18α-Glycyrrhetinic Acid Down-Regulates Expression of Type I and III Collagen via TGF-B1/Smad Signaling Pathway in Human and Rat Hepatic Stellate Cells

Objective: To investigate the effects of 18 alpha-glycyrrhetinic acid (18 alpha-GA) on the expression of type I and III collagen in human and rat hepatic stellate cells (HSC) and to explore the role of TGF-beta 1/Smad signaling pathway involved. Methods: Following 18 alpha-GA treatment, the cell viability and cell growth were detected to determine the optimal concentration of 18 alpha-GA. The expressions of TGF-beta 1/Smad signaling-related genes including type I and III collagen in human and rat HSCs before and after 18 alpha-GA treatment were measured by real time PCR. The expression of related proteins was verified by western blot assay. The phosphorylation level of Smad2 and Smad3 was detected by immunocytochemistry. The DNA binding activities of SP-1, AP-1 and NF-kappa B were measured by both EMSA and ArrayStar transcription factor activity assay. Results: 18 alpha-GA could decrease the mRNA and protein expression of Smad3, type I and III collagen, increase the Smad7 expression in human and rat HSCs (P<0.05), and reduce phosphorylation level of Smad3 at 24 h and 48 h after treatment. The DNA binding activities of transcription factors were suppressed by 18 alpha-GA in human and rat HSCs at 24 h, and the activities reduced in a time dependent manner with the lowest activities at 48 h, especially for SP-1. Conclusion: 18 alpha-GA could inhibit the mRNA and protein expression of type I and III collagen in human and rat HSCs, which may be attributed to down-regulation of Smad3, up-regulation of Smad7, and inhibition of DNA binding activities of SP-1, AP-1 and NF-kappa B.