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）：胆囊化（CCK）是一种主要的胃肠道激素，由离散的肠内分泌细胞（称为I细胞）产生，散布在整个肠道中。 CCK有多种动作，对于调节食物摄入和消化必不可少的营养素很重要。 CCK具体调节胆囊收缩，胰酶分泌，延迟胃排空并诱发饱腹感。与大多数GI激素一样，CCK在摄入一顿饭后将CCK分泌到血液中。摄入的脂肪是CCK分泌的最重要的饮食刺激物。养分如何与肠内分泌细胞相互作用并刺激胃肠道激素的分泌有很大的兴趣。即使人们普遍认为营养直接刺激CCK释放，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尚不清楚。 PI开发了一种分离和表征个体，可行的天然肠道CCK细胞的方法，并且通过高度富集这些细胞，可以在体外研究CCK分泌，鉴定这些细胞上的受体并研究涉及受调节激素分泌的第二递观信号信号传导途径。这些方法一起导致了我们最近在CCK细胞上发现了一种新型受体，即含有受体的免疫球蛋白样结构域（ILDR）。在肠中，ILDR仅在CCK细胞中表达。 PI产生了具有ILDR遗传缺失的小鼠，并证明ILDR缺乏小鼠在脂肪进食后不分泌CCK。 ILDR是蛋白质残留受体家族的新成员，并通过独特的作用机理被激活。这些初步数据表明，CCK细胞表达了一种新型受体，这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）分离和鉴定天然CCK细胞，（2）在体内和体外的CCK分泌测量，（3）定量细胞内钙荧光，以及（4）研究ILDR表达细胞中细胞内信号的方法。新型模型将包括：（1）在CCK细胞（CCK-EGFP小鼠）中表达增强绿色荧光蛋白的转基因小鼠以实现CCK细胞的鉴定和隔离，（2）ILDR基因敲除小鼠（ILDR - / - ），（3）双CCK-EGFP TransgE/ILDRID/ILDRILDR-ILDR-ILDR-ELDR-ELDR-EGFP/CCK-EGFP/CCCK-CCCK-ccck-ccck cccc cccc y cccc y cy theyal and-cccy thecy）肠道CCK细胞特异性ILDR敲除小鼠。这些转基因的小鼠将使我们能够研究完整小鼠的ILDR功能，以及来自正常和ILDR缺乏小鼠的分离的CCK细胞中的ILDR功能。该应用的中心假设是ILDR介导了脂肪刺激的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。以下特定目的将解决ILDR的表征及其与CCK细胞中激素分泌的关系：1。建立脂肪酸和脂蛋白调节ILDR功能的机制。 2。为了表征ILDR在体内小鼠中调节CCK分泌，胆囊功能和进食行为中的作用。这些目标中的每一个都将集中于对ILDR的调节，这是调节脂肪酸刺激的CCK分泌的关键步骤。在全球范围内，这些目标应提供有关GI内分泌细胞受到已知在控制激素分泌很重要的营养调节的机制的大量见解。