SiNiSan alleviates liver injury by promoting hepatic stem cell differentiation via Wnt/β-catenin signaling pathway.

Background: SiNiSan, a Traditional Chinese Medicine containing Radix Bupleuri, Radix Paeoniae Alba, Fructus Aurantii Immaturus, and Radix Glycyrrhizae, has been shown to be clinically effective in treating liver damage, its underlying molecular mechanisms however remains unclear.

Purpose: The aim of the current study was to understand the molecular mechanisms of SiNiSan in the treatment of liver damage utilizing mice and cell culture models.

Methods: Here, mice were gavaged with 0.2% CCl to obtain acute liver injury model and with alcohol to obtain chronic liver injury model. H&E staining was performed to detect liver histomorphology. HPLC-MS was performed to analyze the composition of SiNiSan decoction and SiNiSan-medicated serum (SMS). In addition, western blots were done to analyze the representative protein expression in Wnt/β-catenin signaling. Immunofluorescence staining was done to analyze the protein levels in WB-F344 cells. Finally, in an attempt to measure the influence of SiNiSan on liver regeneration in rats, we constructed a rats partial hepatectomy models.

Results: We demonstrated that SiNiSan treatment mitigated liver damage in mice, as evidenced by the decrease in serum AST and ALT levels, as well as improved liver tissue morphology. HPLC-MS results showed that SMS contained a variety of components from the SiNiSan decoction. Next, our results showed that SMS reduced the expression of α-fetoprotein (AFP) and enhanced the expression of albumin (ALB) and cytokeratin 19 (CK19) in WB-F344 cells. Further, SMS treatment induced the accumulation of β-catenin. After 14 days of SMS treatment, β-catenin protein underwent nuclear translocation and bound to the LEF1 receptor in the nucleus, which regulated c-Myc and Cyclin D1 factors to activate Wnt/β-catenin signaling and promoted differentiation of WB-F344 cells. In addition, we demonstrated that SiNiSan increased liver regeneration in rat hepatectomy.

Conclusion: Collectively, the current study revealed that SiNiSan alleviated the acute liver injury induced by CCl as well as the chronic liver damage triggered by alcohol and sucrose in vitro. Concurrently, SMS treatment induced hepatic stem cell differentiation by activating Wnt/β-catenin signaling in vivo. Further study showed that SiNiSan promoted the regeneration of rats liver. The current study provides a theoretical basis for the clinical treatment of liver-related diseases with SiNiSan.