Mutations in S2 subunit of SARS-CoV-2 Omicron spike strongly influence its conformation, fusogenicity, and neutralization sensitivity.

IMPORTANCE: The Omicron subvariants have substantially evaded host-neutralizing antibodies and adopted an endosomal route of entry. The virus has acquired several mutations in the receptor binding domain and N-terminal domain of S1 subunit, but remarkably, also incorporated mutations in S2 which are fixed in Omicron sub-lineage. Here, we found that the mutations in the S2 subunit affect the structural and biological properties such as neutralization escape, entry route, fusogenicity, and protease requirement. In vivo, these mutations may have significant roles in tropism and replication. A detailed understanding of the effects of S2 mutations on Spike function, immune evasion, and viral entry would inform the vaccine design, as well as therapeutic interventions aiming to block the essential proteases for virus entry. Thus, our study has identified the crucial role of S2 mutations in stabilizing the Omicron spike and modulating neutralization resistance to antibodies targeting the S1 subunit.

Humanized L184Q Mutated Surfactant Protein C Gene Alters Alveolar Type 2 Epithelial Cell Fate.

Alveolar type 2 epithelial (AT2) cells synthesize surfactant protein C (SPC) and repair an injured alveolar epithelium. A mutated surfactant protein C gene (SftpcL184Q, Gene ID: 6440) in newborns has been associated with respiratory distress syndrome and pulmonary fibrosis. However, the underlying mechanisms causing Sftpc gene mutations to regulate AT2 lineage remain unclear. We utilized three-dimensional (3D) feeder-free AT2 organoids in vitro to simulate the alveolar epithelium and compared AT2 lineage characteristics between WT (C57BL/6) and SftpcL184Q mutant mice using colony formation assays, immunofluorescence, flow cytometry, qRT-PCR, and Western blot assays. The AT2 numbers were reduced significantly in SftpcL184Q mice. Organoid numbers and colony-forming efficiency were significantly attenuated in the 3D cultures of primary SftpcL184Q AT2 cells compared to those of WT mice. Podoplanin (PDPN, Alveolar type 1 cell (AT1) marker) expression and transient cell count was significantly increased in SftpcL184Q organoids compared to in the WT mice. The expression levels of CD74, heat shock protein 90 (HSP90), and ribosomal protein S3A1 (RPS3A1) were not significantly different between WT and SftpcL184Q AT2 cells. This study demonstrated that humanized SftpcL184Q mutation regulates AT2 lineage intrinsically. This regulation is independent of CD74, HSP90, and RPS3A1 pathways.

Wnt5a/ß-catenin axis is involved in the downregulation of AT2 lineage by PAI-1.

Failure to regenerate injured alveoli functionally and promptly causes a high incidence of fatality in coronavirus disease 2019 (COVID-19). How elevated plasminogen activator inhibitor-1 (PAI-1) regulates the lineage of alveolar type 2 (AT2) cells for re-alveolarization has not been studied. This study aimed to examine the role of PAI-1-Wnt5a-ß catenin cascades in AT2 fate. Dramatic reduction in AT2 yield was observed in Serpine1Tg mice. Elevated PAI-1 level suppressed organoid number, development efficiency, and total surface area in vitro. Anti-PAI-1 neutralizing antibody restored organoid number, proliferation and differentiation of AT2 cells, and ß-catenin level in organoids. Both Wnt family member 5A (Wnt5a) and Wnt5a-derived N-butyloxycarbonyl hexapeptide (Box5) altered the lineage of AT2 cells. This study demonstrates that elevated PAI-1 regulates AT2 proliferation and differentiation via the Wnt5a/ß catenin cascades. PAI-1 could serve as autocrine signaling for lung injury repair.

Assessment of groundwater salinity using principal component analysis (PCA): a case study from Mewat (Nuh), Haryana, India.

