Molecular Biological Mechanisms of Action of Chrysophanol in Hepatic Stellate Cells Activated by Hepatic B Virus X Based on Network Pharmacology.

INTRODUCTION

Chrysophanol (Cho) is a natural anthraquinone with biological effects such as inducing ferroptosis and anticancer activity. The hepatitis B virus X protein (HBx) is essential for HBV replication. We aimed to identify the key pathways in HBx-induced hepatic stellate cell (HSC) activation and to characterize the potential mechanisms of action of Cho against liver fibrosis.

METHODS

HSC-T6 cells were transfected with FLAG (control group) or FLAG-HBx (HBx group), and RNA sequencing and Western blotting analysis were conducted to assess the effects of HBx and Cho on specific molecular targets and signaling pathways.

RESULTS

Gene ontology and pathway analyses indicated that the genes targeted by HBx participate in immunological responses, chemokine and cytokine activity, cell-substrate adhesion, extracellular matrix organization, growth factor binding, defense responses, and antigen processing and presentation. RNA-seq and Western blotting data revealed that HBx-activated HSC-T6 cells exhibited upregulated expression of mammalian target of rapamycin (mTOR), phosphorylated mTOR (p-mTOR), S6, phosphorylated S6 (p-S6), peroxisome proliferator-activated receptor (PPAR-α), phosphorylated-PPAR-α (p-PPAR-α), CYP27, α-smooth muscle actin (α-SMA), connective tissue growth factor (CTGF), and Integrin-β1, which was reversed after treatment with Cho. These results were also verified in a HBx-activated HSC-T6 and LX-2 cell model and thioacetamide-induced liver fibrosis mouse model.

CONCLUSIONS

Thus, our findings indicate that Cho ameliorates HBx-induced HSC activation and liver fibrosis via inhibition of the mTOR and PPARs signaling pathways, suggesting that Cho is a potential therapeutic for chronic liver inflammation-mediated diseases.