Pulmonary influenza infection promotes the awakening of dormant metastatic breast cancer cells

Breast cancer is the most common form of cancer and the second cancer-causing death in females. Although remission rates are high if detected early, survival rates drop substantially when breast cancer becomes metastatic. The most common sites of metastatic breast cancer are bone, liver and lung. Respiratory viral infections inflict illnesses on countless people. The latest pandemic caused by the respiratory virus, SARS-CoV-2, has infected more than 600 million worldwide, with documented COVID-related death upward of 1 million in the United States alone. Respiratory viral infections result in increased inflammation with immune cell influx and expansion to facilitate viral clearance. Prior studies have shown that inflammation, including through neutrophils, can contribute to dormant cancer cells reawakening and outgrowth. Moreover, inhibition of IL6 has been shown to decrease breast cancer lung metastasis in mouse models. However, how respiratory viral infections contribute to breast cancer lung metastasis remains to be unraveled. Using MMTV/PyMT and MMTV/NEU mouse models of breast cancer lung metastasis and influenza A virus as a model respiratory virus, we demonstrated that acute influenza infection and the accompanying inflammation and immune cell influx awakens and dramatically increased proliferation and expansion of dormant disseminated cancer cells (DCC) in the lungs. Acute influenza infection leads to immune influx and expansion, including neutrophils and macrophages, with increased proportion of MHCII+ macrophages in early time points, and a sustained decrease in CD206+ macrophages starting 6 days post-infection until 28 days after the initial infection. Additionally, we observed a sustained accumulation of CD4+ T cells around expanding tumor cells for as long as 28 days after the infection. Notably, neutrophil depletion or IL6 knockout reversed the flu-induced dormant cell expansion in the lung. Finally, awakened DCC exhibited downregulation of vimentin immunoreactivity, suggesting a role for phenotypic plasticity in DCC outgrowth following viral infection. In conclusion, we show that respiratory viral infections awaken and increase proliferation of dormant breast cancer cells in the lung, and that depletion of neutrophils or blocking IL6 reverses influenza-induced dormant cell awakening and proliferation.

Enhancer deregulation inTET2-mutant clonal hematopoiesis is associated with increased COVID-19 severity and mortality

Background: COVID-19 causes significant morbidity and mortality, albeit with considerable heterogeneity among affected individuals. It remains unclear which host factors determine disease severity and survival. Given the propensity of clonal hematopoiesis (CH) to promote inflammation in healthy individuals, we investigated its effect on COVID-19 outcomes. Method(s): We performed a multi-omics interrogation of the genome, epigenome, transcriptome, and proteome of peripheral blood mononuclear cells from COVID-19 patients (n=227). We obtained clinical data, laboratory studies, and survival outcomes. We determined CH status and TET2-related DNA methylation. We performed single-cell proteogenomics to understand clonal composition in relation to cell phenotype. We interrogated single-cell gene expression in isolation and in conjunction with DNA accessibility. We integrated these multi-omics data to understand the effect of CH on clonal composition, gene expression, methylation of cis-regulatory elements, and lineage commitment in COVID-19 patients. We performed shRNA knockdowns to validate the effect of one candidate transcription factor in myeloid cell lines. Result(s): The presence of CH was strongly associated with COVID-19 severity and all-cause mortality, independent of age (HR 3.48, 95% CI 1.45-8.36, p=0.005). Differential methylation of promoters and enhancers was prevalent in TET2-mutant, but not DNMT3A-mutant CH. TET2- mutant CH was associated with enhanced classical/intermediate monocytosis and single-cell proteogenomics confirmed an enrichment of TET2 mutations in these cell types. We identified celltype specific gene expression changes associated with TET2 mutations in 102,072 single cells (n=34). Single-cell RNA-seq confirmed the skewing of hematopoiesis towards classical and intermediate monocytes and demonstrated the downregulation of EGR1 (a transcription factor important for monocyte differentiation) along with up-regulation of the lncRNA MALAT1 in monocytes. Combined scRNA-/scATAC-seq in 43,160 single cells (n=18) confirmed the skewing of hematopoiesis and up-regulation of MALAT1 in monocytes along with decreased accessibility of EGR1 motifs in known cis-regulatory elements. Using myeloid cell lines for functional validation, shRNA knockdowns of EGR1 confirmed the up-regulation of MALAT1 (in comparison to wildtype controls). Conclusion(s): CH is an independent prognostic factor in COVID-19 and skews hematopoiesis towards monocytosis. TET2-mutant CH is characterized by differential methylation and accessibility of enhancers binding myeloid transcriptions factors including EGR1. The ensuing loss of EGR1 expression in monocytes causes MALAT1 overexpression, a factor known to promote monocyte differentiation and inflammation. These data provide a mechanistic insight to the adverse prognostic impact of CH in COVID-19.

