Induction of insulin secreting cell and its application for transplantation

胰岛移植有望成为治疗胰岛素依赖型糖尿病(IDDM)的有效方法。然而，这一策略尚未被确立为IDDM的标准治疗方法。与供体短缺一样，免疫排斥反应的控制也是胰岛移植的障碍。1.人胰管细胞(HPDC)胰岛素分泌细胞的诱导1)用人乳头瘤病毒早期表达基因建立HPDC细胞系。2)观察细胞外基质刺激后HPDC细胞形态的变化。3)构建PDX-1基因的腺病毒载体。将PDX-1基因导入HPDC后，高血糖素的表达明显高于胰岛素的表达。细胞经体外培养后，形成类似胰岛的团块。2.基因芯片分析和胰岛发育相关基因的检测1)从表达胰岛素的HPDC中提取mRNA，用于DNA微阵列研究。检测胰岛素的表达并筛选出与胰岛素表达模式相同的基因。2)合作小组从小鼠胚胎干细胞(ISSC)建立了胰岛素分泌细胞。我们从发育的5个阶段收集细胞，用基因芯片系统分析其mRNA的表达。对基因表达变化的分析表明，胰岛素相关基因的表达逐渐增加。这些结果表明，致病基因将在进一步研究中被发现。

Islet transplantation has been expected to be effective therapy for insulin dependent DM (IDDM). However, this strategy is not established as a standard treatment for IDDM yet. The control of immuno-rejection is an obstacle for islet transplantation as same as the shortage of donors. The aim of this study is to develop the islet transplantation by using regeneration islet.1.The induction of insulin secreting cells from human pancreatic duct cell (HPDC)1)We established the HPDC cell line by the transfection of early expression gene of papilloma virus.2)The change of shape of this HPDC was observed after the stimulation of extra cellular matrix of tissue culture.3)We constructed the adenovirus vector for the transfection of PDX-1 gene. After transfection of PDX-1 gene into HPDC, the more expression of glucagon was detected than that of insulin. The cells made a mass looking like islet after in vitro culture. These HPDC became neoplastic tumor in vivo transplantation in mice.2.The analysis of genes expression using microarray and the detection of genes relating to islet development1)The mRNA was extracted from HPDC showing the insulin expression and used for DNA microarray study. The exspression of insulin was detected and genes showing the same pattern as insulin were selected.2)The collaborate group has established insulin secreting cell from mouse embryonic stem cell (ISSC). We harvested the cells from 5 stage of this development and analyzed the mRNA expression using microarray system. The analysis of changes in gene expression revealed that the expression of genes relating to insulin was increased step by step. Theses results indicate that the responsible gene would be detected by the further study.