Evaluation of free radical scavenging properties of two classical polyherbal formulations

Two polyherbal formulations of Ayurveda viz., Chandraprabha Vati and Maha yogaraja Guggulu were evaluated for their free radical scavenging properties. Methanolic extracts of the formulations were studied in four different in vitro and ex vivo models. Total phenolic content of Chandraprabha Vati and Maha yogaraja Guggulu was found to be 5.24% and 10.74% respectively. Methanolic extracts of the formulations were good scavengers of all the radicals but there was a difference in the activity of the two formulations in different models. Chandraprabha Vati was a good scavenger of superoxide radical and Maha yogaraja Guggulu was efficient in scavenging nitric oxide (NO), while both inhibited lipid peroxidation efficiently. Free radical scavenging activity of the different extracts can be attributed to the presence of various chemical components including phenolics.     

Type 4 pilus biogenesis and type II-mediated protein secretion by Vibrio cholerae occur independently of the TonB-facilitated proton motive force

In Vibrio cholerae, elaboration of toxin-coregulated pilus and protein secretion by the extracellular protein secretion apparatus occurred in the absence of both TonB systems. In contrast, the cognate putative ATPases were required for each process and could not substitute for each other.     

The flavonoid,quercetin, differentially regulates Th-1 (IFNgamma) and Th-2 (IL4) cytokine gene expression by normal peripheral blood mononuclear cells

Flavonoids are plant metabolites that are dietary antioxidants and exert significant anti-tumor, anti-allergic, anti-inflammatory and anti-viral effects. It is generally accepted that Th-1 derived cytokines such as IL-2, IFNgamma and IL-12 promote cellular immunity while Th-2 derived cytokines such as IL-4, IL-5, IL-6 exert negative immunoregulatory effects on cellular immunity while upregulating humoral immunity. The molecular mechanisms underlying the biological activities of flavonoids have not been elucidated. We hypothesize that the flavonoid, quercetin, exert significant anti-viral and anti-tumor effects possibly by modulating the production of Th-1 and Th-2 derived cytokines. Peripheral blood mononuclear cells (PBMC, 1 x 10(6) cells/ml) from normal subjects were cultured with different concentrations of quercetin (0.5-50 microM) for 24-72 h and supernates were quantitated for IFN-gamma and IL-4 by ELISA and antiviral activity of IFNgamma by bioassay. FACS analysis was done to determine the number of IFN-gamma and IL-4 positive cells and RT-PCR was done to quantitate gene expression. Quercetin significantly induces the gene expression as well as the production of Th-1 derived IFNgamma and the downregulates Th-2 derived IL-4 by normal PBMC. Further, quercetin treatment increased the phenotypic expression of IFNgamma cells and decreased IL-4 positive cells by FACS analysis, which corroborate with protein secretion and gene expression studies. These results suggest that the beneficial immuno-stimulatory effects of quercetin may be mediated through the induction of Th-1 derived cytokine, IFNgamma, and inhibition of Th-2 derived cytokine, IL-4.     

Books received

Publications Received

Books received     

An actin‐like protein is involved in regulation of mitochondrial and flagellar functions as well as in intramacrophage survival of Leishmania donovani

Actin‐related proteins are ubiquitous actin‐like proteins that show high similarity with actin in terms of their amino acid sequence and three‐dimensional structure. However, in lower eukaryotes, such as trypanosomatids, their functions have not yet been explored. Here, we show that a novel actin‐related protein (ORF LmjF.13.0950) is localized mainly in the Leishmania mitochondrion. We further reveal that depletion of the intracellular levels of this protein leads to an appreciable decrease in the mitochondrial membrane potential as well as in the ATP production, which appears to be accompanied with impairment in the flagellum assembly and motility. Additionally, we report that the mutants so generated fail to survive inside the mouse peritoneal macrophages. These abnormalities are, however, reversed by the episomal gene complementation. Our results, for the first time indicate that apart from their classical roles in the cytoplasm and nucleus, actin‐related proteins may also regulate the mitochondrial function, and in case of Leishmania donovani they may also serve as the essential factor for their survival in the host cells.     

Tracking Dengue Virus Intra-host Genetic Diversity during Human-to-Mosquito Transmission

Dengue virus (DENV) infection of an individual human or mosquito host produces a dynamic population of closely-related sequences. This intra-host genetic diversity is thought to offer an advantage for arboviruses to adapt as they cycle between two very different host species, but it remains poorly characterized. To track changes in viral intra-host genetic diversity during horizontal transmission, we infected Aedes aegypti mosquitoes by allowing them to feed on DENV2-infected patients. We then performed whole-genome deep-sequencing of human- and matched mosquito-derived DENV samples on the Illumina platform and used a sensitive variant-caller to detect single nucleotide variants (SNVs) within each sample. >90% of SNVs were lost upon transition from human to mosquito, as well as from mosquito abdomen to salivary glands. Levels of viral diversity were maintained, however, by the regeneration of new SNVs at each stage of transmission. We further show that SNVs maintained across transmission stages were transmitted as a unit of two at maximum, suggesting the presence of numerous variant genomes carrying only one or two SNVs each. We also present evidence for differences in selection pressures between human and mosquito hosts, particularly on the structural and NS1 genes. This analysis provides insights into how population drops during transmission shape RNA virus genetic diversity, has direct implications for virus evolution, and illustrates the value of high-coverage, whole-genome next-generation sequencing for understanding viral intra-host genetic diversity.     

