Predictors of morbidity and mortality in COVID-19.

The mortality of COVID-19 patients is increasing in logarithmic fashion and is mostly observed in older age people and patients having history of chronic ailments like chronic obstructive pulmonary disease (COPD), hypertension, diabetes, cardiovascular & cerebrovascular dysfunction, compromised immunity, renal comorbidities, hepatic, obesity problems etc., and recently investigated thrombotic complications. The molecular underpinnings linking the chronic human diseases with COVID-19 related morbidity and mortality are evolving and poorly understood. The aim of the present review is to discuss the mortality and morbidity in COVID-19 in relation to preexisting comorbidities across the globe, upcoming molecular mechanisms associated with expression profile of ACE2 and viral load, evolving pathophysiology of COVID-19 with special reference to thrombotic complication ('Storm of Blood Clots') and related predictive markers. The levels of plasminogen/plasmin in comorbid diseases of COVID-19 have been elaborated in the framework of risk and benefits of fibrinolysis in COVID-19. We have also attempted to discuss the puzzle of prescribing ARBs and ACEI drugs in COVID-19 management which are routinely prescribed for the management of hypertension in COVID-19 patients. A focused discourse on risk of cardiovascular complications and diabetes in concert with COVID-19 pathogenesis has been presented along with dynamics of SARS-CoV-2 induced immune dysfunctions in COVID-19 patients.

Suggestions on cleavage embryo and blastocyst vitrification/transfer based on expression profile of ACE2 and TMPRSS2 in current COVID-19 pandemic.

An outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is leading to an unprecedented worldwide health crisis. SARS-CoV-2 cell entry depends on ACE2 and TMPRSS2. Our objectives are to analysis the expression profile of ACE2 and TMPRSS2 in human spermatogenic cells, follicle cells, and preimplantation embryos, thereby providing mechanistic insights into viral entry and viral impact on reproduction. We found that ACE2 is mainly expressed during gametogenesis in spermatogonia and oocytes of antral follicles, granulosa cells of antral follicles and pre-ovulatory follicles, while TMPRSS2 almost has no expression in spermatogenic cells, oocytes or granulosa cells. In preimplantation embryos, ACE2 is expressed in early embryos before eight-cell stage, and trophectoderm of late blastocysts, while TMPRSS2 initiates its robust expression in late blastocyst stage. ACE2 and TMPRSS2 only show significant co-expression in trophectoderm of late blastocysts in all above cell types. We speculate that trophectoderm of late blastocysts is susceptible to SARS-CoV-2, and that the chance of SARS-CoV-2 being passed on to offspring through gametes is very low. Therefore, we propose that fertility preservation for COVID-19 patients is relatively safe and rational. We also recommend embryo cryopreservation and embryo transfer into healthy recipient mother at cleavage stage instead of blastocyst stage. Moreover, we unexpectedly found that co-expression pattern of ACE2 and TMPRSS2 in oocytes and preimplantation embryos in human, rhesus monkey and mouse are totally different, so animal models have significant limitations for evaluating transmission risk of SARS-CoV-2 in reproduction.

Age-related differences in the immune response could contribute to determine the spectrum of severity of COVID-19.

Coronavirus disease 2019 (COVID-19), can present with a wide spectrum of severity. Elderly patients with cardiac, pulmonary and metabolic comorbidities are more likely to develop the severe manifestations of COVID-19, which are observed in less than 5% of the pediatric patients. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is able to induce an immune impairment and dysregulation, finally resulting in the massive release of inflammatory mediators, strongly contributing to the pulmonary and systemic manifestations in COVID-19. In children, the immune dysregulation following SARS-CoV-2 can also be responsible of a severe disease phenotype defined as multisystem inflammatory syndrome in children. As the immune system undergoes a complex process of maturation from birth to adult age, differences in the immune and inflammatory response could have a significant impact in determining the spectrum of severity of COVID-19. Indeed, children show a higher ability to respond to viral infections and a reduced baseline pro-inflammatory state compared with elderly patients. Age and comorbidities contribute to disease severity through immune-mediated mechanisms, since they are associated with a chronic increase of pro-inflammatory mediators, and cause an enhanced susceptibility to develop an immune dysregulation following SARS-CoV-2 infection. Also the expression of ACE2, the receptor of SARS-CoV-2, varies with age, and is linked to the immune and inflammatory response through a complex, and not completely elucidated, network. This paper reviews the peculiar immunopathogenic aspects of COVID-19, with a focus on the differences between adult and pediatric patients.

