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STUDY QUESTION What is the functional significance of Tex13b in male germ cell development and differentiation? SUMMARY ANSWER Tex13b regulates male germ cell differentiation by metabolic reprogramming during spermatogenesis. WHAT IS KNOWN ALREADY Studies in mice and humans suggest that TEX13B is a transcription factor and is exclusively expressed in germ cells. STUDY DESIGN, SIZE, DURATION We sequenced the coding regions of TEX13B in 628 infertile men and 427 ethnically matched fertile control men. Further, to identify the molecular function of Tex13b, we created a Tex13b knockout and conditional overexpression system in GC-1spg (hereafter, GC-1) cells. PARTICIPANTS/MATERIALS, SETTING, METHODS Our recent exome sequencing study identified novel candidate genes for male infertility. TEX13B was found to be one of the potential candidates, hence we explored the role of TEX13B in male infertility within a large infertile case–control cohort. We performed functional analyses of Tex13b in a GC-1 cell line using CRISPR-Cas9. We differentially labelled the cell proteins by stable isotope labelling of amino acids in cell culture (SILAC) and performed mass spectrometry-based whole-cell proteomics to identify the differential protein regulation in knockout cells compared to wild-type cells. We found that Tex13b knockout leads to downregulation of the OXPHOS complexes and upregulation of glycolysis genes, which was further validated by western blotting. These results were further confirmed by respirometry analysis in Tex13b knockout cells. Further, we also performed a conditional overexpression of TEX13B in GC-1 cells and studied the expression of OXPHOS complex proteins by western blotting. MAIN RESULTS AND THE ROLE OF CHANCE We identified a rare variant, rs775429506 (p.Gly237Glu), exclusively in two non-obstructive-azoospermia (NOA) men, that may genetically predispose these men for infertility. Further, we demonstrated that Tex13b functions in the transcription regulation of OXPHOS complexes. LARGE SCALE DATA N/A. LIMITATIONS, REASONS FOR CAUTION We examined the function of Tex13b in GC-1 in vitro by knocking out and conditional overexpression, for understanding the function of Tex13b in germ cells. Unfortunately, this could not be replicated in either an animal model or in patient-derived tissue due to the non-availability of an animal model or patient's testis biopsies. WIDER IMPLICATIONS OF THE FINDINGS This study identified that Tex13b plays an important role in male germ cell development and differentiation. The findings of this study would be useful for screening infertile males with spermatogenic failure and counselling them before the implementation of assisted reproduction technique(s). STUDY FUNDING/COMPETING INTEREST(S) Funding was provided by the Council of Scientific and Industrial Research (CSIR) under the network project (BSC0101 and MLP0113) and SERB, the Department of Science and Technology, Government of India (J C Bose Fellowship: JCB/2019/000027). The authors do not have any competing interest. [ABSTRACT FROM AUTHOR]

Introduction: Myopia is most common refractive error leading to visual dysfunctions worldwide. Oxygen deficit due to environmental stress leads to increase in anaerobic glycolysis which results in lactate production. Increase in Reactive Oxygen Species (ROS) and inability of defense system to eliminate them is also responsible for alteration of regulatory pathways. The patient with elevated lactate levels are at significant risk of morbidity and hence, require a prompt treatment. Aim: To estimate and study the lactate levels in tear in various degree of myopia. Materials and Methods: This case-control study was conducted in tertiary care center, Pt. B.D. Sharma PGIMS, Rohtak, Haryana, India. Twenty five patients each with low, medium, high myopia, age and sex matched healthy controls (i.e, Group 1-4). Diagnosis was established with help of detailed history, clinical and ocular examination. Lactate levels were estimated in tear samples by enzymatic method on autoanalyzer within 30 minutes of collection. Independent Student's t-test was used to determine the significance between myopia and controls. Results: Out of total 100 subjects, 68% were males and 32% were females. The mean±SD age was 24.56±6.35, 24.12±5.45, and 23.24±5.17, 21.69±3.95 in the Group 1, Group 2, Group 3 and Group 4 respectively in the control group, group with low, medium and high myopia respectively which was statistically not significant. The results of this study showed that lactate levels in tears were significantly higher (p-value=0.013) in myopia patients as compared to controls (1.34±0.46 versus 1.06±0.50). Additionally, the lactate levels increased with the progression of degree of myopia. Conclusion: Increase in the level of lactate in tear was directly proportional to the degree of myopia. Tear lactate levels can be used as a non invasive method to study oxidative stress in individuals. Further studies with more oxidative markers in tears can also be compared with blood values in larger groups of patients with long-term follow-up to validate the finding. [ABSTRACT FROM AUTHOR]