Respiratory Syncytial Virus (RSV) infections in children: recent concepts in immunopathogenesis and diagnostic dilemma

Respiratory viral infections are important cause of morbidity and mortality in early life. The relative influence of host and viral factors possibly contribute to the disease pathogenesis. Predisposing conditions like prematurity, Low birth weight and congenital heart diseases etc. have been incriminated in the disease progression. The development of cough, wheezing, and tachypnea, usually peaking on days 4 to 5, go parallel with host cytokine responses and viral load. Various host cytokines, chemokines and molecules involved in the immune response against RSV infection might be responsible for the outcome of the disease process. Nasopharyngeal aspirates (NPAs) from children (n = 349) between 2013-2017 were subjected for IL-17A, IFN-gamma, TNF-alpha, IL-10, IL-6 levels by CBA and MMP-9 and TIMP-1 levels by ELISA. The viral load in RSV positive samples and cytokine levels were correlated with the WHO criteria for acute lower respiratory tract illness (ALRTI). RSV viral load, Pro-inflammatory cytokine (TNF-alpha) levels in severe ALRTI patients were significantly higher than the ALRTI patients [p<0.001]. Whereas Th17 cytokine (IL-17) was found to be significantly higher (p<0.05) in ALRTI patients than severe patients. MMP-9 is secreted in higher levels in severe ALRTI patients (n = 77) in comparison to Acute LRTI patients (n = 35) with an increase of thirty seven fold (p<0.001). Thus, the study highlights the role of TNF -alpha, IL-17 and Th2 cytokine biasness in the pathogenesis of RSV disease with the possible contribution of higher MMP-9/TIMP-1 ratio as a bad prognostic marker towards disease severity. To study the gene expression of autophagy and mTOR signalling pathways in RSV infected children with ALRTI. Nasopharyngeal aspirate (NPA) samples (n = 145) from children suffering from ALRTI were subjected for detection of RSV (Oct 2019 to March 2020). Semi-quantitative gene expression analysis for 5 representative genes each of mTOR signalling and autophagy pathway were performed in respiratory tract epithelial cells using 25 RSV positive cases and 10 healthy controls subjects. Autophagy gene expression analysis revealed significant upregulation in NPC1 and ATG3 autophagy genes. mTOR, AKT1 and TSC1 genes of mTOR pathway were significantly down-regulated in RSV positive patients except RICTOR gene which was significantly upregulated. Thus, survival of RSV within autophagosome might have been facilitated by upregulation of autophagy and downregulation of mTOR signalling genes. To assess the impact of SARS-CoV2 pandemic on RSV, samples were collected from children with ALRTIs admitted to emergency, PICU and indoor admissions during pre-pandemic period (October 2019 to February 2020;n = 166) and during COVID-19 Pandemic (July 2021 to July 2022;n = 189, SARS-CoV2 negative). These NP swabs were analyzed for pdm InfA H1N1, InfA H3N2, Inf B, RSV, hMPV, hBoV, hRV, PIV-2 and PIV-3 by PCR. Higher proportion of children with ALRTIs have had virus/es isolated during pre-pandemic period than during pandemic period (p<0.001). During pre-pandemic period, significantly higher proportion of children had RSV positivity (p<0.001);and significantly lower positivity for hRV (p<0.05), hMPV (p<0.05), and hBoV (p <= 0.005). The occurrence of COVID-19 pandemic has significantly impacted the frequency and pattern of detection of RSV among hospitalized children with LRTIs. RSV Fusion protein plays a critical role in the entry of the virus into the host cell by initiating the fusion of host and viral membranes. It happens to be a target of neutralizing antibodies paving the way as a vaccine candidate. Hence effort was made to introduce point mutation in hRSV fusion protein which can confer stability in its prefusion form. In-silico a stable structure of RSV fusion protein was generated making it a potential vaccine candidate. The timely diagnosis of RSV infection in this population is important for initiating therapy and instituting appropriate infection prevention measures. Serological testing is not widely used for the diagnosis of RSV. C ll Cultures including shell vial culture were used for RSV diagnosis. However, culture approaches lack sensitivity, often quite significantly, compared to nucleic acid amplification assays for the diagnosis of RSV infections. Molecular multiplex assays now offer increased sensitivity for a more accurate diagnosis. However issues with the use of these types of commercial panel assays include the requirement for substantial training, quality systems, and infrastructure to maintain and run these assays and many a times identification of viruses where the true pathogenic potential of those multiple viruses are debatable. Studies are available with laboratory- developed nucleic acid amplification test systems for the detection of RSVA and RSVB in clinical specimens either by PCRbased technologies or RT-LAMP. Gene targets of laboratory-developed molecular assays point towards M gene and the N gene in RSVA and -B with the benefits of flexibility to modify assays when targets are under evolutionary pressure to change, as well as a perceived initial low cost to carry out testing.