SARS-CoV-2 infects and replicates in cells of the human endocrine and exocrine pancreas.

Infection-related diabetes can arise as a result of virus-associated ß-cell destruction. Clinical data suggest that the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), causing the coronavirus disease 2019 (COVID-19), impairs glucose homoeostasis, but experimental evidence that SARS-CoV-2 can infect pancreatic tissue has been lacking. In the present study, we show that SARS-CoV-2 infects cells of the human exocrine and endocrine pancreas ex vivo and in vivo. We demonstrate that human ß-cells express viral entry proteins, and SARS-CoV-2 infects and replicates in cultured human islets. Infection is associated with morphological, transcriptional and functional changes, including reduced numbers of insulin-secretory granules in ß-cells and impaired glucose-stimulated insulin secretion. In COVID-19 full-body postmortem examinations, we detected SARS-CoV-2 nucleocapsid protein in pancreatic exocrine cells, and in cells that stain positive for the ß-cell marker NKX6.1 and are in close proximity to the islets of Langerhans in all four patients investigated. Our data identify the human pancreas as a target of SARS-CoV-2 infection and suggest that ß-cell infection could contribute to the metabolic dysregulation observed in patients with COVID-19.

SARS-CoV-2 infects and replicates in cells of the human endocrine and exocrine pancreas.

Infection-related diabetes can arise as a result of virus-associated ß-cell destruction. Clinical data suggest that the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), causing the coronavirus disease 2019 (COVID-19), impairs glucose homoeostasis, but experimental evidence that SARS-CoV-2 can infect pancreatic tissue has been lacking. In the present study, we show that SARS-CoV-2 infects cells of the human exocrine and endocrine pancreas ex vivo and in vivo. We demonstrate that human ß-cells express viral entry proteins, and SARS-CoV-2 infects and replicates in cultured human islets. Infection is associated with morphological, transcriptional and functional changes, including reduced numbers of insulin-secretory granules in ß-cells and impaired glucose-stimulated insulin secretion. In COVID-19 full-body postmortem examinations, we detected SARS-CoV-2 nucleocapsid protein in pancreatic exocrine cells, and in cells that stain positive for the ß-cell marker NKX6.1 and are in close proximity to the islets of Langerhans in all four patients investigated. Our data identify the human pancreas as a target of SARS-CoV-2 infection and suggest that ß-cell infection could contribute to the metabolic dysregulation observed in patients with COVID-19.

Angiotensin-converting enzyme 2 expression in COPD and IPF fibroblasts: the forgotten cell in COVID-19.

The COVID-19 pandemic is associated with severe pneumonia and acute respiratory distress syndrome leading to death in susceptible individuals. For those who recover, post-COVID-19 complications may include development of pulmonary fibrosis. Factors contributing to disease severity or development of complications are not known. Using computational analysis with experimental data, we report that idiopathic pulmonary fibrosis (IPF)- and chronic obstructive pulmonary disease (COPD)-derived lung fibroblasts express higher levels of angiotensin-converting enzyme 2 (ACE2), the receptor for SARS-CoV-2 entry and part of the renin-angiotensin system that is antifibrotic and anti-inflammatory. In preclinical models, we found that chronic exposure to cigarette smoke, a risk factor for both COPD and IPF and potentially for SARS-CoV-2 infection, significantly increased pulmonary ACE2 protein expression. Further studies are needed to understand the functional implications of ACE2 on lung fibroblasts, a cell type that thus far has received relatively little attention in the context of COVID-19.