2-Deoxy-D-glucose (2DG) has recently received emergency approval for the treatment of COVID-19 in India, after a successful clinical trial. SARS-CoV-2 infection of cultured cells is accompanied by elevated glycolysis and decreased mitochondrial function, whereas 2DG represses glycolysis and stimulates respiration, and restricts viral replication. While 2DG has pleiotropic effects on cell metabolism in cultured cells it is not known which of these manifests in vivo. On the other hand, it is known that 2DG given continuously can have severe detrimental effects on the rodent heart. Here, we show that the principal effect of an extended, intermittent 2DG treatment on mice is to augment the mitochondrial respiratory chain proteome in the heart; importantly, this occurs without vacuolization, hypertrophy or fibrosis. The increase in the heart respiratory chain proteome suggests an increase in mitochondrial oxidative capacity, which could compensate for the energy deficit caused by the inhibition of glycolysis. Thus, 2DG in the murine heart appears to induce a metabolic configuration that is the opposite of SARS-CoV-2 infected cells, which could explain the compound's ability to restrict the propagation of the virus to the benefit of patients with COVID-19 disease. [ABSTRACT FROM AUTHOR]

Mechanisms discovered to drive increased glucose metabolism in cancer cells are found to be similar to those in viral‐infected cells. In this mini review, we summarize the major pathways by which the sugar analog, 2‐Deoxy‐d‐glucose, has been shown to exploit increased glucose metabolism in cancer and how this information applies to viral‐infected cells. Moreover, we highlight the relevance of these findings to the emergency approval of 2‐Deoxy‐d‐glucose in India to be used against SARS‐CoV‐2, the virus responsible for COVID‐19. [ABSTRACT FROM AUTHOR]

Head and neck squamous cell carcinoma (HNSCC) is the six most common cancer globally and most common cancer in men in India. The metabolic regulation is highly altered and is considered as a hall mark of HNSCC. TP53-induced glycolysis and apoptosis regulator (TIGAR) plays very important role in the development and progression of HNSCC. The aim of our study is to identify a novel FDA approved anticancer inhibitor against mutated TP53-induced glycolysis and apoptosis regulator (TIGAR) through drug repurposing approach. A library of 105 FDA approved anticancer compounds were screened using molecular docking approach against TIGAR (PDB: 3DCY) both Wild-Type (WT) and mutated (Mut). Specific mutations in TIGAR were identified using cBioPortal, a cancer genomics database and mutated structure was modelled using SWISS-MODEL. Out of 510 sequenced cases/patients samples, 17(3%) patients showed alteration in TIGAR [TIGARWT and TIGARMut (R88W)]. The virtual drug screening showed 45 drugs out of 105 high binding affinity with TIGAR, Trabectedin showed highest binding affinity with both TIGARWT (− 13.3 kcal/mol) as well as TIGARMut (R88W) (− 13.8 kcal/mol). The molecular docking studies were validated using molecular dynamics simulation (MD Simulation) of protein–ligand complex of TIGAR and Trabectedin for 100 ns. The MD Simulation of Trabectedin complex showed more stable with TIGARMut (R88W) compared to TIGARWT. Moreover, the string analysis revealed that metabolic-related genes, HK2, PFKFB1, PFKM, PFKP, PFKL, FBP1 are closely associated with TIGAR in HNSCC. Our findings suggest that Trabectedin can be proposed as an inhibitor for [TIGARMut (R88W)] which can be used to target metabolic signalings in HNSCC. However, further investigation and in vitro and in vivo validation our findings required to understand the molecular mechanisms of regulation of Trabectedin in HNSCC. [ABSTRACT FROM AUTHOR]