Early Inflammatory Profiles Predict Maximal Disease Severity in Covid-19

Background: Better understanding of host inflammatory changes that precede development of severe COVID-19 could improve delivery of available antiviral and immunomodulatory therapies, and provide insights for the development of new therapies. Method(s): In plasma from individuals with COVID-19, sampled <=10 days from symptom onset from the All-Ireland Infectious Diseases Cohort study, we measured 61 biomarkers, including markers of innate immune and T cell activation, coagulation, tissue repair, lung injury, and immune regulation. We used principal component analysis (PCA) and k-means clustering to derive biomarker clusters, and univariate and multivariate ordinal logistic regression to explore association between cluster membership and maximal disease severity, adjusting for risk factors for severe COVID-19, including age, sex, ethnicity, BMI, hypertension and diabetes. Result(s): From March 2020-April 2021, we included 312 individuals, (median (IQR) age 62 (48-77) years, 7 (4-9) days from symptom onset, 54% male) in the analysis. PCA and clustering derived 4 clusters. Compared to cluster 1, clusters 2-4 were significantly older and of higher BMI but there were no significant differences in sex or ethnicity. Cluster 1 had low levels of inflammation, cluster 2 had higher levels of markers of tissue repair and endothelial activation (EGF, VEGF, PDGF, TGFalpha, serpin E1 and p-selectin). Cluster 3 and 4 were both characterised by higher overall inflammation, but compared to cluster 4, cluster 3 had downregulation of growth factors, markers of endothelial activation, and immune regulation (IL10, PDL1), but higher alveolar epithelial injury markers (RAGE, ST2). In univariate analysis, compared to cluster 1, cluster 3 had the highest odds of severe disease (OR (95% CI) 9.02 (4.62-18.31), followed by cluster 4: 5.59 (2.75-11.72) then cluster 2: 4.5 (2.38-8.81), all p < 0.05). Cluster 3 remained most strongly associated with severe disease in fully adjusted analyses;cluster 3: OR(95% CI) 5.99 (2.69-13.35), cluster 2: 3.14 (1.54-6.42), cluster 4: 3.13 (1.36-7.19), all p< 0.05). Conclusion(s): Distinct early inflammatory profiles predicted maximal disease severity independent of known risk factors for severe COVID-19. A cluster characterised by downregulation of growth factor and endothelial markers and early evidence of alveolar injury was associated with highest risk of developing severe COVID19. Whether this reflects a dysregulated inflammatory response that could improve targeted treatment requires further study. Heatmap of biomarker derived clusters and forest plot of association between clusters and disease severity. A: Heatmap demonstrating differences in biomarkers between clusters B: Forest plot demonstrating odds ratio of specific clusters for progressing to moderate or severe disease (reference Cluster 1), calculated using ordinal logistic regression. Odds ratio (95% CI) presented as unadjusted and fully adjusted (for age, sex, ethnicity, BMI, hypertension, diabetes, immunosuppression, smoking and baseline anticoagulant use). Maximal disease severity graded per the WHO severity scale.

Targeting Angiotensin-converting Enzyme 2 (ACE2) for the Discovery of Anticoronaviral Drugs

Coronaviruses are a leading cause of emerging life-threatening diseases, as evidenced by the ongoing coronavirus disease pandemic (COVID-19). According to complete genome sequence analysis reports, severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), which causes COVID-19, has a sequence identity highly similar to the earlier severe acute respiratory syndrome coronavirus (SARS-CoV). The SARS-CoV-2 has the same mode of transmission, replication, and pathogenicity as SARS-CoV. The SARS-CoV-2 spike protein's receptor-binding domain (RBD) binds to host angiotensin-converting enzyme-2 (ACE2). The ACE2 is overexpressed in various cells, most prominently epithelial cells of the lung (surface of type 1 and 2 pneumocytes), intestine, liver, kidney, and nervous system. As a result, these organs are more vulnerable to SARS-CoV-2 infection. Furthermore, renin-angiotensin system (RAS) blockers, which are used to treat cardiovascular diseases, intensify ACE2 expression, leading to an increase in the risk of COVID-19. ACE2 hydrolyzes angioten-sin-II (carboxypeptidase) to heptapeptide angiotensin (1-7) and releases a C-terminal amino acid. By blocking the interaction of spike protein with ACE2, the SARS-CoV-2 entry into the host cell and inter-nalization can be avoided. The pathogenicity of SARS-CoV-2 could be reduced by preventing the RBD from attaching to ACE2-expressing cells. Therefore, inhibition or down-regulation of ACE2 in host cells represents a therapeutic strategy to fight against COVID-19. However, ACE2 plays an essential role in the physiological pathway, protecting against hypertension, heart failure, myocardial infarction, acute respiratory lung disease, and diabetes. Given the importance of ACE's homeostatic role, targeting of ACE2 should be realized with caution. Above all, focusing on the SARS-CoV-2 spike protein and the ACE2 gene in the host cell is an excellent way to avoid viral mutation and resistance. The current review summarises the sequence analysis, structure of coronavirus, ACE2, spike protein-ACE2 complex, essential structural characteristics of the spike protein RBD, and ACE2 targeted approaches for anti-coronaviral drug design and development.Copyright © 2022 Bentham Science Publishers.

