Molecular hydrogen suppresses gut inflammation and pyroptosis in ulcerative colitis through promoting PKM2 lactylation to block NLRP3 inflammasome activation.

Abstract

BACKGROUND: Molecular hydrogen is considered to be able to alleviate the progression of ulcerative colitis (UC), whose underlying molecular mechanism remains largely unclear. METHODS: Lipopolysaccharide (LPS)/adenosine triphosphate (ATP)-stimulated human colonic epithelial cells (HCoEpiC) were used to mimic UC model in vitro and were treated with hydrogen-rich medium (HRM). Dextran sulfate sodium (DSS)-induced mice were used to construct UC model in vivo and were administrated by hydrogen-rich water (HRW). Colon tissues from UC patients and healthy volunteers were collected. Release of inflammatory cytokines was detected by ELISA. Cell viability, gut pathological impairment and pyroptosis were assessed by CCK-8 assay, HE staining and transmission electron microscope (TEM). The protein expressions of NLRP3 inflammasome and pyruvate kinase M2 (PKM2) were analyzed by western blot. The interaction between NLRP3 and PKM2 was verified by co-immunoprecipitation (Co-IP) assay. RESULTS: Molecular hydrogen alleviated cell viability inhibition and cytokines production in HCoEpiC cells, and suppressed gut injury, inflammation, pyroptosis and NLRP3 inflammasome activation in mice. PKM2 lactylation was down-regulated in colon tissues of UC patients. Molecular hydrogen treatment promoted PKM2 lactylation without affecting total PKM2 expression. The lactylation inhibitor, sodium oxamate, abolished molecular hydrogen-mediated protective effects on UC. Additionally, molecular hydrogen facilitated the combination between PKM2 and NLRP3 to down-regulate inflammasome activation. This binding between PKM2 and NLRP3 was abolished by disrupting PKM2 lactylation. Furthermore, the structural interaction between lactylated PKM2 and NLRP3 was predicted. CONCLUSION: Molecular hydrogen can promote PKM2 lactylation to restrain NLRP3 inflammasome-mediated pyroptosis and inflammation, thus ameliorating UC progression.