In the present study, principal component analysis (PCA) is used to investigate the processes controlling groundwater salinity in the Mewat (Nuh) district, Haryana, India. Twenty groundwater samples were collected from salinity-affected areas in the March-April months of years 2018 and 2019 and were analyzed for chemical variables pH, EC, Ca2+, Mg2+, Na+, K+, [Formula: see text], Cl-, SO42-, [Formula: see text], TDS, and total hardness. Three principal components were selected based on the eigen value, which explains 79.58% and 85.08% of the total variation in the years 2018 and 2019, respectively. The first principal component (PC-1) is identified with salinity, the second principal component (PC-2) with alkalinity, and the third principal component (PC-3) described the pollution. When the yearly comparison was made, the samples collected in 2019 were found to have an increased salinity compared to 2018, which shows an increased vulnerability to the aquifer of Mewat on account of the decline in rainfall recharge. It was also evident that declining recharge also triggered the recharge from other sources; thus, the impact of pollution is more pronounced in 2019 compared to 2018.

Structural, functional and biological insights into the role of Mycobacterium tuberculosis VapBC11 toxin-antitoxin system: targeting a tRNase to tackle mycobacterial adaptation.

Toxin-antitoxin (TA) systems are involved in diverse physiological processes in prokaryotes, but their exact role in Mycobacterium tuberculosis (Mtb) virulence and in vivo stress adaptation has not been extensively studied. Here, we demonstrate that the VapBC11 TA module is essential for Mtb to establish infection in guinea pigs. RNA-sequencing revealed that overexpression of VapC11 toxin results in metabolic slowdown, suggesting that modulation of the growth rate is an essential strategy for in vivo survival. Interestingly, overexpression of VapC11 resulted in the upregulation of chromosomal TA genes, suggesting the existence of highly coordinated crosstalk among TA systems. In this study, we also present the crystal structure of the VapBC11 heterooctameric complex at 1.67 Å resolution. Binding kinetic studies suggest that the binding affinities of toxin-substrate and toxin-antitoxin interactions are comparable. We used a combination of structural studies, molecular docking, mutational analysis and in vitro ribonuclease assays to enhance our understanding of the mode of substrate recognition by the VapC11 toxin. Furthermore, we have also designed peptide-based inhibitors to target VapC11 ribonuclease activity. Taken together, we propose that the structure-guided design of inhibitors against in vivo essential ribonucleases might be a novel strategy to hasten clearance of intracellular Mtb.

Culture & differentiation of mesenchymal stem cell into osteoblast on degradable biomedical composite scaffold: In vitro study.

BACKGROUND & OBJECTIVES: There is a significant bone tissue loss in patients from diseases and traumatic injury. The current autograft transplantation gold standard treatment has drawbacks, namely donor site morbidity and limited supply. The field of tissue engineering has emerged with a goal to provide alternative sources for transplantations to bridge this gap between the need and lack of bone graft. The aim of this study was to prepare biocomposite scaffolds based on chitosan (CHT), polycaprolactone (PCL) and hydroxyapatite (HAP) by freeze drying method and to assess the role of scaffolds in spatial organization, proliferation, and osteogenic differentiation of human mesenchymal stem cells (hMSCs) in vitro, in order to achieve bone graft substitutes with improved physical-chemical and biological properties. METHODS: Pure chitosan (100CHT) and composites (40CHT/HAP, 30CHT/HAP/PCL and 25CHT/HAP/PCL scaffolds containing 40, 30, 25 parts per hundred resin (phr) filler, respectively) in acetic acid were freeze dried and the porous foams were studied for physicochemical and in vitro biological properties. RESULTS: Scanning electron microscope (SEM) images of the scaffolds showed porous microstructure (20-300 µm) with uniform pore distribution in all compositions. Materials were tested under compressive load in wet condition (using phosphate buffered saline at pH 7.4). The in vitro studies showed that all the scaffold compositions supported mesenchymal stem cell attachment, proliferation and differentiation as visible from SEM images, [3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide] (MTT) assay, alkaline phosphatase (ALP) assay and quantitative reverse transcription (qRT)-PCR. INTERPRETATION & CONCLUSIONS: Scaffold composition 25CHT/HAP/PCL showed better biomechanical and osteoinductive properties as evident by mechanical test and alkaline phosphatase activity and osteoblast specific gene expression studies. This study suggests that this novel degradable 3D composite may have great potential to be used as scaffold in bone tissue engineering.

