👤 Saket Kumar

Copy number variants (CNVs) are among the main genetic factors identified in schizophrenia (SZ) through genome-scale studies conducted mostly in Caucasian populations. However, to date, there have been no genome-scale CNV reports on patients from India. To address this shortcoming, we generated, for the first time, genome-scale CNV data for 168 SZ patients and 168 controls from South India. In total, 63 different CNVs were identified in 56 patients and 46 controls with a significantly higher proportion of medium-sized deletions (100 kb-1 Mb) after multiple testing (FDR = 2.7E-4) in patients. Of these, 13 CNVs were previously reported; however, when searched against GWAS, transcriptome, exome, and DNA methylation studies, another 17 CNVs with candidate genes were identified. Of the total 30 CNVs, 28 were present in 38 patients and 12 in 27 controls, indicating a significantly higher representation in the former ( Show less

Although VEGF-B was discovered as a VEGF-A homolog a long time ago, the angiogenic effect of VEGF-B remains poorly understood with limited and diverse findings from different groups. Notwithstanding, drugs that inhibit VEGF-B together with other VEGF family members are being used to treat patients with various neovascular diseases. It is therefore critical to have a better understanding of the angiogenic effect of VEGF-B and the underlying mechanisms. Using comprehensive in vitro and in vivo methods and models, we reveal here for the first time an unexpected and surprising function of VEGF-B as an endogenous inhibitor of angiogenesis by inhibiting the FGF2/FGFR1 pathway when the latter is abundantly expressed. Mechanistically, we unveil that VEGF-B binds to FGFR1, induces FGFR1/VEGFR1 complex formation, and suppresses FGF2-induced Erk activation, and inhibits FGF2-driven angiogenesis and tumor growth. Our work uncovers a previously unrecognized novel function of VEGF-B in tethering the FGF2/FGFR1 pathway. Given the anti-angiogenic nature of VEGF-B under conditions of high FGF2/FGFR1 levels, caution is warranted when modulating VEGF-B activity to treat neovascular diseases. Show less

Milk fat composition is an important trait for the dairy industry as it directly influences the nutritional and technological properties of milk and other dairy products. The synthesis of milk fat is a complex process regulated by a network of genes. Thus, understanding the genetic variation and molecular mechanisms regulating milk fat synthesis will help to improve the nutritional quality of dairy products. In this review, we provide an overview of milk fat synthesis in bovines along with the candidate genes involved in the pathway. We also discuss de novo synthesis of fatty acids (ACSS, ACACA, FASN), uptake of FAs (FATP, FAT, LPL), intracellular activation and channelling of FAs (ACSL, FABP), elongation (EVOLV6), desaturation (SCD, FADS), formation of triglycerides (GPAM, AGPAT, LIPIN, DGAT), and milk lipid secretion (BTN1A1, XDH, PLIN2). The genetic variability of individual fatty acids will help to develop selection strategies for obtaining a healthier milk fat profile in bovines. Thus, this review will offer a potential understanding of the molecular mechanisms that regulate milk fat synthesis in bovines. Show less

A linear 16-pole ion trap-based experimental setup has been designed, implemented, and characterized to investigate the photophysics of biomolecules in the gas phase. Electrospray ionization is employed to generate the ions in the gas phase at atmospheric pressure. The voltage configuration on the ion funnel, the ion optic device in the first vacuum interface, is used to control the energy of the ions. A home-built quadrupole mass-filter is utilized for the mass-selection of the ions of interest. A 16-pole ion trap designed and built in-house is implemented for ion trapping. The instrument's versatility and capability are showcased by demonstrating the fragmentation patterns of protonated and deprotonated tryptophan, as well as describing the photodetachment decay of deprotonated indole. Show less

