioxidants in conferring tolerance against accumulated Cr in B. juncea, and thereby signifies the suitability of this plant as one of the potential remediators of Cr.     

Encapsulation of recombinant <Emphasis Type="Italic">E. coli</Emphasis> expressing cyclopentanone monooxygenase in polyelectrolyte complex capsules for Baeyer–Villiger biooxidation of 8-oxabicyclo[3.2.1]oct-6-en-3-one

Recombinant Escherichia coli cells, over-expressing cyclopentanone monooxygenase activity, were immobilized in polyelectrolyte complex capsules, made of sodium alginate, cellulose sulfate, poly(methylene-co-guanidine), CaCl₂ and NaCl. More than 90% of the cell viability was preserved during the encapsulation process. Moreover, the initial enzyme activity was fully maintained within encapsulated cells while it halved in free cells. Both encapsulated and free cells reached the end point of the Baeyer–Villiger biooxidation of 8-oxabicyclo[3.2.1]oct-6-en-3-one to 4,9-dioxabicyclo[4.2.1]non-7-en-3-one at the same time (48 h). Similarly, the enantiomeric excess above 94% was identical for encapsulated and free cells.