In vitro, ex vivo, instrumental and clinical evaluation of a topical cream on the signs of periorbital ageing.

OBJECTIVE: Periorbital skin ageing signs are multidimensional, highly visible and a concern for many. We evaluated the potential efficacy of an eye cream to diminish these signs. METHODS: Biological markers associated with ageing, barrier function and homeostasis were analysed in vitro to determine the effects of topically applied eye cream, compared to those of a placebo using human skin tissue models and/or explants. Collagen IV, elastin and bone morphogenic protein 4 (BMP4) expression was investigated by immunohistochemical labelling, while filaggrin, kallikrein 7 (KLK7) and HB-EGF were evaluated by RT-qPCR. IL-1α and melanin levels in darkly pigmented skin models were also quantified. The protective effect of the cream on glycation was assessed by a non-enzymatic assay. Finally, the benefits of twice-daily applications of the eye cream for 56 days were instrumentally and clinically evaluated on 33 women. RESULTS: Only the eye cream, not the placebo, stimulated collagen IV and BMP4 protein expression, as well as increased elastin fibre length. It also led to higher HB-EGF, filaggrin and KLK7 mRNA levels. The placebo and the eye cream did not induce changes in IL-1α and melanin levels, but both reduced non-enzymatic glycation. When assessing the in vivo effects of the cream, short-term results indicated skin hydration, transepidermal water loss (TEWL) and skin profilometry improvement within 15 min. Instrumental evaluations of wrinkles showed a reduction after 7 days, which was clinically perceivable after 28 or 56 days. The eye-opening angle and eyelid sagging also improved after seven and 28 days, respectively. Finally, dark circles became lighter within 7 days (instrumental measurement) or 28 days (clinical assessment). CONCLUSION: The instrumental and clinical evaluations revealed that the eye cream reduced all periorbital ageing signs evaluated. Its effects are supported by the in vitro and ex vivo analyses of molecular markers.

OBJECTIF: Les signes de vieillissement de la peau périorbitaire sont nombreux, très visibles et préoccupent de nombreuses personnes. Nous avons évalué l'efficacité potentielle d'une crème pour les yeux pour atténuer ces signes. MÉTHODES: Les marqueurs biologiques associés au vieillissement, à la fonction barrière et à l'homéostasie de la peau ont été analysés in vitro pour évaluer l'efficacité d'une crème pour les yeux appliquée localement. Ces effets ont été comparés à ceux d'un placebo, sur des modèles et/ou des explants de tissus cutanés humains. L'expression du collagène IV, de l'élastine et de la protéine morphogénique osseuse 4 (BMP4) a été étudiée par marquage immunohistochimique. Celle de la filaggrine, de la kallikréine 7 (KLK7), et du HB­EGF par RT­qPCR. Les niveaux d'IL­1α et de mélanine dans un modèle de peau pigmentée ont également été quantifiés. L'effet protecteur de la crème sur la glycation a été évalué par un test non enzymatique. Enfin, les bénéfices d'une application biquotidienne de la crème pour les yeux ont été évalués instrumentalement et cliniquement sur 33 femmes pendant 56 jours. RÉSULTATS: Seule la crème pour les yeux a stimulé l'expression du collagène IV et de BMP4 en comparaison avec le placebo. La crème est aussi la seule à augmenter la longueur des fibres d'élastine. Elle a également entraîné une augmentation des niveaux d'ARNm de HB­EGF, de la filaggrine et de KLK7. Le placebo et la crème pour les yeux n'ont pas modifié les niveaux d'IL­1α et de la mélanine, mais ont tous deux réduit la glycation non enzymatique. Lors de l'évaluation des effets in vivo, les résultats à court terme ont montré une amélioration de l'hydratation de la peau, de la Perte Insensible en Eau (PIE) et du profil de la peau en 15 min. Les évaluations instrumentales de la profondeur des rides ont indiqué une réduction après 7 jours d'application, réduction cliniquement perceptible après 28 ou 56 jours. La crème périorbitale induit également une amélioration de l'angle d'ouverture des yeux et de l'affaissement des paupières respectivement après 7 et 28 jours. Enfin, les cernes sont devenus plus clairs après 7 jours (mesure instrumentale) ou 28 jours (évaluation clinique). CONCLUSION: Les évaluations instrumentales et cliniques ont révélé que la crème pour les yeux réduisait tous les signes de vieillissement périorbitaires évalués. Ses effets sont confirmés par les analyses in vitro et ex vivo des marqueurs moléculaires.