COVID-19-Associated Nonocclusive Fibrin Microthrombi in the Heart.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its resultant clinical presentation, coronavirus disease 2019 (COVID-19), is an emergent cause of mortality worldwide. Cardiac complications secondary to this infection are common; however, the underlying mechanisms of such remain unclear. A detailed cardiac evaluation of a series of individuals with COVID-19 undergoing postmortem evaluation is provided, with 4 aims: (1) describe the pathological spectrum of the myocardium; (2) compare with an alternate viral illness; (3) investigate angiotensin-converting enzyme 2 expression; and (4) provide the first description of the cardiac findings in patients with cleared infection. METHODS: Study cases were identified from institutional files and included COVID-19 (n=15: 12 active, 3 cleared), influenza A/B (n=6), and nonvirally mediated deaths (n=6). Salient information was abstracted from the medical record. Light microscopic findings were recorded. An angiotensin-converting enzyme 2 immunohistochemical H-score was compared across cases. Viral detection encompassed SARS-CoV-2 immunohistochemistry, ultrastructural examination, and droplet digital polymerase chain reaction. RESULTS: Male sex was more common in the COVID-19 group (P=0.05). Nonocclusive fibrin microthrombi (without ischemic injury) were identified in 16 cases (12 COVID-19, 2 influenza, and 2 controls) and were more common in the active COVID-19 cohort (P=0.006). Four active COVID-19 cases showed focal myocarditis, whereas 1 case of cleared COVID-19 showed extensive disease. Arteriolar angiotensin-converting enzyme 2 endothelial expression was lower in COVID-19 cases than in controls (P=0.004). Angiotensin-converting enzyme 2 myocardial expression did not differ by disease category, sex, age, or number of patient comorbidities (P=0.69, P=1.00, P=0.46, P=0.65, respectively). SARS-CoV-2 immunohistochemistry showed nonspecific staining, whereas ultrastructural examination and droplet digital polymerase chain reaction were negative for viral presence. Four patients (26.7%) with COVID-19 had underlying cardiac amyloidosis. Cases with cleared infection had variable presentations. CONCLUSIONS: This detailed histopathologic, immunohistochemical, ultrastructural, and molecular cardiac series showed no definitive evidence of direct myocardial infection. COVID-19 cases frequently have cardiac fibrin microthrombi, without universal acute ischemic injury. Moreover, myocarditis is present in 33.3% of patients with active and cleared COVID-19 but is usually limited in extent. Histological features of resolved infection are variable. Cardiac amyloidosis may be an additional risk factor for severe disease.

Structural Variability, Expression Profile, and Pharmacogenetic Properties of TMPRSS2 Gene as a Potential Target for COVID-19 Therapy.

The human serine protease serine 2 TMPRSS2 is involved in the priming of proteins of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and represents a possible target for COVID-19 therapy. The TMPRSS2 gene may be co-expressed with SARS-CoV-2 cell receptor genes angiotensin-converting enzyme 2 (ACE2) and Basigin (BSG), but only TMPRSS2 demonstrates tissue-specific expression in alveolar cells according to single-cell RNA sequencing data. Our analysis of the structural variability of the TMPRSS2 gene based on genome-wide data from 76 human populations demonstrates that a functionally significant missense mutation in exon 6/7 in the TMPRSS2 gene is found in many human populations at relatively high frequencies, with region-specific distribution patterns. The frequency of the missense mutation encoded by rs12329760, which has previously been found to be associated with prostate cancer, ranged between 10% and 63% and was significantly higher in populations of Asian origin compared with European populations. In addition to single-nucleotide polymorphisms, two copy number variants were detected in the TMPRSS2 gene. A number of microRNAs have been predicted to regulate TMPRSS2 and BSG expression levels, but none of them is enriched in lung or respiratory tract cells. Several well-studied drugs can downregulate the expression of TMPRSS2 in human cells, including acetaminophen (paracetamol) and curcumin. Thus, the interactions of TMPRSS2 with SARS-CoV-2, together with its structural variability, gene-gene interactions, expression regulation profiles, and pharmacogenomic properties, characterize this gene as a potential target for COVID-19 therapy.