Evaluation of the protein C pathway in critically ill patients with severe COVID-19 as compared to bacterial sepsis

Introduction Endothelial dysfunction has been shown to play a role in severe COVID-19, the pathophysiology of which may be attributed to a myriad of factors including unmitigated immune and inflammatory response, viral-induced injury to the endothelium, end-stage organ failure, and coagulopathy. In addition, severe COVID-19 is most often seen in patients with multiple comorbidities, which themselves are often associated with endothelial dysfunction (such as myocardial and renal failure, as well as thrombotic disorders). However, the literature is still emerging on this topic and there appears to be no consensus on the extent to which endothelial dysfunction plays a role in severe COVID-19. Method The aim of this study was to assess the functionality of the endothelial protein C pathway in hospitalized patients > 18 years of age with severe COVID-19 as compared to those hospitalized with bacterial sepsis. COVID-19 (n = 31) and sepsis (n = 47) patients who were admitted to the ICU were assessed for rates of thrombin and activated protein C (APC) generation. Indirect markers of thrombin formation, including thrombin-antithrombin (TAT) complex, prothrombin fragment 1 + 2 (F1 + 2), as well as D-dimer, and protein C (PC) were measured additionally. Statistical analysis was performed via the Mann-Whitney test and a p value of < 0.05 was considered statistically significant. Results Plasma levels of free thrombin in COVID-19 and sepsis patients did not differ significantly, with (median, IQR) 0.59 (0.46-1.21) vs 0.57 (0.46-1.10) pmol/L, respectively. TAT was also increased at similar extent in both cohorts (192;111-325 pmol/L in COVID-19 patients, 148;73-213 pmol/L in sepsis patients), whereas F1 + 2 was higher in COVID-19 than in sepsis patients, with 850 (440-1940) vs 380 (130-620) pmol/L (p = 1.3 x 10-5). Interestingly, rates of APC formation did not significantly differ between the two groups, with 7.47 (1.99- 19.14) vs 9.87 (2.08-16.87) pmol/L ( Fig. 1). D-dimer and protein C were significantly higher in the COVID-19 patients than in those with sepsis (14.3 vs 8.1 mg/L, p = 0.01, and 92.9 % vs 58.5 %, p = 3x10-8, respectively). Conclusion We hypothesized that APC formation rates in response to thrombin formation would be significantly lower in patients with severe COVID-19 as compared to those with bacterial sepsis due to the well-known association between severe COVID-19 disease burden and endothelial dysfunction due to the downregulation of thrombomodulin expression. However, our results indicate that this may not be universally true in this patient population, as our observations suggest a largely intact functionality of the protein C pathway. Further studies are warranted to investigate the pathophysiology of severe COVID-19. (Figure Presented).

Research progress on the mechanism of multiple organ dysfunction in patients with COVID-19.

The pathogen of COVID-19 is 2019-nCoV, which belongs to the beta coronavirus. Angiotensin-converting enzyme 2 (ACE2) is the receptor of 2019-nCoV as the same of SARS-CoV. Most of the severe patients were the elderly with underlying diseases, which may be related to the decrease in the number of naive T cells. In addition to pulmonary symptoms, COVID-19 can also cause multiple organ dysfunction and even multiple organ failure (liver, nervous system, heart, kidney, etc.). Pathogenic mechanisms such as direct virus invasion, cytokine storm, endothelial cells damage, and down-regulation of ACE2 may play important roles in the severity of the disease.Copyright © 2021 Chinese Medical Association

Research progress on the mechanism of multiple organ dysfunction in patients with COVID-19.

The pathogen of COVID-19 is 2019-nCoV, which belongs to the beta coronavirus. Angiotensin-converting enzyme 2 (ACE2) is the receptor of 2019-nCoV as the same of SARS-CoV. Most of the severe patients were the elderly with underlying diseases, which may be related to the decrease in the number of naive T cells. In addition to pulmonary symptoms, COVID-19 can also cause multiple organ dysfunction and even multiple organ failure (liver, nervous system, heart, kidney, etc.). Pathogenic mechanisms such as direct virus invasion, cytokine storm, endothelial cells damage, and down-regulation of ACE2 may play important roles in the severity of the disease.Copyright © 2021 Chinese Medical Association

Research progress on the mechanism of multiple organ dysfunction in patients with COVID-19.

The pathogen of COVID-19 is 2019-nCoV, which belongs to the beta coronavirus. Angiotensin-converting enzyme 2 (ACE2) is the receptor of 2019-nCoV as the same of SARS-CoV. Most of the severe patients were the elderly with underlying diseases, which may be related to the decrease in the number of naive T cells. In addition to pulmonary symptoms, COVID-19 can also cause multiple organ dysfunction and even multiple organ failure (liver, nervous system, heart, kidney, etc.). Pathogenic mechanisms such as direct virus invasion, cytokine storm, endothelial cells damage, and down-regulation of ACE2 may play important roles in the severity of the disease.Copyright © 2021 Chinese Medical Association

Effect and mechanism of Qingre Xiaoyanning Tablet in treatment of coronavirus infection.

