Receptor Regulation of CCK Cell Function

CCK 细胞功能的受体调节

• 批准号: 8918605
• 负责人: Rodger A. Liddle
• 金额: $ 34.15万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-18 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8918605
• 关键词: Address; Animals; Apical; Attention; Binding; Blood; Calcium; Cell Line; Cell model; Cell physiology; Cells; Cholecystokinin; Cholecystokinin Receptor; Cholelithiasis; Data; Diet; Digestion; Disease; Distant; Eating; Endocrine; Enteroendocrine Cell; Exhibits; Family; Fatty Acids; Fatty acid glycerol esters; Feeding behaviors; Fluorescence; Fluorescence Microscopy; Fluorescence-Activated Cell Sorting; Food; Food Intake Regulation; Gall Bladder Diseases; Gallbladder; Gastrointestinal Hormones; Gastroparesis; Genetic; Health; Hormonal; Hormones; Human; Immunoglobulin Domain; In Vitro; Individual; Ingestion; Intestines; Knockout Mice; Lead; Lipoprotein Binding; Lipoproteins; Measurement; Mediating; Methods; Modeling; Mus; Normal Cell; Nutrient; Obesity; Pancreatic enzyme; Pattern; Physiological; Property; Protein Family; Receptor Activation; Regulation; Role; Satiation; Second Messenger Systems; Signal Pathway; Signal Transduction; Stream; Study models; Surface; Techniques; Time; Tissues; Transgenic Mice; Transgenic Organisms; absorption; enhanced green fluorescent protein; feeding; gastrointestinal function; hormone regulation; in vivo; increased appetite; insight; interest; intestinal epithelium; lipolysis-stimulated receptor; lipoprotein-remnant receptor; member; mouse model; novel; receptor; receptor function; second messenger; tool

DESCRIPTION (provided by applicant): Cholecystokinin (CCK) is a major gastrointestinal hormone that is produced by discrete enteroendocrine cells (known as I cells) that are scattered throughout the intestine. CCK has several actions that are important for the regulation of food intake and digestion of essential nutrients. Specifically CCK regulates gallbladder contraction, pancreatic enzyme secretion, delays gastric emptying, and induces satiety. As is typical of most GI hormones, CCK is secreted into the blood stream after ingestion of a meal. Ingested fat is the most important dietary stimulant of CCK secretion. There is considerable interest in how nutrients interact with enteroendocrine cells and stimulate gastrointestinal hormone secretion. Even though it is generally believed that nutrients stimulate CCK release directly this has recently come into question and the cellular mechanisms regulating CCK cell function are largely unknown. The PI has developed a method for isolating and characterizing individual, viable, native intestinal CCK cells and by highly enriching these cells it has been possible to study CCK secretion in vitro, identify receptors on these cells and investigate second messenger signaling pathways involved in regulated hormone secretion. Together these approaches led to our very recent discovery of a novel receptor on CCK cells, known as immunoglobulin-like domain containing receptor (ILDR). In the intestine, ILDR is expressed exclusively in CCK cells. The PI has generated mice with genetic deletion of Ildr and demonstrated that ILDR deficient mice do not secrete CCK following fat feeding. ILDR is a novel member of the remnant receptor family of proteins and is activated through a unique mechanism of action. These preliminary data indicate that CCK cells express a novel receptor that is critical for regulating fat-induced CCK secretion. The PI will use a combination of complementary techniques to characterize ILDR in CCK cells and establish how ILDR regulates CCK secretion. These methods include: (1) isolation and identification of native CCK cells, (2) measurements of CCK secretion in vivo and in vitro, (3) quantification of intracellular calcium fluorescence, and (4) approaches to study intracellular signaling in ILDR -expressing cells. Novel models will include: (1) transgenic mice expressing enhanced green fluorescent protein in CCK cells (CCK-eGFP mice) to enable the identification and isolation of CCK cells, (2) ILDR knockout mice (ILDR-/-), (3) dual CCK-eGFP transgenic / ILDR knockout mice (CCK-eGFP/ILDR-/-), and conditional, intestinal CCK cell specific ILDR knockout mice. These genetically modified mice will enable us to study ILDR function in intact mice and in isolated CCK cells from normal and ILDR deficient mice. The central hypothesis of this application is that ILDR mediates fat-stimulated CCK secretion. The overall purpose of this proposal is to understand the mechanism whereby fat and fatty acids control CCK secretion. Characterization of ILDR and its relationship to hormone secretion in CCK cells will be addressed by the following Specific Aims: 1. To establish the mechanism by which fatty acids and lipoproteins regulate ILDR function. 2. To characterize the role of ILDR in regulating CCK secretion, gallbladder function and feeding behavior in mice in vivo. Each of these aims will focus on regulation of ILDR as a critical step in the regulation of fatty acid-stimulated CCK secretion. More globally, these aims should provide considerable insight into the mechanisms by which GI endocrine cells are regulated by nutrients known to be important in the control of hormone secretion.

