The analysis of the mechanism how intestinal stem cells can differentiate into pancreatic beta-cellls

肠道干细胞分化为胰腺β细胞的机制分析

• 批准号: 14570010
• 负责人: NAKAMURA Takaaki
• 金额: $ 2.05万
• 依托单位: Shiga University of Medical Science
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2002
• 资助国家: 日本
• 起止时间: 2002 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14570010/
• 关键词: transcription factor; sumovlation; iRNAs; diabetes mellitus; Pdxl; insulin; post-translation; NFk-B; 小腸幹細胞; インスリン; Pdx1; SUMO-1; 過酸化物質; β細胞死; 膵β細胞; 転写調節因子; 再生

Pancreatic duodenal homeobox-1(Pdxl) is a transcription factor and its phosphorylation is thought to be essential for activation of insulin gene expression. This phosphorylation is related to concomitant shift in molecular mass from 31 to 46 kDa. However, we found that Pdxl was modified by SUMO-1 (small ubiquitin-related modifier 1) in beta TC-6 cells and COS-7 cells, which were transfected with Pdx-1 cDNA. This modification contributed to the increase in molecular mass of Pdxl from 31 to 46 kDa. Additionally, sumoylated Pdxl localized in nucleus. The reduction of SUMO-iRNA protein by use of RNA interference (SUMO-iRNA) resulted in a significant decrease in Pdxl protein in the nucleus. A 34-kDa form of Pdxl was detected by the cells exposed to SUMO-iRNAs in the presence of lactacystin, a proteosome inhibitor.Furthermore, the reduced nuclear sumoylated Pdxl content was associated with significant lower transcriptional activity of insulin gene. Thesse findings indicated that SUMO-1 modif … More ication is associated with both the localization and stability of Pdx l as well as its effect on insulin gene activation.Next, to clarify the effect of dietary lipid hydroperoxide (LPO) on development of glucose intolerance, we fed Sprague-Dawley rats on a diet containing elevated LPO level for 10 weeks and measured both insulin sensitivity and insulin secretion. The contents of LPO in both plasma and skeletal muscle in the LPO-fed rats were significantly higher than those in the controls. Both insulin resistance evaluated by steady-state blood glucose (SSBG) methods and impaired insulin secretion evaluated by oral glucose tolerance test (OGTT) were found in the LPO-fed rats as compared with control rats. Furthermore, the levels of insulin receptor substrate (IRS)-1 protein in the skeletal muscle were significantly lower in the LPO-fed rats. Those impairments were not reversed in LPO-fed rats with supernormal levels of plasma vitamin E following vitamin E supplementation for 5 weeks. Moreover, the immunohistochemical study revealed that NF-kB-p50 protein was found in the nucleus of pancreatic b-cells of the LPO-fed rats, whereas it was not observed in the nucleus of the islets in the control rats. These findings indicate that NF-kB is activated in response to oxidative stress in pancreatic islet cells in LPO-fed rats. In conclusion, our studies reveal that diet high in LPO by vitamin E-deficiency accelerates glucose intolerance through impairments of both sensitivity and secretion of insulin. Less

胰十二指肠同源盒-1（Pdxl）是一种转录因子，其磷酸化被认为是胰岛素基因表达激活的必要条件。这种磷酸化与分子质量从31 kDa到46 kDa的变化有关。然而，我们发现在转染Pdx-1 cDNA的β TC-6细胞和COS-7细胞中，Pdxl被SUMO-1（小泛素相关修饰物1）修饰。该修饰使Pdxl的分子质量从31 kDa增加到46 kDa。此外，sumoylated Pdxl定位于细胞核。使用RNA干扰（SUMO-iRNA）减少SUMO-iRNA蛋白导致细胞核中Pdxl蛋白显著减少。暴露于SUMO-iRNAs的细胞在乳糖蛋白酶（一种蛋白体抑制剂）存在下检测到34 kda形式的Pdxl。此外，核聚合Pdxl含量的降低与胰岛素基因转录活性的显著降低有关。这些结果表明，SUMO-1修饰与胰岛素基因的定位和稳定性以及对胰岛素基因激活的影响有关。接下来，为了阐明膳食脂质过氧化氢（LPO）对葡萄糖耐受不良的影响，我们给Sprague-Dawley大鼠喂食含有高脂质过氧化氢（LPO）水平的饮食10周，并测量胰岛素敏感性和胰岛素分泌。LPO喂养大鼠血浆和骨骼肌中LPO含量均显著高于对照组。与对照大鼠相比，lpo喂养大鼠的胰岛素抵抗（SSBG）和口服糖耐量试验（OGTT）均出现胰岛素分泌障碍。此外，lpo喂养大鼠骨骼肌中胰岛素受体底物(IRS)-1蛋白水平显著降低。在补充维生素E 5周后，血浆维生素E水平超正常的lpo喂养大鼠，这些损伤没有逆转。此外，免疫组化研究显示，在lpo喂养大鼠的胰腺b细胞核中发现NF-kB-p50蛋白，而在对照组大鼠的胰岛核中未发现NF-kB-p50蛋白。这些发现表明，在lpo喂养的大鼠胰岛细胞中，NF-kB被激活以响应氧化应激。综上所述，我们的研究表明，维生素e缺乏导致的高LPO饮食通过损害胰岛素的敏感性和分泌来加速葡萄糖耐受不良。少

项目成果

Akio Kishi, Takaaki Nakamura: "Atsunori Kashiwagi. Sumoylation of Pdx1 Is Essential For Insulin Gene Activation."Am J Physiol, endocrinology and metabolism.. (in press). (2003)

Akio Kishi、Takaaki Nakamura：“Atsunori Kashiwagi。Pdx1 的苏酰化对于胰岛素基因激活至关重要。”Am J 生理学、内分泌学和代谢..（出版中）。

中村高秋, 柏木厚典: "分子糖尿病の進歩 基礎から臨床まで-2003"金原出版. 200 (2003)

Takaaki Nakamura、Atsunori Kashiwagi：“分子糖尿病从基础知识到临床实践的进展 - 2003 年”Kanehara Publishing 200 (2003)。

Katsumasa Tsuzinaka, Takaaki Nakamura: "Dietary lipid hydroperoxide induces insulin resistance and impaired insulin secretion in normal rats."Diabetes Res Clin Pr. (In press).

Katsumasa Tsuzinaka、Takaaki Nakamura：“膳食脂质氢过氧化物会诱导正常大鼠的胰岛素抵抗并损害胰岛素分泌。”Diabetes Res Clin Pr。

中村高秋, 柏木厚典: "小腸上皮幹細胞を用いた膵β細胞の再生"分子糖尿病学の進歩(金原出版). 1. 10-16 (2003)

Takaaki Nakamura、Atsunori Kashiwagi：“利用小肠上皮干细胞再生胰腺 β 细胞”《分子糖尿病学进展》（Kanehara Publishing）（金原出版社）。

Akio Kishi, Takaaki Nakamura, et al.: "Sumoylation of Pdx1 is essential for insulin gene activation"Am J Physiol Endocrinol Metab. 284. E830-E840 (2003)

Akio Kishi、Takaaki Nakamura 等人：“Pdx1 的苏莫化对于胰岛素基因激活至关重要”Am J Physiol Endocrinol Metab。