Biomarkers and molecular endotypes of sarcoidosis: lessons from omics and non-omics studies.

Sarcoidosis is a chronic granulomatous disorder characterized by unknown etiology, undetermined mechanisms, and non-specific therapies except TNF blockade. To improve our understanding of the pathogenicity and to predict the outcomes of the disease, the identification of new biomarkers and molecular endotypes is sorely needed. In this study, we systematically evaluate the biomarkers identified through Omics and non-Omics approaches in sarcoidosis. Most of the currently documented biomarkers for sarcoidosis are mainly identified through conventional "one-for-all" non-Omics targeted studies. Although the application of machine learning algorithms to identify biomarkers and endotypes from unbiased comprehensive Omics studies is still in its infancy, a series of biomarkers, overwhelmingly for diagnosis to differentiate sarcoidosis from healthy controls have been reported. In view of the fact that current biomarker profiles in sarcoidosis are scarce, fragmented and mostly not validated, there is an urgent need to identify novel sarcoidosis biomarkers and molecular endotypes using more advanced Omics approaches to facilitate disease diagnosis and prognosis, resolve disease heterogeneity, and facilitate personalized medicine.

Impact of lockdown due to COVID-19 pandemic on groundwater salinity in Punjab, India: some hydrogeoethics issues.

In the present study, a total of 48 groundwater samples (13 from shallow aquifers depth < 50 m and 35 samples from deep aquifers in the depth range 50-200 m) were collected from three industrial dominant districts (Ludhiana, Jalandhar and Moga) of Punjab after the lockdown period and before the start of southwest monsoon in the month of June, 2020 (pre-monsoon). The values for total dissolved solids (TDS) observed in Monsoon season (August, 2020) and November-December, 2019 (post-monsoon) were compared with the values taken in June, 2020 (pre-monsoon) to see the impact of lockdown due to COVID-19 pandemic on groundwater salinity. ~ 60% of samples were found to have TDS values more than acceptable limit (500 mg/l) before lockdown (post-monsoon season of 2019) period and after or during lockdown period (June, 2020) number of samples more than the acceptable limit (500 mg/l) reduced to 45%. Average TDS values reduced by 25% in shallow aquifers after lockdown and area under TDS values in acceptable limit (500 mg/l) increased by 23% of samples as compared to the TDS values found in monsoon season of year 2019. In deeper aquifers, increase of only 3% area under TDS values in acceptable limit of 500 mg/l was found. Reductions in TDS values in shallow aquifers clearly show that there is an urgent need for proper management of salinity causing elements and regulating these to check groundwater contaminations using the holistic and hydro-geoethical approach.

Tissue-specific mesenchymal stem cell-dependent osteogenesis in highly porous chitosan-based bone analogs.