描述（由申请人提供）：缩胆囊素 (CCK) 是一种主要胃肠道激素，由分散在整个肠道的离散肠内分泌细胞（称为 I 细胞）产生。 CCK 具有多种对于调节食物摄入和必需营养素消化非常重要的作用。具体而言，CCK 调节胆囊收缩、胰酶分泌、延迟胃排空并诱导饱腹感。与大多数胃肠道激素一样，CCK 在进食后分泌到血流中。摄入的脂肪是 CCK 分泌最重要的饮食刺激剂。人们对营养物质如何与肠内分泌细胞相互作用并刺激胃肠激素分泌非常感兴趣。尽管人们普遍认为营养物质直接刺激 CCK 释放，但这最近受到质疑，并且调节 CCK 细胞功能的细胞机制在很大程度上是未知的。 PI 开发了一种分离和表征个体、活的、天然肠道 CCK 细胞的方法，通过高度富集这些细胞，可以在体外研究 CCK 分泌，识别这些细胞上的受体，并研究参与调节激素分泌的第二信使信号传导途径。这些方法共同导致我们最近在 CCK 细胞上发现了一种新型受体，称为包含免疫球蛋白样结构域的受体 (ILDR)。在肠道中，ILDR 仅在 CCK 细胞中表达。 PI 培育出了 Ildr 基因缺失的小鼠，并证明 ILDR 缺陷的小鼠在脂肪喂养后不会分泌 CCK。 ILDR 是蛋白质残余受体家族的新成员，通过独特的作用机制被激活。这些初步数据表明，CCK 细胞表达一种新受体，该受体对于调节脂肪诱导的 CCK 分泌至关重要。 PI 将使用互补技术的组合来表征 CCK 细胞中的 ILDR 并确定 ILDR 如何调节 CCK 分泌。这些方法包括：(1)天然CCK细胞的分离和鉴定，(2)体内和体外CCK分泌的测量，(3)细胞内钙荧光的定量，以及(4)研究表达ILDR的细胞中的细胞内信号传导的方法。新模型将包括：（1）在CCK细胞中表达增强型绿色荧光蛋白的转基因小鼠（CCK-eGFP小鼠），以实现CCK细胞的识别和分离，（2）ILDR敲除小鼠（ILDR-/-），（3）双CCK-eGFP转基因/ILDR敲除小鼠（CCK-eGFP/ILDR-/-），以及条件， 肠道CCK细胞特异性ILDR敲除小鼠。这些转基因小鼠将使我们能够研究完整小鼠以及来自正常和 ILDR 缺陷小鼠的分离 CCK 细胞的 ILDR 功能。本申请的中心假设是 ILDR 介导脂肪刺激的 CCK 分泌。该提案的总体目的是了解脂肪和脂肪酸控制 CCK 分泌的机制。 ILDR 的表征及其与 CCK 细胞激素分泌的关系将通过以下具体目标来解决： 1. 建立脂肪酸和脂蛋白调节 ILDR 功能的机制。 2. 表征ILDR在体内调节CCK分泌、胆囊功能和摄食行为中的作用。这些目标中的每一个都将重点关注 ILDR 的调节，作为调节脂肪酸刺激的 CCK 分泌的关键步骤。在全球范围内，这些目标应该提供对胃肠道内分泌细胞受已知对激素分泌控制很重要的营养物质调节的机制的深入了解。