Objective To study the anti-coronavirus effect of Qingre Xiaoyanning Tablet (), and provide experimental basis for evaluating its prevention and treatment of coronavirus infection. Methods A total of 96 BALB/c mice with half male and half female were randomly divided into control group, model group, Lianhua Qingwen Capsules (, 0.546 g/kg) group and Qingre Xiaoyanning Tablet (8.72, 17.44, 34.89 g/kg) groups with 16 mice in each group. BALB/c mice were infected with ip cyclophosphamide combined with HCoV-229E coronavirus to establish a model of coronavirus infection. The therapeutic effect of Qingre Xiaoyanning Tablet was evaluated by body weight, lung index, viral load, hemagglutination titer and pathological changes in lung tissue of mice;Levels of interleukin-1beta (IL-1beta), IL-4, tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma) and vascular cell adhesion molecule-1 (VCAM-1) in alveolar lavage fluid were detected by ELISA;The proportion of macrophages, lymphocytes (CD3+, CD4+) and NK cells in lung tissue was detected by flow cytometry;Western blotting was used to detect Toll like receptor 4 (TLR4), myeloid differentiation factor 88 (MYD88), inhibitor kappa B kinase-beta (IKK-beta), inhibitor kappa B (IkappaB) and p-IkappaB protein expressions in lung tissue. Results Compared with model group, Qingre Xiaoyanning Tablet significantly increased the body weight of virus infected mice (P < 0.05, 0.01), decreased lung index and hemagglutination titer (P < 0.01), improved lung disease (P < 0.05), and significantly inhibited viral mRNA expression (P < 0.01);TNF-alpha, IL-1 beta and VCAM-1 levels in alveolar lavage fluid were decreased (P < 0.05, 0.01), IFN-gamma level was increased (P < 0.05);The percentage of macrophages was significantly decreased (P < 0.05, 0.01), percentage of CD3+, CD4+ lymphocytes and NK cells was increased (P < 0.01);MYD88, TLR4, IkappaB and IKK-beta protein expressions in lung tissue were significantly down regulated (P < 0.05, 0.01). Conclusion Qingre Xiaoyanning Tablet can inhibit the replication of coronavirus in vivo, reduce inflammatory reaction, protect lung tissue, and has obvious anti-coronavirus effect in vivo. Its mechanism may be related to the regulation of TLR4/MyD88/IKK/IkappaB signal pathway and improving immunity.Copyright © 2023 Editorial Office of Chinese Traditional and Herbal Drugs. All rights reserved.

Genomic regulatory network of human olfactory neuroepithelial cells infected with novel coronavirus (SARS-COV-2).

Objective To explore the genomic changes of human olfactory neuroepithelial cells after the novel coronavirus (SARS-COV-2) infecting the human body, and establish a protein-protein interaction (PPI) network of differentially expressed genes (DEGs), in order to understand the impact of SARS-COV-2 infection on human olfactory neuroepithelial cells, and provide reference for the prevention and treatment of new coronavirus pneumonia. Methods The public dataset GSE151973 was analyzed by NetworkAnalyst. DEGs were selected by conducting Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) signal pathway analysis. PPI network, DEGs-microRNA regulatory network, transcription factor-DEGs regulatory network, environmental chemicals-DEGs regulatory network, and drug-DEGs regulatory network were created and visualized by using Cytoscape 3.7.2. Results After SAR-COV-2 invading human olfactory neuroepithelial cells, part of the gene expression profile was significantly up-regulated or down-regulated. A total of 568 DEGs were found, including 550 up-regulated genes (96.8%) and 18 down-regulated genes (3.2%). DEGs were mainly involved in biological processes such as endothelial development and angiogenesis of the olfactory epithelium, and the expression of molecular functions such as the binding of the N-terminal myristylation domain. PPI network suggested that RTP1 and RTP2 were core proteins. MAZ was the most influential transcription factor. Hsa-mir-26b-5p had the most obvious interaction with DEGs regulation. Environmental chemical valproic acid and drug ethanol had the most influence on the regulation of DEG. Conclusion The gene expression of olfactory neuroepithelial cells is significantly up-regulated or down-regulated after infection with SAR-COV-2. SARS-CoV-2 may inhibit the proliferation and differentiation of muscle satellite cells by inhibiting the function of PAX7. RTP1 and RTP2 may resist SARS-CoV-2 by promoting the ability of olfactory receptors to coat the membrane and enhancing the ability of olfactory receptors to respond to odorant ligands. MAZ may regulate DEGs by affecting cell growth and proliferation. Micro RNA, environmental chemicals and drugs also play an important role in the anti-SAR-COV-2 infection process of human olfactory neuroepithelial cells.Copyright © 2022 Editorial Department of Journal of Shanghai Second Medical University. All rights reserved.

Inhibition of major histocompatibility complex-I antigen presentation by sarbecovirus ORF7a proteins.