Among conventional fabrication techniques, freeze-drying process has widely been investigated for polymeric implants. However, the understanding of the stem cell progenitor-dependent cell functionality modulation and quantitative analysis of early osseointegration of highly porous scaffolds have not been explored. Here, we developed a novel, highly porous, multimaterial composite, chitosan/hydroxyapatite/polycaprolactone (CHT/HA/PCL). The in vitro studies have been performed using mesenchymal stem cells (MSCs) from three tissue sources: human bone marrow-derived MSCs (BM-MSCs), adipose-derived MSCs (AD-MSCs), and Wharton's jelly-derived MSCs (WJ-MSCs). Although cell attachment and metabolic activity [3-4,5-dimethylthiazol-2yl-(2,5 diphenyl-2H-tetrazoliumbromide) assay] were ore enhanced in WJ-MSC-laden CHT/HA/PCL composites, scanning electron microscopy, real-time gene expression (alkaline phosphatase [ALP], collagen type I [Col I], osteocalcin [OCN], and bone morphogenetic protein 4 [BMP-4]), and immunostaining (COL I, ß-CATENIN, OCN, and SCLEROSTIN [SOST]) demonstrated pronounced osteogenesis with terminal differentiation on BM-MSC-laden CHT/HA/PCL composites only. The enhanced cell functionality on CHT/HA/PCL composites was explained in terms of interplay among the surface properties and the optimal source of MSCs. In addition, osteogenesis in rat tibial model over 6 weeks confirmed a better ratio of bone volume to the total volume for BM-MSC-laden composites over scaffold-only and defect-only groups. The clinically conformant combination of 3D porous architecture with pore sizes varying in the range of 20 to 200 µm together with controlled in vitro degradation and early osseointegration establish the potential of CHT/HA/PCL composite as a potential cancellous bone analog.

The guggul for chronic diseases: ancient medicine, modern targets.

Identification of active principles and their molecular targets from traditional medicine is an enormous opportunity for modern drug development. Gum resin from Commiphora wightii (syn C. mukul) has been used for centuries in Ayurveda to treat internal tumors, obesity, liver disorders, malignant sores and ulcers, urinary complaints, intestinal worms, leucoderma (vitiligo), sinuses, edema and sudden paralytic seizures. Guggulsterone has been identified as one of the major active components of this gum resin. This steroid has been shown to bind to the farnesoid X receptor and modulate expression of proteins with antiapoptotic (IAP1, XIAP, Bfl-1/A1, Bcl-2, cFLIP, survivin), cell survival, cell proliferation (cyclin D1, c-Myc), angiogenic, and metastatic (MMP-9, COX-2, VEGF) activities in tumor cells. Guggulsterone mediates gene expression through regulation of various transcription factors, including NF-kappaB, STAT-3 and C/EBPalpha, and various steroid receptors such as androgen receptor and glucocorticoid receptors. Modulation of gene expression by guggulsterone leads to inhibition of cell proliferation, induction of apoptosis, suppression of invasion and abrogation of angiogenesis. Evidence has been presented to suggest that guggulsterone can suppress tumor initiation, promotion and metastasis. This review describes the identification of molecular targets of guggulsterone, cellular responses to guggulsterone, and animal studies and clinical trials of guggulsterone in cancer and other diseases.

Bacillus subtilis HelD, an RNA Polymerase Interacting Helicase, Forms Amyloid-Like Fibrils.

HelD, an RNA polymerase binding protein from Bacillus subtilis, stimulates transcription and helps in timely adaptation of cells under diverse environmental conditions. At present, no structural information is available for HelD. In the current study, we performed size exclusion chromatography coupled to small angle X-ray scattering (SEC-SAXS) which suggests that HelD is predominantly monomeric and globular in solution. Using combination of size exclusion chromatography and analytical ultracentrifugation, we also show that HelD has a tendency to form higher order oligomers in solution. CD experiments suggest that HelD has both α-helical (∼35%) and ß sheet (∼26%) secondary structural elements. Thermal melting experiments suggest that even at 90°C, there is only about 30% loss in secondary structural contents with Tm of 44°C. However, with the increase in temperature, there was a gain in the ß-sheet content and significant irreversible loss of α-helical content. Using a combination of X-ray fiber diffraction analysis, and dye based assays including Thioflavin-T based fluorescence and Congo red binding assays, we discovered that HelD forms amyloid-like fibrils at physiologically relevant conditions in vitro. Using confocal imaging, we further show that HelD forms amyloid inclusions in Escherichia coli. Bioinformatics-based sequence analysis performed using three independent web-based servers suggests that HelD has more than 20 hot-spots spread across the sequence that may aid the formation of amyloid-like fibrils. This discovery adds one more member to the growing list of amyloid or amyloid-like fibril forming cytosolic proteins in bacteria. Future studies aimed at resolving the function of amyloid-like fibrils or amyloid inclusions may help better understand their role, if any, in the bacterial physiology.

