Biology subjects

Mahapatra, N. R.

Publications and source records attributed to Mahapatra, N. R..

5 recordsLinked to original sources

bioRxiv2026

An Efficient Microfluidic Platform for Organ-Specific CyclicMultiaxial Loading and Differentiation of Progenitor Cells

Mechanical loading is a fundamental regulator of tissue development, maturation, and function, yet existing in vitro systems predominantly apply uniaxial load/strain to cell-laden hydrogels, limiting their ability to reproduce physiologically relevant multiaxial on-chip tissue mechanics. Here, we present an engineered system to apply controlled multiaxial mechanical loading to real scaffolds with undifferentiated cells. The system enables programmable cyclic stretching and relaxation at the cell scaffold level and is compatible with both cell-laden native scaffold and engineered soft tissues. We have quantified the strain field experienced by the tissue with distinct mechanical loading periods using computation simulations and validated these with systematic experiments, demonstrating reproducible and various organ-relevant multiaxial strain profiles. As proof of concept, we applied the approach to human cardiac tissues, where cyclic multiaxial mechanical loading of mononuclear cells laden native tissue scaffold promoted cardiac structural organization and supports the emergence of spontaneous contractile activity. This approach provides a generalizable method for reproducing physiologically relevant tissue mechanics in vitro and enables systematic investigation of mechano-regulation across a plethora of human tissues.

bioengineering↗

bioRxiv2026

Dynamic regulation of cystathionine gamma-lyase expression during normoxia, ischemia, and reperfusion in liver: interplay of transcription factors, microRNAs, and histone modifications

Cystathionine gamma-lyase (CTH), a key enzyme in hydrogen sulfide (H2S) biosynthesis, has been studied extensively in models of oxidative stress. Dysregulated CTH has been reported in cardiovascular disease, ischemia-reperfusion (I/R) injury, aging, and xenobiotic stress, where reactive oxygen species are central to pathology. Despite this, how CTH is regulated at the molecular level under basal, ischemic, and reperfusion conditions is largely unknown. We observed that CTH expression is dysregulated during ischemia and reperfusion in Huh7 hepatic cells, mouse liver, and human liver, and therefore systematically dissected its transcriptional, post-transcriptional, and epigenetic control. Promoter-deletion reporter assays identified the -275/+91 bp region as the core human CTH promoter. This region was predicted to harbour binding sites for Sp1, CREB, and NFY, which were confirmed through overexpression, knockdown or inhibition, and ChIP assays in Huh7 cells. Computational and experimental screening identified miR-150-5p and miR-510-5p as novel post-transcriptional regulators that bind directly to the CTH 3'-UTR. We also examined histone marks (H3K9ac, H3K9me3, H3K4me3, and H3K27me3) at the CTH promoter and found that H3K9ac, H3K4me3, and H3K27me3 correlate with CTH expression across basal, ischemic, and reperfusion conditions. Mechanistically, ischemia reduces Sp1 occupancy at the CTH promoter, raises miR-150-5p and miR-510-5p levels, lowers H3K9ac, collectively suppressing CTH. Reperfusion reverses this with greater Sp1 and CREB binding, lower miR-150-5p and miR-510-5p levels, and reduced H3K27me3, restoring CTH expression. Together, these findings reveal novel roles for Sp1, CREB, NFY, miR-150-5p, miR-510-5p, and histone modifications in the regulation of CTH during basal, ischemia and reperfusion conditions.

molecular biology↗

bioRxiv2019

Hypoxia mediated regulation of mitochondrial transcription factors: Implications of hypertensive renal physiology

Kidneys have a high resting metabolic rate and low tissue partial pressure of oxygen due to enhanced mitochondrial oxygen consumption and ATP production for active solute transport. Enhanced mitochondrial activity leads to progressive hypoxia from the renal cortex to renal medulla. Renal tubulointerstitial hypoxia (TiH) is severe in hypertensive rats due to increased sodium reabsorption within their nephrons. Additionally, these rats display increased energy demand and therefore, require healthy mitochondria for adequate salt reabsorption. Hence, we sought to study the regulation of mitochondrial biogenesis and expression of mitochondrial transcription factors (mtTFs, viz. Tfam, Tfb1m and Tfb2m) during hypoxic conditions and in rodent models of genetic hypertension. We report that the expressions of HIF-1 (hypoxia inducible factor-1), PGC-1 (peroxisome proliferator activated receptor-{gamma} co-activator-1), mtTFs and OXPHOS proteins are elevated in hypertensive rats as compared to their normotensive counterparts. Additionally, studies in cultured kidney cells show that acute hypoxia augments the expression of these genes. We also observe a positive correlation between HIF-1 and mtTFs transcripts in human tissues. Furthermore, we report for the first time to our knowledge, that HIF-1 binds to promoters of Tfam, Tfb1m and Tfb2m genes and augments their promoter activities in NRK52e cells subjected to acute hypoxia. Taken together, this study suggests that acute hypoxia may enhance mitochondrial function to meet the energy demand in renal tubular epithelial cells and in young/pre-hypertensive SHR kidneys. Translational StatementOur results suggest that tubulointerstitial hypoxia (TiH) prevailing in prehypertensive rats augments the expression of mitochondrial transcription factors and proteins of electron transport chain. Moreover, previous reports indicate that ATP synthesis in these rats are elevated. Thus, our study provides insights into the molecular mechanism of such enhanced mitochondrial function. We propose that during early stages of kidney diseases (marked by mild TiH) an enhancement of mitochondrial function via stimulation of HIF-1/PGC-1 production may delay renal tubular damage.