Proteomics-based mass spectrometry profiling of SARS-CoV-2 infection from human nasopharyngeal samples.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of the on-going global pandemic of coronavirus disease 2019 (COVID-19) that continues to pose a significant threat to public health worldwide. SARS-CoV-2 encodes four structural proteins namely membrane, nucleocapsid, spike, and envelope proteins that play essential roles in viral entry, fusion, and attachment to the host cell. Extensively glycosylated spike protein efficiently binds to the host angiotensin-converting enzyme 2 initiating viral entry and pathogenesis. Reverse transcriptase polymerase chain reaction on nasopharyngeal swab is the preferred method of sample collection and viral detection because it is a rapid, specific, and high-throughput technique. Alternate strategies such as proteomics and glycoproteomics-based mass spectrometry enable a more detailed and holistic view of the viral proteins and host-pathogen interactions and help in detection of potential disease markers. In this review, we highlight the use of mass spectrometry methods to profile the SARS-CoV-2 proteome from clinical nasopharyngeal swab samples. We also highlight the necessity for a comprehensive glycoproteomics mapping of SARS-CoV-2 from biological complex matrices to identify potential COVID-19 markers.

Profound N-glycan remodelling accompanies MHC-II immunopeptide presentation.

Immunopeptidomics, the study of peptide antigens presented on the cell surface by the major histocompatibility complex (MHC), offers insights into how our immune system recognises self/non-self in health and disease. We recently discovered that hyper-processed (remodelled) N-glycans are dominant features decorating viral spike immunopeptides presented via MHC-class II (MHC-II) molecules by dendritic cells pulsed with SARS-CoV-2 spike protein, but it remains unknown if endogenous immunopeptides also undergo N-glycan remodelling. Taking a multi-omics approach, we here interrogate published MHC-II immunopeptidomics datasets of cultured monocyte-like (THP-1) and breast cancer-derived (MDA-MB-231) cell lines for overlooked N-glycosylated peptide antigens, which we compare to their source proteins in the cellular glycoproteome using proteomics and N-glycomics data from matching cell lines. Hyper-processed chitobiose core and paucimannosidic N-glycans alongside under-processed oligomannosidic N-glycans were found to prevalently modify MHC-II-bound immunopeptides isolated from both THP-1 and MDA-MB-231, while complex/hybrid-type N-glycans were (near-)absent in the immunopeptidome as supported further by new N-glycomics data generated from isolated MHC-II-bound peptides derived from MDA-MB-231 cells. Contrastingly, the cellular proteomics and N-glycomics data from both cell lines revealed conventional N-glycosylation rich in complex/hybrid-type N-glycans, which, together with the identification of key lysosomal glycosidases, suggest that MHC-II peptide antigen processing is accompanied by extensive N-glycan trimming. N-glycan remodelling appeared particularly dramatic for cell surface-located glycoproteins while less remodelling was observed for lysosomal-resident glycoproteins. Collectively, our findings indicate that both under- and hyper-processed N-glycans are prevalent features of endogenous MHC-II immunopeptides, an observation that demands further investigation to enable a better molecular-level understanding of immune surveillance.

Assessing water, sanitation and hygiene (WASH) practices and their association with diarrhoea in under-five children in urban Chandernagore: Community-based evidence from a small municipal corporation in Hooghly District of West Bengal, India.

The study aims to understand the relationship of childhood diarrhoea (under-five children) with water, sanitation and hygiene factors in the light of other contextual factors in an urban setting in the district of Hooghly in West Bengal, India. This primary study was carried out by SIGMA Foundation, Kolkata from 4 to 24 January 2023 across 404 households having at least one under-five child. The findings suggested that the water score was 'good' in 85.1% of the households whereas the hand hygiene score was 'good' in 14.6% of households. More than 90% of the households had piped water supply. Less than half of them treated water before consumption among which 45.3% used cloths for straining water; 59.2% of the caregivers followed safe disposal of child's faeces; 66.8% of households had no handwashing arrangement, and 30.5% had taps and wash basins for handwashing; 20.3% of the under-five children had suffered from diarrhoea in the last month before the survey and its prevalence was higher in children aged 12-23 months. Multivariate results suggested diarrhoea prevalence was lower in households that were pucca and had good water and hand hygiene scores, lower in children that had received iron fortification and whose caregivers followed safe child's stool disposal.

Utilizing coordination chemistry through formation of a CuII-quinalizarin complex to manipulate cell biology: An in vitro, in silico approach.

