Glycogen Synthase Kinase 3 β Hepatocyte Deletion Attenuates Ferroptosis and Metabolic Dysfunction-associated Steatohepatitis in Mice.

Abstract

BACKGROUND & AIMS: Metabolic dysfunction-associated steatohepatitis (MASH) is characterized by toxic lipid-induced cellular stress (lipotoxicity), which culminates in lethal and sublethal hepatocyte injury and a sterile fibroinflammatory response. We previously reported that pharmacological inhibition of glycogen synthase kinase 3 (GSK3) ameliorates murine MASH. However, the hepatocyte-specific role of GSK3β in lipotoxic injury and the fibroinflammatory response in MASH remains unclear. METHODS: We generated hepatocyte-specific Gsk3β knockout (Gsk3β) mice by crossing Gsk3βmice with albumin-Cre mice. Mice were fed either a choline-deficient high-fat diet (CDHFD) or a high-fat, fructose, and cholesterol (FFC) diet to induce MASH. RESULTS: Metabolic parameters and hepatic lipidomic were similar between FFC-fed Gsk3βand Gsk3βmice. The NanoString Metabolic Pathways Panel on liver tissues showed upregulation of NAD, mitochondrial function, and oxidative phosphorylation signaling pathways in FFC-fed Gsk3βmice compared with Gsk3βmice. In vitro studies in palmitate-treated hepatocytes showed that mitochondrial morphology, biogenesis, contact with lipid droplets, and respiration were improved, whereas mitochondrial DNA release and Ferroptosis Suppressor Protein 1 (FSP1) phase separation were reduced with pharmacological GSK3 inhibition or in hepatocytes isolated from Gsk3βmice. Similarly, liver injury, lipid peroxidation, ferroptosis markers, and circulating mitochondrial DNA levels were reduced in Gsk3βmice with MASH. Furthermore, Gsk3βmice with MASH had reduced hepatic expression of proinflammatory genes, myeloid cell infiltration, NETosis, and showed significant downregulation of fibrosis signaling pathways. CONCLUSIONS: Gsk3βreduced liver injury, mitochondrial DNA release, inflammation, and fibrosis in mice with MASH, secondary to improved mitochondrial bioenergetics and reduced ferroptosis. Therefore, GSK3β may be a potential therapeutic target for human MASH.