Viruses employ a variety of strategies to escape or counteract immune responses, including depletion of cell surface major histocompatibility complex class I (MHC-I), that would ordinarily present viral peptides to CD8+ cytotoxic T cells. As part of a screen to elucidate biological activities associated with individual severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) viral proteins, we found that ORF7a reduced cell surface MHC-I levels by approximately fivefold. Nevertheless, in cells infected with SARS-CoV-2, surface MHC-I levels were reduced even in the absence of ORF7a, suggesting additional mechanisms of MHC-I down-regulation. ORF7a proteins from a sample of sarbecoviruses varied in their ability to induce MHC-I down-regulation and, unlike SARS-CoV-2, the ORF7a protein from SARS-CoV lacked MHC-I downregulating activity. A single amino acid at position 59 (T/F) that is variable among sarbecovirus ORF7a proteins governed the difference in MHC-I downregulating activity. SARS-CoV-2 ORF7a physically associated with the MHC-I heavy chain and inhibited the presentation of expressed antigen to CD8+ T cells. Specifically, ORF7a prevented the assembly of the MHC-I peptide loading complex and caused retention of MHC-I in the endoplasmic reticulum. The differential ability of ORF7a proteins to function in this way might affect sarbecovirus dissemination and persistence in human populations, particularly those with infection- or vaccine-elicited immunity.

Genomic regulatory network of human olfactory neuroepithelial cells infected with novel coronavirus (SARS-COV-2)

Objective To explore the genomic changes of human olfactory neuroepithelial cells after the novel coronavirus (SARS-COV-2) infecting the human body, and establish a protein-protein interaction (PPI) network of differentially expressed genes (DEGs), in order to understand the impact of SARS-COV-2 infection on human olfactory neuroepithelial cells, and provide reference for the prevention and treatment of new coronavirus pneumonia. Methods The public dataset GSE151973 was analyzed by NetworkAnalyst. DEGs were selected by conducting Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) signal pathway analysis. PPI network, DEGs-microRNA regulatory network, transcription factor-DEGs regulatory network, environmental chemicals-DEGs regulatory network, and drug-DEGs regulatory network were created and visualized by using Cytoscape 3.7.2. Results After SAR-COV-2 invading human olfactory neuroepithelial cells, part of the gene expression profile was significantly up-regulated or down-regulated. A total of 568 DEGs were found, including 550 up-regulated genes (96.8%) and 18 down-regulated genes (3.2%). DEGs were mainly involved in biological processes such as endothelial development and angiogenesis of the olfactory epithelium, and the expression of molecular functions such as the binding of the N-terminal myristylation domain. PPI network suggested that RTP1 and RTP2 were core proteins. MAZ was the most influential transcription factor. Hsa-mir-26b-5p had the most obvious interaction with DEGs regulation. Environmental chemical valproic acid and drug ethanol had the most influence on the regulation of DEG. Conclusion The gene expression of olfactory neuroepithelial cells is significantly up-regulated or down-regulated after infection with SAR-COV-2. SARS-CoV-2 may inhibit the proliferation and differentiation of muscle satellite cells by inhibiting the function of PAX7. RTP1 and RTP2 may resist SARS-CoV-2 by promoting the ability of olfactory receptors to coat the membrane and enhancing the ability of olfactory receptors to respond to odorant ligands. MAZ may regulate DEGs by affecting cell growth and proliferation. Micro RNA, environmental chemicals and drugs also play an important role in the anti-SAR-COV-2 infection process of human olfactory neuroepithelial cells.

Steroids - Exploitation

Corticosteroids, more specifically glucocorticoids are one of the most prescribed drugs. Corticosteroids are adrenal hormones that serve significant physiologic activities such as modulating glucose metabolism, protein catabolism, calcium metabolism, bone turnover control, immunosuppression, and down-regulation of inflammatory cascade. Corticosteroids are regarded life-saving due to their various effects and have been used therapeutically to treat broad range of auto-immune, rheumatologic, inflammatory, neoplastic, and viral illnesses.However, the therapeutic benefits of glucocorticoids are restricted by the adverse effects. The most serious side effects of corticosteroids are associated with the use of higher doses for longer periods and OTC availability in specific pharmacies, which leads to dependency, as well as its usage in mild and moderate server instances, which is contrary to guidelines. In the recent times the use of corticosteroids has been multiplied with the emergence of the Covid -19 pandemic. WHO and the standard guidelines has recommended the usage of corticosteroids in critically ill covid-19 patients but their usage in mild and moderate cases caused more harm than benefit. This illicit usage has resulted in the development of opportunistic fungal illnesses such as mucormycosis, posing an extra risk to patients in terms of quality of life and finances. Other adverse effects of systemic corticosteroids include morphological changes, increased blood sugar levels, delayed wound healing, infections, decreased bone density, truncal obesity, cataracts, glaucoma, blood pressure abnormalities, and muscle fibre atrophy.In this review we want to discuss the significance and detrimental effects of corticosteroids emphasizing on the recent times i.e., COVID-19.