Quinalizarin, an analogue of anthracycline anticancer agents, is an anticancer agent itself. A CuII complex was prepared and characterized by elemental analysis, UV-Vis & IR spectroscopy, mass spectrometry, EPR and DFT. The intention behind the preparation of the complex was to increase cellular uptake, compare its binding with DNA against that of quinalizarin, modulation of semiquinone formation, realization of human DNA topoisomerase I & human DNA topoisomerase II inhibition and observation of anticancer activity. While the first two attributes of complex formation lead to increased efficacy, decrease in semiquinone generation could results in a compromise with efficacy. Inhibition of human DNA topoisomerase makes up this envisaged compromise in free radical activity since the complex shows remarkable ability to disrupt activities of human DNA topoisomerase I and II. The complex unlike quinalizarin, does not catalyze flow of electrons from NADH to O2 to the extent known for quinalizarin. Hence, decrease in semiquinone or superoxide radical anion could make modified quinalizarin [as CuII complex] less efficient in free radical pathway. However, it would be less cardiotoxic and that would be advantageous to qualify it as a better anticancer agent. Although binding to calf thymus DNA was comparable to quinalizarin, it was weaker than anthracyclines. Low cost of quinalizarin could justify consideration as a substitute for anthracyclines but the study revealed IC50 of quinalizarin/CuII-quinalizarin was much higher than anthracyclines or their complexes. Even then, there is a possibility that CuII-quinalizarin could be an improved and less costly form of quinalizarin as anticancer agent.

Glycoproteome remodeling and organelle-specific N-glycosylation accompany neutrophil granulopoiesis.

Neutrophils store microbicidal glycoproteins in cytosolic granules to fight intruding pathogens, but their granule distribution and formation mechanism(s) during granulopoiesis remain unmapped. Herein, we comprehensively profile the neutrophil N-glycoproteome with spatiotemporal resolution by analyzing four key types of intracellular organelles isolated from blood-derived neutrophils and during their maturation from bone marrow-derived progenitors using a glycomics-guided glycoproteomics approach. Interestingly, the organelles of resting neutrophils exhibited distinctive glycophenotypes including, most strikingly, highly truncated N-glycans low in α2,6-sialylation and Lewis fucosylation decorating a diverse set of microbicidal proteins (e.g., myeloperoxidase, azurocidin, neutrophil elastase) in the azurophilic granules. Excitingly, proteomics and transcriptomics data from discrete myeloid progenitor stages revealed that profound glycoproteome remodeling underpins the promyelocytic-to-metamyelocyte transition and that the glycophenotypic differences are driven primarily by dynamic changes in protein expression and less by changes within the glycosylation machinery. Notable exceptions were the oligosaccharyltransferase subunits responsible for initiation of N-glycoprotein biosynthesis that were strongly expressed in early myeloid progenitors correlating with relatively high levels of glycosylation of the microbicidal proteins in the azurophilic granules. Our study provides spatiotemporal insights into the complex neutrophil N-glycoproteome featuring intriguing organelle-specific N-glycosylation patterns formed by dynamic glycoproteome remodeling during the early maturation stages of the myeloid progenitors.

Hydathode immunity protects the Arabidopsis leaf vasculature against colonization by bacterial pathogens.

Plants prevent disease by passively and actively protecting potential entry routes against invading microbes. For example, the plant immune system actively guards roots, wounds, and stomata. How plants prevent vascular disease upon bacterial entry via guttation fluids excreted from specialized glands at the leaf margin remains largely unknown. These so-called hydathodes release xylem sap when root pressure is too high. By studying hydathode colonization by both hydathode-adapted (Xanthomonas campestris pv. campestris) and non-adapted pathogenic bacteria (Pseudomonas syringae pv. tomato) in immunocompromised Arabidopsis mutants, we show that the immune hubs BAK1 and EDS1-PAD4-ADR1 restrict bacterial multiplication in hydathodes. Both immune hubs effectively confine bacterial pathogens to hydathodes and lower the number of successful escape events of an hydathode-adapted pathogen toward the xylem. A second layer of defense, which is dependent on the plant hormones' pipecolic acid and to a lesser extent on salicylic acid, reduces the vascular spread of the pathogen. Thus, besides glands, hydathodes represent a potent first line of defense against leaf-invading microbes.

Understanding the gap between knowledge and practice of handwashing in rural India: evidence from a cross-sectional study.

