【作者】 张钰鹏； 李非； 孙海晨； 崔叶青； 孙家邦；

【Author】 Zhangyupeng Lifei Sunjiabang Sunhaichen Cuiyeqing. Department of General surgery,XuanWu Hospital,Beijing 100053,China

【机构】 首都医科大学宣武医院普外科；

【摘要】 目的:模拟微重力方法培养大鼠原代肝细胞,初步分析其形态学特点及其意义。方法:改良 Seglen 原位胶原酶灌注法获得大鼠肝脏单细胞悬液,2.2×10~5细胞/ml 加微载体 Cytodex-3(4g/l)接种,采用旋转细胞培养系统(RCCS)进行模拟微重力培养。第0、6、24、72、120、168h 取样,相差、体式显微镜观察活细胞形态,第120h 标本 H-E 染色观察组织学形态,电镜观察超微结构。结果:模拟微重力状态下肝细胞24h 内贴附微载体并出现三维结构,48-72h 发展为独特的肝细胞.微载体聚球体结构。电镜下可见细胞膜的三种不同形态,其分布与功能相一致。结论:本实验证实模拟微重力培养方法能使肝细胞形成分化的三维类组织结构,是一种有希望的人工肝生物成分制备技术。更多还原

【Abstract】 Objective:culture primary,rat hepatocytes by means of simulated microgravity,describe their basic morphological characteristics and illustrate the significance primarily.Methods.Separate SD rat hepetocytes by the modified Seglen’s two step in-situ collagenase perfusion method.Inoculate cells at 2.2×10~5 ceils/ml with microcarrier Cytodex-3(4g/l)as the solidial support materials.Creating simulated microgravity environment by Rotary Cell Culture System(RCCS Synthecon.com U.S.A).Take samples on 0,6,24,72,120 and 168 hour, make morphological studies by means of phase contrast microscope,hematoxylin and eosin(H.E)dyed paraffin sections and electron microscope(TEM).Result:In the RCCS,hepatocytes start to grow on the surface of microcarrier in the first 24 hour and then forming three-dimensiongal polylayer structure during 24 to 72 hours.TEM shows three different kinds of membranes among simulated microgravity cultured cells,whos distribution in accordance with their functions.Conclusion:In this study,we succeesfully cultured primary rat hepatocytes in simulated microgravity condition.Initial morphological study shows that this new culture technology can establish a three-dimensional structure in vivo.Suggested that this method may be potentially valuable in areas of bioartificial liver.更多还原