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目的探讨心衰心室重构中血管紧张素Ⅱ(AngⅡ)对人α1(Ⅰ)胶原基因启动子活性的调控作用和中药单体丹参酮ⅡA(tanshinoneⅡA,TSN)药物干预的影响环节。方法将人α1(I)胶原基因上游-2.5 Kb长度的启动子片段与氯霉素乙酰基转移酶(CAT)报告基因组成重组体pCOLH2.5,采用脂质体法转染至培养的大鼠心肌成纤维细胞和ELISA法测定CAT活性的方法,观察不同浓度TSN对pCOLH2.5活性及由AngⅡ诱导的pCOLH2.5活性的影响。结果AngⅡ可剂量依赖性地促进pCOLH2.5的活性,表现为其剂量增大时重组体CAT活性相对增加,与对照组比较有显著性差异(P<0.05)。TSN能抑制pCOLH2.5的活性,并随着药物浓度的增加,其抑制作用亦增强。与对照组、低浓度组比较,差异有显著性(P<0.05)。而且在与AngⅡ共同作用时,pCOLH2.5 CAT的升高幅度明显减少(P<0.01)。结论在心肌成纤维细胞中,AngⅡ具有促进人α1(Ⅰ)胶原基因启动子的转录活性作用。TSN能下调人α1(Ⅰ)胶原基因启动子的激活,并可部分拮抗AngⅡ的促进作用。
Objective To investigate the effect of angiotensin II (Ang II) on the promoter activity of human α1(I) collagen gene in heart failure ventricular remodeling and the influence of intervention of traditional Chinese medicine monomer tanshinone IIA (TSN). Methods The recombinant plasmid pCOLH2.5 was constructed by using a 2.5-kb-long promoter fragment and a chloramphenicol acetyltransferase (CAT) reporter gene upstream of the human α1(I) collagen gene and transfected into cultured rat by liposome method. The method of measuring CAT activity by myocardial fibroblasts and ELISA was used to observe the effect of different concentration of TSN on the activity of pCOLH2.5 and the activity of pCOLH2.5 induced by AngII. Results AngII could promote the activity of pCOLH2.5 in a dose-dependent manner. The expression of CAT activity was increased when the dose was increased, which was significantly different from that of the control group (P<0.05). TSN can inhibit the activity of pCOLH2.5, and with the increase of drug concentration, its inhibitory effect is also enhanced. Compared with the control group and low concentration group, the difference was significant (P<0.05). In addition, the increase of pCOLH2.5 CAT was significantly reduced when combined with AngII (P<0.01). Conclusion In cardiac fibroblasts, AngII can promote the transcriptional activity of human α1(I) collagen gene promoter. TSN can down-regulate the activation of human α1(I) collagen gene promoter and partially antagonize the promotion of AngII.