Deep urban groundwater vulnerability in India revealed through the use of emerging organic contaminants and residence time tracers.

Demand for groundwater in urban centres across Asia continues to rise with ever deeper wells being drilled to avoid shallow contamination. The vulnerability of deep alluvial aquifers to contaminant migration is assessed in the ancient city of Varanasi, India, using a novel combination of emerging organic contaminants (EOCs) and groundwater residence time tracers (CFC and SF6). Both shallow and intermediate depth private sources (<100 m) and deep (>100 m) municipal groundwater supplies were found to be contaminated with a range of EOCs including pharmaceuticals (e.g. sulfamethoxazole, 77% detection frequency, range <0.0001-0.034 µg L-1), perfluoroalkyl substances (e.g. PFOS, range <0.0001-0.033 µg L-1) as well as a number of pesticides (e.g. phenoxyacetic acid, range <0.02-0.21 µg L-1). The profile of EOCs found in groundwater mirror those found in surface waters, albeit at lower concentrations, and reflect common waste water sources with attenuation in the subsurface. Mean groundwater residence times were found to be comparable between some deep groundwater and shallow groundwater sources with residence times ranging from >70 to 30 years. Local variations in aquifer geology influence the extent of modern recharge at depth. Both tracers provide compelling evidence of significant inputs of younger groundwater to depth >100 m within the aquifer system.

Bicuspidization of the tricuspid valve for the treatment of posterior leaflet endocarditis: a case report.

Patients who require surgical therapy for the treatment of tricuspid valve regurgitation can avoid undergoing tricuspid valve replacement if valve-sparing repair techniques are employed. Tricuspid valvular endocarditis frequently requires valvectomy, leaving the right side of the heart and pulmonary system vulnerable to unregulated blood flow. We present a case of complete posterior leaflet excision and plication of the involved portion of the tricuspid annulus, which resulted in "bicuspidization" of the valve, for the treatment of tricuspid valve endocarditis localized to the posterior leaflet.

Groundwater quality in the alluvial aquifer system of northwest India: New evidence of the extent of anthropogenic and geogenic contamination.

Groundwater depletion has been widely studied in northwest India, but water quality concerns are still poorly constrained. In this study, we explore the hydrochemistry of the top 160m of the aquifer system, through detailed field studies in the Bist-Doab region, considering both anthropogenic and geogenic controls. A detailed comparison is made between sites dominated by urban and agricultural landuse. Salinity, nitrate, chloride and lead concentrations are significantly higher in the shallow (0-50m) groundwater system due to surface anthropogenic contaminant loading from agricultural and urban sources. The widespread occurrence of oxic groundwater within the aquifer system means that denitrification potential is limited and also enhances the mobility of selenium and uranium in groundwater. Geogenic trace elements (e.g. As, Se, F), are generally found at concentrations below WHO guideline drinking water values, however elevated U concentrations (50-70µg/L) are found within the deeper part of the aquifer and shallow urban aquifers associated with higher bicarbonate waters. Higher concentration of Se (10-40µg/L) are found exclusively in the shallow groundwater system where Se is mobilised from soils and transported to depth in the shallow aquifer due to the prevailing oxidising aquifer conditions. New evidence from a range of environmental tracers shows elevated concentrations of anthropogenic contaminants in the deeper part of the aquifer (50-160m deep) and demonstrates vulnerability to vertical migration of contaminants. Continued intensive groundwater abstraction from >100m deep means that water quality risks to the deep aquifer system need to be considered together with water quantity constraints.

Spurts in India's population growth during the twentieth century: a spatial view.

