Annexin A5 Maintains Mitochondrial Integrity by Inhibiting mPTP Opening to Protect Against Acetaminophen-Induced Acute Liver Injury.

Acetaminophen (APAP) represents a major cause of drug-induced liver injury (DILI), and effective pharmacological interventions remain limited. Annexin A5 (AnxA5), a Ca2+-dependent phospholipid-binding protein, participates in diverse biological processes related to tissue repair. In this study, we evaluated AnxA5 expression in APAP-challenged mouse livers and clinical samples from patients with liver injury. Using hepatic cell lines AML12 and HepG2, we performed overexpression-based functional assays to assess the cytoprotective effects of AnxA5 against APAP toxicity. Co-immunoprecipitation assays were applied to characterize protein interactions, and mitochondrial functional parameters were measured to dissect the underlying molecular mechanism. We found that AnxA5 was significantly upregulated in both APAP exposed mice and APAP DILI patients. Cellular functional assays showed that AnxA5 overexpression mitigated APAP triggered cytotoxicity in AML12 and HepG2 cells. Mechanistically, AnxA5 bound to voltage dependent anion channel 1 (VDAC1), restrained VDAC1 mediated mitochondrial Ca2+ influx, and suppressed VDAC1 oligomerization, which further inhibited mitochondrial permeability transition pore (mPTP) opening. In an APAP-induced liver injury mouse model, exogenous recombinant AnxA5 treatment maintained mitochondrial integrity and ameliorated hepatic inflammation and liver damage. Collectively, our data reveal AnxA5 as an endogenous mitochondrial protective factor and support its therapeutic potential against APAP-induced liver injury.