Neutral lipid dysregulation of the pancreatic beta-cell

胰腺β细胞的中性脂质失调

• 批准号: 6517794
• 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 2005-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6517794
• 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：(摘自申请者的描述)2型糖尿病的特征是慢性高血糖，通常与 血脂水平升高。这项提议的总体目标是 确定长期暴露在高脂肪水平下的机制 酸(FA)影响2型糖尿病患者的胰岛β细胞功能。 此前，我们已经证明，长期接触FA会损害胰岛素 只有在高糖存在的情况下才能表达基因，而这是 与中性脂肪合成增加有关。具体目标一：确定 沿着中性脂途径产生的代谢中间体(S) 胰岛素分泌和基因损伤的合成 在长期暴露于FA时的表达。分离的大鼠胰岛，HIT-T15，和 将培养1~7天的BetaHC-L3细胞置于增殖期 葡萄糖和FA的浓度。将使用药理工具来抑制 或刺激中性脂肪合成的每一步，以识别 沿着酯化途径产生的代谢中间体(S， 长链酰辅酶A、二酰甘油或三酰甘油)负责 甲醛对胰岛β细胞功能的损伤。具体目标二：评估是否 长期暴露于高浓度甲醛的血糖依赖性有害效应 对β细胞功能的影响是葡萄糖专一性的，以及这些机制是否 影响是转录的、转录后的或翻译的。β细胞 在下列情况下的实验中，将区分精疲力竭和真实毒性 二氮嗪将用于抑制胰岛素的释放。糖蛋白的葡萄糖特异性 PA的影响将通过使用非葡萄糖促分泌剂来研究 刺激胰岛素分泌和胰岛素基因表达。依赖葡萄糖的 FA对胰岛素原生物合成、胰岛素信使核糖核酸稳定性的影响 内源性胰岛素基因转录将被评估。甲醛对人体健康的影响 胰岛素启动子活性将在HIT-T15和BetaHC-13细胞中表征 并在原代胰岛中使用重组腺病毒进行了研究 系统。具体目标三：确定高脂肪饲料是否 高血糖Goto-Kakizaki(GK)大鼠胰岛素分泌受损 生物合成和胰岛素基因表达，以及这些影响是否 通过血糖水平的正常化来预防。GK或对照大鼠将被 喂高脂饮食6周后，胰岛素分泌、胰岛素原 将评估胰岛素的生物合成和胰岛素基因的表达。血糖 GK大鼠体内的血药浓度将通过给药使其正常化。 预防高脂饮食对胰岛β细胞有害影响的尝试 功能。这些实验将为我们提供对 2型糖尿病胰岛β细胞功能障碍的病理生理学研究 对这种疾病的治疗的影响。