Regulation and Function of Urocortins and their Receptors

尿皮质素及其受体的调节和功能

• 批准号: 8586661
• 负责人: Aditi Bhargava
• 金额: $ 0.15万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8586661
• 关键词: Abbreviations; Adrenal Glands; Affect; Agonist; Anxiety; Arrestins; Brain region; Cell Line; Cell membrane; Cells; Chronic; Clostridium difficile; Colitis; Colon; Confocal Microscopy; Corticotropin-Releasing Hormone; Corticotropin-Releasing Hormone Receptors; Crohn' s disease; Data; Development; Disease; Dose; Double-Stranded RNA; Down-Regulation; Dynamin; Employee Strikes; Endocytosis; Exhibits; Extravasation; Family; Feeding behaviors; Financial compensation; Gastrointestinal Motility; Gastrointestinal tract structure; Genes; Genetic; Genetic Models; Glucocorticoids; Hormonal; Hypothalamic structure; Ileitis; Immune; Immune response; Immune system; Immunofluorescence Immunologic; Immunohistochemistry; Inflammation; Inflammation Mediators; Inflammatory; Inflammatory Bowel Diseases; Inflammatory disease of the intestine; Interleukin-10; Intestines; Knock-out; Knockout Mice; Ligands; Link; Liquid substance; Measures; Mediating; Mediator of activation protein; Messenger RNA; Modeling; Modification; Molecular; Moods; Mus; Neuraxis; Neurons; Neuropeptides; Neurosecretory Systems; Organ; Outcome; Pathway interactions; Patients; Peptide Receptor; Peptides; Piroxicam; Pituitary Hormones; Plasma; Play; Process; Production; Protein Isoforms; Proteins; Quality of life; RNA Interference; Recycling; Regulation; Research; Role; Scaffolding Protein; Severities; Signal Transduction; Stress; Sulfonic Acids; System; Testing; Therapeutic; Tissues; Toxin; Trinitrobenzenes; Ubiquitin; Weight Gain; autonomic nerve; biological adaptation to stress; cytokine; defined contribution; desensitization; gastrointestinal; gastrointestinal function; hypothalamic-pituitary-adrenal axis; ileum; immunoreactivity; improved; innovation; life time cost; mortality; psychologic; receptor; receptor function; receptor recycling; response; single molecule; stressor; trafficking; urocortin

PROJECT SUMMARY/ABSTRACT The corticotropin-releasing factor (CRF) family of neuropeptides (CRF and urocortins [Ucn] 1-3) and their receptors (CRFR1, CRFR2) are essential mediators of stress in the central nervous system. Therefore a systemic inhibition of their function is not an attractive therapeutic model. Components of this peptide/receptor family are prominently expressed within the intestine, where their local functions remain to be defined. Our preliminary data from CRFR2 heterozygous mice suggests that perturbation of the CRFR2 system renders the mice more susceptible to sudden stress and immune challenges. Our results suggest that new treatments for intestinal inflammatory diseases may require either antagonists or agonists of CRF receptors, depending upon the affected intestinal region and the type of inflammatory disease. A striking feature we observed during colitis was reciprocal changes in mRNA and protein levels of Ucn ligands and their receptors. These differential changes can alter Ucn-induced trafficking of receptors from and to the plasma membrane. However, the molecular mechanisms of this divergent trafficking to recycling or degradatory pathways remain unexplored. Thus, our hypothesis is that Ucn isoforms, differentially signaling through CRF receptors, govern intestinal inflammation at the molecular, cellular and organ levels. The aims of this proposal are to: Specific Aim 1. To define the systemic role of the Ucn/CRFR system in intestinal inflammation. We will use pharmacological and genetic approaches to define the systemic role of the Ucn/CRFR system in intestinal inflammation. We will determine if CRFR2 knockout and heterozygous mice exhibit differential sensitivity to inflammation and agonist treatment as compared with their wild type littermates. We will determine alterations in colitis severity by measuring changes in body weight gain, mortality, histological damage, and fluid leak. Specific Aim 2. To define the contributions of colon-specific Ucns and CRFRs in intestinal inflammation using RNA interference (RNAi). To determine the colon-specific role of the Ucn/CRFR system, we will establish the use RNAi to knockdown expression of Ucns and CRFR2 locally in the colon. We will determine the spatial and temporal effects of RNAi in the colon, and assess whether colon-specific knockdown of Ucns and CRFRs is sufficient to ameliorate or exacerbate colitis, using end points defined in Specific Aim 1. Specific Aim 3. To define the mechanism and function of Ucn1-induced trafficking of CRFR1. We will examine the influence of Ucn1 stimulation on the subcellular localization of CRFR1 using confocal microscopy and immunofluorescence. We will explore the ability of the adaptor/scaffolding proteins (¿-arrestins and dynamin) to desensitize and resensitize CRFRs. We will assess Ucn1 regulation of CRFR1 activity by measuring Ucn1-stimulated intracellular Ca2+ mobilization. Thus, manipulation of the urocortins and their receptors provides an effective starting point for understanding their GI functions, which could contribute to the development of new treatments for inflammatory bowel disease.

