Glucose Regulation of Pancreatic Islet Gene Experession

描述（由申请人提供）：许多证据表明，长期暴露于高血糖症对2型糖尿病特征性胰岛细胞功能的持续下降有重要作用。我们和其他人在2型糖尿病的实验室模型（动物，分离的胰岛，b细胞系）中已经证明了这种现象的机制，称为细胞的葡萄糖毒性，涉及两种蛋白质PDX-1和MafA的水平降低，这两种蛋白质是胰岛素启动子活性的关键调节剂，从而是胰岛素基因表达的关键调节剂。我们的新研究分为3个具体目标：具体目标#1。确定谷胱甘肽过氧化物酶（GPx-1）的β细胞特异性过表达是否可防止C57 BLKS小鼠中链脲霉素诱导的高血糖症、C57 BU 6小鼠中高脂饮食诱导的高血糖症以及db/db和NOD小鼠中自发观察到的胰岛素分泌和葡萄糖耐量恶化。具体目标#2。确定从特定目的#1中体内研究的转基因小鼠胰腺中分离的胰岛是否可防止慢性暴露于高血糖症时发现的特征性异常。这些包括（1）胰岛素和PDX-1基因和MafA蛋白的表达降低;（2）胰岛素含量和胰岛素分泌减少;和（3）细胞凋亡增加和β细胞质量降低。具体目标#3确定MafA、PDX-1或两者联合稳定重建是否足以恢复葡萄糖毒性β细胞中内源性胰岛素基因表达的正常水平。实验将涉及体内生理学研究和体外分离胰岛的分子生物学研究，以及使用MafA和PDX-1的逆转录病毒过表达的研究。在这些实验的结论中，我们将确定在db/db和NOD小鼠的β细胞中特异性地增强内源性抗氧化酶活性是否将提供针对β细胞功能障碍和死亡的保护作用，所述β细胞功能障碍和死亡是这些自发性糖尿病动物模型的特征。我们还将确定MafA、PDX-1或两者组合在葡萄糖毒性β细胞中胰岛素基因表达缺陷中发挥机制作用的程度。外行语言：2型糖尿病发病后，多年的高葡萄糖水平会对制造胰岛素的细胞造成持续损害。这被认为是由于氧化应激，因为高葡萄糖水平增加了这些细胞中的氧自由基水平。在1型糖尿病发作时，引起氧化应激的细胞因子引起β细胞死亡。我们将研究是否增加抗氧化保护保护这些细胞，并防止糖尿病血糖控制恶化。

DESCRIPTION (provided by applicant): Many lines of evidence suggest that chronic exposure to hyperglycemia contributes importantly to the relentless decline in pancreatic islet ¿-cell function characteristic of type 2 diabetes. The mechanisms of this phenomenon, termed glucose toxicity of the ¿-cell, has been shown by us and by others in laboratory models of type 2 diabetes (animals, isolated islets, b-cell lines) to involve decreased levels of two proteins, PDX-1 and MafA, which are critical regulators of insulin promoter activity and thereby insulin gene expression. Our new studies are organized into 3 specific aims: Specific Aim #1. To determine whether beta cell-specific overexpression of glutathione peroxidase (GPx-1) will protect against worsening of insulin secretion and glucose tolerance characteristically observed in streptozotocin-induced hyperglycemia in C57BLKS mice, in high fat diet- induced hyperglycemia in C57BU6 mice, and spontaneously in db/db and NOD mice. Specific Aim #2. To determine whether islets isolated from pancreases of the transgenic mice studied in vivo in Specific Aim #1 are protected against abnormalities that are characteristically found with chronic exposure to hyperglycemia. These include (1) decreased expression of the insulin and PDX-1 genes and MafA protein; (2) decrements in insulin content and insulin secretion; and (3) increased apoptosis and decreased beta cell mass. Specific Aim #3. To determine whether stable reconstitution of MafA, PDX-1, or both in combination are sufficient to restore normal levels of endogenous insulin gene expression in glucotoxic beta cells. Experiments will involve physiologic studies in vivo and molecular biologic studies with isolated islets in vitro, as well as studies using retroviral overexpression of MafA and PDX-1. At the conclusion of these experiments we will ascertain whether enhancing endogenous antioxidant enzyme activity specifically in ¿- cells of db/db and NOD mice will provide protective effects against the beta cell dysfunction and death that are characteristic of these animal models of spontaneous diabetes. We also will determine to what degree MafA, PDX-1, or both in combination play mechanistic roles in defective insulin gene expression in glucotoxic beta cells. Lay language: After onset of type 2 diabetes, high glucose levels over many years cause continuing damage to the cells that make insulin. This is thought to be due to oxidative stress because high glucose levels increase oxygen radical levels in these cells. At the time of onset of type 1 diabetes, cytokines, which cause oxidative stress, cause beta cell death. We will study whether increasing antioxidant protection protects these cells and prevents deterioration of glucose control in diabetes.