Lung Expression of Human Angiotensin-Converting Enzyme 2 Sensitizes the Mouse to SARS-CoV-2 Infection.

Preclinical mouse models that recapitulate some characteristics of coronavirus disease (COVID-19) will facilitate focused study of pathogenesis and virus-host responses. Human agniotensin-converting enzyme 2 (hACE2) serves as an entry receptor for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to infect people via binding to envelope spike proteins. Herein we report development and characterization of a rapidly deployable COVID-19 mouse model. C57BL/6J (B6) mice expressing hACE2 in the lung were transduced by oropharyngeal delivery of the recombinant human adenovirus type 5 that expresses hACE2 (Ad5-hACE2). Mice were infected with SARS-CoV-2 at Day 4 after transduction and developed interstitial pneumonia associated with perivascular inflammation, accompanied by significantly higher viral load in lungs at Days 3, 6, and 12 after infection compared with Ad5-empty control group. SARS-CoV-2 was detected in pneumocytes in alveolar septa. Transcriptomic analysis of lungs demonstrated that the infected Ad5-hACE mice had a significant increase in IFN-dependent chemokines Cxcl9 and Cxcl10, and genes associated with effector T-cell populations including Cd3 g, Cd8a, and Gzmb. Pathway analysis showed that several Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were enriched in the data set, including cytokine-cytokine receptor interaction, the chemokine signaling pathway, the NOD-like receptor signaling pathway, the measles pathway, and the IL-17 signaling pathway. This response is correlative to clinical response in lungs of patients with COVID-19. These results demonstrate that expression of hACE2 via adenovirus delivery system sensitized the mouse to SARS-CoV-2 infection and resulted in the development of a mild COVID-19 phenotype, highlighting the immune and inflammatory host responses to SARS-CoV-2 infection. This rapidly deployable COVID-19 mouse model is useful for preclinical and pathogenesis studies of COVID-19.

Neonatal hyperoxia enhances age-dependent expression of SARS-CoV-2 receptors in mice.

The severity of COVID-19 lung disease is higher in the elderly and people with pre-existing co-morbidities. People who were born preterm may be at greater risk for COVID-19 because their early exposure to oxygen (hyperoxia) at birth increases the severity of respiratory viral infections. Hyperoxia at birth increases the severity of influenza A virus infections in adult mice by reducing the number of alveolar epithelial type 2 (AT2) cells. Since AT2 cells express the SARS-CoV-2 receptors angiotensin converting enzyme (ACE2) and transmembrane protease/serine subfamily member 2 (TMPRSS2), their expression should decline as AT2 cells are depleted by hyperoxia. Instead, ACE2 was detected in airway Club cells and endothelial cells at birth, and then AT2 cells at one year of age. Neonatal hyperoxia stimulated expression of ACE2 in Club cells and in AT2 cells by 2 months of age. It also stimulated expression of TMPRSS2 in the lung. Increased expression of SARS-CoV-2 receptors was blocked by mitoTEMPO, a mitochondrial superoxide scavenger that reduced oxidative stress and DNA damage seen in oxygen-exposed mice. Our finding that hyperoxia enhances the age-dependent expression of SARS-CoV-2 receptors in mice helps explain why COVID-19 lung disease is greater in the elderly and people with pre-existing co-morbidities.

Gender Disaggregation in COVID-19 and Increased Male Susceptibility.

Novel coronavirus disease 2019 (COVID-19) is a growing public health crisis. Despite initial focus on the elderly population with comorbidities, it seems that large studies from the worst affected countries follow a sex-disaggregation pattern. Analysis of available data showed marked variations in reported cases between males and females among different countries with higher mortality in males.  At this early stage of the pandemic, medical datasets at the individual level are not available; therefore, it is challenging to conclude how different factors have impacted COVID-19 susceptibility. Thus, in the absence of patients' level data, we attempted to provide a theoretical description of how other determinants have affected COVID-19 susceptibility in males compared to females.  In this article, we have identified and discussed possible biological and behavioral factors that could be responsible for the increased male susceptibility. Biological factors include - an absence of X-chromosomes (a powerhouse for immune-related genes), a high level of testosterone that inhibits antibody production, and the presence of Angiotensin-converting enzyme 2 (ACE2) receptors that facilitate viral replication. Similarly, behavioral factors constitute - higher smoking and alcohol consumptions, low level of handwashing practices, and high-risk behavior like non-adherence to health services and reluctance to follow public health measures in males. Keywords: COVID-19; gender; males; sex disaggregation; susceptibility.