molecular biology↗

bioRxiv2019

Functional promoter polymorphisms govern the differential expression of HMG-CoA Reductase gene in rat models of essential hypertension

3-Hydroxy-3-methyl glutaryl-coenzyme A reductase (Hmgcr) encoding the rate-limiting enzyme in the cholesterol biosynthesis pathway is a candidate gene for essential hypertension (EH). However, the regulation of Hmgcr in rat models of hypertension (viz. Spontaneously Hypertensive Rats [SHR] and its normotensive control Wistar/Kyoto [WKY] strain) is unknown. Here, we show that Hmgcr transcript and protein levels are diminished in liver tissues of SHR as compared to WKY. Consistently, a number of other rat models of hypertension display diminished cholesterol levels as compared to corresponding control strains. Sequencing of the Hmgcr promoter in SHR/WKY reveals three variations: A-405G, C-62T and a 11 bp insertion (-393_-382insTGCGGTCCTCC) in SHR. Moreover, SHR-Hmgcr promoter displays higher activity than WKY-Hmgcr promoter in various cell lines. Transient transfections of Hmgcr-promoter mutants and in silico analysis suggest altered binding of Runx3 and Srebf1 across A-405G and -393_-382insTGCGGTCCTCC sites. Indeed, chromatin immunoprecipitation assays confirm differential binding of Runx3/Srebf1 to Hmgcr promoter leading to diminished expression of Hmgcr in SHR as compared to WKY under basal/cholesterol-modulated conditions. Taken together, this study provides mechanistic insights for the altered Hmgcr expression in these models of EH, thereby unravelling the links of this gene to hypertension and associated cardiovascular disease states.

molecular biology↗

bioRxiv2019

A common tag nucleotide variant in MMP7 promoter increases risk for hypertension via enhanced interactions with CREB transcription factor

MMP7 (Matrilysin), a potent extracellular matrix degrading enzyme with wide substrate specificity, is emerging as a new regulator of cardiovascular diseases including coronary artery disease and atherosclerosis. However, potential contributions of MMP7 genetic variations to hypertension remain unknown. In this study, we first probed for the association of a tag single nucleotide polymorphism (SNP) in the MMP7 gene promoter (-181A/G; rs11568818) with hypertension in an urban south Indian population (n=1517). The heterozygous A/G genotype showed a strong association with hypertension as compared to the A/A wild-type genotype (OR=1.641, 95% CI=1.276-2.109; p=1x10-4); AG genotype carriers also displayed significantly higher diastolic blood pressure and mean arterial pressure than AA genotype subjects. The study was replicated in a north Indian population (n=977) as well (OR=1.520, 95% CI =1.106-2.090; p=0.01). Transient transfection experiments using MMP7 promoter-luciferase reporter constructs revealed that the variant -181G allele conferred greater promoter activity than the -181A allele. Computational prediction and structure-based conformational and molecular dynamics simulation studies suggested higher binding affinity for the transcription factor CREB to the -181G promoter. In corroboration, over-expression/down-regulation of CREB and chromatin immunoprecipitation experiments provided convincing evidence for stronger binding of CREB with the -181G promoter. Further, the -181G promoter also displayed an enhanced response to hypoxia and epinephrine-treatment. The higher promoter activity of -181G allele also translated to increased MMP7 protein levels. Indeed, MMP7-181A/G heterozygous individuals displayed elevated plasma MMP7 levels which positively correlated with blood pressure. In conclusion, the MMP7 A-181G promoter SNP increased expression of MMP7 under pathophysiological (such as hypoxic stress and catecholamine excess) conditions via increased interactions with the transcription factor CREB and enhanced the risk for hypertension in its carriers.

genetics↗