[Study on the mechanism of Fuzheng Huayu formula against biliary fibrosis in mice]

Objective: To investigate the effect and mechanisms of Fuzheng huayu formula (FZHY) on biliary fibrosis in mice. Methods: Mdr2 gene knowout (Mdr2^-/-) mice were randomly divided into model group, FZHY-treated group and obeticholic acid (OCA)-treated group. Wide type C57BL/6J (WT) mice of the same age were used as the control group. Mdr2^-/- mice were treated with the corresponding drugs by gavage from the first day of the 9^th week old. The WT and model groups were given 0.3% sodium carboxymethyl cellulose by gavage, and the mice were sacrificed at the end of 12^th week old. The activities of serum alkaline phosphatase (ALP) and alanine aminotransferase (ALT) were detected by high-speed biochemical analyzer. H&E staining and sirius red staining were used to observe the pathological changes of liver tissues. The hepatic hydroxyproline content was determined. The hepatic expressions of Col-Ⅰ and α-SMA, ductular reaction markers Epcam, CK7, CK19, and the expression of Pcna, Mki67 and Ccnd1; as well as the expression of inflammatory cytokines Ccl2, Ccl5, Tnf-α, Il10 and Cxcl4 were detected by immunohistochemical staining, western blot and qRT-PCR, respectively. Furthermore, the expression of PPARα and the phosphorylation NF-κB were detected by western blot. Results: Compared with WT mice, the serum ALT and ALP activities of Mdr2^-/- mice were significantly increased (P<0.001). The percentage of SR positive stained area and hepatic Hyp content were significantly increased (P<0.01). The hepatic expression of Col-Ⅰ and α-SMA; Epcam, CK7, CK19, Pcna, Mki67 and Ccnd1; Ccl2, Ccl5, Tnf-α, Il10 and Cxcl4 were significantly increased (P<0.01). However, both FZHY and OCA significantly reversed the elevation of these aforementioned indicators (P<0.05; P<0.01). Further study showed that the hepatic expression of PPARα in Mdr2^-/- mice was significantly lower than that of WT mice, however the phosphorylation of NF-κB was significantly enhanced (P<0.01). The expression of PPARα in liver tissues of FZHY mice was significantly increased (P<0.05), and the NF-κB phosphorylation was significantly inhibited compared with Mdr2^-/- mice (P<0.05). Conclusion: FZHY significantly ameliorated liver fibrosis, ductular reaction and inflammation in Mdr2^-/- spontaneous biliary fibrosis mice, and its mechanism was related to the regulation of PPARα/NF-κB pathway.