Limited studies in India had captured the gap in knowledge and practice of handwashing in the community. This study assesses the gap in knowledge and practice of handwashing in rural India. The study was conducted across 10 districts in five states of India - Andhra Pradesh, Assam, Maharashtra, Odisha and West Bengal from December 2021 to January 2022 by the SIGMA Foundation, Kolkata in collaboration with UNICEF India. Descriptive statistics, bivariate analysis, creation of indices and multinomial logistic regression were employed. Findings demonstrated that both knowledge of different aspects of hand hygiene and practice of handwashing with soap and water (HWWS) at critical times varied by socio-economic groups and also across the districts/states. Half of the respondents used only water to wash their hands after taking meals, before serving food, whenever their hands seemed dirty and before eating or cooking. Overall, the 'HWWS knowledge index' was 0.46, whereas the 'HWWS practice index' was 0.36. The correlation coefficient between the two was 0.36. The HWWS practice index was lower than the HWWS knowledge index for 50% of the sampled households. Both HWWS knowledge and practice indices were higher among females, higher educated and younger population. The gap between handwashing practice and knowledge was also higher among females and higher educated.

N-acetyl-ß-D-hexosaminidases mediate the generation of paucimannosidic proteins via a putative noncanonical truncation pathway in human neutrophils.

We recently discovered that human neutrophils express immunomodulatory glycoproteins carrying unusual and highly truncated paucimannosidic N-glycans (Man1-3GlcNAc2Fuc0-1), but their biosynthesis remains elusive. Guided by the well-characterized truncation pathway in invertebrates and plants in which the N-acetyl-ß-D-hexosaminidase (Hex) isoenzymes catalyze paucimannosidic protein (PMP) formation, we here set out to test if the homologous human Hex α and ß subunits encoded by HEXA and HEXB drive a similar truncation pathway in human neutrophils. To this end, we performed quantitative glycomics and glycoproteomics of several CRISPR-Cas9-edited Hex-disrupted neutrophil-like HL-60 mutants (HEXA-KO and HEXB-KO) and matching unedited cell lines. Hex disruption was validated using next-generation sequencing, enzyme-linked immunosorbent assay (ELISA), quantitative proteomics and Hex activity assays. Excitingly, all Hex-disrupted mutants displayed significantly reduced levels of paucimannosylation, particularly Man2-3GlcNAc2Fuc1, relative to unedited HL-60 suggesting that both HEXA and HEXB contribute to PMP formation via a hitherto unexplored truncation pathway in neutrophils. Quantitative N-glycomics indeed demonstrated reduced utilization of a putative noncanonical truncation pathway in favor of the canonical elongation pathway in all Hex-disrupted mutants relative to unedited controls. Quantitative glycoproteomics recapitulated the truncation-to-elongation switch in all Hex-disrupted mutants and showed a greater switch for N-glycoproteins cotrafficking with Hex to the azurophilic granules of neutrophils such as myeloperoxidase. Finally, we supported the Hex-PMP relationship by documenting that primary neutrophils isolated from an early-onset Sandhoff disease patient (HEXB-/-) displayed dramatically reduced paucimannosylation relative to neutrophils from an age-matched unaffected donor. We conclude that both human Hex α and ß mediate PMP formation via a putative noncanonical truncation pathway in neutrophils.

Fertility transition and socioeconomic development in districts of India, 2001-2016.

The fertility-development relationship is bi-directional, context-specific, multi-phased and inconsistent over time. Indian districts provide an ideal setting to study this association due to their size, diversity and disparity in socioeconomic development. The objective of this study was to understand the association of fertility and socioeconomic development among the 640 districts of India. Data were drawn from multiple sources: Censuses of India 2001 and 2011; DLHS-2; NFHS-4; and other published sources. A district-level data file for Total Fertility Rate (TFR) and a set of developmental indices were prepared for the 640 districts for 2001 and 2016. Computation of a composite index (District Development Index, DDI), Ordinary Least Squares, Two Stage Least Squares and panel regressions were employed. By 2016, almost half of all Indian districts had attained below-replacement fertility, and 15% had a TFR of above 3.0. The DDI of India increased from 0.399 in 2001 to 0.511 by 2016 and showed large variations across districts. The correlation coefficient between TFR and DDI was -0.658 in 2001 and -0.640 in 2016. Districts with a DDI of between 0.3 and 0.6 in 2001 had experienced a fertility decline of more than 20%. The fertility-development relationship was found to be strongly negative, convex and consistent over time, but the level of association varied regionally. For any given level of DDI, fertility in 2016 was lower than in 2001; and the association was stronger in districts with a DDI below 0.45. The negative convex association between the two was prominent in the northern, central and eastern regions and the curves were flatter in the west, south and north-east. The increasing number of districts with low fertility and low development draws much attention. Some outlying districts in the north-eastern states had high TFR and high DDI (>0.6). Based on the findings, a multi-layered strategy in districts with low socioeconomic development is recommended. Additional investment in education, child health, employment generation and provisioning of contraceptives would improve the human development to achieve India's demographic goals.

