Mincle-GSDMD-mediated release of IL-1β containing small extracellular vesicles contributes to ethanol-induced liver injury

Abstract

Background & AimsMacrophage inducible C-type lectin (Mincle) is expressed on Kupffer cells and senses ethanol-induced danger signals released from dying hepatocytes and promotes IL-1{beta} production. However, it remains unclear what and how ethanol-induced Mincle ligands activate downstream signaling events to mediate IL-1{beta} release and contribute to alcohol-associated liver disease (ALD). In this study, we investigated the association of circulating {beta}-glucosylceramide ({beta}-GluCer), an endogenous Mincle ligand, with severity of ALD and examined the mechanism by which {beta}-GluCer engages Mincle on Kupffer cells to release IL-1{beta} in the absence of cell death and exacerbates ALD. Approach and ResultsConcentrations of {beta}-GluCer were increased in serum of patients with severe AH and correlated with disease severity. Challenge of Kupffer cells with LPS and {beta}-GluCer induced formation of a Mincle and Gsdmd-dependent secretory complex containing chaperoned full-length GSDMD (Hsp90-CDC37-NEDD4) with polyubiquitinated pro-IL-1{beta} and components of the Casp8-NLRP3 inflammasome loaded as cargo in small extracellular vesicles (sEV). Gao-binge ethanol exposure to wild-type, but not Mincle-/- and Gsdmd-/-, mice increased release of IL-1{beta} containing sEVs from liver explant cultures. Myeloid-specific deletion of Gsdmd similarly decreased the formation of sEVs by liver explant cultures and protected mice from ethanol-induced liver injury. sEVs collected from ethanol-fed wild-type, but not Gsdmd-/-, mice promoted injury of cultured hepatocytes and, when injected into wild-type mice, aggravated Gao-binge ethanol-induced liver injury. Conclusion{beta}-GluCer functions as a DAMP activating Mincle-dependent GSDMD-mediated formation and release of IL-1{beta}-containing sEVs, which in turn exacerbate hepatocyte cell death and contribute to the pathogenesis of ALD.