Impact of Neprilysin on Islet Function

脑啡肽酶对胰岛功能的影响

• 批准号: 8595010
• 负责人: Sakeneh Zraika
• 金额: $ 29万
• 依托单位: SEATTLE INST FOR BIOMEDICAL/CLINICAL RES
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-01 至 2018-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8595010
• 关键词: Ablation; Address; Binding; Biological Preservation; Blood Circulation; C-terminal; Calcium; Capsaicin; Cell Culture Techniques; Cell membrane; Cell physiology; Cells; Cleaved cell; Data; Deafferentation procedure; Diabetes Mellitus; Diet; Dietary Fats; Dipeptidyl Peptidases; Dose; Exposure to; Failure; Fatty acid glycerol esters; Functional disorder; Gastric Inhibitory Polypeptide; Genetic; Glucagon; Glucose; Half-Life; Hepatic; Human; Hyperglycemia; Hyperlipidemia; In Vitro; Insulin; Insulin Resistance; Intestines; Islet Cell; L Cells; Link; Maintenance; Measures; Mediating; Modeling; Mus; Neprilysin; Neurons; Non-Insulin-Dependent Diabetes Mellitus; Oral; Oral Administration; Palmitates; Pancreas; Peptide Hydrolases; Peptides; Peripheral; Production; Relative (related person); Role; Signal Transduction; Site; Somatostatin; Streptozocin; Testing; Therapeutic; Transplantation; Vagotomy; Wit; blood glucose regulation; comparative efficacy; diabetic; feeding; glucagon-like peptide; glucagon-like peptide 1; glucose tolerance; improved; in vivo; in vivo Model; inhibitor/antagonist; insulin secretion; islet; neuromechanism; novel; overexpression; paracrine; public health relevance; receptor; receptor binding; relating to nervous system; research study; response; therapeutic development; villin

DESCRIPTION (provided by applicant): Type 2 diabetes is characterized by islet ¿-cell failure, contributed to by hyperglycemia and hyperlipidemia. Neprilysin (NEP) is a widely expressed plasma membrane peptidase that we have shown is increased in islets with prolonged exposure to elevated glucose and fat. Like dipeptidyl peptidase-4 (DPP-4), NEP can degrade and inactivate glucagon-like peptide-1 (GLP-1), a glucose-dependent insulinotropic peptide secreted by intestinal L cells and islet ? cells. Our preliminary data show that under conditions of increased dietary fat, NEP ablation in mice enhances active GLP-1 levels thereby contributing to increased glucose-stimulated insulin secretion (GSIS) and improved glucose tolerance. Thus in this proposal, we will test the hypothesis that increased NEP activity limits the beneficial effect of GLP-1 on insulin secretion and glucose homeostasis. To address this hypothesis, the following specific aims have been identified: 1) To determine the contribution of islet NEP to reduced GSIS via its ability to degrade active GLP-1. First, an in vitro culture model will be utilized to measure active GLP-1 levels in human and mouse islets cultured with or without palmitate or high glucose to determine whether GLP-1 levels are reduced in conditions associated with increased NEP activity. Islets with pharmacological inhibition or genetic ablation of NEP will be used for comparison. Second, an in vivo model will be utilized in which wild-type or NEP-/- islets will be transplanted into streptozocin-diabetic wild-type or NEP-/- syngeneic recipients to evaluate the contribution of islet NEP to the maintenance of active GLP-1 levels and normal ¿-cell function following transplantation. 2) To determine the contribution of intestinal NEP to reduced GSIS via its ability to degrade active GLP- 1. GLP-1 released from L cells facilitates GSIS directly via the systemic circulation and indirectly via neural signals from vagal afferents. We will assess whether selective reduction of NEP activity in mouse intestine using pharmacologic and genetic (NEPflox ? Villin-Cre) approaches increases portal and peripheral active GLP- 1 levels and thereby enhances GSIS. The contribution of neural signaling to enhanced GSIS will be determined using selective hepatic vagotomies or capsaicin to block activation of pancreatic vagal efferents. 3) To compare the efficacy of NEP versus DPP-4 inhibition in enhancing active GLP-1 levels and improving fat-induced insulin secretory dysfunction in vivo. NEP, DPP-4 or both will be pharmacologically inhibited in wild-type control mice fed a low or high fat diet for 18 weeks. Active GLP-1 levels and GSIS will be measured to determine the relative contributions of each peptidase to preservation of ¿-cell function in mice fed a high fat diet. Dual peptidase inhibition will test whether any beneficial effect observed wit NEP inhibition can be augmented with concurrent DPP-4 inhibition. These studies will delineate a novel role for islet and intestinal NEP in modulating ¿-cell function, and will have significant implications for development of therapeutics to treat ¿-cell dysfunction in diabetes.

描述(申请人提供)：2型糖尿病的特征是胰岛细胞衰竭，由高血糖和高脂血症引起。Neprilysin(NEP)是一种广泛表达的质膜多肽酶，我们已经发现，在长期暴露于高糖和高脂肪的胰岛中，NEP增加。与二肽基肽酶-4(DPP-4)一样，NEP也能降解和失活肠L细胞和胰岛分泌的葡萄糖依赖型胰岛素样多肽-1。细胞。我们的初步数据显示，在饮食脂肪增加的条件下，消融NEP可以提高小鼠体内活性GLP-1的水平，从而有助于增加葡萄糖刺激的胰岛素分泌(GSIS)和改善葡萄糖耐量。因此，在这个提案中，我们将检验NEP活性增加限制了GLP-1对胰岛素分泌和葡萄糖稳态的有益影响的假设。为了解决这一假说，已经确定了以下具体目的：1)确定胰岛NEP通过其降解活性GLP-1的能力而降低GSIS的贡献。首先，将利用体外培养模型来测量在有或没有棕榈酸盐或高葡萄糖的情况下培养的人和小鼠胰岛中活性GLP-1的水平，以确定在与NEP活性增加相关的条件下GLP-1水平是否降低。NEP的药物抑制或基因消融的胰岛将用于比较。其次，将利用体内模型，将野生型或NEP-/-胰岛移植到链脲佐菌素-糖尿病野生型或NEP-/-同基因受体中，以评估胰岛NEP在移植后维持活性GLP-1水平和正常细胞功能方面的贡献。从L细胞释放的GLP-1直接通过体循环促进GSIS，并通过神经信号间接促进GSIS。 迷走神经传入。我们将评估是否使用药物和遗传方法选择性降低小鼠肠道中的NEP活性(NEP？维林-CRE治疗可增加门静脉和外周血中活性GLP-1的水平，从而增强GSIS。神经信号对增强GSIS的作用将通过选择性肝迷走神经切断术或辣椒素阻断胰腺迷走神经传出的激活来确定。3)比较NEP和DPP-4抑制剂在体内提高GLP-1活性和改善脂肪诱导的胰岛素分泌功能障碍的效果。在喂食低脂或高脂饮食18周的野生型对照组小鼠中，NEP、DPP-4或两者都会受到药物抑制。将测量活性GLP-1水平和GSIS，以确定每种多肽酶对高脂饮食小鼠细胞功能保存的相对贡献。双肽酶抑制将测试在抑制NEP的情况下观察到的任何有益效果是否可以与同时抑制DPP-4一起增强。这些研究将描绘出胰岛和肠道NEP在调节细胞功能方面的新作用，并将对开发治疗糖尿病细胞功能障碍的药物具有重要意义。