Trends in oligomannosylation and α1,2-mannosidase expression in human cancers.

Aberrant protein glycosylation is a prominent cancer feature. While many tumour-associated glycoepitopes have been reported, advances in glycoanalytics continue to uncover new associations between glycosylation and cancer. Guided by a comprehensive literature survey suggesting that oligomannosylation (Man5-9 GlcNAc2) is a widespread and often regulated glycosignature in human cancers, we here revisit a valuable compilation of nearly 500 porous graphitized carbon LC-MS/MS N-glycomics datasets acquired across 11 human cancer types to systematically test for oligomannose-cancer associations. Firstly, the quantitative glycomics data obtained across 34 cancerous cell lines demonstrated that oligomannosylation is a pan-cancer feature spanning in a wide abundance range. In keeping with literature, our quantitative glycomics data of tumour and matching control tissues and new MALDI-MS imaging data of tissue microarrays showed a strong cancer-associated elevation of oligomannosylation in both basal cell (p = 1.78 × 10-12) and squamous cell (p = 1.23 × 10-11) skin cancer and colorectal cancer (p = 8.0 × 10-4). The glycomics data also indicated that some cancer types including gastric and liver cancer exhibit unchanged or reduced oligomannose levels, observations also supported by literature and MALDI-MS imaging data. Finally, expression data from public cancer repositories indicated that several α1,2-mannosidases are regulated in tumour tissues suggesting that these glycan-processing enzymes may contribute to the cancer-associated modulation of oligomannosylation. This omics-centric study has compiled robust glycomics and enzyme expression data revealing interesting molecular trends that open avenues to better understand the role of oligomannosylation in human cancers.

Serum N-Glycomics Stratifies Bacteremic Patients Infected with Different Pathogens.

Bacteremia-i.e., the presence of pathogens in the blood stream-is associated with long-term morbidity and is a potential precursor condition to life-threatening sepsis. Timely detection of bacteremia is therefore critical to reduce patient mortality, but existing methods lack precision, speed, and sensitivity to effectively stratify bacteremic patients. Herein, we tested the potential of quantitative serum N-glycomics performed using porous graphitized carbon liquid chromatography tandem mass spectrometry to stratify bacteremic patients infected with Escherichia coli (n = 11), Staphylococcus aureus (n = 11), Pseudomonas aeruginosa (n = 5), and Streptococcus viridans (n = 5) from healthy donors (n = 39). In total, 62 N-glycan isomers spanning 41 glycan compositions primarily comprising complex-type core fucosylated, bisecting N-acetylglucosamine (GlcNAc), and α2,3-/α2,6-sialylated structures were profiled across all samples using label-free quantitation. Excitingly, unsupervised hierarchical clustering and principal component analysis of the serum N-glycome data accurately separated the patient groups. P. aeruginosa-infected patients displayed prominent N-glycome aberrations involving elevated levels of fucosylation and bisecting GlcNAcylation and reduced sialylation relative to other bacteremic patients. Notably, receiver operating characteristic analyses demonstrated that a single N-glycan isomer could effectively stratify each of the four bacteremic patient groups from the healthy donors (area under the curve 0.93-1.00). Thus, the serum N-glycome represents a new hitherto unexplored class of potential diagnostic markers for bloodstream infections.

Structural and functional diversity of neutrophil glycosylation in innate immunity and related disorders.

