大鼠胰岛素原基因转染肠道上皮细胞构建类胰岛B细胞系的研究.pdfVIP

· 2053· · 论著 · 大鼠胰岛素原基因转染肠道上皮细胞构建 类胰岛B细胞系的研究 蒋琳，刘超，刘翠萍，季昊琚，吴海玮，张洪慧 【摘要】 目的 观察重组大鼠胰岛素原基因转染小鼠肠道上皮细胞 (IEC一6)后，IEC一6的胰岛素表达情况。 方法 将IEC一6分为重组大鼠胰岛素原基因pCMV／proinsulin转染组 (A组)、空载质粒pCMV转染组 (B组)和未转 染组 (c组)。利用脂质体法将pCMV／proinsulin和pCMV分别转染IEC一6，用RT—PCR法检测转染后IEC一6胰岛素 原基因mRNA的表达，并用超敏放免法测定转染后IEC一6的胰岛素表达水平。结果 pCMV／proinsulin转染IEC一6 后，IEC一6能表达胰岛素原基因mRNA。3组IEC一6表达的胰岛素水平间差别有统计学意义 (P0．05)。A组IEC 一 6胰岛素表达水平与B组、c组比较，差别有统计学意义 (P0．001)；B组IEC一6胰岛素表达水平与c组比较， 差别无统计学意义 (P0．05)。结论 重组大鼠胰岛素原基因能成功转染小鼠肠道细胞并分泌胰岛素，为开展糖尿 病基因治疗的研究奠定了实验基础。 【关键词】 转染；肠道上皮细胞；类胰岛B细胞系 【中图分类号】R 349．65 R 329．457 【文献标识码】A 【文章编号】1007—9572(2007)24—2053—03 Construction of Pseudo—B Cell Line by Transfecting Rat Proinsulin Gene to Mouse Intestinal Epithelial Cell JIANG Lin，LIU Chao，LIU Cui—ping，et a1．Department of Endocrinology，the First Affiliated Hospital to Nanjing Medical University， Nanjing 210029，China 【Abstract】 Objective To investigate the expression of rat intestinal epithelial ceHs(IEC一6)insulin after construc— tion of pseudo—line by transfecting rat proinsulin gene to mouse IEC一6．Methods IEC一6 were divided into three groups： Group A with recombinant rat proinsulin gene(pCMV／proinsulin)，Group B with no—load plasmid(pCMV)and Group C with— out transfection．The pCMV／proinsuin and pCMV were transfected to IEC一6 by lipofetin—mediated DNA transfection．RT— PCR was used to determine expression of proinsuin gene mRNA in modified IEC一6 of insulin gene，and hypersensitive radioim— munity was applied to test insulin expression in IEC一6 after transfection．Results After transfection，proinsulin gene mRNA could be detected in IEC一6．There was significant difference in insulin level among the three groups(P0．05)；Group A was significantly different from Group B and Group C in expression of IEC一6 in