Selenium Downregulates RAGE and NFκB Expression in Diabetic Rats

Diabetes is one of the leading causes of death in developed and developing countries. Oxidative stress has been proposed to play a crucial role in the pathogenesis of diabetic vascular complications. In recent years, selenium has been shown to mediate a number of insulinlike actions in a dose-dependent fashion both in vitro and in vivo. In this study, the effect of selenium as sodium selenite was investigated in streptozotocin-induced diabetic rats at the dose of 1?μg sodium selenite/kg body weight. Selenium supplementation restored the streptozotocin-induced alterations in the activities of antioxidant enzymes, decreased the serum glucose level, glycated hemoglobin content as well as the levels of lipid peroxidation products, and downregulated the expressions of both NFkB and RAGE. The histopathological studies also reinforce our findings. Hence, selenium has a protective role in streptozotocin-induced diabetes mellitus.     

A novel catalytic mechanism for ATP hydrolysis employed by the N-terminal nucleotide-binding domain of Cdr1p, a multidrug ABC transporter of Candida albicans

Although essentially conserved, the N-terminal nucleotide-binding domain (NBD) of Cdr1p and other fungal transporters has some unique substitutions of amino acids which appear to have functional significance for the drug transporters. We have previously shown that the typical Cys193 in Walker A as well as Trp326 and Asp327 in the Walker B of N-terminal NBD (NBD-512) of Cdr1p has acquired unique roles in ATP binding and hydrolysis. In the present study, we show that due to spatial proximity, fluorescence resonance energy transfer (FRET) takes place between Trp326 of Walker B and MIANS [2-(4-maleimidoanilino) naphthalene-6-sulfonic acid] on Cys193 of Walker A motif. By exploiting FRET, we demonstrate how these critical amino acids are positioned within the nucleotide-binding pocket of NBD-512 to bind and hydrolyze ATP. Our results show that both Mg²⁺ coordination and nucleotide binding contribute to the formation of the active site. The entry of Mg²⁺ into the active site causes the first large conformational change that brings Trp326 and Cys193 in close proximity to each other. We also show that besides Trp326, typical Glu238 in the Q-loop also participates in coordination of Mg²⁺ by NBD-512. A second conformational change is induced when ATP, but not ADP, docks into the pocket. Asn328 does sensing of the γ-phosphate of the substrate in the extended Walker B motif, which is essential for the second conformational change that must necessarily precede ATP hydrolysis. Taken together our results imply that the uniquely placed residues in NBD-512 have acquired critical roles in ATP catalysis, which drives drug extrusion.     

Identification of marker genes in Alzheimer's disease using a machine-learning model

Alzheimer''s Disease (AD) is one of the most common causes of dementia, mostly affecting the elderly population. Currently, there is no proper diagnostic tool or method available for the detection of AD. The present study used two distinct data sets of AD genes, which could be potential biomarkers in the diagnosis. The differentially expressed genes (DEGs) curated from both datasets were used for machine learning classification, tissue expression annotation and co-expression analysis. Further, CNPY3, GPR84, HIST1H2AB, HIST1H2AE, IFNAR1, LMO3, MYO18A, N4BP2L1, PML, SLC4A4, ST8SIA4, TLE1 and N4BP2L1 were identified as highly significant DEGs and exhibited co-expression with other query genes. Moreover, a tissue expression study found that these genes are also expressed in the brain tissue. In addition to the earlier studies for marker gene identification, we have considered a different set of machine learning classifiers to improve the accuracy rate from the analysis. Amongst all the six classification algorithms, J48 emerged as the best classifier, which could be used for differentiating healthy and diseased samples. SMO/SVM and Logit Boost further followed J48 to achieve the classification accuracy.     

Identification of a peroxide-sensitive redox switch at the CXXC motif in the human mitochondrial branched chain aminotransferase