项目摘要/摘要 神经肽蛋白蛋白释放因子（CRF）家族（CRF和尿道素[UCN] 1-3）及其家族 受体（CRFR1，CRFR2）是中枢神经系统应力的基本介体。因此 系统性抑制其功能不是一个有吸引力的治疗模型。这个胡椒/受体的成分 家庭在肠内占据着突出的表达，那里的局部功能仍有待定义。我们的 来自CRFR2杂合小鼠的初步数据表明，CRFR2系统的扰动使得 小鼠更容易受到突然的压力和免疫挑战。我们的结果表明，新的治疗方法 肠道炎症性疾病可能需要CRF受体的拮抗剂或激动剂，具体取决于 受影响的肠道区域和炎症性疾病的类型。我们在结肠炎期间观察到的惊人特征 是UCN配体及其受体的mRNA和蛋白质水平的相互变化。这些差异 变化可以改变UCN引起的接收器从质膜和到质膜的贩运。但是， 这种发散运输到回收或降解途径的分子机制仍然出乎意料。 这是我们的假设是，通过CRF受体的信号不同的UCN同工型控制 分子，细胞和器官水平的肠炎。该提案的目的是： 具体目的1。定义UCN/CRFR系统在肠道注射中的系统作用。我们将 使用药物和遗传方法来定义UCN/CRFR系统在肠道中的系统作用 炎。我们将确定CRFR2敲除和杂合小鼠是否暴露于 与野生型同窝仔相比，炎症和激动剂治疗。我们将确定更改 通过衡量体重增加，死亡率，组织学损害和液体泄漏的变化，在结肠炎的严重程度中。 特定目的2。定义肠中结肠特异性UCN和CRFR的贡献 使用RNA干扰（RNAI）的炎症。为了确定UCN/CRFR系统的结肠特异性作用， 我们将建立RNAi在结肠中局部的UCN和CRFR2的表达。我们将 确定RNAi在结肠中的空间和临时效应，并评估结肠特异性敲低 使用特定目标1中定义的终点，UCN和CRFR足以改善或恶化结肠炎。 具体目的3。定义UCN1诱导的CRFR1运输的机理和功能。我们将 使用共聚焦显微镜检查UCN1模拟对CRFR1亚细胞定位的影响 和免疫荧光。我们将探讨适配器/脚手架蛋白（�-arrestins and脚手架）的能力 dynamin）脱敏和恢复CRFR。我们将通过 测量UCN1刺激的细胞内Ca2+动员。那是对尿道素及其的操纵 接收者为理解其GI功能提供了有效的起点，这可能有助于 开发炎症性肠病的新疗法。