Obacunone attenuates Cholestatic liver disease via PPARα activation and gut microbiota remodeling.

BACKGROUND: Cholestatic liver disease (CLD) comprises a collection of disorders marked by impaired bile formation or flow, which can progress to fibrosis, cirrhosis, and liver failure if left untreated. Although there have been significant advances in understanding its pathogenesis, effective pharmacotherapies are still limited; ursodeoxycholic acid (UDCA), the first-line treatment, exhibits an incomplete response in approximately 30-40% of patients. Obacunone (OBA), a naturally occurring limonoid triterpenoid, has shown potent anti-inflammatory and antioxidant effects in various models of liver injury; however, its role in CLD has yet to be investigated.

PURPOSE: To investigate the mechanism by which OBA ameliorates α-naphthyl isothiocyanate (ANIT)/ethinyl estradiol (EE) -induced CLD through the regulation of peroxisome proliferator-activated receptor α (PPARα).

METHODS: Cholestatic liver injury was induced in mice via ANIT/EE treatment. UDCA served as a positive control, while OBA was administered at 1.5, 3, and 6 mg/kg as low, medium, and high dose intervention groups, respectively. Serum and liver tissues were collected to evaluate liver function, cholestasis-related markers, and hepatic histology using assay kits, quantitative polymerase chain reaction (qPCR), and Western blotting. Bile acid profiles were quantified by ultra performance liquid chromatography-mass spectrometry (UPLC-MS/MS), and transcriptomic sequencing identified PPARα as a key target involved in bile acid metabolism. A PPARα inhibitor (GW6471) was used to verify the regulatory role of OBA via PPARα. Additionally, 16S rRNA high-throughput sequencing was performed to explore the effects of OBA on gut microbiota within the gut-liver axis.

RESULTS: ANIT/EE induced severe cholestatic liver injury in mice, characterized by elevated Aspartate aminotransferase (AST), Alanine aminotransferase (ALT), Alkaline phosphatase (AKP), total bile acid (TBA), total bilirubin (TBIL), total cholesterol (TC) and triglyceride(TG) levels, decreased aenosine tiphosphate (ATP) and ketone body (KB) levels, obvious liver pathological damage, enhanced inflammation, disordered fatty acid oxidation, abnormal bile acid synthesis, reduced beneficial bacteria (Akkermansia, Dubosiella, Lactobacillus) and increased harmful bacteria (Helicobacter, Escherichia-Shigella). High-dose OBA markedly ameliorated these abnormalities, inhibited hepatic inflammatory cytokines (tumor necrosis factor-alpha (Tnf-α), Interleukin 6 (Il6), Interleukin-1 beta (Il-1β)), regulated bile acid synthesis, absorption and efflux, and restored gut microbiota balance. In contrast, PPARα inhibition significantly attenuated the protective effects of OBA against cholestasis.

CONCLUSION: This study demonstrates that OBA activates PPARα, thereby regulating bile acid metabolism, fatty acid oxidation, inflammatory responses, and gut microbiota to alleviate CLD, providing a theoretical basis for targeted therapy targeting the PPARα-bile acid-microbiota axis in CLD.