Protecting mitochondria via inhibiting VDAC1 oligomerization alleviates ferroptosis in acetaminophen-induced acute liver injury

Acetaminophen (APAP) overdose is a common cause of drug-induced liver injury (DILI). Ferroptosis has been recently implicated in APAP-induced liver injury (AILI). However, the functional role and underlying mechanisms of mitochondria in APAP-induced Ferroptosis are unclear. In this study, the voltage-dependent anion channel (VDAC) oligomerization inhibitor VBIT-12 and Ferroptosis inhibitors were injected via tail vein in APAP-injured mice. Targeted metabolomics and untargeted lipidomic analyses were utilized to explore underlying mechanisms of APAP-induced mitochondrial dysfunction and subsequent Ferroptosis. As a result, APAP overdose led to characteristic changes generally observed in Ferroptosis. The use of Ferroptosis inhibitor ferrostatin-1 (or UAMC3203) and iron chelator deferoxamine further confirmed that Ferroptosis was responsible for AILI. Mitochondrial dysfunction, which is associated with the tricarboxylic acid cycle and fatty acid β-oxidation suppression, may drive APAP-induced Ferroptosis in hepatocytes. APAP overdose induced VDAC1 oligomerization in hepatocytes, and protecting mitochondria via VBIT-12 alleviated APAP-induced Ferroptosis. Ceramide and cardiolipin levels were increased via UAMC3203 or VBIT-12 in APAP-induced Ferroptosis in hepatocytes. Knockdown of Smpd1 and Taz expression responsible for ceramide and cardiolipin synthesis, respectively, aggravated APAP-induced mitochondrial dysfunction and Ferroptosis in hepatocytes, whereas Taz overexpression protected against these processes. By immunohistochemical staining, we found that levels of 4-hydroxynonenal (4-HNE) protein adducts were increased in the liver biopsy samples of patients with DILI compared to that in those of patients with autoimmune liver disease, chronic viral hepatitis B, and non-alcoholic fatty liver disease (NAFLD). In summary, protecting mitochondria via inhibiting VDAC1 oligomerization attenuated hepatocyte Ferroptosis by restoring ceramide and cardiolipin content in AILI.