Mechanistic study of anti-hepatitis B virus and gluconeogenesis of Scutellariae radix in human hepatoma cells

• Main Authors: Chia-I Shen, 沈佳怡
• Other Authors: Sheau-Farn Yeh
• Format: Others
• Language: zh-TW
• Published: 2010
• Online Access: http://ndltd.ncl.edu.tw/handle/85881416143091150940

碩士 === 國立陽明大學 === 生化暨分子生物研究所 === 98 === The Chinese herbal medicine, Xiao-Chai-Hu-Tang, is a seven components prescription used for liver disease treatment. Our laboratory has found Scutellaria radix (1S), a major component of Xiao-Chai-Hu-Tang, is in possession of suppressing hepatitis B virus surface antigen (HBsAg) secretion in HepG2/A2 cells. In this thesis, human hepatoma cell lines 1.3ES2, HepG2/A2 and Hep3B/T2 were used to elucidate the mechanism of anti-HBV effect of Scutellaria radix including two of its components, Chrysin and Wogonin. In this study, it was found that 1S, Chrysin and Wogonin not only suppressed HBsAg production in cultured HepG2/A2 cells but also decrease HBV transcripts of RNA, core protein and viral particles in 1.3ES2 cells. To further understand the molecular mechanism of suppression of HBV gene expression by 1S, Chrysin and Wogonin, luciferase reporter assay was used for examining the viral promoter activity. The results showed that 1S, Chrysin and Wogonin suppressed viral core promoter expression. Such suppressive effects are liver-specific because the repressive effect was not observed in human embryonic kidney 293T cells. Furthermore, ectopic expression of liver-specific transcription factors in HepG2/A2 cells could partially reverse the suppression effect of 1S, Chrysin and Wogonin on core promoter. Using western blotting, 1S, Chrysin and Wogonin not only showed the decrease of endogenous PPARγ protein but also 1S suppress endogenous HNF4α protein expression, therefore, these three compounds may inhibit hepatitis B virus through different transcription factors of core promoter in molecular mechanisms. Using the treatment of 8-Br-cAMP and dexamethasone of 3B/T2 cells to mimic the starvation state of cells, it was found the HBV core promoter activity was stimulated. Gluconeogenesis genes, glucose-6-phosphatase (G6Pase) and phosphenolpyruvate carboxykinase (PEPCK) were activated. The expression of transcription coactivator PGC1α proteins were elevated in starvation state of Hep3B/T2 cells. Results showed that 1S effectively suppressed 8-Br-cAMP and dexamethasone induced G6Pase mRNA expression and HBV core promoter activity, however Chrysin and Wogonin had no effect on G6Pase and PEPCK. Taken together, 1S, Chrysin and Wogonin may act through modulating hepatic transcriptional machinery to suppress HBV viral gene expression and virus production. However, turns on the gluconeogenic program, only 1S showed suppression of G6Pase.