原代肝细胞的分离和移植  

Isolation and transplantation of primary hepatocytes

作　　者：冯渊[1] 李德卫[1] 杨晓波[1] 陈睿[1] 樊斌[1] 

机构地区：[1]重庆医科大学附属第一医院肝胆外科,重庆市400016

出　　处：《中国组织工程研究》2012年第18期3314-3317,共4页Chinese Journal of Tissue Engineering Research

摘　　要：背景:肝细胞移植可以作为一种桥梁作用,帮助肝功能衰竭患者渡过肝衰期,并提高患者生存率和预后。目的:采用Seglen改良的原位灌注探讨SD大鼠原代肝细胞分离的影响因素和方法的改进,同时分析原代肝细胞治疗急性肝功能衰竭大鼠的疗效。方法:两步法分离大鼠肝细胞。D-氨基半乳糖诱导大鼠急性肝衰竭,24h后分2组:细胞移植组经脾脏移植约2×107个肝细胞;对照组经脾脏注射0.4mLHank’s液。观察不同时间点大鼠的生存率和血清中转氨酶、总胆红素变化情况、脾内白蛋白分泌作用及脾内肝细胞分布情况。结果与结论:大鼠肝细胞分离存活率达85%～95%。细胞移植组14d存活率(75%)显著高于对照组组(30%)(P=0.01),且肝功能改善情况明显优于对照组。移植30d后,脾内有肝细胞白蛋白绿色荧光信号;移植15d后,可以看到肝细胞在脾脏红髓中簇集在一起并定植。结果说明胶原酶、pH值、灌注液、灌注方法等均是影响肝细胞分离存活率的因素;经脾脏移植的肝细胞能提高急性肝衰竭大鼠的生存率和改善肝功能。BACKGROUND：Hepatocyte transplantation as a role of bridge can help the liver failure patients to pass through the liver failure phase and improve patient survival rate and prognosis.OBJECTIVE：To research the improvement of method and the influencing factors of primary hepatocytes isolation technology in Sprague Dawley rats by Seglen modified in situ perfusion method,and to evaluate the treatment effects of primary hepatocytes transplantation on rats with acute hepatic failure.METHODS：Rats＇ hepatocytes were isolated by the two step collagenese digestion,and acute hepatic failure was induced by D-galactosamine in rats.After 24 hours,the acute hepatic failure rats were divided into two groups randomly.The rats in the transplantation group received 2×107 hepatocytes through intrasplenic injection,the rats in the control group received 0.4 mL Hank＇s solution by the same way.The survival rate,level of transaminase and total bilirubin in serum,secretion of albumin and the distribution of hepatocytes in spleen were observed at different time points.RESULTS AND CONCLUSION：The survival rate of rat hepatocytes was 85-95%.At 14 days,the survival rate of transplantation group（75%） was significantly higher than that of control group（30%）（P=0.01）.Hepatic function improvement of transplantation group was significantly better than that of control group.At 30 days after transplantation,immunofluorescent staining of albumin could be seen in spleen;at 15 days after transplantation,transplanted hepatocytes were found in spleen.There were many influence factors in the success of hepatocyte separation,such as collagen enzyme,pH value,perfusion fluid and perfusion method.Hepatocyte intraplenic transplantation could improve the survival rate and ameliorate the hepatic function of rats with acute hepatic failure.

关 键 词：肝细胞分离 移植 急性 肝功能衰竭 存活率 

分 类 号：R617[医药卫生—外科学]