摘要
背景:转化生长因子β1通过Smads信号通路刺激心脏成纤维细胞增殖与分化是心肌纤维化最重要的发生机制之一。前期研究证实丹参酮ⅡA能有效抑制心肌纤维化,但是否通过阻断转化生长因子β1/Smads信号通路起作用尚不清楚。目的:观察丹参酮ⅡA对大鼠心脏成纤维细胞内转化生长因子β1信号转导的影响。方法:采用胰酶消化法和差速贴壁法获取新生SD大鼠心脏成纤维细胞,应用5μg/L转化生长因子β1刺激及不同浓度丹参酮ⅡA(10-6,10-5和10-4mol/L)。用反转录聚合酶链反应法和免疫蛋白印迹法分别检测转化生长因子β1刺激后6,12和24h纤维连接蛋白的表达,免疫蛋白印迹法检测转化生长因子β1刺激后15,30,60和120min的Smads蛋白表达。结果与结论:纤维连接蛋白mRNA和蛋白表达量在转化生长因子β1刺激6h后开始呈现上升趋势,至作用24h时分别增加1.3倍和1.8倍(P<0.01);磷酸化Smad2/3蛋白表达量在转化生长因子β1刺激15min后开始上升,1h达到高峰,2h后虽有所下降,但仍较刺激前增加3.9倍(P<0.01)。丹参酮ⅡA(10-5和10-4mol/L)预处理可下调纤维连接蛋白和磷酸化Smad2/3表达(P<0.05或P<0.01),而且效应呈剂量依赖性。由此可知,转化生长因子β1在一定范围内以时间依赖方式诱导纤维连接蛋白及其mRNA和磷酸化Smad2/3表达。丹参酮ⅡA抗心肌纤维化作用可能与其抑制转化生长因子β1诱导的Smad2/3磷酸化,阻断心脏成纤维细胞内转化生长因子β1/Smads信号通路有关。
BACKGROUND：One of important mechanisms underlying myocardial fibrosis is that transforming growth factor β1（TGF-β1） stimulates the proliferation and differentiation of cardiac fibroblasts via Smads signaling pathway.Previous studies have confirmed that tanshinone ⅡA can effectively inhibit myocardial fibrosis.But whether blockage of TGF-β1/Smads signaling pathway is involved in this process remains unclear.OBJECTIVE：To investigate the effects of tanshinone ⅡA on TGF-β1 signal transduction in rat cardiac fibroblasts.METHODS：Neonatal rat cardiac fibroblasts were harvested by trypsin digestion and differential attachment and treated with 5 μg/L TGF-β1 and different concentrations of tanshinone ⅡA（10-6,10-5 and 10-4 mol/L）.At 6,12,and 24 hours after TGF-β1 application,fibronectin expression was detected by reverse transcription-polymerase chain reaction and Western blot analysis.At 15,30,60,and 120 minutes after TGF-β1 application,Smads protein expression was determined by Western blot analysis.RESULTS AND CONCLUSION：Fibronectin mRNA and protein expression began to increase at 6 hours after TGF-β1 application and was 1.3 and 1.8 times higher than initial level,respectively（P〈0.01）,at 24 hours after TGF-β1 application.Phosphorylated Smad2/3 protein expression began to increase at 15 minutes after TGF-β1 application,peaked at 1 hour,decreased at 2 hours,but it was still 3.9 times higher than initial level（P〈0.01）.Tanshinone ⅡA（10-5 and 10-4 mol/L） pretreatment downregulated fibronectin and phosphorylated Smad2/3 expression（P 0.05 or P〈0.01） in a dose-dependent manner.These findings demonstrate that TGF-β1 induced fibronectin protein and mRNA expression and Smad2/3 protein expression in a time-dependent manner.Tanshinone ⅡA against myocardial fibrosis was likely related to its inhibition of TGF-β1-induced Smad2/3 phosphorylation and blockage of TGF-β1/Smads signaling pathways within cardiac fibroblasts.
出处
《中国组织工程研究与临床康复》
CAS
CSCD
北大核心
2010年第37期7021-7025,共5页
Journal of Clinical Rehabilitative Tissue Engineering Research
基金
the Natural Science Foundation of Hubei Province,No.2007ABA272
2009CDB092~~
