Neutral lipid dysregulation of the pancreatic beta-cell

胰腺β细胞的中性脂质失调

• 批准号: 6635296
• 负责人: VINCENT POITOUT
• 金额: $ 26.25万
• 依托单位: PACIFIC NORTHWEST RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-06-01 至 2005-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6635296
• 关键词: acyl coA; blood glucose; diacylglycerols; esterification; fatty acid biosynthesis; fatty acids; gel mobility shift assay; gene expression; genetic promoter element; genetic transcription; genetic translation; glucose metabolism; hyperglycemia; insulin; insulin sensitivity /resistance; laboratory rat; messenger RNA; noninsulin dependent diabetes mellitus; northern blottings; pancreatic islet function; pancreatic islets; posttranscriptional RNA processing; proinsulin; tissue /cell culture; transfection

DESCRIPTION: (Scanned from the applicant's description) Type 2 diabetes mellitus is characterized by chronic hyperglycemia and is often associated with elevated plasma lipid levels. The overall objective of this proposal is to ascertain the mechanisms whereby prolonged exposure to elevated levels of fatty acids (FA) affects pancreatic beta-cell function in Type 2 diabetes. Previously, we have demonstrated that prolonged exposure to FA impairs insulin gene expression only in the presence of high glucose, and that this is associated with increased neutral lipid synthesis. Specific Aim I: To identify the metabolic intermediate(s) generated along the pathway of neutral lipid synthesis responsible for the impairment of insulin secretion and gene expression upon prolonged exposure to FA. Isolated rat islets, HIT-T15, and betaHC-l3 cells will be cultured for 1 to 7 days in the presence of increasing concentrations of glucose and FA. Pharmacological tools will be used to inhibit or stimulate each step of neutral lipid synthesis, in order to identify the metabolic intermediate(s) generated along the esterification pathway (i.e., long-chain Acyl-CoA, diacyiglycerols, or triacylglycerols) responsible for the FA-induced impairment of beta-cell function. Specific Aim II: To assess whether the glucose-dependent deleterious effects of prolonged exposure to elevated FA on beta-cell function are glucose-specific, and whether the mechanisms of these effects are transcriptional, post-transcriptional, or translational. beta-cell exhaustion will be distinguished from bona fide toxicity in experiments where diazoxide will be used to inhibit insulin release. The glucose-specificity of PA effects will be investigated by using a non-glucose secretagogue to stimulate insulin secretion and insulin gene expression. The glucose-dependent effects of FA on proinsulin biosynthesis, insulin mRNA stability, and endogenous insulin gene transcription will be assessed. The effects of FA on insulin promoter activity will be characterized in HIT-Tl5 and betaHC-13 cells and also investigated in primary islets using the recombinant adenovirus system. Specific Aim III: To determine whether high-fat feeding in hyperglycemic Goto-Kakizaki (GK) rats impairs insulin secretion, insulin biosynthesis, and insulin gene expression, and whether these effects are prevented by normalization of blood glucose levels. GK or control rats will be fed a high-fat diet for 6 weeks, after which insulin secretion, proinsulin biosynthesis, and insulin gene expression will be assessed. Blood glucose levels will be normalized in GK rats by phloridzin administration, in an attempt to prevent the deleterious effects of high-fat diet on beta-cell function. These experiments will provide important insights into the pathophysiology of beta-cell dysfunction of type 2 diabetes, and have clear implications for the treatment of this disease.

描述：（从申请人的描述中扫描）类型2糖尿病的特征是慢性高血糖，并且通常与 血浆脂质水平升高。该提议的总体目的是 确定长期暴露于脂肪水平的机制 酸（FA）影响2型糖尿病中的胰腺β细胞功能。 以前，我们已经证明了长时间暴露于FA会损害胰岛素 基因表达仅在高葡萄糖的情况下，这就是 与中性脂质合成增加有关。特定目的I：确定 沿中性脂质途径产生的代谢中间体 负责胰岛素分泌和基因损害的合成 长时间暴露于FA时表达。孤立的大鼠胰岛，HIT-T15和 在存在增加的情况下，将培养BetAHC-L3细胞1至7天 葡萄糖和FA的浓度。药理工具将用于抑制 或刺激中性脂质合成的每个步骤，以识别 沿酯化途径生成的代谢中间体（即 长链酰基-COA，二酰甘油甘油或三酰基甘油）负责 FA诱导的β细胞功能受损。特定目标II：评估是否 长时间暴露于升高的FA的葡萄糖依赖性有害作用 在β细胞功能上是葡萄糖特异性的，以及这些机制是否具有 效果是转录，转录后或翻译。 beta细胞 在实验中，精疲力尽将与真正的毒性区分开 二氮杂将用于抑制胰岛素释放。葡萄糖特异性 PA效应将通过使用非葡萄糖秘密来研究 刺激胰岛素分泌和胰岛素基因表达。葡萄糖依赖性 FA对蛋白质生物合成，胰岛素mRNA稳定性和 内源性胰岛素基因转录将被评估。 FA的影响 胰岛素启动子活性将在HIT-TL5和BETAHC-13细胞中进行表征 并使用重组腺病毒在主要胰岛进行研究 系统。特定目标III：确定是否进食高脂 高血糖goto-kakizaki（GK）大鼠损害胰岛素分泌，胰岛素 生物合成和胰岛素基因表达，以及这些作用是否为 通过血糖水平归一化阻止。 GK或控制大鼠将是 喂高脂饮食6周，然后胰岛素分泌，促硫素 将评估生物合成和胰岛素基因表达。血糖 菲洛津给药将在GK大鼠中标准化水平 尝试防止高脂饮食对β细胞的有害影响 功能。这些实验将为您提供重要的见解 2型糖尿病的β细胞功能障碍的病理生理学，并具有清晰的 对治疗这种疾病的影响。