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目的筛选联合用药作用大鼠肝星状细胞(HSC-T6)后差异表达的蛋白质,以探讨联合用药抑制肝星状细胞增殖的机制。方法以牛磺酸、表没食子儿茶素没食子酸酯(EGCG)和三羟基异黄酮三者组成的联合用药作用HSC-T6后,提取细胞总蛋白,运用同位素标记的i TRAQ技术结合高效液相色谱-电喷雾串联质谱(LC-ESI-MS/MS)的定量蛋白质组学方法,分析和鉴定差异表达的蛋白质,运用生物信息学分析差异表达的蛋白质功能。结果质谱共分析鉴定到727种蛋白质。3次实验分别鉴定到的差异表达蛋白质数量分别为85,102,94个,在3次重复均鉴定到的差异表达蛋白为52个,其中上调蛋白24个,下调蛋白28个。生物信息学分析表明,差异表达的蛋白参与翻译后修饰、转录、重组和信号传导等过程,在这些蛋白质中既有与肝纤维化发生、发展直接相关的蛋白质如基质金属蛋白酶抑制剂1、层黏连蛋白、前纤维蛋白2等,也有在蛋白质的相互作用中处于功能网络交叉点的蛋白质如细胞周期素依赖性激酶4、肝癌衍生生长因子、谷氨酸脱氢酶1和丝氨酸/精氨酸剪接因子9等。结论联合用药抑制肝星状细胞增殖的作用,可能是通过调控多种蛋白质的表达来实现,所获得的差异表达蛋白可能在肝纤维化发生发展和药物抗肝纤维化作用过程中扮演重要角色。
Objective To screen differentially expressed proteins of rat hepatic stellate cells (HSC-T6) in combination therapy to explore the mechanism of combined inhibition of hepatic stellate cell proliferation. Methods HSC-T6 cells were treated with a combination of taurine, epigallocatechin gallate (EGCG) and trihydroxyisoflavone (HSC-T6), and the total cellular protein was extracted. Using isotope labeled iTRAQ technology combined with high performance liquid chromatography Chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS / MS) was used to analyze and identify differentially expressed proteins. Bioinformatics analysis of differentially expressed proteins was performed. Results A total of 727 proteins were identified by mass spectrometry. The number of differentially expressed proteins identified in three experiments were 85, 102, and 94, respectively. Among the three replicates, 52 differentially expressed proteins were identified, including 24 up-regulated proteins and 28 down-regulated proteins. Bioinformatics analysis showed that the differentially expressed proteins involved in post-translational modification, transcription, recombination and signal transduction process, among these proteins are both directly related to the occurrence and development of liver fibrosis, such as matrix metalloproteinase inhibitor 1, layer Such as cyclin-dependent kinase 4, HCC-derived growth factor, glutamate dehydrogenase 1, and serine / arginine, as well as proteins that cross the functional network in protein interactions Acid splicing factor 9 and so on. Conclusions The effect of combination therapy on the inhibition of hepatic stellate cell proliferation may be achieved by regulating the expression of various proteins. The differentially expressed proteins may play an important role in the development of hepatic fibrosis and anti-hepatic fibrosis.