Elevated Myl9 reflects the Myl9-containing microthrombi in SARS-CoV-2-induced exudative vasculitis and predicts COVID-19 severity

The mortality of coronavirus disease 2019 (COVID-19) is strongly correlated with pulmonary vascular pathology accompanied by SARS-CoV-2 infection-triggered immune dysregulation and aberrant activation of platelets. We combined histological analyses using field emission scanning electron microscopy with energy-dispersive X-ray spectroscopy analyses of the lungs from autopsy samples and single-cell RNA sequencing of peripheral blood mononuclear cells to investigate the pathogenesis of vasculitis and immunothrombosis in COVID-19. We found that SARS-CoV-2 accumulated in the pulmonary vessels, causing exudative vasculitis accompanied by the emergence of thrombospondin-1 (THBS-1)-expressing non-canonical monocytes and the formation of Myosin light chain 9 (Myl9)-containing microthrombi in the lung of COVID-19 patients with fatal disease. The amount of plasma Myl9 in COVID-19 was correlated with the clinical severity and measuring plasma Myl9 together with other markers allowed us to predict the severity of the disease more accurately. This study provides detailed insight into the pathogenesis of vasculitis and immunothrombosis, which may lead to optimal medical treatment for COVID-19. Overall design: PBMC samples recovered from COVID-19 patients at admission (n=13) or within 10 days after admission (n=8), were stained with anti-human CD45 antibody. Propidiumiodine was also added to samples to remove dead cells. Live CD45+ cells were sorted using a SONY SH800S (4,000-10,000 cells each, cell viability >98%) and then encapsulated into droplets. Libraries were prepared using Chromium Single Cell 3' Reagent Kits v3.1 following the manufacturer's protocol (10X Genomics, CA,USA). The generated scRNA-seq libraries were sequenced using a total of 127 cycles (paired-end reads) with a NovaSeq 6000 (Illumina, Inc., CA, USA).