三明治培养新生实验小型猪肝细胞的功能与形态  

Function and morphological characteristics of Chinese experimental suckling mini-pig hepatocytes cultured in sandwich

作　　者：杨杰[1] 王英杰[1] 刘鸿凌[1] 刘俊[1] 王宇明[1] 

机构地区：[1]第三军医大学附属西南医院全军感染病研究所,重庆400038

出　　处：《第三军医大学学报》2003年第6期475-477,共3页Journal of Third Military Medical University

基　　金：国家自然科学基金资助项目 ( 39970 2 14 ;30 0 2 70 0 1) ;全国优秀博士基金资助项目 ( 19994 7)

摘　　要：目的　探索双层胶原夹心培养猪肝细胞的方法 ,观察培养猪肝细胞的功能与形态特征。方法　将两步法分离的新生中国实验小型猪肝细胞接种于胶原被覆的培养板常规培养 ,2 4h后 ,肝细胞上再加一层胶原形成双层胶原夹心(三明治 )培养 ,观察培养猪肝细胞的白蛋白分泌、形态学特征和乳酸脱氢酶 (LDH)漏出量。结果　成功将新生实验小型猪肝细胞固定于双层胶原中 ,形成三明治状。培养 1周内 ,用SDS/PAGE可检测出猪肝细胞分泌的白蛋白 ,倒置相差显微镜下观察到典型的形态特征 ,培养期内LDH漏出量较少。结论　可用双层胶原对新生乳猪肝细胞进行三明治样培养 ,为肝细胞提供更接近体内的培养环境 。Objective To study the method for the culture of porcine hepatocytes between 2 layers of gelled collagen and to observe the function and morphological characteristics of the cultured porcine hepatocytes. Methods Porcine hepatocytes isolated from Chinese experimental suckling minipigs by two step perfusion with collagenase using an extracorporeal perfusion apparatus were inoculated onto collagen coated dishes for culture. After culture for 24 h, a layer of gelled collagen was added on the porcine hepatocytes to form sandwich for culture. The albumin excretion, cellular morphological characteristics and release of lactate dehydrogenase(LDH) content were examined within 1 week in the collagen sandwich culture. Results The freshly isolated hepatocytes of Chinese experimental suckling minipigs were cultured between 2 layers of gelled collagen as sandwich configuration with success. For up to 7 days of culture, the porcine albumin could be estimated by SDS/PAGE and the typical cellular morphological characteristics of cultured hepatocytes could be observed. With hepatocytes cultured in the sandwich configuration, a slight release of the LDH into the culture medium could also be found. Conclusion The hepatocytes of Chinese experimental suckling minipigs can be sandwich cultured by two layers of collagenous matrix, which provides an environment that more closely resembles the one in vivo and allows the maintenance of certain liver specific functions and morphological characteristics of hepatocytes.

关 键 词：肝细胞 小型猪 培养 三明治 

分 类 号：Q813.11[生物学—生物工程] R322.47[医药卫生—人体解剖和组织胚胎学]