肝纤维化-肝癌病理机制与MAPK通路调控下的TGF-β/Smad信号相关，肝癌时Smad3连接区磷酸化（pSmad3L）增强。复方芪参提取物通过促进Smad3 C末端磷酸化（pSmad3C）和减少pSmad3L延缓肝癌发展。丹酚酸B抗肝纤维化亦与TGF-β/Smad相关。推测：pSmad3C/pSmad3L转换可能是促肝癌发展的关键机制；丹酚酸B可能影响pSmad3L/3C及下游靶基因而抑制肝癌发生。故拟采用ES打靶技术建立的Smad3C/3L磷酸化位点突变小鼠并诱导其肝纤维化→肝癌；Smad3C/3L磷酸化位点突变质粒（3-SA和EPSM）转染HepG2细胞，免疫沉淀、WB、qRT-PCR等测蛋白和靶基因，观察Smad3磷酸化位点突变对肝纤维化-肝癌发展进程的调控及丹酚酸B的干预。阐明肝纤维化-肝癌进程中的pSmad3C/p21//pSmad3L/c-Myc转换机制以及丹酚酸B作用机制。

The pathomechanism of hepatofibrosis–hepatocarcinogenesis is related with TGF-β/Smad signal modulated by MAPK pathway. The phosphorylation of Smad3 at linker region (pSmad3L) increased in hepatocarcinogenesis period. Our previous results showed that CASE could suppress hepatocellular carcinoma progression by increasing the phosphorylation of Smad3 at C-terminal (pSmad3C) and inhibiting pSmad3L. Furthermore Salvianolic acid B has also the action of anti-hepatic fibrosis via TGF-β/Smad signal. We presume that the shift of pSmad3C/pSmad3L might be the important mechanisms of hepatocarcinogenesis, and Salvianolic acid B might effect hepatocarcinogenesis via pSmad3L/pSmad3C and their downstream target gene. Therefore, we propose to investigate the regulation of phosphorylation sites mutation of Smad3 at C-terminal and linker region in hepatofibrosis-hepatocarcinogenesis progression and the action of Salvianolic acid B by using pSmad3C and/or pSmad3L mutant gene knock-in mice in vivo and by using transfer Smad 3 mutant plasmids (3-SA or EPSM) into HepG2 cells in vitro. The expressing level of the protein and gene in the signaling pathway will be detected by using immunoprecipitate, western blot and qRT-PCR methods. This study attemps to elucidate the transducer mechanism of pSmad3C / p21 // pSmad3L / c-Myc in hepatofibrosis-hepatocarcinogenesis progression and Salvianolic acid B’s anti- hepatocarcinogenesis effect.