大黄酸对人肾小管上皮细胞血小板反应蛋白1和转化生长因子β1表达的影响

Effect of rhein on expression of thrombospondin-1 and transforming growth factor β1 in renal tubular cells

目的:探讨大黄酸对人肾小管上皮细胞(HK-2)血小板反应蛋白1(TSP-1)和转化生长因子β1(TGF-β1)表达的影响。方法:体外培养HK-2细胞,观察AngⅡ(1×10-7mol/L)诱导下低、中和高浓度的大黄酸(10μg/ml、20μg/ml和40μg/ml)对HK-2TSP-1、TGF-β1的mRNA和蛋白表达的影响。采用Real Time PCR检测细胞TSP-1、TGF-β1mRNA表达,采用Westernb lot检测TSP-1、TGF-β1蛋白表达。培养上清液中TGF-β1和活性TGF-β1检测采用ELISA法。结果:与空白对照组比较,AngⅡ(1×10-7mol/L)明显上调了HK-2TSP-1的mRNA表达(3.84±0.48vs1.32±0.19;P<0.05)、伴有TGF-β1的mRNA表达增加(3.68±0.46vs1.18±0.13;P<0.05)。同时AngⅡ(1×10-7mol/L)也上调TSP-1蛋白表达(0.5246±0.0936 vs 0.1015±0.0768;P<0.01),伴有TGF-β1蛋白表达的增加(0.5948±0.0624 vs 0.1264±0.0811;P<0.01)。与AngⅡ阳性对照组比较,中浓度和高浓度大黄酸明显抑制了AngⅡ(1×10-7mol/L)诱导的TSP-1、TGF-β1的mRNA(P<0.05,P<0.01)和蛋白表达(P<0.05,P<0.01)。中浓度大黄酸干预组总的TGF-β1含量和活性TGF-β1含量均减少,(P<0.05);高浓度大黄酸干预组总的TGF-β1含量和活性TGF-β1含量均明显减少(P<0.01)。结论:大黄酸能抑制人肾小管上皮细胞TSP-1、TGF-β1的mRNA和蛋白表达,抑制TGF-β分泌。

Objective：To investigate the effect of rhein on expression of thrombospondin-1 and transforming growth factor β1（TGF-β1） in renal tubular cells. Methodology ： HK-2 cells were stimulated by different concentrations of rhein （10,20,40μg/ml） in the presence of angiotensin-Ⅱ（AngⅡ, 10^-7mol/L）, untreated HK-2 cells were acted as normal control group,HK-2 cells treated by AngⅡ（10^-7 mol/L） were acted as positive group. Real Time PCR was used to detect the expression of TSP-1 mRNA and TGF-β1 mRNA. Western blot analysis was used to detect the expression of TSP-1 and TGF-β1 protein. ELISA was used to detect the levels of total and active TGF-β1. Results ：The expression of TSP-1 mRNA （3.84±0.48 vs 1.32±0.19, P 〈 0.05） and TGF-β1 mRNA （3.68±0. 46 vs 1.18±0. 13, P 〈 0. 05 ） was upregulated obviously when stimulated by AngII （10^-7 mol/L）. The protein expression of TSP-1 （0.5246±0.0936 vs 0.1015±0.0768, P 〈0.01 ）and TGF-β1（0.5948±0.0624 vs 0.1264±0.0811, P 〈0. 01 ）were also up-regulated simul- taneously when stimulated by AngⅡ（ 10^-7 mol/L）. The expression of TSP-1 and TGF-β1 induced by AngII was down-reg-ulated markedly with 20μg/ml and 40μg/ml of rhein in mRNA and protein levels（P 〈0. 05, P 〈0.01 ）. And rhein （20μg/ml and 40μg/ml） could reduce the levels of total and active TGF-β1（ P 〈 0. 05, P 〈0.01 ）. Conclusion： Rhein can inhibit the expression of TSP-1 and TGF-β1 in mRNA and protein levels and down-regulate the levels of total and active TGF-β1.