The genome is pervasively transcribed to produce a vast array of non-coding RNAs (ncRNAs). Long non-coding RNAs (lncRNAs) are transcripts of >200 nucleotides and are best known for their ability to regulate gene expression. Enhancer RNAs (eRNAs) are subclass of lncRNAs that are synthesized from enhancer regions and have also been shown to coordinate gene expression. The biological function and significance of most lncRNAs and eRNAs remain to be determined. Epithelial to mesenchymal transition (EMT) is a ubiquitous cellular process that occurs during cellular migration, homeostasis, fibrosis, and cancer-cell metastasis. EMT-transcription factors, such as SNAI1 induce a complex transcriptional program that coordinates the morphological and molecular changes associated with EMT. Such complex transcriptional programs are often subject to coordination by networks of ncRNAs and thus can be leveraged to identify novel functional ncRNA loci. Here, using a genome-wide CRISPR activation (CRISPRa) screen targeting ∼10,000 lncRNA loci we identified ncRNA loci that could either promote or attenuate EMT. We discovered a novel locus that we named Show less

Systemic sclerosis is a fibrotic disease that initiates in the skin and progresses to internal organs, leading to a poor prognosis. Unraveling the etiology of a chronic, multifactorial disease such as systemic sclerosis has been aided by various animal models that recapitulate certain aspects of the human pathology. We found that the transcription factor SNAI1 is overexpressed in the epidermis of patients with systemic sclerosis, and a transgenic mouse recapitulating this expression pattern is sufficient to induce many clinical features of the human disease. Using this mouse model as a discovery platform, we have uncovered a critical role for the matricellular protein Mindin (SPON2) in fibrogenesis. Mindin is produced by SNAI1 transgenic skin keratinocytes and aids fibrogenesis by inducing early inflammatory cytokine production and collagen secretion in resident dermal fibroblasts. Given the dispensability of Mindin in normal tissue physiology, targeting this protein holds promise as an effective therapy for fibrosis. Show less

Genetics play a significant role in coagulation phenotype and venous thromboembolism risk. Resistance to the anticoagulant activated protein C (APC) is an established risk for thrombosis. Herein, we explored the genetic determinants of thrombin generation (TG) and thrombomodulin (TM)-modulated TG using plasma from the Human Functional Genomics Project. Calibrated TG was measured both in absence and presence of TM using tissue factor as trigger. Genetic determinants of TG parameters and protein C pathway function were assessed using genome-wide single-nucleotide polymorphism (SNP) genotyping. Plasma samples were supplemented with purified apolipoprotein A-IV, prekallikrein, or kallikrein to test their influence on the anticoagulant function of TM and APC in TG. Thrombin generation data from 392 individuals were analyzed. Genotyping showed that the KLKB1 gene (top SNP: rs4241819) on chromosome 4 was associated with the normalized sensitivity ratio of endogenous thrombin potential to TM at genome-wide level (nETP-TMsr, P = 4.27 × 10 Our results suggest that kallikrein plays a role in the regulation of the anticoagulant protein C pathway in TG, which may provide a novel mechanism for the previously observed association between the KLKB1 gene and venous thrombosis. Show less

Alzheimer's disease (AD) is an irreversible, progressive neurological disorder characterized by amyloid plaques, hyperphosphorylated tau protein (hyper p-tau), neuronal damage, memory loss, etc. Various factors, such as age, lifestyle, family history, environmental factors, and gene mutation, cause AD. BACE-1 is an interesting target to prevent or reverse AD progression. BACE-1 cleaves amyloid precursor protein (APP) into soluble amyloid precursor protein β (sAPPβ) and membrane-bound C-terminal fragment called C99, a rate-limiting step, and C99 is further cleaved by gamma-secretase to generate neurotoxic amyloid β (Aβ). Discovery and development of selective β amyloid precursor protein cleavage enzyme 1 (BACE-1) inhibitors have a great potential for the treatment and maintenance of Alzheimer's disease. In this review, we have compiled literature pertaining to guanidine-based novel BACE-1 inhibitors for the treatment and maintenance of AD. We have also discussed role of BACE-1 substrates, and its crystal structure, BACE-1 inhibitors in the clinical trial, and essential points to overcome challenges associated with selective development of BACE-1 inhibitors. This paper provides valuable information for the design and discovery of selective new BACE-1 inhibitors against other aspartyl protease enzymes to treat AD. Show less