PIP: The demographic transition in twentieth-century India is analyzed using census data. The authors first examine the extent to which 1921 and 1951 can be identified as the two critical points in the demographic transition process. Particular attention is paid to regional differences in the pace of the transition process. The results confirm the overall validity of the two selected years as critical points in the transition although the timing of the transition was significantly different when examined from a regional perspective. They also indicate that the demographic transition model is appropriate to the Indian experience although the threshold levels at which changes occurred were different.^ieng

Dose-dependent facilitation of peripheral nerve regeneration by bone marrow-derived mononuclear cells: a randomized controlled study: laboratory investigation.

OBJECT: Bone marrow-derived stem cells enhance the rate of regeneration of neuronal cells leading to clinical improvement in nerve injury, spinal cord injury, and brain infarction. Recent experiments in the local application of bone marrow-derived mononuclear cells (BM-MNCs) in models of sciatic nerve transection in rats have suggested their beneficial role in nerve regeneration, although the effects of variable doses of stem cells on peripheral nerve regeneration have never been specifically evaluated in the literature. In this paper, the authors evaluated the dose-dependent role of BM-MNCs in peripheral nerve regeneration in a model of sciatic nerve transection in rats. METHODS: The right sciatic nerve of 60 adult female Wistar rats (randomized into 2 test groups and 1 control group, 20 rats in each group) underwent transection under an operating microscope. The cut ends of the nerve were approximated using 2 epineural microsutures. The gap was filled with low-dose (5 million BM-MNCs/100 µl phosphate-buffered saline [PBS]) rat BM-MNCs in one group, high-dose (10 million BM-MNCs/100 µl PBS) rat BM-MNCs in another group, and only PBS in the control group, and the approximated nerve ends were sealed using fibrin glue. Histological assessment was performed after 30 days by using semiquantitative and morphometric analyses and was done to assess axonal regeneration, percentage of myelinated fibers, axonal diameter, fiber diameter, and myelin thickness at distal-most sites (10 mm from site of repair), intermediate distal sites (5 mm distal to the repair site), and site of repair. RESULTS: The recovery of nerve cell architecture after nerve anastomosis was far better in the high-dose BM-MNC group than in the low-dose BM-MNC and control groups, and it was most evident (p < 0.02 in the majority of the parameters [3 of 4]) at the distal-most site. Overall, the improvement in myelin thickness was most significant with incremental dosage of BM-MNCs, and was evident at the repair, intermediate distal, and distal-most sites (p = 0.001). CONCLUSIONS: This study emphasizes the role of BM-MNCs, which can be isolated easily from bone marrow aspirates, in peripheral nerve injury and highlights their dose-dependent facilitation of nerve regeneration.

Effect of parental consanguinity on anthropometric measurements among the Sheikh Sunni Muslim boys of Delhi.

The study of consanguineous marriage is an efficient way to elucidate the genetic structure of human populations. Such matings give an opportunity for recessive genes to manifest themselves by becoming homozygous. The present attempt examines the effects of parental consanguinity on various anthropometric measurements among the Sheikh Sunni Muslim boys of old Delhi between the ages of 11 and 16 years. A slight inbreeding depression has been observed for all eight anthropometric measurements, i.e., stature, span, sitting height, head length, head circumference, chest girth, and calf circumference. The results support earlier studies in regard to the effect of consanguinity on anthropometric measurements.

Is India overurbanised?

This paper questions the frequently posed thesis of India being overurbanized. The diverse connotations of the term "overurbanization" have been taken into account and tested against the Indian reality. It is concluded that on a net balance of positive and negative effects emanating from urbanization, there is no reason to believe that India is urbanized to an extravagant degree. Any continued adherence to the "overurbanization" thesis is likely to distort our perceptions about the vital role which Indian urbanization is playing in economic growth.

Map Series: 3. Infant mortality in India.

PIP: The author assesses the extent of infant mortality in India, with a focus on geographical variations.^ieng