Targeted disruption of Reg gene : A new animal model of diabetes and pancreatitis.

1.为了产生缺乏Reg I基因的突变小鼠，在胚胎干细胞中电穿孔突变的Reg I基因，其外显子2-3被新霉素耐药基因取代，并获得突变了Reg I等位基因的胚胎干细胞克隆。雄性嵌合体将突变的等位基因传给后代。异种小鼠明显正常，并生出与突变的Reg I位点同源的小鼠。regi缺陷小鼠在包括胰腺细胞在内的各种组织中均未出现形态学改变。regi缺陷小鼠的体重、血糖和血清胰岛素水平与对照组小鼠几乎相同。甲氧基葡萄糖治疗诱导小鼠肥胖并伴有胰岛β细胞增生。金硫葡萄糖处理的regi缺陷小鼠的胰岛明显小于对照组，表明regi缺陷小鼠的胰岛在成年期对β细胞增殖的需求中具有较低的增殖活性。因此，regi缺陷小鼠在肥胖过程中容易发生非胰岛素依赖型糖尿病（NIDDM）的可能性较大，为研究非胰岛素依赖型糖尿病在肥胖中的发展提供了良好的模型。在乙硫氨酸诱导的慢性胰腺炎模型中，Reg i缺失小鼠发生了严重的胰腺炎，突变小鼠的体重低于对照组。提示Reg I参与了慢性胰腺炎的耐药过程，Reg I缺陷小鼠是一种新的慢性胰腺炎模型。我们发现所有小鼠的Reg基因构成一个多基因家族，Reg家族包括3个亚型（I型、II型和III型），并且所有的Reg基因都分布在6C/D.6染色体的邻近位点。我们通过Northern blot检测了Reg家族其他成员（Reg II、Reg iii α、Reg iii β和Reg IIIgamma）在小鼠胰腺中的表达，发现RegI缺失小鼠中Reg iii α和Reg IIIgamma的表达增加，提示RegI的某些功能可能通过残余Reg家族表达的增加而得到补偿。为了澄清这种可能性，我们现在正在培养缺乏所有Reg家族基因的突变小鼠。少

1.To generate mutant mice lacking Reg I gene, the mutated Reg I gene, in which exon 2-3 was replaced by neomycin resistant gene, was electroporated in ES cells, and an ES cell clone mutated the Reg I allele was obtained. A male chimera transmitted the mutant allele to its offspring. Heterogous mice were apparently normal, and gave birth to mice homologous for mutant Reg I locus.2.Reg I deficient mice showed no morphological changes in various tissues including pancreatic beta cells. Body weight, blood glucose and serum insulin levels in Reg I-deficient mice were almost the same as those in control mice.3.Aurothioglucose treatment induced obesity with hyperplasia of islet beta-cells in control mice. The islets in aurothioglucose-treated Reg I-deficient mice were significantly smaller than those of control, indicating that islets of Reg I-deficient mice have lower proliferation activity in response to a demand of beta-cell proliferation in adulthood. Therefore, it is quite possible to as … More sume that Reg I-deficient mice easily develop non-insulin dependent diabetes mellitus (NIDDM) when they become obese and be a good model for studying development of NIDDM in obesity.4.In the ethionine-induced chronic pancreatitis model, Reg I-deficient mice developed severe pancreatitis and the body weight of the mutant mice was lower than that of control mice. This result suggests that Reg I is involved in resistance of chronic pancreatitis and that Reg I-deficient mice is a new model of chronic pancreatitis.We revealed that all the mouse Reg genes constitute a multigene family, Reg family, which consists of three subtypes (type I,II and III), and that all the Reg genes assigned to the adjacent site of chromosome 6C/D.6.We examined expression of the other members of Reg family (Reg II,Reg IIIalpha, RegIIIbeta and Reg IIIgamma) in mouse pancreas by Northern blot analyzes, and revealed that expressions of Reg IIIalpha and Reg IIIgamma were increased in Reg I-deficient mice, suggesting a possibility that some functions of Reg I were compensated by the increased expression of remnant Reg family. To clarify this possibility, we are now generating mutant mice deficient in all the Reg family genes. Less