Natural small molecule hinokitone mitigates NASH fibrosis by targeting regulation of FXR-mediated hepatocyte apoptosis

Introduction

Liver fibrosis is the common fate of NASH and poses a major health threat with very limited pharmacological treatments.

Objectives

This study aims to investigate the preventive effect of hinokitone (HO), an isolated compound from Agathis dammara, on NASH fibrosis and its underlying mechanism.

Methods

To investigate the effect of HO on NASH fibrosis, C57BL/6 mice were either fed a high-fat diet (HFD) in conjunction with intraperitoneal injection of CCl₄ for 8 weeks or single CCl₄ for 14 days to establish mouse liver fibrosis model, and HO was administered by gavage simultaneously. To elucidate the underlying mechanisms, HepG2 cells were stimulated by palmitic acid (PA) or tumor necrosis factor α plus actinomycin-D (Act-D + TNFα) to induce hepatocyte apoptosis model. Furthermore, hepatocyte Farnesoid-X-receptor (FXR) specifically knocked out mice were established by the albumin-Cre-loxP recombination enzyme system to ascertain the role of FXR in the anti-NASH fibrosis effects of HO.

Results

The results showed that HO presented dose-dependent anti-liver fibrosis efficacy in NASH mice induced by HFD + CCl₄ and CCl₄-induced mouse liver fibrosis. Cellularly, HO significantly inhibited PA-induced lipotoxic apoptosis and Act-D + TNFα-induced exogenous apoptosis in hepatocytes, which in turn prevented HSC activation. Mechanistically, bioinformatics analysis and surface plasmon resonance assay had identified hepatocyte FXR as a target of HO. Specifically, HO directly bound to FXR and upregulated its protein level by inhibiting proteasomal degradation. In turn, HO attenuated hepatocyte lipid deposition through upregulating the FXR’s downstream target genes SHP and CES1, and reduced cleaved-CASP8 level, thereby inhibiting hepatocyte apoptosis. Furthermore, HO lost its function in the inhibition of hepatocyte apoptosis and liver fibrosis when knockout hepatocyte FXR.

Conclusion

In conclusion, HO has an inhibitory effect on NASH fibrosis. This effect is mediated by targeting upregulation of hepatocyte FXR, which in turn attenuates hepatocyte apoptosis and thus indirectly inhibits the activation of HSCs.