The human mitochondrial branched chain aminotransferase isoenzyme (hBCATm) must be stored in a reducing environment to remain active. Oxidation or labeling of hBCATm with sulfhydryl reagents results in enzyme inhibition. In this study, we investigated both the structural and biochemical basis for the sensitivity of hBCATm to these reagents. In its native form, hBCATm has two reactive cysteine residues which were identified as Cys315 and Cys318 using iodinated beta-(4-hydroxyphenyl)ethyl maleimide. These are located in the large domain of the homodimer, about 10 A from the active site. The crystal structures show evidence for a thiol-thiolate hydrogen bond between Cys315 and Cys318. Under oxidizing conditions, these cysteine residues can reasonably form a disulfide bond because of the short distance between the sulfur atoms (3.09-3.46 A), requiring only a decrease of 1.1-1.5 A. In addition to Cys315 playing a structural role by anchoring Tyr173, which in the ketimine form increases access to the active site, our evidence indicates that these cysteine residues act as a redox switch in hBCATm. Electrospray ionization mass spectrometry analysis and UV-Vis spectroscopic studies of 5,5'-dithiobis(2-nitrobenzoic acid) labeled hBCATm showed that during labeling, an intrasubunit disulfide bond was formed in a significant portion of the protein. Furthermore, it was established that reaction of hBCATm with H2O2 abolished its activity and resulted in the formation of an intrasubunit disulfide bond between Cys315 and Cys318. Addition of dithiothreitol completely reversed the oxidation and restored activity. Therefore, the results demonstrate that there is redox-linked regulation of hBCATm activity by a peroxide sensitive CXXC center. Future studies will determine if this center has an in vivo role in the regulation of branched chain amino acid metabolism.     

Suppression,disaggregation, and modulation of γ‐Synuclein fibrillation pathway by green tea polyphenol EGCG

Oligomerization of γ‐Synuclein is known to have implications for both neurodegeneration and cancer. Although it is known to co‐exist with the fibrillar deposits of α‐Synuclein (Lewy bodies), a hallmark in Parkinson's disease (PD), the effect of potential therapeutic modulators on the fibrillation pathway of γ‐Syn remains unexplored. By a combined use of various biophysical tools and cytotoxicity assays we demonstrate that the flavonoid epigallocatechin‐3‐gallate (EGCG) significantly suppresses γ‐Syn fibrillation by affecting its nucleation and binds with the unstructured, nucleus forming oligomers of γ‐Syn to modulate the pathway to form α‐helical containing higher‐order oligomers (~158 kDa and ~ 670 kDa) that are SDS‐resistant and conformationally restrained in nature. Seeding studies reveal that these oligomers although “on‐pathway” in nature, are kinetically retarded and rate‐limiting species that slows down fibril elongation. We observe that EGCG also disaggregates the protofibrils and mature γ‐Syn fibrils into similar SDS‐resistant oligomers. Steady‐state and time‐resolved fluorescence spectroscopy and isothermal titration calorimetry (ITC) reveal a weak non‐covalent interaction between EGCG and γ‐Syn with the dissociation constant in the mM range (K_d ~ 2–10 mM). Interestingly, while EGCG‐generated oligomers completely rescue the breast cancer (MCF‐7) cells from γ‐Syn toxicity, it reduces the viability of neuroblastoma (SH‐SY5Y) cells. However, the disaggregated oligomers of γ‐Syn are more toxic than the disaggregated fibrils for MCF‐7cells. These findings throw light on EGCG‐mediated modulation of γ‐Syn fibrillation and suggest that investigation on the effects of such modulators on γ‐Syn fibrillation is critical in identifying effective therapeutic strategies using small molecule modulators of synucleopathies.     

Burden of Complicated Malaria in a Densely Forested Bastar Region of Chhattisgarh State (Central India)

Background

A prospective study on severe and complicated malaria was undertaken in the tribal dominated area of Bastar division, Chhattisgarh (CG), Central India, with an objective to understand the clinical epidemiology of complicated malaria in patients attending at a referral hospital.

Methods

Blood smears, collected from the general medicine and pediatric wards of a government tertiary health care facility located in Jagdalpur, CG, were microscopically examined for malaria parasite from July 2010 to December 2013. The Plasmodium falciparum positive malaria cases who met enrollment criteria and provided written informed consent were enrolled under different malaria categories following WHO guidelines. PCR was performed to reconfirm the presence of P.falciparum mono infection among enrolled cases. Univariate and multivariate logistic regression analysis was done to identify different risk factors using STATA 11.0.

Results

A total of 40,924 cases were screened for malaria. The prevalence of malaria and P.falciparum associated complicated malaria (severe and cerebral both) in the hospital was 6% and 0.81%, respectively. P.falciparum malaria prevalence, severity and associated mortality in this region peaked at the age of>4–5 years and declined with increasing age. P.falciparum malaria was significantly more prevalent in children than adults (P<0.00001). Among adults, males had significantly more P.falciparum malaria than females (P<0.00001). Case fatality rate due to cerebral malaria and severe malaria was, respectively, 32% and 9% among PCR confirmed mono P.falciparum cases. Coma was the only independent predictor of mortality in multivariate regression analysis. Mortality was significantly associated with multi-organ complication score (P = 0.0003).

Conclusion

This study has revealed that the pattern of morbidity and mortality in this part of India is very different from earlier reported studies from India. We find that the peak morbidity and mortality in younger children regardless of seasonality. This suggests that this age group needs special care for control and clinical management.