Development and Internal Validation of a Predictive Model Including Pulse Oximetry for Hospitalization of Under-Five Children in Bangladesh

Background

The reduction in the deaths of millions of children who die from infectious diseases requires early initiation of treatment and improved access to care available in health facilities. A major challenge is the lack of objective evidence to guide front line health workers in the community to recognize critical illness in children earlier in their course.

Methods

We undertook a prospective observational study of children less than 5 years of age presenting at the outpatient or emergency department of a rural tertiary care hospital between October 2012 and April 2013. Study physicians collected clinical signs and symptoms from the facility records, and with a mobile application performed recordings of oxygen saturation, heart rate and respiratory rate. Facility physicians decided the need for hospital admission without knowledge of the oxygen saturation. Multiple logistic predictive models were tested.

Findings

Twenty-five percent of the 3374 assessed children, with a median (interquartile range) age of 1.02 (0.42–2.24), were admitted to hospital. We were unable to contact 20% of subjects after their visit. A logistic regression model using continuous oxygen saturation, respiratory rate, temperature and age combined with dichotomous signs of chest indrawing, lethargy, irritability and symptoms of cough, diarrhea and fast or difficult breathing predicted admission to hospital with an area under the receiver operating characteristic curve of 0.89 (95% confidence interval -CI: 0.87 to 0.90). At a risk threshold of 25% for admission, the sensitivity was 77% (95% CI: 74% to 80%), specificity was 87% (95% CI: 86% to 88%), positive predictive value was 70% (95% CI: 67% to 73%) and negative predictive value was 91% (95% CI: 90% to 92%).

Conclusion

A model using oxygen saturation, respiratory rate and temperature in combination with readily obtained clinical signs and symptoms predicted the need for hospitalization of critically ill children. External validation of this model in a community setting will be required before adoption into clinical practice.     

The JH2 domain and SH2-JH2 linker regulate JAK2 activity: A detailed kinetic analysis of wild type and V617F mutant kinase domains

JAK2 tyrosine kinase regulates many cellular functions. Its activity is controlled by the pseudokinase (JH2) domain by still poorly understood mechanisms. The V617F mutation in the pseudokinase domain activates JAK2 and causes myeloproliferative neoplasms. We conducted a detailed kinetic analysis of recombinant JAK2 tyrosine kinase domain (JH1) and wild-type and V617F tandem kinase (JH1JH2) domains using peptide microarrays to define the functions of the kinase domains. The results show that i) JAK2 follows a random Bi–Bi reaction mechanism ii) JH2 domain restrains the activity of the JH1 domain by reducing the affinity for ATP and ATP competitive inhibitors iii) V617F decreases affinity for ATP but increases catalytic activity compared to wild-type and iv) the SH2–JH2 linker region participates in controlling activity by reducing the affinity for ATP.     

Characterisation of the nuclear proteome of a dehydration‐sensitive cultivar of chickpea and comparative proteomic analysis with a tolerant cultivar

Water deficit or dehydration hampers plant growth and development, and shrinks harvest size of major crop species worldwide. Therefore, a better understanding of dehydration response is the key to decipher the regulatory mechanism of better adaptation. In recent years, nuclear proteomics has become an attractive area of research, particularly to study the role of nucleus in stress response. In this study, a proteome of dehydration‐sensitive chickpea cultivar (ICCV‐2) was generated from nuclei‐enriched fractions. The LC‐MS/MS analysis led to the identification of 75 differentially expressed proteins presumably associated with different metabolic and regulatory pathways. Nuclear localisation of three candidate proteins was validated by transient expression assay. The ICCV‐2 proteome was then compared with that of JG‐62, a tolerant cultivar. The differential proteomics and in silico analysis revealed cultivar‐specific differential expression of many proteins involved in various cellular functions. The differential tolerance could be attributed to altered expression of many structural proteins and the proteins involved in stress adaptation, notably the ROS catabolising enzymes. Further, a comprehensive comparison on the abiotic stress‐responsive nuclear proteome was performed using the datasets published thus far. These findings might expedite the functional determination of the dehydration‐responsive proteins and their prioritisation as potential molecular targets for better adaptation.     

Small peptides derived from somatotropin domain-containing proteins inhibit blood and lymphatic endothelial cell proliferation, migration, adhesion and tube formation

Angiogenesis is thoroughly balanced and regulated in health; however, it is dysregulated in many diseases including cancer, age-related macular degeneration, cardiovascular diseases such as coronary and peripheral artery diseases and stroke, abnormal embryonic development, and abnormal wound healing. In addition to angiogenesis, lymphangiogenesis is pivotal for maintaining the immune system, homeostasis of body fluids and lymphoid organs; dysregulated lymphangiogenesis may cause inflammatory diseases and lymph node mediated tumor metastasis. Anti-angiogenic or anti-lymphangiogenic small peptides may play an important role as therapeutic agents normalizing angiogenesis or lymphangiogenesis in disease conditions. Several novel endogenous peptides derived from proteins containing a conserved somatotropin domain have been previously identified with the help of our bioinformatics-based methodology. These somatotropin peptides were screened for inhibition of angiogenesis and lymphangiogenesis using in vitro proliferation, migration, adhesion and tube formation assays with blood and lymphatic endothelial cells. We found that the peptides have the potential for inhibiting both angiogenesis and lymphangiogenesis. Focusing the study on the inhibition of lymphangiogenesis, we found that a peptide derived from the somatotropin conserved domain of transmembrane protein 45A human was the most potent lymphangiogenesis inhibitor, blocking lymphatic endothelial cell migration, adhesion, and tube formation.