Dysregulation of xenobiotic metabolism and mitochondrial dysfunction exacerbate acetaminophen-induced hepatotoxicity in human antigen R-deficient male mice

Acetaminophen (APAP) overdose is a leading cause of acute liver failure worldwide. The RNA-binding protein Human antigen R (HuR) is a multifunctional post-transcriptional regulator that plays a pivotal role in cellular stress responses, including those triggered by APAP toxicity. This study investigated the mechanisms by which HuR protects against APAP-induced hepatotoxicity in male mice. Hepatocyte-specific HuR-deficient (HuRHep-/-) male mice on a C57BL/6N background and wild-type (WT) littermates were treated with 200 mg/kg APAP, and liver tissues were collected at 2, 6, and 24 hours post-treatment. APAP administration increased hepatic HuR mRNA expression and induced HuR cleavage and the formation of a higher-molecular weight HuR-immunoreactive band, with the latter two correlating with injury severity. Compared with WT controls, HuRHep-/- mice exhibited markedly increased susceptibility to hepatotoxicity at both 2 and 6 hours. Metabolite profiling revealed altered APAP metabolism and reduced glutathione S-transferase (Gst) expression in HuRHep-/- livers, consistent with impaired APAP detoxification and increased APAP-protein adduct formation. Fourier-transform infrared (FTIR) spectroscopy further identified early biochemical differences between WT and HuRHep-/- livers as early as 2 hours after APAP exposure. Additionally, HuR deficiency resulted in pronounced mitochondrial structural abnormalities and dysfunction at 2 and 6 hours, accompanied by reduced expression of the mitochondrial fission and fusion proteins Drp1 and Mfn2, increased mitochondrial protein release, and enhanced hepatocyte death. Although pro-inflammatory cytokine levels were elevated in HuRHep-/- mice relative to WT controls at 24 hours, hepatocyte proliferation was similarly blunted in both genotypes, consistent with severe liver injury and delayed recovery. Collectively, these findings identify hepatocyte HuR as a critical regulator of xenobiotic metabolism and mitochondrial integrity and establish its essential role in early protection against APAP-induced hepatotoxicity in male mice.