Mitigation of SARS-CoV2-Mediated Endothelial Injury via Suppression of the Epigenetic Enzyme KMT2A/MLL1 in Macrophages

Severe infection with SARS-CoV2 results in COVID-associated coagulopathy, which features macrophage-mediated systemic inflammation (macrophage activation syndrome), endothelial damage, and hypercoagulability. COVID-associated coagulopathy predisposes patients to macro- and microvascular thrombosis, thereby culminating in organ dysfunction and death. We hypothesized that COVID-associated macrophage activation is dependent on the inflammation related epigenetic enzyme, KMT2A/MLL1, and results in inflammatory cytokine release and promotes endothelial injury. Plasma specimens were obtained from critically ill patients with primary diagnosis of SARS-CoV2 pneumonia who required mechanical ventilation. CD14+ monocytes were isolated from hospitalized SARS-CoV2 positive [hCOV(+)] patients and hospitalized controls without SARS-CoV2 [hCOV(–)]. Murine experiments utilized intranasal inoculation of murine hepatitis virus A59 (MHVA59), an established model of coronavirus infection. Mice harboring myeloid specific knockout of Mll1 (LysMCre Mll1fl/fl; Cre+) or littermate control (Cre–) animals were inoculated with 2 × 105 plaque forming units of MHVA59 and plasma was collected from infected and sham (received intranasal phosphate-buffered saline) mice. Immortalized macrophages (RAW264.7 cells) were infected with MHVA59. Quantitative polymerase chain reaction (IL6, IL1β, tumor necrosis factor α) and enzyme-linked immunosorbent assays (soluble E-selectin: sEsel—marker of endothelial injury; IL6, IL1β, tumor necrosis factor α) were performed. hCOV(+) patients had an increase in mRNA levels of MLL1 (3.2-fold; P < .001), as well as IL6, IL1β, and tumor necrosis factor α in circulating monocytes relative to hCOV(–) patients (Fig 1, A). Further showing an association between MLL1 levels and expression of inflammatory cytokines, silencing of Mll1 in RAW264.7 cells infected with MHVA59 resulted in blunted transcription of IL6 (by >65%; P < .01), IL1β (by >65%; P < .05), and tumor necrosis factor α (by >50%; P < .05) mRNA (Fig 1, B). In Cre– mice inoculated with MHVA59, there was a concurrent rise of sEsel levels and plasma levels of IL6, IL1β, and tumor necrosis factor α in a time-dependent fashion, and the induction of these factors was blunted Cre+ (MLL1 knockout) mice (Fig 2, A). Finally, among critically ill patients with confirmed SARS-CoV2 infection, average circulating sEsel levels were higher in nonsurvivors compared to survivors (126.7 ng/mL vs 87.1 ng/mL; P = .0004) (Fig 2, B). Endothelial injury as measured by sEsel is associated with mortality in patients with SARS-CoV2. In a murine model, suppression of MLL1 in macrophages suppresses inflammatory cytokine expression and decreases endothelial injury as measured by sEsel. Targeting macrophage activation syndrome has potential to improve endothelial dysfunction in SARS-CoV2.Fig 2Critically ill patients with confirmed SARS-CoV2 infection, average circulating sEsel levels.View Large Image Figure ViewerDownload Hi-res image Download (PPT)