Elucidation of the mechanism of pancreas development and its application for the regenerative medicine

1)小鼠胰岛β细胞特异性Smad 2基因的阻断为了研究Smad 2在胰岛β细胞中的作用，我们利用Cre-loxP系统特异性阻断胰岛β细胞中的Smad 2基因。我们将Smad 2的floxed等位基因纯合的小鼠（Smad 2 ^<loxC/loxC>）与在大鼠胰岛素启动子控制下表达Cre重组酶的Ins-Cre小鼠杂交，以获得Smad 2 ^<loxC/loxC>Ins-Cre（Smad 2 βKO）Amad 2 βKO小鼠显示出对葡萄糖的响应的胰岛素分泌缺陷和葡萄糖耐量的进行性损害。有趣的是，组织学分析显示，Smad 2 βKO中的胰腺胰岛比对照同窝小鼠中的更大，这表明Smad 2不仅参与胰岛素分泌，还参与β细胞的增殖。2）小鼠胰腺β细胞中激活素IB型受体的条件性表达。我们首先产生了携带转基因构建体的转基因小鼠，其中绿色荧光蛋白（GFP）基因侧翼直接重复 ...更多信息 将loxP序列置于鸡β-肌动蛋白启动子的控制下，然后是组成型活性形式的激活素IB型受体（ActRIBCA）基因和IRES βgeo盒（ActRIBCATg）。转基因指导ActRIBCA在Cre介导的loxP侧翼GFP基因切除后的表达。为了获得R细胞特异性ActRIBCA转基因小鼠（ActRIBCAβS-Tg），我们将ActRIBCATg小鼠与Ins-Cre小鼠杂交。所得小鼠表现出响应于葡萄糖的胰岛素分泌缺陷和葡萄糖耐量的进行性损害。分离的β细胞的电生理分析表明，<ATP>突变小鼠β细胞的K通道功能明显减弱。突变小鼠的胰岛素分泌功能障碍可能与K通道功能改变有关<ATP>，提示ActRIB-Smad 2在2型糖尿病的葡萄糖敏感和胰岛素分泌中起重要作用。少

1) Pancreas beta cell-specific gene disruption of Smad2 in miceTo investigate the role of Smad2 in pancreatic β cells, we have used the Cre-loxP system to specifically disrupt Smad2 gene in the β cells. We crossed the mice homozygous for a floxed allele of Smad2 (Smad2^<loxC/loxC>) with Ins-Cre mice that express the Cre recombinase under the control of the rat insulin promoter to obtain Smad2^<loxC/loxC>Ins-Cre (Smad2βKO).Smad2βKO mice show a defective insulin secretion in response to glucose and a progressive impairment of glucose tolerance. Interestingly, histological analysis revealed that pancreas islets in Smad2βKO were bigger that those in control littermates, suggesting that Smad2 is involved in not only insulin secretion but also proliferation of β cells.2) Conditional expression of activin type IB receptor in pancreas β cells in mice.We first generated the transgenic mice carrying a transgene construct in which green fluorescence protein (GFP) gene flanked by directly repeated … More loxP sequences was placed under the control of chicken β-actin promoter and followed by the constitutive active form of activin type IB receptor (ActRIBCA) gene and an IRES βgeo cassette (ActRIBCATg). The transgene directs the expression of ActRIBCA upon the Cre-mediated excision of the loxP-flanked GFP gene. To achieve R cell specific ActRIBCA transgenic mouse (ActRIBCAβS-Tg), we crossed ActRIBCATg mice with Ins-Cre mice. The resultant mice exhibited a defective insulin secretion in response to glucose and a progressive impairment of glucose tolerance. The electrophysiological analysis performed with isolated β cells revealed that the function of K_<ATP> channel of β cells of the mutant mice was dramatically attenuated. The impairment of insulin release in the mutant mice may be explained, at least in part, by the alteration of the function of K_<ATP> channels.Our results suggested an important role for signaling by the ActRIB-Smad2 in both glucose sensing and insulin secretion in type 2 diabetes. Less