摘要
目的:观察核转录因子-肌细胞增强因子2A(MEF2A)在肝星状细胞(HSC)活化过程中的动态变化及与HSC活化指标α-平滑肌肌动蛋白(α-SMA)的关系,探讨MEF2A在HSC活化过程中可能作用。方法:从SD大鼠肝脏分离HSC并培养在塑料细胞培养皿中,在体外培养过程中HSC逐步活化,分别收集新分离(0d)和经培养(1、2、3、4、5、6、7、8d)的HSC,采用实时荧光定量PCR检测各期细胞中MEF2A mRNA表达;Western blotting检测各期细胞中MEF2A和α-SMA的含量;凝胶阻滞实验(EMSA)方法检测MEF2ADNA结合活性。结果:实时荧光定量PCR结果表明新分离的HSC内有少量MEF2AmRNA表达,随着HSC培养逐步活化,MEF2A mRNA表达逐步增加;Western blotting检测结果发现新分离的HSC内有少量MEF2A及α-SMA,随着HSC培养逐步活化,细胞内MEF2A及α-SMA逐步增加,二者呈正相关;EMSA结果表明MEF2ADNA结合活性逐步增高。结论:随着HSC激活,MEF2A基因和蛋白表达量逐渐增加,且MEF2ADNA结合活性同步增高,MEF2A可能参与了HSC活化过程。
AIM ： To observe the changes in expression and activity of the transcription factor myocyte enhancer factor 2A （MEF2A） during hepatic stellate cells （HSC） activation, and to study the roles of MEF2A in the process of HSC activation. METHODS ： Cultured HSC was isolated from male sprague - dawley rat liver on plastic dishes and were used as model of activation. The freshly isolated （0 day） and cultured HSC at time points of 1st, 2nd, 3rd, 4th, 5th, 6th, 7th and 8th day were collected. Expression of MEF2A mRNA was detected by real - time quantitative PCR. MEF2A and α -smooth muscle actin （α -SMA, a marker for activated HSC） were tested by Western blotting. Meanwhile, the MEF2A DNA binding activity was determined by electrophoretic mobility shift assays （EMSA）. RESULTS ： The expression of MEF2A mRNA was small amounts in the freshly isolated HSC and increased gradually after culture on plastic dishes. Western blotting revealed that the freshly isolated HSC expressed very low levels of MEF2A and α- SMA. The proteins of MEF2A and ot - SMA were increased gradually in the process of HSC activation. Increased MEF2A protein was correlated with α- SMA. EMSA revealed that MEF2A DNA binding activity was increased gradually during HSC activation. CONCLUSION： In the process of HSC activation, expression and activity of MEF2A are increased gradually, indicating a role in HSC activation.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2010年第2期333-336,共4页
Chinese Journal of Pathophysiology
基金
浙江省中医药管理局科研基金资助项目(No.2006C129)
关键词
肌细胞增强因子2
肝星状细胞
Myocyte enhancer factor 2
Hepatic stellate cells
