THE ROLE OF NUTRIENT SENSITIVE PROTEIN O-GLCNACYLATION IN DEVELOPMENTAL CORTICAL NEUROGENESIS

The nutrient responsive O-GlcNAcylation is a dynamic, posttranslational protein modification present on many nucleocytoplasmic and mitochondrial proteins. Previous research has indicated that hyperglycaemia increases the levels of total O-GlcNAcylation within cells. Transcription factors and histones are among hundreds of proteins that have been reported to be O-GlcNAcylated and have importance in cell fate determination during cell growth, proliferation, and differentiation. However, the role of protein O-GlcNAcylation in epigenome control in response to nutritional perturbations is poorly understood. Hyperglycaemia induced protein O-GlcNAcylation have been linked to several pathologies, including obesity, diabetes, cancer, cardiovascular and neurodegenerative diseases. Given that maternal hyperglycaemia during pregnancy is linked to adverse neurodevelopmental outcomes in the offspring, it is interesting to identify the impact of elevated protein O-GlcNAcylation on embryonic neurogenesis. Herein, we investigate and confirm the implications of protein O-GlcNAcylation by pharmacological induction of O-GlcNAc during embryonic neurogenesis by directed differentiation of human embryonic stem cells (hESCs) into cortical neurons, which precisely mimics early events of human embryonic corticogenesis. The presence of total O-GlcNAc levels during neural differentiation was determined by immunocytochemistry and western blotting techniques. Investigation of several regulatory transcription factor genes and histones needed for stem cell fate during neurogenesis was carried out through different molecular biology techniques including, western blotting, qPCR, and immunocytochemistry. The impact of elevated O-GlcNAc on transcriptional/epigenetic mechanisms was investigated through high-throughput RNA sequencing and ChIP-qPCR. Resulted that increased O-GlcNAcylation is associated with decreased neural progenitor proliferation, premature cortical neurogenesis, reduced AKT signaling, induced apoptosis and defective expression of several genes essential for neural differentiation. This also led to increased expression of key neurogenic transcription factor (TF) genes. Further investigation has shown that de-repression of neurogenic TFs is associated with increased H3K4me3 and decreased H3K27me3 (promoter bi-valency) levels at the promoter of these genes. Increased O-GlcNAc levels also increased Pol IISer5 phosphorylation whereas, levels of H2BS112O-GlcNAc and H2BK120Ub1 were inconsistently affected at different gene promoters. Also studied the effect of elevated O-GlcNAc levels on embryonic neurogenesis in a rat model of maternal hyperglycaemia. We observed similar epigenetic defects including changes in promoter bivalency and induced Pol IISer5p, H2BS112O-GlcNAc and H2BK120Ub1 in the developing embryo brain cortex due to hyperglycaemia. Resulted findings show O-GlcNAc regulated chromatin sensitivity based on maternal nutritional status on neurodevelopment and suggest that metabolic dysregulations can affect stem cell fate decisions via O-GlcNAc mediated epigenetic gene regulating mechanisms during development. These results may have implications in neurodevelopmental disorders associated with metabolically compromised pregnancies

Synergistic association of STX1A and VAMP2 with cryptogenic epilepsy in North Indian population

Introduction “Common epilepsies”, merely explored for genetics are the most frequent, nonfamilial, sporadic cases in hospitals. Because of their much debated molecular pathology, there is a need to focus on other neuronal pathways including the existing ion channels. Methods For this study, a total of 214 epilepsy cases of North Indian ethnicity comprising 59.81% generalized, 40.19% focal seizures, and based on epilepsy types, 17.29% idiopathic, 37.38% cryptogenic, and 45.33% symptomatic were enrolled. Additionally, 170 unrelated healthy individuals were also enrolled. Here, we hypothesize the involvement of epilepsy pathophysiology genes, that is, synaptic vesicle cycle, SVC genes (presynapse), ion channels and their functionally related genes (postsynapse). An interactive analysis was initially performed in SVC genes using multifactor dimensionality reduction (MDR). Further, in order to understand the influence of ion channels and their functionally related genes, their interaction analysis with SVC genes was also performed. Results A significant interactive two-locus model of STX1A_rs4363087|VAMP2_rs2278637 (presynaptic genes) was observed among SVC variants in all epilepsy cases (P1000-value = 0.054; CVC = 9/10; OR = 2.86, 95%CI = 1.88–4.35). Further, subgroup analysis revealed stronger interaction for the same model in cryptogenic epilepsy patients only (P1000-value = 0.012; CVC = 10/10; OR = 4.59, 95%CI = 2.57–8.22). However, interactive analysis of presynaptic and postsynaptic genes did not show any significant association. Conclusions Significant synergistic interaction of SVC genes revealed the possible functional relatedness of presynapse with pathophysiology of cryptogenic epilepsy. Further, to establish the clinical utility of the results, replication in a large and similar phenotypic group of patients is warranted