Earlier investigations on biological methods of wastewater treatment have revealed that algal based wastewater treatment could be a green, cost effective and efficient approach for the removal of heavy metals. So, this study aimed to assess the potential of microalga Chlorella pyrenoidosa for remediation of heavy metals (Cr, Cu, Pb, Zn, Cd, Mn, and Ni) from varying concentration (25%, 50%, 75 and 100%) of wastewater collected from Common Effluent Treatment Plant. Heavy metals such as Cr, Cu, Pb, Zn, Cd, Mn, and Ni have been removed significantly from the wastewater, with percentage removal ranging from 73%, 60%, 75%, 66%, 87%, 83%, and 74% with 50% test solution, 57%, 59%, 70%, 56%, 72%, 66%, and 62% with 75% test solution, and 47%, 55%, 56%, 71%, 61%, 77%, and 72% with 100% test solution respectively. Studies on biochemical assay (protein, carbohydrate, and pigment) of Chlorella pyrenoidosa were also an important part of the present investigation to understand the interaction of heavy metals with algal biochemical compounds using Pearson correlation co-efficient. Biomass grown in CETP wastewater can be used for synthesis of various fruitful value-added end products like bio-diesel, pharmaceutical products, cosmetic products, bio-adsorbent etc. Show less

The metabolically promiscuous pentachlorophenol (PCP) hydroxylating Phe4MO (represented as CpsB) was detected, amplified (from the genome of Bacillus tropicus strain AOA-CPS1), cloned, overexpressed, purified and characterized here. The 1.755-kb gene cloned in the pET15b vector expressed a ≅ 64 kDa monomeric protein which was purified to homogeneity by single-step affinity chromatography, with a total yield of 82.1%. The optimum temperature and pH of the enzyme were found to be 30 °C and 7.0, respectively. CpsB showed functional stability between pH 6.0-7.5 and temperature 25-30 °C. The enzyme-substrate reaction kinetic studies showed the allosteric nature of the enzyme and followed pre-steady state using NADH as a co-substrate with apparent v Show less

High-grade serous epithelial ovarian carcinoma (HGSOC) is an immunogenic tumor with a unique tumor microenvironment (TME) that extends to the peritoneal cavity. The immunosuppressive nature of TME imposes the major challenge to develop effective treatment options for HGSOC. Interaction of immune cells in TME is an important factor. Hence, a better understanding of immune profile of TME may be required for exploring alternative treatment options. Immune profiling of peritoneal fluid (PF), tumor specimens, and blood were carried out using flowcytometry, ELISA, and Procartaplex immunoassay. The frequency of CD56 Show less

Rheumatoid arthritis (RA) is a chronic autoimmune disease which is characterized primarily by synovial hyperplasia and accumulation of several types of immune infiltrates that promote progressive destruction of the articular structure. Glucocorticoids are often prescribed to treat RA because of their strong anti-inflammatory and immunosuppressive effects. However, their application must be limited to the short-term due to a risk of adverse events. In the present study, we examined the potential combination of low-dose prednisone with gene delivery of an agent of promising and complementary effectiveness in RA, interleukin (IL)-27. IL-27 has been shown to have anti-inflammatory potential, while also acting as an effective bone-normalization agent in prior reports. The present report examined a version of IL-27 targeted at the C-terminus with a short 'peptide L' (pepL, LSLITRL) that binds the interleukin 6 receptor α (IL-6Rα) upregulated during inflammation. By focusing on this targeted form, IL-27pepL or 27pL, we examined whether the anti-inflammatory potential of prednisone (at a relatively low dose and short duration) could be further enhanced in the presence of 27pL as a therapy adjuvant. Our results indicate that 27pL represents a novel tool for use as an adjuvant with current therapeutics, such as prednisone, against inflammatory conditions. Show less

