乳鼠窦房结细胞的原代培养和形态学特征

目的比较差速贴壁技术和常规培养方法在体外获取窦房结自律细胞的密度,并观察其形态学特征。旨在建立可靠的窦房结自律细胞体外取材分离、纯化培养及形态学鉴定。方法取24 h内新生Wistar乳鼠1 20只,随机分为实验组(差速贴壁技术培养方法)和对照组(常规培养方法),每组60只。每次随机各取5只乳鼠行窦房结取材,分别用2种方法进行原代细胞纯化培养。通过光、电镜观察比较2种方法体外窦房结自律细胞的密度比及形态学特征。结果体外培养窦房结细胞有3种不同形态,梭形细胞、三角形细胞和不规则形细胞,其中梭形细胞最多,搏动频率最快;三角形细胞与心房肌三角形细胞特征相似;实验组窦房结梭形细胞比例明显高于对照组(65±4)%vs(45±3)%,P<0.01]。结论采用差速贴壁技术纯化培养方法较常规培养可更明显提高窦房结梭形细胞的比例,是一种可靠的窦房结自律细胞培养、纯化技术。在培养的乳鼠窦房结细胞中,细胞体积小、搏动频率快的梭形细胞即是窦房结的起搏细胞。

Primary culture and morphologic characteristics of sinoatrial node cells in neonatal rats

Objective To establish a set of reliable culture methods for isolation,purification and identifcation of sinoatrial node(SAN) autorhythmic cells by comparing their density and observing their morphologic characteristics.Methods One hundred and twenty 24 h-old neonatal Wistar rats were randomly divided into experiment group and control group,60 in each group.SAN autorhythmic cells,isolated from 5 mice,were cultured and purified with differential adherent culture technique and routine culture method,respectively.Their density and morphologic characteristics were observed and compared by light microscopy and transmission electron microscopy,respectively. Results The in vitro cultured spindle,triangle and irregular SAN cells were observed.The number of spindle SAN cells was the greatest and their pulsating frequency was the fastest.The charateristics of triangle SAN cells were similar with those of atrial muscle triangle cells.The proportion of SAN autorhythmic cells was significantly higher in experiment group than in control group(65%±4%vs 45%±3%,P<0.01).Conclusion Differential adherent purification and culture technique can increase the proportion of spindle SAN cells more significantly than routine culture method,and is thus a reliable culture and purification technique for SAN autorhythmic cells.Spindle SAN cells with a small volume and a rapid pulsating frequency in cultured SAN cells from neonatal rats are the pacemaker cells in SAN.