Molecular Profiling of COVID-19 Autopsies Uncovers Novel Disease Mechanisms (preprint)

Background: Current understanding of COVID-19 pathophysiology is limited by disease heterogeneity, complexity, and a paucity of studies evaluating patient tissues with advanced molecular tools. Methods: Autopsy tissues from two COVID-19 patients, one of whom died after a month-long hospitalization with multi-organ involvement while the other died after a few days of respiratory symptoms, were evaluated using multi-scale RNASeq methods (bulk, single-nuclei, and spatial RNASeq next-generation sequencing) to provide unprecedented molecular resolution of COVID-19 induced damage. Findings: Comparison of infected/uninfected tissues revealed four major regulatory pathways. Effectors within these pathways could constitute novel therapeutic targets, including the complement receptor C3AR1, calcitonin-like receptor or decorin. Single-nuclei RNA sequencing of olfactory bulb and prefrontal cortex highlighted remarkable diversity of coronavirus receptors. Angiotensin I converting enzyme 2 was rarely expressed, while Basignin showed diffuse expression, and alanyl aminopeptidase was associated with vascular/mesenchymal cell types. Comparison of lung and lymph node tissues from patients with different symptomatology with Digital Spatial Profiling resulted in distinct molecular phenotypes. Interpretation: COVID-19 is a far more complex and heterogeneous disease than initially anticipated. Evaluation of COVID-19 rapid autopsy tissues with advanced molecular techniques can identify pathways and effectors at play in individual patients, measure the staggering diversity of receptors in specific brain areas and other well-defined tissue compartments at the single-cell level, and help dissect differences driving diverging clinical courses among patients. Extension of this approach to larger datasets will substantially advance the understanding of the mechanisms behind COVID-19 pathophysiology. Funding: No external funding was used in this study.

Gastrointestinal involvement attenuates COVID-19 severity and mortality (preprint)

Given that gastrointestinal (GI) symptoms are a prominent extrapulmonary manifestation of coronavirus disease 2019 (COVID-19), we investigated the impact of GI infection on disease pathogenesis in three large cohorts of patients in the United States and Europe. Unexpectedly, we observed that GI involvement was associated with a significant reduction in disease severity and mortality, with an accompanying reduction in key inflammatory proteins including IL-6, CXCL8, IL-17A and CCL28 in circulation. In a fourth cohort of COVID-19 patients in which GI biopsies were obtained, we identified severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) within small intestinal enterocytes for the first time in vivo but failed to obtain culturable virus. High dimensional analyses of GI tissues confirmed low levels of cellular inflammation in the GI lamina propria and an active downregulation of key inflammatory genes including IFNG, CXCL8, CXCL2 and IL1B among others. These data draw attention to organ-level heterogeneity in disease pathogenesis and highlight the role of the GI tract in attenuating SARS-CoV-2-associated inflammation with related mortality benefit.

SARS-CoV-2 Viral Load Predicts COVID-19 Mortality (preprint)

The need for reliable and widely available SARS-CoV-2 testing is well recognized, but it will be equally necessary to develop quantitative methods that determine viral load in order to guide patient triage and medical decision making. We are the first to report that SARS-CoV-2 viral load at the time of presentation is an independent predictor of COVID-19 mortality in a large patient cohort (n=1,145). Viral loads should be used to identify higher-risk patients that may require more aggressive care and should be included as a key biomarker in the development of predictive algorithms.

Pathophysiology of SARS-CoV-2: targeting of endothelial cells renders a complex disease with thrombotic microangiopathy and aberrant immune response. The Mount Sinai COVID-19 autopsy experience (preprint)

BACKGROUND Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) and its associated clinical syndrome COVID-19 are causing overwhelming morbidity and mortality around the globe, disproportionately affecting New York City. A comprehensive, integrative autopsy series that advances the mechanistic discussion surrounding this disease process is still lacking. METHODS Autopsies were performed at the Mount Sinai Hospital on 67 COVID-19 positive patients and data from the clinical records were obtained from the Mount Sinai Data Warehouse. The experimental design included a comprehensive microscopic examination carried out by a team of expert pathologists, along with transmission electron microscopy, immunohistochemistry, RNA in situ hybridization, as well as immunology and serology assays. RESULTS Laboratory results of our COVID-19 cohort show elevated inflammatory markers, abnormal coagulation values, and elevated cytokines IL-6, IL-8 and TNF. Autopsies revealed large pulmonary emboli in four cases. We report microthrombi in multiple organ systems including the brain, as well as conspicuous hemophagocytosis and a secondary hemophagocytic lymphohistiocytosis-like syndrome in many of our patients. We provide electron microscopic, immunofluorescent and immunohistochemical evidence of the presence of the virus and the ACE2 receptor in our samples. CONCLUSIONS We report a comprehensive autopsy series of 67 COVID-19 positive patients revealing that this disease, so far conceptualized as a primarily respiratory viral illness, also causes endothelial dysfunction, a hypercoagulable state, and an imbalance of both the innate and adaptive immune responses. Novel findings reported here include an endothelial phenotype of ACE2 in selected organs, which correlates with clotting abnormalities and thrombotic microangiopathy, addressing the prominent coagulopathy and neuropsychiatric symptoms. Another original observation is that of macrophage activation syndrome, with hemophagocytosis and a hemophagocytic lymphohistiocytosis-like disorder, underlying the microangiopathy and excessive cytokine release. We discuss the involvement of critical regulatory pathways.