In People with HIV (PWH), chronic immune activation and systemic inflammation are associated with increased risk to develop comorbidities including bone loss. Numerous cells of the immune system, namely, T cells are involved in the regulation of the bone homeostasis and osteoclasts (OCs) activity. IL-27, a cytokine that belongs to the IL-12 family can regulate the secretion of pro- and anti-inflammatory cytokines by T cells, however its role in the setting of HIV is largely unknown. In the present study, we determined the impact of OCs in T cell secretion of cytokines and whether IL-27 can regulate this function. We found that the presence of OCs in the T cell cultures significantly enhanced secretion of IFNγ, TNFα, IL-17, RANKL, and IL-10 in both PWH and healthy controls. In PWH, IL-27 inhibited IL-17 secretion and downregulated surface expression of RANKL in CD4 T cells. All together these results suggest that in the context of HIV infection IL-27 may favor IFNγ and TNFα secretion at the sites of bone remodeling. Show less

An emerging approach in treating skeletal malignancies utilizes osteoimmunology to investigate new multifunctional immune-stimulatory agents that can simultaneously combat tumor growth and promote bone repair. We have hypothesized that cytokine Interleukin-27 (IL-27) is an excellent candidate biologic to help rebalance the prostate tumor cells and bone cell environment. In this work, we examined the proof of principle for a short, secreted luciferase (Nanoluc or Nluc) fusion with IL-27 to produce a novel cytokine-based biologic (Nluc-27), whereby we examined its efficacy in vitro in reducing prostate tumor growth and rebalancing bone cell proliferation and differentiation. This work demonstrates the targeting and anti-tumor efficacy of the Nluc-27 fusion cytokine in cancer and bone cell models. The fusion cytokine is detectable in conditioned media, and bioactive in different cell systems. This novel Nluc-27 cytokine will allow flexible incorporation of other targeting domains and may serve as flexible tool to augment IL-27's bioactivity and reengineer its efficacy against prostate tumor or bone cells, and may prove applicable to several other cell types for targeted gene therapy applications. Show less

Fibrosis is a leading cause of morbidity and mortality worldwide. Although fibrosis may involve different organ systems, transforming growth factor-β (TGFβ) has been established as a master regulator of fibrosis across organs. Pirfenidone and Nintedanib are the only currently-approved drugs to treat fibrosis, specifically idiopathic pulmonary fibrosis, but their mechanisms of action remain poorly understood. To identify novel drug targets and uncover potential mechanisms by which these drugs attenuate fibrosis, we performed an integrative 'omics analysis of transcriptomic and proteomic responses to TGFβ1-stimulated lung fibroblasts. Significant findings were annotated as associated with pirfenidone and nintedanib treatment in silico via Coremine. Integrative 'omics identified a co-expressed transcriptomic and proteomic module significantly correlated with TGFβ1 treatment that was enriched (FDR-p = 0.04) with genes associated with pirfenidone and nintedanib treatment. While a subset of genes in this module have been implicated in fibrogenesis, several novel TGFβ1 signaling targets were identified. Specifically, four genes (BASP1, HSD17B6, CDH11, and TNS1) have been associated with pirfenidone, while five genes (CLINT1, CADM1, MTDH, SYDE1, and MCTS1) have been associated with nintedanib, and MYDGF has been implicated with treatment using both drugs. Using the Clue Drug Repurposing Hub, succinic acid was highlighted as a metabolite regulated by the protein encoded by HSD17B6. This study provides new insights into the anti-fibrotic actions of pirfenidone and nintedanib and identifies novel targets for future mechanistic studies. Show less

HIV-specific T cells have diminished effector function and fail to control/eliminate the virus. IL-27, a member of the IL-6/IL-12 cytokine superfamily has been shown to inhibit HIV replication. However, whether or not IL-27 can enhance HIV-specific T cell function is largely unknown. In the present manuscript, we investigated the role of IL-27 signaling in human T cells by evaluating the global transcriptional changes related to the function of HIV-specific T cells. We found that T cells from people living with HIV (PLWH), expressed higher levels of STAT1 leading to enhanced STAT1 activation upon IL-27 stimulation. Observed IL-27 induced transcriptional changes were associated with IFN/STAT1-dependent pathways in CD4 and CD8 T cells. Importantly, IL-27 dependent modulation of T-bet expression promoted IFNγ secretion by TIGIT Show less

