长爪沙鼠肝星状细胞的分离培养与鉴定  

Establishment of a primary culture protocol of Mongolian gerbil hepatic stellate cells

作　　者：楼琦[1] 李巍[1] 石巧娟[1] 卢领群[1] 郭红刚[1] 杜江涛[1] 萨晓婴[1] 

机构地区：[1]浙江省医学科学院实验动物中心浙江省实验动物与安全性研究重点实验室,杭州310010

出　　处：《中国比较医学杂志》2016年第3期29-34,共6页Chinese Journal of Comparative Medicine

基　　金：国家自然科学基金(31301933);浙江省科技计划项目(2013C37012;2013C37013;2014C27013)

摘　　要：目的探索长爪沙鼠稳定、经济的肝星状细胞分离和培养方法,为深入探讨长爪沙鼠肝纤维化的细胞机制提供技术支撑。方法取成年雄性长爪沙鼠,用链蛋白酶、胶原酶及DNA酶体内门静脉灌注消化长爪沙鼠肝脏细胞,经Nycodenz密度梯度离心,分离肝星状细胞。台盼蓝拒染实验鉴定细胞活力,α-SMA,Desmin免疫细胞化学染色鉴定细胞性质。结果肝星状细胞得率为0.5-1×10^7/肝。肝星状细胞存活率在90%以上。原代培养3 dα-SMA阳性细胞达75%以上,传代培养后,α-SMA,Desmin阳性达100%。结论成功建立了稳定可靠的长爪沙鼠肝星状细胞分离培养方法,为肝脏相关疾病研究和防治药物的开发提供了技术支持。Objective To investigate the method to isolate and culture hepatic stellate cells（ HSCs） for studying the cellular mechanisms of hepatic frbrosis. Methods HSCs were isolated by nycodenz density gradient centrifugation after the hepatocytes obtained from adult male gebils were digested with pronase,collagenase and DNase,infused via portal vein. The cell viability was determined by trypan blue exclusion test. The purity of HSCs was identified by detectingα-SMA,desmin immunohistochemical staining. Results The yield rate of HSCs was 0. 5 ~ 1 × 10^7 per gerbil liver,and the cell viability was more than 90%. The percentage of α-SMA-positive cells was more than 75% after 3 days primary culture and almost 100% cells were α-SMA and desmin positive in passage culture. Conclusion The successful protocol of primary culture of Mongolian gerbil HSC provide a technical support for research of relevant liver diseases and drug development in the future.

关 键 词：长爪沙鼠 肝星状细胞 细胞分离 原代培养 

分 类 号：R-332[医药卫生]