改良细胞灌流分离装置提高心肌细胞分离效率  

Modified cell perfusion appliance for improving the separation efficacy of cardiomyocytes

作　　者：张琳[1] 纵静[2] 张庆涛[3] 

机构地区：[1]徐州医学院临床学院,江苏省徐州市221002 [2]徐州医学院附属医院,江苏省徐州市221002 [3]徐州医学院附属第三医院,江苏省徐州市221003

出　　处：《中国组织工程研究》2015年第51期8300-8304,共5页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金(81400178);徐州市科技计划项目(XM13B062)~~

摘　　要：背景:成熟心肌细胞分离过程复杂,获得数量稳定、高质量的心肌细胞迄今仍无统一、高效、活性好、简便的方法可循。目的:通过改良Langendorff细胞灌流分离装置,探讨一种更高效的成熟心肌细胞分离方法。方法:采用细胞灌流分离装置,用Ⅱ型胶原酶经主动脉逆行灌流法分离成年SD大鼠心肌细胞。对照组采用传统Langendorff细胞灌流分离装置;实验组采用改良Langendorff细胞灌流分离装置。消化结束即刻在显微镜下观察细胞形态、杆状细胞比率并计算活细胞产量;通过锥虫蓝染色评价心肌细胞活性。结果与结论:实验组活细胞产量为(3.7±0.5)×107,显著高于对照组(2.1±0.4)×107,差异有显著性意义(P<0.05)。实验组锥虫蓝染色阴性的杆状细胞比率(85.6±5.5)%,显著高于对照组(71.8±5.7)%,差异有显著性意义(P<0.05)。提示采用改良Langendorff离体心脏灌流系统可获得高产量、高活性的心肌细胞,提高了成熟心肌细胞的分离效率,为成熟心肌细胞培养及研究奠定了基础。BACKGROUND： Separating mature cardiomyocytes is is a complex process. Until now, there is no uniform, high-yield, simple method to harvest cardiomyocytes with high viability. OBJECTIVE： To explore a more effective method of isolating adult rat cardiomyocytes by modifying the cell perfusion appliance. METHODS： Cardiomyocytes from adult Sprague-Dawley rats were isolated by collagenase type II perfusion method via a cell perfusion appliance. Traditional Langendorff cell perfusion appliance was used in control group, and modified Langendorff cell perfusion appliance used in experimental group. Under an optical microscopy, morphology of cardiomyocytes was observed and rod-shaped cell ratio was detected to calculate the yield of viable cells. Cardiomyocyte viability was evaluated by trypan blue exclusion test. RESULTS AND CONCLUSION： The yield of viable cells per rat was significantly higher in the experimental group than in the control group [（3.7±0.5）×107 vs.（2.1±0.4）×107, P 〈 0.05]. The ratio of trypan blue test-negative cells was also significantly higher in the experimental group than in the control group [（85.6±5.5）% vs.（71.8±5.7）%, P 〈 0.05]. These findings indicate that high-yield and high-viability cardiomyocytes can be harvestedusing the modified Langendorff cell perfusion appliance, which improves the separation efficiency of mature cardiomyocytes and lays the foundation for mature cardiomyocyte culture.

关 键 词：肌细胞 心脏 细胞分离 组织构建 组织工程 心肌细胞 大鼠 分离装置 细胞活性 国家自然科学基金 

分 类 号：R446[医药卫生—诊断学]