The gene for receptor tyrosine kinase ErbB2 is amplified in breast and ovarian tumours. The linear pathway by which signals are transduced through ErbB2 are well known. However, second generation questions that address spatial aspects of signaling remain. To address this, we have undertaken a mass spectrometry approach to identify phosphoproteins specific for ErbB2 using the inhibitors Lapatinib and CP724714 in ovarian cancer cells. The ErbB2 specific proteins identified in SKOV-3 cells were Myristoylated alanine-rich C-kinase substrate, Protein capicua homolog, Protein peptidyl isomerase G, Protein PRRC2C, Chromobox homolog1 and PRP4 homolog. We have evaluated three phosphoproteins PKM2, Aldose reductase and MARCKS in SKOV-3 cells. We observed that PKM2 was phosphorylated by EGF but was not inhibited by Lapatinib and CP724714. The activity of aldose reductase in reducing NADPH as a substrate was significantly higher in EGF stimulated cells which was inhibited by Lapatinib and CP724714 but not by Geftinib (EGFR inhibitor). MARCKS was phosphorylated on stimulation of SKOV-3 cells with EGF that was inhibited by Lapatinib and CP724714 which was dependent on the kinase activity of ErbB2. These results have identified phosphoproteins that are specific to ErbB2 which have not been previously reported and sets the basis for future experiments. Show less

Triple-negative subtype of breast cancer (TNBC) is hallmarked by frequent disease relapse and shows highest mortality rate. Although PD-1/PD-L1 immune checkpoint blockades have recently shown promising clinical benefits, the overall response rate remains largely insufficient. Hence, alternative therapeutic approaches are warranted. Given the immunosuppressive properties of CD73-mediated adenosine release, CD73 blocking approaches are emerging as attractive strategies in cancer immunotherapy. Understanding the precise mechanism regulating the expression of CD73 is required to develop effective anti-CD73-based therapy. Our previous observations demonstrate that the transcription factors driving epithelial-to-mesenchymal transition (EMT-TF) can regulate the expression of several inhibitory immune checkpoints. Here we analyzed the role of the EMT-TF SNAI1 in the regulation of CD73 in TNBC cells. We found that doxycycline-driven SNAI1 expression in the epithelial -like TNBC cell line MDA-MB-468 results in CD73 upregulation by direct binding to the CD73 proximal promoter. SNAI1-dependent upregulation of CD73 leads to increased production and release of extracellular adenosine by TNBC cells and contributes to the enhancement of TNBC immunosuppressive properties. Our data are validated in TNBC samples by showing a positive correlation between the mRNA expression of CD73 and SNAI1. Overall, our results reveal a new CD73 regulation mechanism in TNBC that participates in TNBC-mediated immunosuppression and paves the way for developing new treatment opportunities for CD73-positive TNBC. Show less

Preservation of a small population of cancer stem cells (CSCs) within a heterogeneous carcinoma serves as a paradigm to understand how select cells in a tissue maintain their undifferentiated status. In both embryogenesis and cancer, Snail has been correlated with stemness, but the molecular underpinning of this phenomenon remains largely ill-defined. In models of cutaneous squamous cell carcinoma (cSCC), we discovered a non-epithelial-mesenchymal transition function for the transcription factor Snail in maintaining the stemness of epidermal keratinocytes. Snail-expressing cells secrete the matricellular protein Mindin, which functions in an autocrine fashion to activate a Src-STAT3 pathway to reinforce their stem/progenitor phenotype. This pathway is activated by the engagement of Mindin with the leukocyte-specific integrin, CD11b (ITGAM), which is also unexpectedly expressed by epidermal keratinocytes. Interestingly, disruption of this signaling module in human cSCC attenuates tumorigenesis, suggesting that targeting Mindin would be a promising therapeutic approach to hinder cancer recurrence. Show less