A feeding trial was conducted to study the effect of dietary protein (DP) levels on the enzyme activities of different stocks of Macrobrachium rosenbergii juveniles, fed two diets prawn type diets either at optimum (ODP, 32%) or sub‐optimum (SDP, 27%) dietary protein. The activity of key enzymes: glycolysis, gluconeogenesis, amino acid catabolism, pentose phosphate pathway, and energy metabolism was examined on different stocks of M. rosenbergii juveniles in various tissues (muscle, hepatopancreas, and gill). Wild juveniles of M. rosenbergii were collected from the west coast of India, Gujarat (G), Maharashtra (M) and from the east coast of India, Andhra Pradesh (A), and raised in culture ponds of 200 m2 at 1 juvenile m‐2. The experiment was conducted for a period of 60 days. A Visible Implant Elastomer (VIE) Tag was injected into different prawn juveniles' stocks A, (3.06–3.10 g), M, (0.80–1.01 g) and G (0.90–1.06 g) for the individual identification of each class. All the animals were acclimatized for 7 days before being released into the pond. Each of the two diets, the first with 27% SDP, termed the sub‐optimum level (S), and the second 32% DP, termed the optimum level (O), was fed in triplicate ponds. The weight gain of stocked prawn was significantly higher for G stock (3.03 ± 0.18 g) compared to M stock (2.53 ± 0.25 g) and A stock (1.33 ± 0.10 g). In addition weight gain in ODP was significantly higher (3.05 ± 0.22 g) compared to SDP (2.11 ± 0.17 g). Furthermore, both protein level and stock type had a significant (p ≤ 0.05) effect on the survival rate, specific growth rate, and feed conversion ratio of the prawn. A significant interaction was recorded between metabolic enzyme activities and the variation of dietary protein levels and differences in M. rosenbergii stocks, in which stock "G" and "M" exhibited healthier enzyme physiological status than stock "A". The findings presented here may conclude that the enzymes quickly respond to dietary protein manipulations of cultured shrimp. The results can be useful in identifying the optimum stock (genotypes) in a given culture condition and subsequently higher production may be achieved. [ABSTRACT FROM AUTHOR]

Objectives: Glucose-6-phosphate isomerase (GPI) deficiency is an autosomal recessive genetic disorder causing hereditary non-spherocytic hemolytic anemia (HNSHA) coupled with a neurological disorder. The aim of this study was to identify GPI genetic defects in a cohort of Indian patients with HNSHA coupled with neurological dysfunction., Methods: Thirty-five patients were screened for GPI deficiency in the HNSHA patient group; some were having neurological dysfunction. Enzyme activity was measured by spectrophotometric method. The genetic study was done by single-stranded conformation polymorphism (SSCP) analysis, restriction fragment length polymorphism (RFLP) analysis by the restriction enzyme AciI for p.Arg347His (p.R347H) and confirmation by Sanger's sequencing., Results: Out of 35 patients, 15 showed 35% to 70% loss of GPI activity, leading to neurological problems with HNSHA. Genetic analysis of PCR products of exon 12 of the GPI gene showed altered mobility on SSCP gel. Sanger's sequencing revealed a homozygous c1040G > A mutation predicting a p.Arg347His replacement which abolishes AciI restriction site. The molecular modeling analysis suggests p.Arg347 is involved in dimerization of the enzyme. Also, this mutation generates a more labile enzyme which alters its three-dimensional structure and function., Conclusions: This report describes the high prevalence of p.Arg347His pathogenic variant identified in Indian GPI deficient patients with hemolytic anemia and neuromuscular impairment. It suggests that neuromuscular impairment with hemolytic anemia cases could be investigated for p.Arg347His pathogenic variant causing GPI deficiency because of neuroleukin activity present in the GPI monomer which has neuroleukin action at the same active site and generates neuromuscular problems as well as hemolytic anemia.

Glucose-6-phosphate isomerase (GPI) deficiency is an autosomal recessive genetic disorder causing congenital haemolytic anaemia (CHA). Diagnosis of GPI deficiency by the biochemical method is unpredicted. Molecular diagnosis by identifying genetic mutation is the gold standard method for confirmation of disease, but causative genes involved in CHA are numerous, and identifying a gene-by-gene approach using Sanger sequencing is also cumbersome, expensive and labour intensive. Recently, next-generation targeted sequencing is more useful in the diagnosis of unexplained haemolytic anaemia. We used targeted next-generation sequencing (NGS) clinical panel for diagnosis of unexplained haemolytic anaemia in two Indian patients which were pending for a long time. All possible causes of haemolytic anaemia were found within normal limit. NGS by clinical exome panel revealed homozygous novel missense mutation in exon 12, c.1009G>A (p.Ala337Thr) in both patients. We further confirm by measuring red blood cell GPI activity in the patients and showed deficiency whereas parents were having intermediate activity. c.1009G>A mutation was also confirmed by Sanger sequencing of exon 12 of GPI gene. The structural-functional analysis by bioinformatics software like Swiss PDB, PolyPhen-2 and PyMol suggested that this pathogenic variant has a direct impact on the structural rearrangement at the region near the active site of the enzyme. This rapid and high-performance targeted NGS assay can be configured to detect specific CHA mutations unique to an individual defect, making it a potentially valuable method for diagnosis of unexplained haemolytic anaemia., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2019. No commercial re-use. See rights and permissions. Published by BMJ.)