MSCs-exosomes in regeneration medicine: Current evidence and future perspectives

Exosomes, especially from mesenchymal stem cells, have attracted extensive attention in regeneration medicine. Mesenchymal stem cells derived exosomes (MSCs-exosomes) have shown anti-inflammatory, anti-oxidant, antiapoptosis and tissue regeneration effects in a variety of tissue injury repair models. MSCs-exosomes hold many excellent properties such as low immunogenicity, biocompatibility, and targeting capability. With the in-depth study on the generation and function of exosomes, MSCs-exosomes are considered to be the bright stars in the field of regenerative medicine. However, there are still many obstacles to overcome in terms of exosomes isolation, clinical trials and safety evaluation. In this article, what we should focus on about MSCs-exosomes in regeneration medicine will be discussed. Copyright © 2022 Centro Regional de Invest. Cientif. y Tecn.. All rights reserved.

In-Vitro and in-Vivovascular Endothelial Growth Factor Subtypes a & B in Severe Acute Respiratory Syndrome Pathogenesis

BACKGROUND AND AIM: Increased Vascular Endothelial Growth Factor (VEGFA) Gene Expression (GE) has been documented in SARS-CoV2 infection. We wished to understand the relationship of VEGFA and VEGF B GE in both Murine SARS-CoV and Human SARS-CoV-2 in-vitro models of infection. METHOD(S): Secondary analysis of datasets from mice given nasal installation of SARS-CoV (MA15), MS1 (GSE33266MCV-1) and MS2 (GSE68820) from pulmonary tissues was undertaken. This allowed viral dose and temporal response analysis, respectively. Also studied were In-vitro Human hACE2 cells infected with SARS-CoV2 (dataset INV, GSE169158). Gene expression (GE) VEGF sub-types were analysed using Qlucore Omics Explorer (QOE) and gene enrichment functional profiling through the g:Profiler online platform. RESULT(S): For Murine studies, MA15 instillation compared to controls in MS1, lead to down-regulation of both VEGFB (MA15 10

The Prognostic Role of Genotypic Variants of Ace2 and Tmprss2 Polymorphisms in Sars-Cov2 / Covid-19 Infection

Background: to date, more than 243 million COVID-19 cases have been diagnosed globally, with 4.94 million deaths, 489.000 new cases and 8.474 deaths per day. In Italy there are currently 4.73 million cases and 132.000 deaths. It is well known that the entry of the SARS- CoV-2 virus into cells is mediated by the binding between the virus Spike-glycoprotein (S) and the membrane ACE2-receptor (ACE2-R). When SARS-CoV-2 binds to ACE2-R, with subsequent membrane fusion and virus entry into the cell, a down-regulation of these receptors occurs. ACE2 -R downregulation plays a crucial role in the pulmonary and systemic inflammatory response. A serious clinical course appears to be associated with some factors such as age, previous pathologies and comorbidities. However, also a dysregulation of the RAA system linked to a different expression of ACE-2 R and TMPRSS2 gene polymorphisms and different serum levels of soluble ACE2 (sACE2), could be associated with abnormal inflammatory and immune response to SARS-CoV-2 infection. Aim of the Study: we aimed to verify whether there is an association between the clinical course of COVID-19 patients (pts) and the presence of more frequent ACE2 and TMPRSS2 single-nucleotide polymorphisms (SNPs) and if sACE2 levels are related to specific ACE2 and TMPRSS2 polymorphic variants. Method(s): we consecutively enrolled subjects with previous documented SARS-CoV-2 infection and divided our sample into three groups: pts with asymptomatic course;pts with symptomatic course but without the need for hospitalization for COVID-19;pts with severe symptomatic course requiring hospitalization in intensive care unit. Data about age, clinical course, comorbidities, and therapies were collected. Blood samples were taken for the genetic analysis of the most frequent SNPs of the ACE2-R and TMPRSS2 detected in Italian population, in particular genotypic variants TTand CC of ACE2 SNPs 1 and 5 (rate of 5% and 14% respectively) and genotypic variants TTand CC of TMPRSS2 SNPs 2 and 3 (rate of 50% and 30% respective). Result(s): among 178 pts enrolled up to March 2022, we have so far analyzed the genetic polymorphisms of 74 pts.;21 (28%) were hospitalized for COVID-19, 38 (51%) had symptomatic course without hospitalization and 15 (21%) were completely pauci-asymptomatics. Serum concentrations of sACE2 and distribution of polymorphic variants in the three groups are summarized in Table 1. We found that sACE2 levels were higher in genotypic variant CC of SNP 1 of TMPRSS2 gene (Table 2). Considering that a high concentration of sACE2 outlines a proinflammatory condition, it could be hypothesized that the CC genotype may be a predisposing condition to the cytokine storm of COVID-19. Perspective(s): Genetic analysis of ACE2 and TMPRSS2 SNPs will help to clarify the relationship between these polymorphic variant, sACE2 levels, risk of SARS-CoV2 infection and severity of clinical presentation of COVID-19 in patients with or without CV diseases.