Suitability assessment for pluvial flood mitigation in dense urban areas, using nature-based solutions: a case of Delhi

Over the past years, pluvial floods have become a major concern for Indian cities. The factors that exacerbate the flood conditions include climate change, urbanisation, outdated stormwater infrastructure, decreasing capacity of natural wetlands, and natural stormwater sinks. As India's metropolitan cities are at risk due to their high rate of urbanisation, the study aims to develop a methodology to identify areas at risk due to pluvial floods, at a watershed scale. Also, it aims to support flood mitigation through nature-based solutions (NBS) and geospatial analysis, by considering the case of Delhi. Based on local flood data, the settlements at risk of pluvial floods were mapped using GIS, and the "pluvial flood susceptibility map" has been developed to assess the risk of flooding, followed by the selection of the most vulnerable local action area with the help of parameters like susceptibility (based on past pluvial flooding incidences) and urbanisation. Further, the "NBS suitability map" for the milli watershed (area of about 1000 to 10,000 ha) has been generated using the geospatial overlay technique of GIS. It can help identify potential areas within the study region at a sub-watershed level where NBS can be integrated, considering suitable and unsuitable conditions, based on physical, environmental, and planning parameters

Effects of Aminoglycoside Antibiotics on Human Embryonic Stem Cell Viability during Differentiation In Vitro

Human embryonic stem cells (hESCs) are being used extensively in array of studies to understand different mechanisms such as early human embryogenesis, drug toxicity testing, disease modeling, and cell replacement therapy. The protocols for the directed differentiation of hESCs towards specific cell types often require long-term cell cultures. To avoid bacterial contamination, these protocols include addition of antibiotics such as pen-strep and gentamicin. Although aminoglycosides, streptomycin, and gentamicin have been shown to cause cytotoxicity in various animal models, the effect of these antibiotics on hESCs is not clear. In this study, we found that antibiotics, pen-strep, and gentamicin did not affect hESC cell viability or expression of pluripotency markers. However, during directed differentiation towards neural and hepatic fate, significant cell death was noted through the activation of caspase cascade. Also, the expression of neural progenitor markers Pax6, Emx2, Otx2, and Pou3f2 was significantly reduced suggesting that gentamicin may adversely affect early embryonic neurogenesis whereas no effect was seen on the expression of endoderm or hepatic markers during differentiation. Our results suggest that the use of antibiotics in cell culture media for the maintenance and differentiation of hESCs needs thorough investigation before use to avoid erroneous results

Role of Umbilical Artery Doppler in Prediction of Adverse Perinatal outcomes in Hypertensive Disorders of Pregnancy

