信号转导及转录激活因子3对心肌细胞基质金属蛋白酶-10活性的影响

Effect of Signal Transducer and Activator of Transcription 3 on Up-Regulation of Matrix Metalloproteinase-10 Activity in Neonatal Rat Cardiomyocytes

目的:探讨信号转导及转录激活因子3(STAT3)对心肌细胞基质金属蛋白酶-10(MMP-10)活性的影响。方法:以炎症细胞因子C反应蛋白(CPP)诱导基质金属蛋白酶-10上调的原代乳鼠心肌细胞为研究对象。使用STAT3的特异磷酸化抗体,用蛋白免疫印迹杂交技术检查STAT3的磷酸化水平。提取细胞核蛋白,应用滤板法检测核内转录因子STAT3的脱氧核糖核酸(DNA)结合活性,并转染STAT3小分子干扰核糖核酸(siRNA),进一步分析转录因子STAT3对基质金属蛋白酶-10的调节作用。结果:磷酸化的STAT3在C反应蛋白刺激后5 min即可出现高表达,一直持续至2 h,而总的STAT3不随C反应蛋白作用的时间而改变。C反应蛋白作用不同时间,核内STAT3的DNA结合活性均可明显增加,且在12 h达到高峰(P<0.01)。加入STAT3 siRNA后,基质金属蛋白酶-10的活性明显减弱(P<0.01),STAT3的蛋白表达以及核内STAT3的DNA结合活性(P<0.01)也均降低。结论:STAT3作为转录因子能够促进C反应蛋白诱导基质金属蛋白酶-10活性的增加。

Objective ： To investigate the effect of signal transducer and activator of transcription 3 （ STA33 ） on up-regulation of matrix metalloproteinase-10（MMP-10） activity induced by C-reactive protein （CRP） in neonatal rat cardiomyocytes. Methods：We studied the neonatal rat cardiomyocytes with elevated MMP-10 level which was induced by CRP. Phospho- specific antibody for STA33 was used to determine the expression of STA33 by Western blot analysis. Cardiomyocytes nuclear pro- tein was extracted for filter assay to monitor STAT3 DNA binding activity in MMP-10 promoter. The cells were further transfected with small infereing RNA （siRNA） of STAT3 to observe the role of STA33 in up-regulation of MMP-10 activity induced by CRP. Results：The phosphorylated STAT3 level increased 5 minutes after CRP treatment and lasted for 2 hours, while the total STAT3 level remained the same. Nuclear STA33 DNA-binding activity was significantly increased after CRP treatment at different time points, and the maximal level was obtained at 12 hours,P〈0. 01. With STAT3 siRNA transfection,MMP-10 activity was sig- nificantly inhibited,P〈0. 01, STArI3 protein expression and DNA binding activity were also inhibited. Conclusion： STAT3 can mediate up-regulation of CRP-induced MMP-10 activity in neonatal rat cardiomyocytes.