摘要
目的优化SD乳鼠心肌细胞和心肌成纤维细胞的体外分离培养的条件和方法。方法分别用胰酶、Ⅱ型胶原酶依次消化分离心肌组织,再通过两次差速贴壁分离法分别收集、培养乳鼠心肌细胞和心肌成纤维细胞。光学显微镜下分别观察心肌细胞和心肌成纤维细胞的基本形态特征的变化,原代心肌细胞行心肌肌钙蛋白免疫荧光鉴定,而心肌成纤维细胞则用第2代行波形蛋白免疫荧光鉴定。结果心肌细胞在2~4 h开始贴壁生长,12~24 h贴壁速率大幅增加,并出现自发性搏动,48~72 h心肌细胞粘合成簇,细胞成活率,纯度均达95%以上;心肌成纤维细胞第2~4代细胞生长良好,纯度高达98%以上。结论此种方法可得到产量大,纯度高,活力好的心肌细胞和心肌成纤维细胞,为心血管疾病的基础与临床研究提供了理想的实验模型。
Aim To optimize the conditions and methods of SD rat cardiomyocytes and cardiac fibroblasts in vitro culture. Method Digested ventricular tissue with trypsin first,and then collagenaseⅡ. Collected and cultured neonatal rat cardiac myocytes and fibroblasts through the way of differential adhesion for two times. The basic shape change of cardiac myocytes and cardiac fibroblasts were observed under the opitical microscope. Cardiac myocytes were assayed by cardiac troponin immunofluorescenee and the second generation cardiac fibroblasts were assayed by vimentin immunofluorescence. Results Cardiomyocytes began to adhere and grow after 2 ~ 4 hours; The adherent rate increased substantially and the cells beated spontaneously after 12 ~ 24 hours; 48 ~ 72 hours later cardiomyocytes adhered to be clustered; Both of the cells' survival rate and purity reached more than 95%. The second generation to the fourth generation cardiac fibroblasts grew wel1 and its purity was as high as 98%. Conclusion A great quantity,purity and high survival rate of cardiocytes and cardiac fibroblasts can be effectively cultured by this method. And it provides an ideal experimental model for the cardiovascular disease and clinical studies.
出处
《中国动脉硬化杂志》
CAS
北大核心
2015年第1期90-93,共4页
Chinese Journal of Arteriosclerosis
基金
国家自然科学基金-地区科学基金项目(81260028)
石河子大学科学技术研究发展计划"自然科学与技术创新"团队创新项目(2011ZRKXTD-07)
关键词
心肌细胞
心肌成纤维细胞
原代培养
Cardiomyocytes
Cardiac Fibroblasts
Primary Cell Culture
