Biology subjects

Sharma, J. B.

Publications and source records attributed to Sharma, J. B..

3 recordsLinked to original sources

bioRxiv2021

Hypoxia induces early neurogenesis in human fetal neural stem cells by activating the WNT pathway.

Fetal neural stem cells (FNSCs) are present in the brain of human fetuses that differentiate into cells of neuronal and glial lineages. Difference in oxygen concentration between maternal and fetal circulation indicates that the developing fetus may be exposed to lower oxygen concentrations compared to adults. This physiological hypoxia may influence the growth and differentiation of the FNSCs. This study aimed to evaluate the effect of hypoxia on the differentiation potential of human FNSCs isolated from the sub-ventricular zone of aborted fetal brains (n=5). FNSCs were isolated, expanded, and characterized by Nestin and Sox2 expression, using immunocytochemistry and flowcytometry respectively. These FNSCs were exposed to 20% oxygen (normoxia) and 0.2% oxygen (hypoxia) concentrations for 48 hours, and hypoxia exposure (n=5) was evaluated by a panel of markers (CA9, PGK1 and VEGF). Whole human genome transcriptomic analyses (Genespring GX13) of FNSCs exposed to hypoxia (Agilent 4x44K human array slides), highlighted that genes associated with neurogenesis were getting enriched. The pathway analysis of these enriched genes (using Metacore) showed that WNT signaling played a role in determining the cell fate of FNSCs exposed to hypoxic environment. Microarray analyses was validated using neuronal and glial lineage commitment markers such as Ngn1, Ngn2, ASCL1, DCX, GFAP, Olig2 and Nkx2.2 using qPCR (n=9). This demonstrated upregulation of the neuronal commitment markers on hypoxia exposure, while no change was observed in astrocytic and oligodendrocyte lineage commitment markers. Increased expression of downstream targets of the WNT signaling pathway, TCF4 and ID2, by qPCR (n=9), indicated its involvement in mediating neuronal differentiation on exposure to hypoxia.

neuroscience↗

bioRxiv2021

Human fetal neural stem cell-derived astrocytes maintain glutamate transport after hypoxic injury in vitro

Astrocytes are the most abundant glial cells that play many critical roles in the central nervous system physiology including the uptake of excess glutamate from the synapse by Excitatory Amino Acid Transporters (EAATs). Among the EAATs, EAAT2 are predominantly functional, astrocyte-specific glutamate transporters in the forebrain. Hypoxic brain injury is a pathological phenomenon seen in various clinical conditions including stroke and neonatal hypoxic ischemic encephalopathy. Glutamate excitotoxicity is an important cause of neuronal cell death in disorders involving hypoxic brain injury. As findings from rodent models cannot always be reliably extrapolated to humans, we aimed to develop a homogenous population of primary human astrocytes to study the effect of hypoxic injury on astrocyte function, especially glutamate uptake. We successfully isolated, established and characterized cultures of human fetal neural stem cells (FNSCs) from aborted fetal brains. FNSCs were differentiated into astrocytes, and characterized by increased expression of the astrocyte markers, glial fibrillary acidic protein (GFAP), EAAT1 and EAAT2. A concomitant decrease in neural stem cell marker, Nestin, was observed. Differentiated astrocytes were exposed to various oxygen concentrations mimicking normoxia (20% and 6%), moderate and severe hypoxia (2% and 0.2% respectively). Interestingly, no change was observed in the expression of glutamate transporter, EAAT2 and glutamate uptake by astrocytes, even after exposure to hypoxia. Our novel model of human FNSC derived astrocytes exposed to hypoxic injury, establishes that astrocytes are able to maintain glutamate uptake even after exposure to severe hypoxia for 48 hours, and thus provides evidence for the neuroprotective role of astrocytes in hypoxic injury.

neuroscience↗

bioRxiv2020

Genotypic and antigenic study of SARS-CoV-2 from an Indian isolate.

Coronaviruses (CoVs) are one of the largest groups of positive-sense RNA virus families within the Nidovirales order, which are further classified into four genera: alpha, beta, gamma, and delta. Coronaviruses have an extensive range of natural hosts and are known to be responsible for a broad spectrum of diseases in multiple species. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the ongoing coronavirus disease 2019 (COVID-19) that has unleashed a global threat to public health and the economy. Coronaviruses are extensively present in birds and mammals, with horseshoe bats (Rhinolophus affinis), being the reservoir for the ongoing SARS-CoV-2 that seems to have resulted from a zoonotic spillover to the human host, causing respiratory infections, lung injury and Acute Respiratory Distress Syndrome(ARDS). About six coronavirus serotypes are linked with the disease in humans, namely HCoV-229E, HCoV-NL63, HCoV-OC43, HCoV-HKU1, SARS-CoV, SARS-CoV-2, and MERS-CoV. SARS-CoV-2 is the seventh CoV to infect humans. We analyzed the genome sequence of CoV-2 from isolates derived from China as well from India and encountered minute variations in their sequence. A cladogram analysis revealed the predominant strain circulating in India belongs to the A2a clad. We took one such strain (MT012098) and performed a rigorous in-silico genotypic and antigenic analysis to identify its relatedness to other strains. Further, we also performed a detailed prediction for B and T cell epitopes using BepiPred 2.0 server and NetCTL 1.2 server (DTU Bioinformatics), respectively. We hope this information may assist in an effective vaccine designing program against SARS-CoV-2.

biochemistry↗