普通大鼠心肌细胞的分离培养和收缩舒张观察

Isolation and Culture of Adult Rat Cardiomyocytes and Measurement of Systolic/Diastolic Function

目的:建立稳定的成年大鼠心肌细胞分离和培养方法,以及利用IonOptixTM系统观察成年大鼠离体心肌细胞收缩/舒张功能的变化。方法:将成年大鼠心脏挂于Langendorff装置上灌流,胶原酶消化分离成年大鼠心肌,Laminin附壁无血清培养心肌细胞。光镜观察单个心肌细胞的形态学特点。用含氧台氏液灌流并予电场刺激(0.5Hz,3ms),利用IonOptixTM系统检测其收缩/舒张功能。结果:平均单个心脏所分离的心肌细胞的收获量为(5～7)×106个,长杆状细胞横纹清晰,无血清培养72h后,细胞保持正常完整的形态结构。采用IonOptixTM系统检测心肌细胞收缩/舒张功能变化,测定其收缩幅度为(11.84±2.21)%。结论:本方法可以对成年大鼠心肌细胞进行良好的分离培养。IonOptixTM系统可实时同步观察和记录心肌细胞机械功能的变化,因而可直接反映各种不同病理情况下细胞水平的心肌功能改变。

Objective： To establish a stable method for isolation and culture of adult rat cardiomyocytes, and to observe single cardiomyocytes with a IonOptix^TM system. Methods： The isolated adult rat heart was hanged on to the Langendorff apparatus for perfusion and collagenase digestion. The single cardiomyocytes were cultured in serum-free medium with laminin-covered dishes. The morphological features of cardiomyocytes were observed with microscope. With perfusion and field-stimulation（0.5 Hz, 3 ms）, the cardiomyocytes contraction were Simultaneously recorded by IonOptix^TM. Results： The total of freshly isolated an adult rat eardiomyocytes was （5-7）×10^6, most of them were rod-shaped cells with clear cross-striations. 72 h after cultured with serum-free medium, the shape of cells didn＇t change. The amplitude of shortening/relengthening was （11.84±2.21）% with stimulation. Conclusion： Adult rat eardiomyoeytes can be isolated and cultured well with the stated method. Using the IonOptix^TM, one may real-time beat-to-beat observe and record the change of isolated eardiomyoeytes mechanics. These data can describe the change of myocardial function induced by different diseases from cell level.