Kidney ACE2 expression: Implications for chronic kidney disease.

Angiotensin-converting enzyme 2 (ACE2) has been implicated in the pathogenesis of chronic kidney disease (CKD) and is a membrane receptor for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus responsible for coronavirus disease (COVID-19), whereas transmembrane protease, serine 2 (TMPRSS2) is involved in viral attachment. Together, tissue expression of ACE2 and TMPRSS2 may determine infection. Sex, age, body mass index (BMI), and CKD are clinical risk factors for COVID-19 severity, but the relationships between kidney ACE2 and TMPRSS2 expression and these clinical variables are unknown. Accordingly, we obtained renal tubulointerstitial and glomerular microarray expression data and clinical variables from healthy living donors (HLD) and patients with CKD from the European Renal cDNA Bank. ACE2 expression was similar in the tubulointerstitium of the two groups, but greater in females than males in HLD (P = 0.005) and CKD (P < 0.0001). ACE2 expression was lower in glomeruli of CKD patients compared to HLD (P = 0.0002) and lower in males than females. TMPRSS2 expression was similar in the tubulointerstitium but lower in glomeruli of CKD patients compared to HLD (P < 0.0001). There was a strong relationship between ACE2 and TMPRSS2 expression in the glomerulus (r = 0.51, P < 0.0001). In CKD, there was a relationship between tubulointerstitial ACE2 expression and estimated glomerular filtration rate (r = 0.36, P < 0.0001) and age (r = -0.17, P = 0.03), but no relationship with BMI. There were no relationships between TMPRSS2 expression and clinical variables. Genes involved in inflammation (CCL2, IL6, and TNF) and fibrosis (COL1A1, TGFB1, and FN1) were inversely correlated with ACE2 expression. In summary, kidney expression of ACE2 and TMPRSS2 differs in HLD and CKD. ACE2 is related to sex and eGFR. ACE2 is also associated with expression of genes implicated in inflammation and fibrosis.

Tissue-specific and interferon-inducible expression of nonfunctional ACE2 through endogenous retroelement co-option.

Angiotensin-converting enzyme 2 (ACE2) is an entry receptor for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and a regulator of several physiological processes. ACE2 has recently been proposed to be interferon (IFN) inducible, suggesting that SARS-CoV-2 may exploit this phenomenon to enhance viral spread and questioning the efficacy of IFN treatment in coronavirus disease 2019. Using a recent de novo transcript assembly that captured previously unannotated transcripts, we describe a new isoform of ACE2, generated by co-option of intronic retroelements as promoter and alternative exon. The new transcript, termed MIRb-ACE2, exhibits specific expression patterns across the aerodigestive and gastrointestinal tracts and is highly responsive to IFN stimulation. In contrast, canonical ACE2 expression is unresponsive to IFN stimulation. Moreover, the MIRb-ACE2 translation product is a truncated, unstable ACE2 form, lacking domains required for SARS-CoV-2 binding and is therefore unlikely to contribute to or enhance viral infection.

GB-2 inhibits ACE2 and TMPRSS2 expression: In vivo and in vitro studies.

