非酒精性脂肪性肝炎大鼠肝细胞和库普弗细胞同时分离及鉴定  

Simultaneously isolation and identification of hepatocytes and kupffer cells from nonalcohlic steatohepatitis rat

作　　者：冯高飞[1] 杨钦河[1] 王文晶[1] 何秀敏[1] 张玉佩[1] 纪桂元[2] 李光秋[1] 赵亚凤[1] 沈灿宏[1] 杨晓蕾[1] 

机构地区：[1]暨南大学医学院中医系,广州510632 [2]中山大学公共卫生学院营养系,广州510080

出　　处：《广东医学》2012年第1期40-43,共4页Guangdong Medical Journal

基　　金：国家自然科学基金资助项目(编号:30973694);国家大学生创新性实验计划资助项目(编号:091055930)

摘　　要：目的建立一种稳定、高效、同时分离非酒精性脂肪性肝炎(nonalcohlic steatohepatitis,NASH)大鼠肝细胞(hepatocytes,HCs)和库普弗细胞(Kupffer cells,KCs)的新方法。方法采用离体循环灌注Ⅳ型胶原酶,低速离心获取HCs;差速离心、密度梯度离心和选择性贴壁获得KCs。用Typan blue染色和流式细胞术(FCM)分别进行活性和纯度鉴定。结果每只大鼠获得纯化的HCs[(4.0～4.5)×108]、KCs[(1.5～2.0)×107],经Typanblue染色活力均在95%以上,FCM检测纯度均在90%以上。结论此实验建立同时分离NASH大鼠HCs和KCs的方法高效、稳定,细胞数量和纯度都符合进一步实验的要求。Objective To establish an effective and stable method for isolation and identification of hepatocytes（HCs） and Kupffer cells（KCs） from nonalcohlic steatohepatitis（NASH） rats simultaneously. Methods Rat liver in vitro was perfused with collagenase Ⅳ and centrifuged in low-speed isolation of HCs.Subsequently,differential centrifugation,density gradient centrifugation and selective adherence were carried out for isolation of KCs.Typan blue stain and Flow cytometry（FCM） were applied for tests of activity and purity. Results The yields of purified cell of HC and KC in each rat were（4.0~4.5）×108 and（1.5~2.0）×107,respectively.The viability of HCs and KCs isolated were higher than 95%,with purity over 90%. Conclusion The method for isolation and identification of HCs and KCs from NASH rat simultaneously is effective and stable.Both quantity and purity are qualified for further experiments.

关 键 词：非酒精性脂肪性肝炎大鼠 肝细胞 库普弗细胞 同时分离 鉴定 

分 类 号：R575.5[医药卫生—消化系统]