丹参酮ⅡA抑制心肌细胞的纤维化  

Tanshinone ⅡA inhibits cardiac fibrosis

作　　者：周代星[1] 李智慧[1] 占成业[1] 张莉伟[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院急诊科,湖北省武汉市430030

出　　处：《中国组织工程研究》2013年第20期3715-3722,共8页Chinese Journal of Tissue Engineering Research

基　　金：湖北省自然科学基金资助项目(2009CDB092;2007ABA272)~~

摘　　要：背景:转化生长因子β-Smads信号通路是心肌纤维化的关键通路;丹参酮ⅡA具有抑制心肌纤维化作用。目的:验证丹参酮ⅡA对转化生长因子β1致心肌纤维化的作用并探讨其机制。方法:取出生24h内的SD大鼠心脏,利用酶消化法结合差速贴壁体外培养心肌成纤维细胞。取上述第3代心肌成纤维细胞,用5μg/L转化生长因子β1培养15,30,60,120min或6,12,24h后收集细胞,用不同浓度(10-5mol/L、10-4mol/L)丹参酮ⅡA预处理2h后再加入5μg/L转化生长因子β1培养120min或24h后收集细胞,并设空白对照组。免疫细胞化学染色法进行细胞鉴定,反转录聚合酶链反应检测结缔组织生长因子及Ⅰ型胶原蛋白mRNA表达,Western Blot检测Smad7及磷酸化Smad3蛋白表达。免疫组织化学染色及免疫荧光法检测磷酸化Smad3、结缔组织生长因子及Ⅰ型胶原蛋白表达。结果与结论:转化生长因子β1在一定范围内以时间依赖方式诱导结缔组织生长因子、Ⅰ型胶原、磷酸化Smad3及Smad7的表达,刺激终末结缔组织生长因子、Ⅰ型胶原mRNA的表达量明显上升(均P<0.01);磷酸化Smad3及Smad7蛋白表达量在刺激后1h达到峰值,表达量显著增加(均P<0.01)。高浓度丹参酮ⅡA预处理可显著下调磷酸化Smad3、结缔组织生长因子及Ⅰ型胶原表达(均P<0.01)。两种浓度的丹参酮ⅡA预处理均可上调Smad7蛋白表达(P<0.05,P<0.01)。提示丹参酮ⅡA对心肌纤维化有抑制作用,可能与其上调Smad7蛋白表达,抑制转化生长因子β1诱导的Smad3磷酸化,部分阻断转化生长因子β1-Smads信号通路有关。BACKGROUND： Transforming growth factor β-Smads is the critical path of cardiac fibrosis, and tanshinone ⅡA can limit cardiac fibrosis. OBJECTIVE： To explore the effect of tanshinone ⅡA on the cardiac fibrosis induced by transforming growth factor-β1 and the possible mechanisms. METHODS： Cardiac fibroblasts were isolated from cardiac tissues of neonatal Sprague-Dawley rats by the trypsin digestion and differential adhesion method. Passage 3 cel s were treated with 5 μg/L transforming growth factor β1 alone or pretreated with tanshinone ⅡA at different concentrations （10-5 mol/L and 10-4 RESULTS AND CONCLUSION： Transforming growth factor-β1 induced the expression of connective tissue growth factor, type Ⅰ col agen, phosphorylated Smads and Smad7 in a time-dependant manner. The mRNA expression of connective tissue growth factor and type Ⅰ col agen was significantly increased 24 hours after transforming growth factor-β1 stimulation （P 〈 0.01 for al ）. The protein expression of phosphorylated Smad3 and Smad7 reached a peak 1 hour after transforming growth factor-β1 stimulation, much higher than the baseline level （P 〈 0.01 for al ）. Pretreatment with high concentration of tanshinone ⅡA resulted in a decrease in the expression of phosphorylated Smad3, connective tissue growth factor and type Ⅰ col agen （P 〈 0.01）. The protein expression of Smad7 was substantial y upregulated after pretreatment with two concentrations of tanshinone ⅡA as compared with that in the cel s at 2 hours post transforming growth factor-β1 stimulation （P 〈 0.05, P 〈 0.01）. Tanshinone ⅡA may exert an inhibitory effect on cardiac fibrosis by upregulating the expression of Smad7, suppressing the transforming growth factor-β1-induced phosphorylated Smad3 and partial y blocking the transforming growth factor-β1-Smads signaling pathway. mol/L）. Immunocytochemistry was used for cel identification, reverse transcription-PCR for detection of the mRNA expression of connective tissue growth

关 键 词：组织构建 组织构建与中医药 丹参酮ⅡA 心肌成纤维细胞 心肌纤维化 转化生长因子Β1 SMAD3 SMAD7 结缔组织生长因子 Ⅰ型胶原 信号通路 细胞培养 省级基金 

分 类 号：R318[医药卫生—生物医学工程]