After the first case of Coronavirus disease 2019 (COVID-19) was reported in Wuhan, COVID-19 has rapidly spread to almost all parts of world. Angiotensin converting enzyme 2 (ACE2) receptor can bind to spike protein of SARS-CoV-2. Then, the spike protein of SARS-CoV-2 can be cleaved and activated by transmembrane protease, serine 2 (TMPRSS2) of the host cells for SARS-CoV-2 infection. Therefore, ACE2 and TMPRSS2 are potential antiviral targets for treatment of prevention of SARS-CoV-2 infection. In this study, we discovered that 10-250 µg/mL of GB-2, from Tian Shang Sheng Mu of Chiayi Puzi Peitian Temple, can inhibit ACE2 mRNA expression and ACE2 and TMPRSS2 protein expression in HepG2 and 293 T cells without cytotoxicity. GB-2 treatment could decrease ACE2 and TMPRSS2 expression level of lung tissue and kidney tissue without adverse effects, including nephrotoxicity and hepatotoxicity, in animal model. In the compositions of GB-2, we discovered that 50 µg/mL of theaflavin could inhibit protein expression of ACE2 and TMPRSS2. Theaflavin could inhibit the mRNA expression of ACE2. In conclusion, our results suggest that GB-2 and theaflavin could act as potential compounds for ACE2 and TMPRSS2 inhibitors in the further clinical study.

ACE2 & TMPRSS2 Expressions in Head & Neck Tissues: A Systematic Review.

To review the data regarding the expression of angiotensin converting enzyme-2 (ACE2) and transmembrane protease serine-2 (TMPRSS2) in head and neck tissue. Scopus, Cochrane Library, Medrxiv, Google Scholar and PubMED/MEDLINE were searched by four independent investigators for studies investigating ACE2 or TMPRSS2 expressions in head and neck tissues. The following outcomes were considered: sample origin (animal versus human); detection method; anatomical location and cell types. PRISMA checklist and modified population, intervention, comparison, outcome, timing and setting (PICOTS) framework were used to perform the review. Of the 24 identified studies, 17 met our inclusion criteria. Thirteen studies were conducted during the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) pandemic. ACE2 and TMPRSS2 were expressed in oral, pharyngeal, sinusonasal human mucosa. The following cell types expressed ACE2: basal, apical, goblet, minor salivary, and endothelial cells. TMPRSS2 was found in goblet and apical respiratory cells. ACE2 and TMPRSS2 were found in the olfactory region, especially in sustentacular non-neural and neural stem cells. Animal studies suggested that ACE2 expression may vary regarding age. There was an important heterogeneity between studies in the methods used to detect ACE2 and TMPRSS2, leading to a potential identification bias. The SARS-CoV-2 receptors, ACE2 and TMPRSS2, are both expressed in many head and neck tissues, enabling the viral entry into the host organism.

COVID-19: the role of excessive cytokine release and potential ACE2 down-regulation in promoting hypercoagulable state associated with severe illness.

The novel coronavirus disease (COVID-19) has become a universally prevalent infectious disease. The causative virus of COVID-19 is severe acute respiratory syndrome coronavirus type 2. Recent retrospective clinical studies have established a significant association between the incidence of vascular thrombotic events and the severity of COVID-19. The enhancement in serum levels of markers that reflect a hypercoagulable state has been suggested to indicate a poor prognosis. Therefore, at present, it is crucial to understand the mechanisms that foster the hypercoagulable state in COVID-19. Over-activated inflammatory response, which is manifested as excessive cytokine release in COVID-19 patients, is also associated with COVID-19 severity. This review discusses the immuno-pathological basis of the excessive cytokine release in COVID-19. Besides, this article reviews the role of pro-inflammatory or anti-inflammatory cytokines, whose significant elevations in their serum levels have been consistently detected in multiple different clinical studies, in promoting the hypercoagulable state. Since the expression of angiotensin-converting enzyme 2 (ACE2) is potentially down-regulated in COVID-19, as proposed by a recent bio-informatic analysis, mechanisms through which reduced ACE2 expressions promote vascular thrombosis are summarized. In addition, the reciprocal-enhancing effects of the excessive cytokine release and the downregulated ACE2 expression on their pro-thrombotic activities are further discussed. Here, based on currently available evidence, we review the pathogenic mechanisms of the hypercoagulable state associated with severe cases of COVID-19 to give insights into prevention and treatment of the vascular thrombotic events in COVID-19.