The granulated neutrophils are abundant innate immune cells that utilize bioactive glycoproteins packed in cytosolic granules to fight pathogenic infections, but the neutrophil glycobiology remains poorly understood. Facilitated by technological advances in glycoimmunology, systems glycobiology and glycoanalytics, a considerable body of literature reporting on novel aspects of neutrophil glycosylation has accumulated. Herein, we summarize the building knowledge of the structural and functional diversity displayed by N- and O-linked glycoproteins spatiotemporally expressed and sequentially brought-into-action across the diverse neutrophil life stages during bone marrow maturation, movements to, from and within the blood circulation and microbicidal processes at the inflammatory sites in peripheral tissues. It transpires that neutrophils abundantly decorate their granule glycoproteins including neutrophil elastase, myeloperoxidase and cathepsin G with peculiar glyco-signatures not commonly reported in other areas of human glycobiology such as hyper-truncated chitobiose core- and paucimannosidic-type N-glycans and monoantennary complex-type N-glycans. Sialyl Lewisx, Lewisx, poly-N-acetyllactosamine extensions and core 1-/2-type O-glycans are also common neutrophil glyco-signatures. Granule-specific glycosylation is another fascinating yet not fully understood feature of neutrophils. Recent literature suggests that unconventional biosynthetic pathways and functions underpin these prominent neutrophil-associated glyco-phenotypes. The impact of glycosylation on key neutrophil effector functions including extravasation, degranulation, phagocytosis and formation of neutrophil extracellular traps during normal physiological conditions and in innate immune-related diseases is discussed. We also highlight new technologies that are expected to further advance neutrophil glycobiology and briefly discuss the untapped diagnostic and therapeutic potential of neutrophil glycosylation that could open avenues to combat the increasingly prevalent innate immune disorders.

Hyper-truncated Asn355- and Asn391-glycans modulate the activity of neutrophil granule myeloperoxidase.

Myeloperoxidase (MPO) plays essential roles in neutrophil-mediated immunity via the generation of reactive oxidation products. Complex carbohydrates decorate MPO at discrete sites, but their functional relevance remains elusive. To this end, we have characterised the structure-biosynthesis-activity relationship of neutrophil MPO (nMPO). Mass spectrometry demonstrated that nMPO carries both characteristic under-processed and hyper-truncated glycans. Occlusion of the Asn355/Asn391-glycosylation sites and the Asn323-/Asn483-glycans, located in the MPO dimerisation zone, was found to affect the local glycan processing, thereby providing a molecular basis of the site-specific nMPO glycosylation. Native mass spectrometry, mass photometry and glycopeptide profiling revealed significant molecular complexity of diprotomeric nMPO arising from heterogeneous glycosylation, oxidation, chlorination and polypeptide truncation variants and a previously unreported low-abundance monoprotomer. Longitudinal profiling of maturing, mature, granule-separated and pathogen-stimulated neutrophils demonstrated that nMPO is dynamically expressed during granulopoiesis, unevenly distributed across granules and degranulated upon activation. We also show that proMPO-to-MPO maturation occurs during early/mid-stage granulopoiesis. While similar global MPO glycosylation was observed across conditions, the conserved Asn355-/Asn391-sites displayed elevated glycan hyper-truncation, which correlated with higher enzyme activities of MPO in distinct granule populations. Enzymatic trimming of the Asn355-/Asn391-glycans recapitulated the activity gain and showed that nMPO carrying hyper-truncated glycans at these positions exhibits increased thermal stability, polypeptide accessibility and ceruloplasmin-mediated inhibition potential relative to native nMPO. Finally, molecular modelling revealed that hyper-truncated Asn355-glycans positioned in the MPO-ceruloplasmin interface are critical for uninterrupted inhibition. Here, through an innovative and comprehensive approach, we report novel functional roles of MPO glycans, providing new insight into neutrophil-mediated immunity.

Recent updates on COVID-19: A holistic review.

Coronaviruses are large positive-sense RNA viruses with spike-like peplomers on their surface. The Coronaviridae family's strains infect different animals and are popularly associated with several outbreaks, namely SARS and MERS epidemic. COVID-19 is one such recent outbreak caused by SARS-CoV-2 identified first in Wuhan, China. COVID-19 was declared a pandemic by WHO on 11th March 2020. Our review provides information covering various facets of the disease starting from its origin, transmission, mutations in the virus to pathophysiological changes in the host upon infection followed by diagnostics and possible therapeutics available to tackle the situation. We have highlighted the zoonotic origin of SARS-CoV-2, known to share 96.2% nucleotide similarity with bat coronavirus. Notably, several mutations in SARS-CoV-2 spike protein, nucleocapsid protein, PLpro, and ORF3a are reported across the globe. These mutations could alter the usual receptor binding function, fusion process with the host cell, virus replication, and the virus's assembly. Therefore, studying these mutations could help understand the virus's virulence properties and design suitable therapeutics. Moreover, the aggravated immune response to COVID-19 can be fatal. Hypertension, diabetes, and cardiovascular diseases are comorbidities substantially associated with SARS-CoV-2 infection. The review article discusses these aspects, stating the importance of various comorbidities in disease outcomes. Furthermore, medications' unavailability compels the clinicians to opt for atypical drugs like remdesivir, chloroquine, etc. The current diagnostics of COVID-19 include qRT-PCR, CT scan, serological tests, etc. We have described these aspects to expose the information to the scientific community and to accelerate the research.