The Serum Ace2 Molecule in Sars-Cov2 / Covid-19 Infection and Its Potential Prognostic and Therapeutic Role (Accept Study)

Background: It is well known that the entry of the SARS- CoV-2 virus into cells is mediated by the binding between the virus Spike-glycoprotein (S) and the membrane ACE2-receptor (ACE2R). When SARS-CoV-2 binds to ACE2R, with subsequent membrane fusion and virus entry into the cell, a down-regulation of these receptors occurs. ACE2R downregulation plays a crucial role in the pulmonary and systemic inflammatory response. Also ACE2 deficiency is thought to play an important role in the pathogenesis of SARS-CoV-2 infection. The down-regulation of ACE2 induced by viral infection could be particularly harmful in subjects with pre-existing ACE2 deficiency, for example due to advanced age, the presence of DM, arterial hypertension or pre-existing heart disease, including HF. Literature data suggest an association between the patient's age and comorbidities and a greater risk of severe clinical course and with a worse prognosis. However, serious clinical pictures requiring hospitalization or leading to death have also been observed in young subjects or subjects without comorbidities. Aim of the Study: we aimed to identify predicting factors of a higher risk in terms of severity of the clinical course and worse prognosis in the population of the Brescia area, affected by a large number of cases in the first period of COVID-19 outbreak. In particular, we aimed to verify whether there is correlation between levels of serum ACE2 (sACE2) and the risk of SARS-CoV-2 infection, as well as between sACE2 and the different severity of the clinical manifestations of COVID-19 in patients with and without CV diseases. Method(s): we consecutively enrolled subjects with previous documented SARS-CoV-2 infection and divided our sample into three groups: pts with asymptomatic course;pts with symptomatic course but without the need for hospitalization for COVID-19;pts with severe symptomatic course requiring hospitalization in intensive care unit. Blood samples were taken for sACE2 dosage. We compared the concentrations of sACE2 in these groups in relation to the age, clinical course, comorbidities, and ongoing therapies. Result(s): at March 2022, we enrolled 178 pts, 51 (28%) were hospitalized for COVID-19, whereas 78 (44%) had symptomatic course without hospitalization and 49 (28%) were completely pauci-asymptomatic. Only 6 pts (4%) had myocarditis or pericarditis SARS-CoV-2-related. Between hospitalized pts, male sex (67%), older age and higher BMI were most frequent. Moreover, chronic heart failure (20%), a diagnosis of cardiopathy (29%) and AF or atrial flutter (22%) were most frequent (Table 1). Plasma concentrations of sACE2 will be dosed and analyzed in relation to the clinical characteristics of each patient. Perspective(s): ACCEPTstudy will help to clarify the relationship between ACE2 molecule, the risk of SARS-CoV2 infection and the severity of clinical presentation of COVID-19 in pts with or without CV diseases.

Hypercoagulability in COVID-19 and its putative association with the downregulation of protein C signalling

Background: Hematological complications associated with prothrombotic events with extrapulmonary manifestations have been demonstrated in autopsies of patients affected by coronavirus disease 2019 (COVID-19). Based on the close relationship of coagulation and immune response, we hypothesized that hypercoagulability in COVID-19 could result from the activation of tissue factor (F3) and subsequent alterations in Activated Protein C (APC) signaling (Figure 1). Aim(s): We aimed to identify changes in the expression of APC signaling network in liver, peripheral blood and nasal epithelium of COVID-19 patients that may contribute to local and systemic disarrangement of hemostasis. Method(s): For the expression of PROC and receptor genes public single-cell- RNA- sequencing datasets were analyzed from COVID-19 patients and healthy individuals, using the toolkit Scanpy 1.7.2 in Phyton. Result(s): The key compounds of Protein C (PC) activation and signaling;PROCR, F2R, THBD, S1PR1 and PROC were downregulated in COVID-19 patients;a greater expression of F3 in all COVID-19 tissues analyzed and upregulation of AGTR1, NFKB1, PTPN1, THBS1, PTGS2, PLAU, SERPINE1 and F5 pro-inflammatory and procoagulant genes in the liver of COVID-19 patients compared to control (Figure 2B, E and G). The hepatocyte PROC expression was changed in COVID-19 patients from hepatocyte 4 ADH1B+ PCK1+ in healthy liver (Figure 2F) to hepatocyte 3 CYP2A6+ in the liver of COVID-19 patients (Figure 2A). The ACE2 expression was increased in all COVID-19 tissues (Figure 2B, E and G) overlapping the PROC expression in the epithelium (Figure 2D) and liver tissues (Figure 2A). There was a co-expression of ACE2, PROC, PROS1, RHOA, and RAC1 in ciliated cells of COVID-19 patients (Figure 2C-D). Conclusion(s): The results provide evidence indicating a deficient synthesis and activation of PC and its receptors in COVID-19 patients that might contribute to a pronounced hypercoagulable state in response to endothelial COVID-19- related injury.