<p><strong>Background:</strong> Using Umbilical Artery Doppler indices to detect fetoplacental compromise early and to understand the predictive significance of each index in predicting perinatal outcome and therapeutic methods for these patients. <strong>Methods:</strong> The present prospective study was conducted at Department of Obstetrics and Gynaecology, Bokaro General Hospital from 1<sup>st</sup> September 2018 to 31<sup>st</sup> March 2020 (18 months) on 100 women with hypertensive  disorder of pregnancy. Umbilical artery doppler evaluation done in all the patients at (28-32) weeks, (33-36) weeks and (37-40) weeks of gestation and more frequently in those patients having deranged  Doppler. Patients divided into two groups women with abnormal Umbilical artery indices and normal indices. Perinatal outcome of both the groups were compared, analyzed statistically using Chi-square test. Multiple   pregnancy, chronic hypertension, fetal congenital anomalies, systemic disease and those lost to follow up till delivery were excluded from study. <strong>Results: </strong>A total of 100 women with pregnancy induced hypertension were taken into this study after considering inclusion and exclusion criteria. After last Doppler study, patients were divided into two groups, study group and the control group. Out of 100 the study group contain 38 women with abnormal umbilical artery Doppler indices and the control group contain 62 women with abnormal umbilical artery Doppler indices. Hypertension in pregnancy affects women of all ages. In our study the mean maternal age was 27.76 years. <strong>Conclusion:</strong> The most accurate way to anticipate unfavorable prenatal outcomes and assist in choosing the right moment for intervention to enhance perinatal outcomes is with umbilical artery PI.</p> <p> </p> <p> </p><p><strong>Background:</strong> Using Umbilical Artery Doppler indices to detect fetoplacental compromise early and to understand the predictive significance of each index in predicting perinatal outcome and therapeutic methods for these patients. <strong>Methods:</strong> The present prospective study was conducted at Department of Obstetrics and Gynaecology, Bokaro General Hospital from 1<sup>st</sup> September 2018 to 31<sup>st</sup> March 2020 (18 months) on 100 women with hypertensive  disorder of pregnancy. Umbilical artery doppler evaluation done in all the patients at (28-32) weeks, (33-36) weeks and (37-40) weeks of gestation and more frequently in those patients having deranged  Doppler. Patients divided into two groups women with abnormal Umbilical artery indices and normal indices. Perinatal outcome of both the groups were compared, analyzed statistically using Chi-square test. Multiple   pregnancy, chronic hypertension, fetal congenital anomalies, systemic disease and those lost to follow up till delivery were excluded from study. <strong>Results: </strong>A total of 100 women with pregnancy induced hypertension were taken into this study after considering inclusion and exclusion criteria. After last Doppler study, patients were divided into two groups, study group and the control group. Out of 100 the study group contain 38 women with abnormal umbilical artery Doppler indices and the control group contain 62 women with abnormal umbilical artery Doppler indices. Hypertension in pregnancy affects women of all ages. In our study the mean maternal age was 27.76 years. <strong>Conclusion:</strong> The most accurate way to anticipate unfavorable prenatal outcomes and assist in choosing the right moment for intervention to enhance perinatal outcomes is with umbilical artery PI.</p> <p> </p> <p> </p&gt

Saturated fatty acid regulated lncRNA dataset during in vitro human embryonic neurogenesis

Human embryonic stem cells (hESCs) were used as a model of embryonic neurogenesis to identify the effect of excess fat uptake on neurodevelopment (Ardah et al., 2018). Herein, by directed differentiation of hESCs into neurons using established protocols, this data was generated for expression profiles of select lncRNAs during in vitro embryonic neurogenesis and their differential expression due to excess fat (palmitate) uptake. The undifferentiated hESCs were treated with 250 µM palmitate after identifying it as the highest concentration which is non-toxic to these cells. The palmitate treated hESCs were differentiated towards neurons keeping the levels of palmitate consistent throughout the differentiation process and fat uptake was confirmed by Oil Red O staining. The expression analysis of lncRNAs was performed by RT-qPCR on vehicle control and palmitate treated cells from 4 stages of differentiation, D0 (undifferentiated hESCs), D12 (neural stem cells), D44 (neural progenitors) and D70 (neurons) using lncRNAs array plates from Arraystar Inc. which contains 372 functionally identified lncRNAs found to be associated with lipid metabolism and other pathways (Cat# AS-NR-004)

Higher O-GlcNAc Levels Are Associated with Defects in Progenitor Proliferation and Premature Neuronal Differentiation during in-Vitro Human Embryonic Cortical Neurogenesis

The nutrient responsive O-GlcNAcylation is a dynamic post-translational protein modification found on several nucleocytoplasmic proteins. Previous studies have suggested that hyperglycemia induces the levels of total O-GlcNAcylation inside the cells. Hyperglycemia mediated increase in protein O-GlcNAcylation has been shown to be responsible for various pathologies including insulin resistance and Alzheimer's disease. Since maternal hyperglycemia during pregnancy is associated with adverse neurodevelopmental outcomes in the offspring, it is intriguing to identify the effect of increased protein O-GlcNAcylation on embryonic neurogenesis. Herein using human embryonic stem cells (hESCs) as model, we show that increased levels of total O-GlcNAc is associated with decreased neural progenitor proliferation and premature differentiation of cortical neurons, reduced AKT phosphorylation, increased apoptosis and defects in the expression of various regulators of embryonic corticogenesis. As defects in proliferation and differentiation during neurodevelopment are common features of various neurodevelopmental disorders, increased O-GlcNAcylation could be one mechanism responsible for defective neurodevelopmental outcomes in metabolically compromised pregnancies such as diabetes