Liu Ping, Liu Cheng-Hai, Wang Hai-Nan, Hu Yi-Yang, Liu Cheng
Institute of Liver Disease, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China.
Acta Pharmacol Sin. 2002 Aug;23(8):733-8.
To investigate the mechanism of salvianolic acid-B (SA-B) action against liver fibrosis relating to mediating hepatic stellate cell (HSC) activation and transforming growth factor-beta1(TGF-beta1) intracellular signal transduction.
HSC was isolated from normal rat through in situ perfusion of liver with pronase E and density-gradient centrifugation with 11 % nycondenz, then cells were subcultured. Cell proliferation was observed by [3H]TdR uptake. Cellular collagen deposition was measured with Ponceau S stain and semi-quantified with image analytic system. Type I collagen secretion in the supernatant was detected with ELISA. The gene expression of type I pro-collagen was analyzed by RT-PCR. The supernatant was acidified and active TGF-beta1 contents were assayed with ELISA. Mitogen-activated protein kinase (MAPK) activity was analyzed with immunoprecipitation and Western blot.
SA-B 0.1, 1, 10, and 100 micromol/L suppressed HSC proliferation concentration-dependently as determined by [3H]TdR uptake by 94.1 %, 82.4 %, 62.7 %, and 4 % of the control respectively (P<0.05 or P<0.01). SA-B 1, 10, and 100 micromol/L inhibited soluble type I collagen secretion by 75.3 %, 69.8 %, and 63.5 % of the control and decreased the matrix collagen deposition to 86.2 %, 75.4 %, and 73.4 % (P<0.05 or P<0.01). SA-B 1 and 10 micromol/L decreased the cell active TGF-beta1 secretion by 63.3 % and 15.6 % of the control, down-regulated pro-collgen alpha1(I) mRNA expression to 77.0 % and 51.8 % respectively (P<0.05). SA-B 1 and 10 micromol/L also inhibited MAPK activity by 1 to 2 fold respectively.
SA-B inhibited HSC proliferation and collagen production as well as decreased the cells' TGF-beta1 autocrine and MAPK activity, which might contribute to the mechanism of SA-B action against hepatic fibrosis.
探讨丹酚酸B（SA-B）抗肝纤维化的作用机制，涉及介导肝星状细胞（HSC）活化及转化生长因子-β1（TGF-β1）细胞内信号转导。
通过用链霉蛋白酶E原位灌注肝脏及用11% Nycodenz进行密度梯度离心从正常大鼠分离HSC，然后进行细胞传代培养。通过[3H]TdR摄取观察细胞增殖。用丽春红S染色测量细胞胶原沉积并用图像分析系统进行半定量。用ELISA检测上清液中I型胶原分泌。通过RT-PCR分析I型前胶原的基因表达。将上清液酸化并用ELISA测定活性TGF-β1含量。用免疫沉淀和Western印迹分析丝裂原活化蛋白激酶（MAPK）活性。
通过[3H]TdR摄取测定，0.1、1、10和100 μmol/L的SA-B分别以浓度依赖方式抑制HSC增殖，抑制率分别为对照组的94.1%、82.4%、62.7%和4%（P<0.05或P<0.01）。1、10和100 μmol/L的SA-B分别抑制可溶性I型胶原分泌至对照组的75.