SARS-CoV-2, an Underestimated Pathogen of the Nervous System.

Numerous clinical studies have reported neurological symptoms in COVID-19 patients since the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), apart from the atypical signs of pneumonia. Angiotensin-converting enzyme-2 (ACE-2), a potential receptor for SARS-CoV-2 entry, is expressed on various brain cells and cerebral parts, i.e., subfornical organ, paraventricular nucleus, nucleus of the tractus solitarius, and rostral ventrolateral medulla, as well as in non-cardiovascular areas such as the motor cortex and raphe. The resident CNS cells like astrocytes and microglia also express ACE-2, thus highlighting the vulnerability of the nervous system to SARS-CoV-2 infection. Additionally, transmembrane serine protease 2 (TMPRSS2) and furin facilitate virus entry into the host. Besides, the probable routes of virus entry into the nervous system include the hematogenic pathway, through the vagus, the olfactory nerve, or the enteric nervous system. However, the trajectory of SARS-CoV-2 to the brain needs investigation. Furthermore, a Th17-mediated cytokine storm is seen in COVID-19 cases with higher levels of IL-1ß/2/7/8/9/10/17, GM-CSF, IFN-γ, TNF-α, CXCL-10, MCP1, and MIP1α/ß. Some cytokines can cross the blood-brain barrier and activate the brain's immune cells to produce neural cytokines, leading to neuronal dysfunctions. Nonetheless, most of the neurological conditions developed due to viral infections may not have effective and registered treatments. Although, some antivirals may inhibit the virus-mediated pathogenesis and prove to be suitable in COVID-19 treatment. Therefore, clinicians' and researchers' collective expertise may unravel the potential of SARS-CoV-2 infection to prevent short-term and long-term CNS damage.

Comorbidity Assessment Is Essential During COVID-19 Treatment.

Coronavirus disease 2019 (COVID-19) caused by SARS-CoV2 is associated with various comorbidities; cardiovascular diseases, hypertension, diabetes, liver, lung diseases, and neurological ailments. The majority of the dysfunctions mentioned above are often associated with endothelial deterioration, indicating that endothelium can be the target of SARS-CoV2. Our study is an exclusive observational study that quantitatively analyses COVID-19 related comorbidities. We retrieved the data of % population of COVID-19 hospitalized and deceased patients with associated comorbidities from publicly accessible portals of the five European countries. A two tailed t-test enabled us to determine the significant proportions of deaths compared to hospitalized patients with associated comorbidity. Our study revealed that deaths associated with cardiovascular diseases and diabetes are highly significant (p < 0.0001) compared to hospitalized in countries like Italy, France, and Spain unlike the Netherlands. Deaths from kidney diseases (Italy- p < 0.0001; Sweden- p < 0.0001; Netherlands- p = 0.0001; France- p = 0.0033) and neurological ailments (France- p = 0.0001; Netherlands- p < 0.0001) are significantly higher than the total hospitalized patients affected by the particular comorbidity. We have noted that deaths due to liver diseases are least associated with COVID-19 among all comorbidities. Intriguingly, immunodeficiency shows mixed outcomes in death proportions compared to the hospital admitted individuals. Besides, the treatment regime involves drugs like losartan, ACE inhibitors, angiotensin-receptor blockers, Remdesivir, Chloroquine, Hydroxychloroquine, etc. may modulate the severity of the comorbidities. These comorbidities can create chaos in the existing healthcare system and may worsen the disease outcome.

Unintended effects of Janani Suraksha Yojana on maternal care in India.

