结缔组织生长因子诱导大鼠心肌细胞肥大的作用及其与细胞外信号调节激酶1/2的关系

Connective Tissue Growth Factor Induce Cardiac Myocytes Hypertrophy in Rats and Its Relation with ERK1/2

目的观察结缔组织生长因子(CTGF)诱导大鼠心肌细胞肥大的作用,探讨其作用与细胞外信号调节激酶1/2(ERK1/2)的关系。方法以培养的新生Sprague-Dawley(SD)大鼠心肌细胞为实验模型,用图象分析法测定心肌细胞表面积,用[3H]亮氨酸掺入法测定心肌细胞蛋白合成速率,用考马斯亮兰法测定心肌细胞蛋白含量,用蛋白免疫印迹法(Westernblot)测定心肌细胞总ERK1/2(t-ERK1/2)与磷酸化ERK1/2(p-ERK1/2)的蛋白表达水平。结果(1)随着CTGF浓度的增加,心肌细胞表面积呈剂量依赖性增加,其中10、25、50、100μg/L的CTGF组心肌细胞表面积分别为(929·9±132·2)、(1411·3±129·2)、(1732·0±153·0)、(2040·6±205·4)μm2,均明显高于对照组心肌细胞表面积[(606·3±72·7)μm2,P均<0·01];100μmol/L的PD98059明显减少CTGF诱导的心肌肥大(P<0·01)。(2)随着CTGF浓度的增加,心肌细胞蛋白合成速率与蛋白含量呈剂量依赖性增加,CTGF组心肌细胞[3H]亮氨酸掺入率明显高于对照组(P<0·01);ERK1/2抑制剂PD98059明显减少CTGF诱导的心肌细胞[3H]亮氨酸掺入率及蛋白含量(P<0·01)。(3)随着CTGF浓度的增加,心肌细胞p-ERK1/2表达呈剂量依赖性增高,5、10、25、50、100μg/L的CTGF组的心肌细胞p-ERK1/2表达明显高于对照组,而t-ERK1/2在各组表达差异不明显。结论CTGF可诱导心肌细胞肥大,该作用可能是通过ERK1/2的磷酸化来实现的。

Objective To study the effects of connective tissue growth factor （CTGF） on the hypertrophy of cultured rat cardiac myocytes and investigate whether extracellular regulated protein kinases 1/2 （ERK1/2） is involved in myocytes hypertrophy induced by CTGF. Methods Neonatal cardiomyocytes were obtained from Sprague-Dawley（SD） rats. Image analysis system was used to measure cell surface area; protein synthesis of myocytes was measured via [^3H]-leucine incorporation rate, Coomassie Brilliant blue method was used to measure cardiac myocytes protein content and Western blot was used to study the protein expression level of t-ERK1/2 and p-ERK1/2. Results （ 1 ） CTGF increased cardiac myocytes surface area in a dose dependent manner. The cell surface of cardiac myocytes stimulated by 10, 25, 50, 100 μg/L CTGF was （929.9±132.2）, （1411.3±129.2）, （1732. 0±153. 0）, （2040. 6 ±205. 4 ）μm^2 , respectively, which were significantly higher than that of control group （P〈0. 01 ）. PD98059 substantially inhibited the increase of cardiac myocytes surface area induced by CTGF （P〈0.01 ）. （ 2 ）CTGF increased cardiac myocytes protein synthesis and protein content in a dose dependent manner. PD98059 substantially inhibited the increase of [^3 H]-leucine incorporation rate induced by CTGF （P〈 0. 01 ）. （3）CTGF increased the protein level of p-ERK1/2 in a dose and time dependent manner. The protein level of p-ERK1/2 was significantly higher than that of control group, while the protein level of t-ERK1/2 in all groups was not significantly different. Conclusion Cardiac myocytes hypertrophy induced by CTGF probably was mediated via ERK1/2 phosphorylation.