The epithelial to mesenchymal transition (EMT) is crucial for cancer progression and chemoresistance. EMT is a dynamic process with multiple phases that change cell migration and invasion activity. We used pan-cancer expression data to find 14-LncRNAs that had a high correlation with the EMT markers VIM, CDH1, FN1, SNAI1, and SNAI2. The expression of 14 EMT-associated LncRNA, which also showed high cancer specificity, was used to calculate the pan-cancer EMT score. The EMT score was then applied to the 32 cancer types to classify them as epithelial, epithelial-mesenchymal, mesenchymal-epithelial, or mesenchymal tumors. We discovered that the EMT score is a poor prognostic predictor and that as tumor mesenchymal nature increased, patient survival decreased. We also showed that the cell of origin did not influence the EMT nature of tumors. Pathway analysis employing protein expression data revealed that the PI3K pathway is the most crucial in determining the EMTness of tumors. Further, we divided CCLE-cell lines into EMT classes and discovered that mesenchymal cells, which exhibited higher PI3K pathway activation, were more sensitive to PI3K inhibitors than epithelial cells. We identified Linc01615 as a mesenchymal LncRNA whose expression significantly correlated with survival in several cancer types. We showed that Linc01615 is regulated by the TGFβ-STAT3 pathway in a feedback loop. Knockdown of Linc01615 inhibited cell proliferation and migration by regulating the PI3K pathway and mesenchymal markers. We also identified RP4-568C11.4 as an epithelial cancer marker. We showed that knocking down RP4-568C11.4 decreased cell growth but not migration. In addition, we discovered that ESR1 regulates RP4-5681C11.4 in breast cancer. Taken together, we have developed a pan-cancer EMT signature. Also, we found two new LncRNAs that have different effects on cancer development and EMT. Show less

Parkinson's disease (PD) is a genetically heterogeneous neurodegenerative disease with poorly defined environmental influences. Genomic studies of PD patients have identified disease-relevant monogenic genes, rare variants of significance, and polygenic risk-associated variants. In this study, whole genome sequencing data from 90 young onset Parkinson's disease (YOPD) individuals are analyzed for both monogenic and polygenic risk. The genetic variant analysis identifies pathogenic/likely pathogenic variants in eight of the 90 individuals (8.8%). It includes large homozygous coding exon deletions in PRKN and SNV/InDels in VPS13C, PLA2G6, PINK1, SYNJ1, and GCH1. Eleven rare heterozygous GBA coding variants are also identified in 13 (14.4%) individuals. In 34 (56.6%) individuals, one or more variants of uncertain significance (VUS) in PD/PD-relevant genes are observed. Though YOPD patients with a prioritized pathogenic variant show a low polygenic risk score (PRS), patients with prioritized VUS or no significant rare variants show an increased PRS odds ratio for PD. This study suggests that both significant rare variants and polygenic risk from common variants together may contribute to the genesis of PD. Further validation using a larger cohort of patients will confirm the interplay between monogenic and polygenic variants and their use in routine genetic PD diagnosis and risk assessment. Show less

Parkinson's disease may be caused by a single pathogenic variant (monogenic) in 5-10% of cases, but investigation of these disorders provides valuable pathophysiological insights. In this review, we discuss each genetic form with a focus on genotype, phenotype, pathophysiology, and the geographic and ethnic distribution. Well-established Parkinson's disease genes include autosomal dominant forms ( Show less

Receptors for the peptide hormones glucagon-like peptide-1 (GLP-1R), glucose-dependent insulinotropic polypeptide (GIPR), and glucagon (GCGR) are important regulators of insulin secretion and energy metabolism. GLP-1R agonists have been successfully deployed for the treatment of type 2 diabetes, but it has been suggested that their efficacy is limited by target receptor desensitization and downregulation due to recruitment of β-arrestins. Indeed, recently described GLP-1R agonists with reduced β-arrestin-2 recruitment have delivered promising results in preclinical and clinical studies. We therefore aimed to determine if the same phenomenon could apply to the closely related GIPR and GCGR. In HEK293 cells depleted of both β-arrestin isoforms the duration of G protein-dependent cAMP/PKA signaling was increased in response to the endogenous ligand for each receptor. Moreover, in wildtype cells, "biased" GLP-1, GCG, and GIP analogs with selective reductions in β-arrestin-2 recruitment led to reduced receptor endocytosis and increased insulin secretion over a prolonged stimulation period, although the latter effect was only seen at high agonist concentrations. Biased GCG analogs increased the duration of cAMP signaling, but this did not lead to increased glucose output from hepatocytes. Our study provides a rationale for the development of GLP-1R, GIPR, and GCGR agonists with reduced β-arrestin recruitment, but further work is needed to maximally exploit this strategy for therapeutic purposes. Show less

