卵圆细胞诱导分化为胰岛素分泌细胞  

Differentiation of oval cells into insulin-producing cells

作　　者：缴富斌[1] 张波 孙晓艳[3] 罗军 季建华 

机构地区：[1]解放军军医进修学院,北京市100853 [2]解放军总参警卫局保健处,北京市100017 [3]解放军总医院基础医学研究部,北京市100853

出　　处：《中国组织工程研究与临床康复》2008年第21期4007-4011,共5页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：国家自然科学基金(2004B34001007)~~

摘　　要：背景:原代分离的肝卵圆细胞具有双向分化的能力和横向分化的可塑性。目的:分离大鼠肝脏卵圆细胞,在化学诱导剂作用下使其定向分化为胰岛素分泌细胞。设计、时间及地点:对照观察细胞实验,于2007-05/2008-01在解放军总医院基础研究部,国家重点实验室完成。材料:SPF级SD大鼠,体质量(120±20)g。方法:采用改良梯度离心法分离、培养大鼠肝卵圆细胞,并行免疫组织化学鉴定。随后将卵圆细胞置于含有诱导剂Nicotimide和高浓度葡萄糖的RPMI1640培养液继续培养,同时以不含诱导剂的RPMI1640培养液培养的卵圆细胞作为阴性对照。主要观察指标:采用SDS-聚丙烯酰胺凝胶电泳、酶联免疫吸附实验、放射免疫分析检测两组卵圆细胞胰岛素的表达。结果:改良的梯度离心方法分离的细胞具干细胞特点,可形成集落,并且OV6、CD34、Nestin染色呈阳性。分离的大鼠肝卵圆细胞,加入诱导剂Nicotimide和高浓度葡萄糖诱导培养1周后,SDS-聚丙烯酰胺凝胶电泳、酶联免疫吸附、RIA检测均显示胰岛素分泌量远高于不含诱导剂细胞(P<0.05)。结论:化学诱导剂Nicotimide和高浓度葡萄糖可以在短时期内作用肝卵圆细胞可启动胰十二指肠同源盒基因-l基因,使之定向分化为胰岛素分泌细胞。BACKGROUND： Primarily isolated hepatic oval cells have the potential of bidirectional differentiation and the ductility of transversal differentiation. OBJECTIVE： To induce the differentiation of rat hepatic oval cells into insulin-producing cells with a chemical inducer. DESIGN, TIME AND SETTING： A control observational cellular experiment was performed at the State Key Laboratory of Basic Research Department of General Hospital of Chinese PLA from May 2007 to January 2008. MATERIALS： Specific pathogen free （SPF） SD rats weighting （120±20） g were selected in this study. METHODS： Hepatic oval cells were isolated using modified gradient centrifugation, and then identified by immunohistochemistry. Subsequently, hepatic oval cells were cultured in RPMI 1640 medium containing Nicotimide and a high concentration of glucose. Simultaneously, hepatic oval cells were cultured in RPMI 1640 medium without Nicotirnide or high concentration glucose as a negative control. MAIN OUTCOME MEASURES： SDS-polyacrylamide gel electrophoresis （PAGE）, enzyme linked immunosorbent assay （ELISA） and radioimmunoassay （RIA） were used to detect the insulin expression in oval cells from both groups. RESULTS： The oval cells isolated by modified gradient centrifugation had the features of stem cells, could self-assemble to form colony. These cells were positive for OV6, CD34 and Nestin. SDS-PAGE, ELISA and R1A demonstrated that insulin levels were significantly higher in the cells cultured in the RPMI 1640 medium containing Nicotimide and high concentration glucose than in the negative controls 1 week after culture （P 〈 0.05）. CONCLUSION： Nicotimide and a high concentration of glucose can induce hepatic oval cells to directionally differentiate into insulin-producing cells by activating the expression of PDX-1 in a short period.

关 键 词：肝脏干细胞 卵圆细胞 PDX-1 梯度离心 RIA 

分 类 号：R394.2[医药卫生—医学遗传学]