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目的 研究中药复方861对血管紧张素Ⅱ(AngⅡ)引起的肝星状细胞增殖、细胞内钙浓度及胶原合成的影响。方法 体外实验对象为肝星状细胞系(HSC-T6)。细胞增殖采用甲基-~3H胸腺嘧啶核苷(~3H-TdR)掺入法,细胞内钙浓度测定用Fluo-3/AM(乙酰氧甲基酯)标记后共聚焦显微镜下测定荧光光密度值。培养上清液中Ⅰ型胶原(CoL-Ⅰ)用酶联免疫吸附(ELISA)法测定,Ⅲ型前胶原肽(PⅢP)的浓度用放免法检测。HSC-T6细胞CoL-Ⅰ及Ⅲ型胶原(CoL-Ⅲ)mRNA的水平用RT-PCR方法检测。结果 AngⅡ可促进HSC-T6细胞的增殖和细胞内钙浓度的增加,复方861可阻断这些变化。复方861也可降低HSC-T6细胞CoL-Ⅰ及PⅢP的分泌和CoL-Ⅰ及CoL-ⅢmRNA的水平。结论 复方861通过作用于血管紧张素Ⅱ1型受体(AT1R)抑制肝星状细胞的增殖及胶原蛋白的合成,这可能是其抗纤维化的机理之一。
Objective To study the effect of Chinese Herbal Compound 861 on hepatic stellate cell proliferation, intracellular calcium concentration and collagen synthesis induced by angiotensin II (Ang II). Methods The in vitro test subject was a hepatic stellate cell line (HSC-T6). Cell proliferation was performed using methyl-~3H thymidine (~3H-TdR) incorporation method, intracellular calcium concentration was measured with Fluo-3/AM (acetoxymethyl ester) label, and fluorescence intensity was measured under a confocal microscope. value. The type I collagen (CoL-I) in the culture supernatant was determined by enzyme-linked immunosorbent assay (ELISA), and the concentration of type III procollagen peptide (PIIIP) was detected by radioimmunoassay. The levels of CoL-I and type III collagen (CoL-III) mRNA in HSC-T6 cells were detected by RT-PCR. Results AngII can promote the proliferation of HSC-T6 cells and increase the intracellular calcium concentration. Compound 861 can block these changes. Compound 861 also reduced CoL-I and PIIIP secretion and CoL-I and CoL-III mRNA levels in HSC-T6 cells. Conclusion Compound 861 inhibits the proliferation of hepatic stellate cells and the synthesis of collagen by acting on angiotensin II type 1 receptor (AT1R), which may be one of the mechanisms of anti-fibrosis.