ABSTRACT
Background. Predictors of SARS-CoV-2 RNA levels and changes over time during early COVID-19 are not well characterized. Methods. ACTIV-2 is a phase II/III randomized, placebo-controlled, platform trial to evaluate investigational agents for treatment of COVID-19 in non-hospitalized adults. Participants enrolled within 10 days of symptom onset. Nasopharyngeal samples were collected for SARS-CoV-2 RNA testing on Days 0, 3, 7, 14 and 28;RNA was quantified with qPCR assay. SARS-CoV-2 seropositivity was defined as detectable IgG to any of nucleocapsid, receptor binding domain, S1 and S2 antigens by Bio-Plex multiplex assay. Censored linear regression and repeated measures Poisson models evaluated predictors of RNA including age, sex, race, ethnicity, risk of severe COVID-19, diabetes, BMI, obesity (BMI > 35 kg/m2) and serostatus. Results. The study enrolled 537 participants from Aug 2020 to July 2021 at US sites. Median age was 48 years;49% were female sex, >99% cis-gender, 83% white, 29% Hispanic/Latino, and 21% had BMI > 35 kg/m2. At Day 0, median symptom duration was 6 days, 50% were seropositive (2 were vaccinated) and 17% had RNA below the lower limit of quantification (LLoQ). Higher Day 0 RNA was associated with shorter symptom duration (Spearman correlation = -0.40, p< 0.001), as well as older age, white race, lower BMI and seronegativity, even when adjusting for symptom duration (all p< 0.03). Among the 203 on placebo with Day 0 RNA >= LLoQ, female sex had larger decreases in RNA at Day 3 vs male sex (difference in mean change: -0.8 log10 copies/mL (95% CI: -1.2, -0.4), p< 0.001) when adjusted for symptom duration and Day 0 RNA;this difference was also observed when evaluating the proportion with RNA < LLoQ at Day 3 (Risk Ratio (95% CI): 2.38 (1.11, 5.09)). Seropositivity at Day 0 was associated with higher probability of RNA < LLoQ at Days 3 and 7 (p< 0.001) in adjusted models. Seropositivity at Day 0 did not differ by sex. Conclusion. In this well characterized clinical trial cohort, shorter symptom duration, older age, white race, lower BMI and seronegativity were associated with higher RNA in early infection. Female sex and seropositivity were associated with earlier viral clearance. Further research is needed to determine if viral decay differences mediated by these host factors influence clinical outcomes.
ABSTRACT
Background: SARS-CoV-2 infection can compromise respiratory function and cause thrombotic events. SARS-CoV-2 binds to and mediates downregulation of angiotensin converting enzyme 2 (ACE2) on infected cells. Diminished enzymatic activity of ACE2 could result in increased concentrations of the pro-inflammatory molecules angiotensin II and bradykinin, contributing to SARS-CoV-2 pathology. Methods: Immunofluorescence microscopy and digital image data quantification, Computer assisted molecular docking analyses, Western blot. Results: Using immunofluorescence microscopy of lung tissues from uninfected and SARS-CoV-2 infected individuals, we find evidence that ACE2 is highly expressed in the pulmonary alveolar epithelium and is significantly reduced along the alveolar lining of SARS-CoV-2 infected lungs. Ex vivo analyses indicate that ACE2 is readily detected on primary human pulmonary alveolar epithelial and primary human aortic endothelial cells (HAoECs). Exposure of these cells to recombinant SARS-CoV-2 spike protein was sufficient to reduce surface ACE2 expression. Moreover, exposure of HAoECs to spike protein induced endothelial dysfunction (increased expression of von Willebrand Factor and decreased expression of Krüppel-like Factor 2), caspase activation, and apoptosis. Exposure of HAoECs to bradykinin (BK, 10μ M) induced calcium signaling and endothelial dysfunction but did not adversely affect viability. Computer assisted analyses of molecules with potential to bind bradykinin receptor B2 (BKRB2) suggested a potential role for aspirin as a bradykinin antagonist. When tested in our in vitro model, we found that aspirin (1μM) could significantly blunt cell signaling, and endothelial dysfunction caused by bradykinin in these cells. Conclusion: SARS-CoV-2 causes complex effects on microvascular homeostasis that potentially contribute to organ dysfunction and coagulopathies. Reduced ACE2 enzymatic activity could contribute to inflammation and pathology in the lung. Our studies add to this understanding by providing evidence that spike protein alone can mediate adverse effects on vascular cells. Understanding these mechanisms of pathogenesis may provide rationale for interventions, such as interference with the interactions of spike protein or bradykinin with endothelial cells, that could limit microvascular events associated with SARS-CoV-2 infection and stabilize microvascular homeostasis in COVID-19 disease.