BACKGROUND: The Janani Suraksha Yojana (JSY) is the largest ever conditional cash transfer programme worldwide. It primarily aimed to reduce the maternal and child mortality by increasing the facility based delivery in India. Besides, the JSY has resulted in reduction of out-of-pocket expenditure for delivery care and increased antenatal care. Though studies have examined the direct outcome of JSY, limited studies have attempted to understand the unintended effects (indirect) of the programme. The aim of this study is to examine the effect of JSY on contraceptive use, initiation of breast feeding and postnatal check-up in India. DATA & METHODS: Data from the National Family Health Survey 4, 2015-16 was used in the analyses. A total of 148,746 institutional births in five years preceding the survey were analysed and the analyses were carried out for Low Performing States (LPS) and High Performing States (HPS). Descriptive statistics and the propensity score matching were used to understand the unintended effects of JSY. RESULTS: In India, the use of contraception, early initiation of breastfeeding and postnatal check up was consistently higher among JSY beneficiaries compared to non-JSY beneficiaries. Among JSY beneficiaries, about 45% of the mothers breastfed their child within one hour compared to 42% of the JSY non-beneficiaries. The pattern was almost similar for postnatal check-up. The variations in contraceptive use, breastfeeding practice and postnatal check-up among JSY beneficiaries were higher in LPS states compared to HPS. For instance, in LPS, among JSY beneficiaries, about 58% mothers breastfed their child within one hour of delivery compared to 46% in HPS. Controlling for socio-economic covariates, the JSY beneficiaries in LPS were 12% more likely to use contraception, 8% were more likely to initiate the breast feeding within one hour of child delivery and 6% were more likely to get their postnatal check-up than their counterparts in HPS. DISCUSSION: The unintended effects of JSY were strong and significant in the low performing states. The coverage of JSY should be further extended and the programme needs to be continued.

Glycan analysis of human neutrophil granules implicates a maturation-dependent glycosylation machinery.

Protein glycosylation is essential to trafficking and immune functions of human neutrophils. During granulopoiesis in the bone marrow, distinct neutrophil granules are successively formed. Distinct receptors and effector proteins, many of which are glycosylated, are targeted to each type of granule according to their time of expression, a process called "targeting by timing." Therefore, these granules are time capsules reflecting different times of maturation that can be used to understand the glycosylation process during granulopoiesis. Herein, neutrophil subcellular granules were fractionated by Percoll density gradient centrifugation, and N- and O-glycans present in each compartment were analyzed by LC-MS. We found abundant paucimannosidic N-glycans and lack of O-glycans in the early-formed azurophil granules, whereas the later-formed specific and gelatinase granules and secretory vesicles contained complex N- and O-glycans with remarkably elongated N-acetyllactosamine repeats with Lewis epitopes. Immunoblotting and histochemical analysis confirmed the expression of Lewis X and sialyl-Lewis X in the intracellular granules and on the cell surface, respectively. Many glycans identified are unique to neutrophils, and their complexity increased progressively from azurophil granules to specific granules and then to gelatinase granules, suggesting temporal changes in the glycosylation machinery indicative of "glycosylation by timing" during granulopoiesis. In summary, this comprehensive neutrophil granule glycome map, the first of its kind, highlights novel granule-specific glycosylation features and is a crucial first step toward a better understanding of the mechanisms regulating protein glycosylation during neutrophil granulopoiesis and a more detailed understanding of neutrophil biology and function.

Glycoengineered hepatitis B virus-like particles with enhanced immunogenicity.

Virus-like particles (VLP) represent biological platforms for the development of novel products such as vaccines and delivery platforms for foreign antigenic sequences. VLPs composed of the small surface antigen (HBsAgS) derived from the hepatitis B virus (HBV) are the immunogenic components of a licensed, preventative vaccine, which contains aluminum hydroxide as adjuvant. Herein, we report that glycoengineering of N-glycosylated HBsAgS to generate hyper-glycosylated VLPs display an enhanced immunogenicity relative to the wild type (WT) HBsAgS VLPs when expressed in FreeStyle HEK 293F cells. Comparative mass spectrometry-based N-glycan profiling, gel electrophoresis, and immunoassays demonstrated that WT and hyper-glycosylated HBsAgS VLPs contain the same type and distribution of N-glycan structures, but the latter shows a higher glycan abundance per protein mass. The antigenic integrity of the modified VLPs was also shown to be retained. To assess whether hyper-glycosylated VLPs induce an enhanced immune response in the presence of the adjuvant aluminum hydroxide, the anti-HBV surface antigen (anti-HBsAgS) antibody response was monitored in BALB/c mice, subcutaneously injected with different VLP derivatives. In the absence and presence of adjuvant, hyper-glycosylated VLPs showed an enhanced immunogenicity compared to WT VLPs. The ability of hyper-glycosylated VLPs to promote potent anti-HBsAgS immune responses compared to VLPs with a native N-glycan level as well as non-glycosylated, yeast-derived HBsAgS VLPs opens exciting avenues for generating more efficacious VLP-based vaccines against hepatitis B and improved HBsAgS VLP carrier platforms using glycoengineering.