The association of melanocortin receptor 4 (MC4R) gene with adiposity measures is widely studied in European populations. Only six studies have investigated the role of MC4R gene with adiposity measures among Indian populations. We have evaluated the role of MC4R (rs17782313) gene polymorphism in influencing adiposity measures in India among children and adults. The present population based cross sectional study was conducted among 303 individuals (208 children and 95 adults) of age group 10-30 years, belonging to Rajasthan. Somatometric measurements (standing height, weight, and waist and hip girths) and blood samples were taken after obtaining written informed consent. Genotyping of MC4R rs17782313 single nucleotide polymorphism was done using restriction fragment length polymorphism method for polymerase chain reaction amplified fragments. We examined association between rs17782313 and different adiposity measures (height, weight, BMI, WHR, and waist and hip girths) using linear regression models. The MC4R variant (rs17782313) predicted increased body weight (0.15 kg, S.E ± 0.076, P = 0.043) among children. In combined population, the rs17782313 variant was moderately associated with body weight (0.13 kg, S.E ± 0.070, P = 0.057). This variant was not found to be associated with any other adiposity measure. Further studies are needed to evaluate the association of MC4R variants through sequencing and functional genomics with different adiposity measures in Indian populations for understanding the genetic underpinnings of adiposity in India. Show less

The melanocortin-4 receptor (MC4R), a critical G-protein-coupled receptor (GPCR) regulating energy homeostasis, activates multiple signalling pathways, including mobilisation of intracellular calcium ([Ca2+]i). However, very little is known about the physiological significance of MC4R-induced [Ca2+]i since few studies measure MC4R-induced [Ca2+]i. High-throughput, read-out assays for [Ca2+]i have proven unreliable for overexpressed GPCRs like MC4R, which exhibit low sensitivity mobilising [Ca2+]i. Therefore, we developed, optimised, and validated a robust quantitative high-throughput assay using Fura-2 ratio-metric calcium dye and HEK293 cells stably transfected with MC4R. The quantitation enables direct comparisons between assays and even between different research laboratories. Assay conditions were optimised step-by-step to eliminate interference from stretch-activated receptor increases in [Ca2+]i and to maximise ligand-activated MC4R-induced [Ca2+]i. Calcium imaging was performed using a PheraStar FS multi-well plate reader. Probenecid, included in the buffers to prevent extrusion of Fura-2 dye from cells, was found to interfere with the EGTA-chelation of calcium, required to determine Rmin for quantitation of [Ca2+]i. Therefore, we developed a method to determine Rmin in specific wells without probenecid, which was run in parallel with each assay. The validation of the assay was shown by reproducible α-melanocyte-stimulating hormone (α-MSH) concentration-dependent activation of the stably expressed human MC4R (hMC4R) and mouse MC4R (mMC4R), inducing increases in [Ca2+]i, for three independent experiments. This robust, reproducible, high-throughput assay that quantitatively measures MC4R-induced mobilisation of [Ca2+]i in vitro has potential to advance the development of therapeutic drugs and understanding of MC4R signalling associated with human obesity. Show less

