大鼠H9c2心肌细胞缺氧/复氧损伤模型肌浆网钙调控相关蛋白表达及当归补血汤的干预作用  

Danggui Buxue Decoction on expression of sarcoplasmic reticulum calcium regulatory protein in H9c2 myocardial cell model with hypoxia / reoxygenation damage

作　　者：周春刚[1] 汤加 李卿[1] 徐辰[1] 张志斌[1] 

机构地区：[1]无锡市中医医院中心实验室,江苏无锡214070

出　　处：《吉林中医药》2016年第6期601-604,共4页Jilin Journal of Chinese Medicine

基　　金：江苏省无锡市科技局科技支撑发展项目(CSEOIN1224)

摘　　要：目的建立缺氧/复氧大鼠H9c2心肌细胞损伤模型,观察心肌细胞肌浆网钙调控相关蛋白表达及当归补血汤对H9c2细胞缺氧/复氧损伤钙超载的影响。方法 JC-1染色流式细胞仪检测细胞线粒体膜电位,Fluo-3 AM钙离子荧光探针检测细胞内钙离子浓度,RT-PCR荧光相对定量检测大鼠心肌肌浆网Ca转运ATP酶(SERCA2a)、L型钙通道(CAV1.3)、兰尼碱受体(RyR1,RyR2)、受磷蛋白(PLB)、肌集钙蛋白(CASQ)的mRNA表达水平。结果与正常对照组比较,模型组细胞早期凋亡显著增加(P<0.05),细胞内钙离子浓度增加明显(P<0.05),CAV1.3mRNA表达水平显著升高(P<0.05),RyR1和CASQmRNA表达明显降低(P<0.05)。SERCA2a和PLB mRNA表达水平亦降低但差异无统计学意义。当归补血汤干预后,与模型组比较,能够显著降低复氧损伤细胞的早期凋亡率(P<0.05),细胞内钙离子浓度降低,差异具有统计学意义(P<0.05)。SERCA2a和CASQ mRNA水平显著增加(P<0.05),PLB和RyR1 mRNA虽有不同程度的增加,CAV1.3mRNA水平降低,但差异均无统计学意义(P>0.05)。结论 H9c2心肌细胞缺氧/复氧损伤后出现明显的钙超载及肌浆网钙调节蛋白调节紊乱现象,当归补血汤能够通过升高肌浆网钙调节蛋白SERCA2a和CASQ mRNA的表达,减轻复氧后钙超载,降低复氧损伤H9c2心肌细胞早期凋亡率。Objective To establish the model of hypoxia / reoxygenation damage in H9c2 rat myocardial cell and observe the expression of myocardial sarcoplasmic reticulum calcium regulatory protein and the effect of Danggui Buxue Decoction on calcium overload in H9c2 cells with hypoxia / reoxygenation injury. Methods JC-1 staining was used to detect the mitochondrial membrane potential by flow cytometry,Fluo-3 AM calcium fluorescence probe for the detection of intracellular calcium concentration,The expression level of sarcoplasmic reticulum Ca transport ATP enzyme（ SERCA2a）,L type calcium channel（ CAV1. 3）,ryanodine receptor（ RyR1,RyR2）,phospholamban（ PLB）,calsequestrin（ CASQ） mRNA were detected by RT-PCR. Results Compared with the normal control group,the early apoptosis rate and intracellular calcium concentration in model group increased significantly（ P〈0. 05）. The expression level of CAV1. 3 mRNA increased significantly（ P〈0. 05）,RyR1 and CASQ mRNA expression decreased obviously in model group cells（ P〈0. 05）. The expression level of SERCA2 a and PLB mRNA were also lower but the difference was not obviously significant. The early apoptosis rate and intracellular calcium concentration in Danggui Buxue Decoction intervention group decreased obviously（ P〈0. 05）. The expression level of SERCA2 a and CASQ mRNA increased significantly（ P〈0. 05）. RyR1 and PLB mRNA levels are increased in different degree,CAV1. 3mRNA expression decrease,but the differences were not significant（ P〈0. 05）,compared with the model group. Conclusion There are obvious calcium overload and sarcoplasmic reticulum calcium regulating proteins adjustment disorder in the model of H9c2 rat myocardial cell with hypoxia / reoxygenation damage,Danggui Buxue Decoction can increase the expression of the sarcoplasmic reticulumcalcium regulating protein SERCA2 a and CASQ mRNA after reoxygenation,alleviate calcium overload,and significantly reduce the early apoptosis rate of reoxygenation damage cells.

关 键 词：H9C2心肌细胞 肌浆网 钙超载 当归补血汤 

分 类 号：R289.9[医药卫生—方剂学]