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目的检测TNNI3K基因过表达对成年大鼠心肌细胞肌丝钙敏感度变化的影响,为进一步研究TNNI3K基因在心肌收缩功能、心肌肥厚、心衰及其它心脏疾病中的作用提供了可靠有效的研究模型。方法恒压Langedoff灌流装置逆向灌流法分离成年大鼠心肌细胞,之后对该心肌细胞进行逐级复钙。然后,将心肌细胞接种于改良的M199培养基中。分别感染Ad.GFP病毒和Ad.TNNI3K病毒,24小时后,以IonOptix单细胞可视化动缘系统检测心肌细胞肌丝钙敏感度。结果成功分离培养了成年大鼠心肌细胞,并在该细胞中成功过表达了TNNI3K基因。钙敏感度测定结果表明,TNNI3K基因能够增强心肌细胞肌丝钙敏感度。结论在本实验成功分离培养的成年大鼠心肌细胞中以腺病毒载体过表达TNNI3K基因,可增强心肌细胞肌丝的钙敏感度,该模型为进一步研究TNNI3K基因在心肌收缩功能中的作用及机制奠定了基础。
Objective To detect the effect of TNNI3K overexpression on changes of myofilament calcium sensitivity in adult rat myocardial cells and to provide a reliable and effective research model for the further study on the role of TNNI3K in myocardial systolic function, cardiac hypertrophy, heart failure and other heart diseases . Methods Myocardial cells were isolated from cardiomyocytes of adult rat by reverse perfusion with Langedoff perfusion device. Cardiomyocytes were then seeded in modified M199 medium. The Ad.GFP and Ad.TNNI3K viruses were respectively infected. After 24 hours, the myofilament calcium sensitivity of cardiomyocytes was detected by IonOptix single cell visualization kinetic system. Results The adult rat cardiomyocytes were successfully isolated and cultured, and TNNI3K gene was overexpressed successfully in this cell line. The results of calcium sensitivity test showed that TNNI3K gene can enhance myofilament calcium sensitivity in cardiomyocytes. Conclusion The overexpression of TNNI3K in adult rat cardiomyocytes successfully isolated in this experiment can enhance the calcium sensitivity of myofilament in cardiac myocytes. This model is to further investigate the role and mechanism of TNNI3K in myocardial systolic function Foundation.