Children exposed to antiretroviral therapy (ART) are at risk of developing metabolic complications. The association between gene polymorphisms and the development of dyslipidemia in children post ART initiation was studied. Children initiating first-line ART were followed for 2 years at the National Institute for Research in Tuberculosis (Chennai, India), and St. John's Medical College Hospital (Bangalore, India). Clinical examination and fasting serum lipid profiles were measured every 6 months. Participants were genotyped for the polymorphisms in the APOC3 gene (rs2854116; rs2854117, and rs5128). Changes in lipid levels from baseline to months 6, 12, and 24, and the difference between the various genotype variants were analyzed using a modified analysis of variance test. Study enrolled 393 ART-naive HIV-infected children (mean age: 7.6 ± 3 years, mean weight: 18 ± 6) of whom 289 (75%) were started on nevirapine (NVP)-based ART and the remaining 96 (25%) were started on efavirenz-based ART. Only children carrying the GG allele of rs5128 genotype showed a decrease in CD4% and serum triglycerides pre-ART. An increasing trend of total cholesterol, high-density lipoprotein cholesterol (HDL-c), and low-density lipoprotein cholesterol were seen at 6 months in both EFZ and NVP groups, which subsequently stabilized by 12 months irrespective of genotype variants. Genotype variants of APOC3 (rs2854116 and rs2854117 polymorphism) did not show significant changes in serum lipid levels after 24 months of ART, whereas rs5128 polymorphism with "G" allele showed an association with HDL-c levels when on NVP-based ART. Our results suggest that ART plays a major role in normalizing lipid levels in HIV-infected children and APOC3 polymorphisms may not play a significant role in ART-induced dyslipidemia. Show less

In this article, monomers (tannic acid (TA) and m- phenylenediamine (MPD)) were used in the fabrication of a novel PES based thin-film composite nanofiltration (TFC-NF) membrane for the treatment of a common effluent treatment plant (CETP) of textile industrial wastewater. PES support sheets and TFC layers were fabricated via non-solvent induced phase inversion and in-situ interfacial polymerization (IP) process. The ultra-thin active layer was synthesized via the IP process with monomers such as tannic acid (TA) and m- phenylenediamine (MPD). T and M series membranes correspond to (PES/x wt% TA, x = 2, 4, 6) as T1, T2, T3 -TA and (PES/x wt% MPD, x = 2, 4, 6) as M1, M2, M3-MPD respectively. M0 corresponds to PES which is the virgin membrane. The chemical structure, surface morphology, surface roughness and surface properties were explored using fourier transform infrared (FTIR) spectroscopy, field emission scanning electron microscopy (FESEM), atomic force microscopy (AFM) and contact angle, respectively. The filtration performance of the thin-film composite nanofiltration (TFC-NF) membranes was investigated by various properties like pure water flux, salt rejection, porosity, mean pore radius and antifouling analysis. T1-TA membrane showed better water permeability, high salt rejection and better industrial effluent rejection with 94.4% of TDS that are suitable for industrial reuse and agricultural irrigation. Moreover, for T1-TA membrane, the water flux, porosity, mean pore radius, salt rejection, surface roughness and contact angle of 43.5lm The online version contains supplementary material available at 10.1007/s40201-021-00624-x. Show less

Identifying the microbial community and their functional potential from different stages of common effluent treatment plants (CETP) can enhance the efficiency of wastewater treatment systems. In this study, wastewater metagenomes from 8 stages of CETP were screened for microbial diversity and gene profiling along with their corresponding degradation activities. The microbial community displayed 98.46% of bacterial species, followed by Eukarya (0.10%) and Archaea 0.02%. At the Phylum level, Proteobacteria (28.8%) was dominant, followed by Bacteroidetes (16.1%), Firmicutes (11.7%), and Fusobacteria (6.9%) which are mainly capable of degrading the aromatic compounds. Klebsiella pneumoniae, Wolinella succinogenes, Pseudomonas stutzeri, Desulfovibrio vulgaris, and Clostridium sticklandii were the most prevalent species. The functional analysis further demonstrated the presence of enzymes linked with genes/pathways known to be involved in the degradation/metabolization of aromatic compounds like benzoate, bisphenol, 1,2-dichloroethane phenylalanine. This information was further validated with the whole genome analysis of the bacteria isolated from the CETP. We anticipate that integrating both shotgun and whole-genome analyses can reveal the rich reservoir for novel enzymes and genes present in CETP effluent that can contribute to designing efficient bioremediation strategies for the environment in general CETP system, in particular. Show less