Mechanisms of mechanotransduction in the enterochromaffin cells

肠嗜铬细胞中的机械转导机制

• 批准号: 9317486
• 负责人: Arthur Beyder
• 金额: $ 16.91万
• 依托单位: MAYO CLINIC ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-16 至 2020-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9317486
• 关键词: Absenteeism; Acute; Address; Affect; Alpha Cell; Animals; Area; Basic Science; Biology; Biophysics; Cartoons; Cations; Cell Line; Cell physiology; Cells; Chronic; Clinic; Colon; Consumption; Coupled; Couples; Cytoskeleton; Data; Development; Direct Expenditure; Disease; Doctor of Philosophy; Electric Capacitance; Electrochemistry; Electrophysiology (science); Elements; Enterochromaffin Cells; Environment; Epithelial; Epithelial Cells; Epithelium; Esthesia; Foundations; Functional Gastrointestinal Disorders; Functional disorder; Gastroenterology; Gastrointestinal Motility; Gastrointestinal tract structure; Goals; Health; Healthcare; Hepatology; Human; Image; Immunohistochemistry; International; Intestines; Ion Channel; Irritable Bowel Syndrome; K-Series Research Career Programs; Knock-out; Link; Manometry; Measurement; Mechanics; Mentors; Mentorship; Merkel Cells; Methods; Microscopy; Molecular; Morbidity - disease rate; Mus; Organoids; Outcome; Pharmacology; Physicians; Physiological; Physiology; Population; Postdoctoral Fellow; Productivity; Research; Resolution; Role; Scientist; Serotonin; Signal Transduction; Skin; Stimulus; Symptoms; Techniques; Testing; Therapeutic; Tissues; Training; Training Programs; Transgenic Animals; Visceral; Visceral Afferents; Work; authority; career; cell motility; cell type; cellular targeting; experimental study; gastrointestinal; gastrointestinal epithelium; in vivo; knock-down; mechanical force; mechanotransduction; new therapeutic target; novel; novel diagnostics; optogenetics; public health relevance; serotonin receptor; skills; targeted treatment; tool

 DESCRIPTION (provided by applicant): This Mentored Career Development Award (K08) proposal describes a five year training program for the physician scientist candidate with a long-term goal of becoming an academic authority in cellular and molecular mechanosensitivity in the gastrointestinal (GI) tract in health and disease. The candidate acquired a strong basic science foundation in ion channel biophysics during his PhD and post-doctoral work. The candidate now proposes to solidify his foundation and acquire a set of novel complementary skills necessary for an independent research career in GI tract mechanosensitivity. Mechanosensitivity is important for normal GI tract function and abnormal mechanosensitivity leads to disease. Enterochromaffin (EC) cells are mechanosensitive GI epithelial cells that synthesize and release serotonin (5-HT), which regulates GI motility and sensation. Mechanical forces are a major stimulus for 5-HT release by the EC cell, but the molecular mechanisms of EC cell mechanosensation are unclear. Acute cellular mechanosensation involves mechanosensitive ion channels (MSCs). A recently cloned MSC PIEZO2 is critical for Merkel cell mechanosensation, which is a mechanosensitive cell of the skin epithelium with developmental and functional similarities to the EC cell. The central hypothesis of this application is that PIEZO2 ion channels are the primary mechanosensors that transduce mechanical energy into 5-HT release by the EC cells. We will test the central hypothesis in 2 AIMs. The experiments in AIM 1 will determine the localization and mechanical activation of PIEZO2 in EC cells. The experiments in AIM 2 will determine how PIEZO2 activation is coupled to 5-HT release in single EC cells and intestinal organoids and how PIEZO2 activation couples to GI motility. The AIMs are supported by strong preliminary data which show that PIEZO2 is specific to human and mouse colon EC cells, that mechanical stimuli activate PIEZO2 in primary EC cells, leading to release of 5-HT, while the block of PIEZO2 in the colon epithelium decreases motility. To test the central hypothesis, we will use immunohistochemistry and super resolution imaging, electrophysiology and optogenetics in parallel with mechanostimulation of single cells and intestinal organoids, 5-HT measurements by electrochemistry and novel techniques, novel ex vivo and in vivo recordings of colonic motility in wild type and transgenic animals. This work will be performed in an academically nurturing environment within Mayo Clinic and with full support of the Division of Gastroenterology & Hepatology. The candidate will be supported by several consultants who are experts in the areas that the candidate identified as necessary training. He will be guided by a strong mentorship committee (Drs. Gianrico Farrugia and Nicholas LaRusso). Both are international authorities - Dr. Farrugia in ion channels in GI motility, and Dr. LaRusso in epithelial biology and mechanosensation. As a result of this work, the candidate will significantly advance our understanding of EC cell mechanosensitivity mechanisms in human physiology with a goal of providing a novel platform for therapeutic strategies as well as obtain data for a R01 application.

 描述(由申请人提供)：这份指导职业发展奖(K08)提案描述了一项针对内科科学家候选人的为期五年的培训计划，其长期目标是成为胃肠道(GI)细胞和分子机械敏感性方面的健康和疾病方面的学术权威。这位候选人在他的博士和博士后工作中获得了离子通道生物物理学方面的坚实的基础科学基础。候选人现在建议巩固他的基础，并获得一套新的补充技能，这是在胃肠道机械敏感性方面的独立研究生涯所必需的。机械敏感性对正常的胃肠道功能很重要，机械敏感性异常会导致疾病。肠嗜铬细胞(EC)是机械敏感的胃肠道上皮细胞，能合成和释放5-羟色胺(5-羟色胺)，调节胃肠动力和感觉。机械力是EC细胞释放5-羟色胺的主要刺激因素，但EC细胞机械感觉的分子机制尚不清楚。急性细胞机械感觉涉及机械敏感离子通道(MSCs)。最近克隆的MSC Piezo2是Merkel细胞机械感觉的关键，Merkel细胞是皮肤上皮细胞的机械敏感细胞，在发育和功能上与EC细胞相似。这一应用的中心假设是，Piezo2离子通道是EC细胞将机械能转化为5-羟色胺释放的主要机械感受器。我们将在两个目标中检验中心假设。AIM 1中的实验将确定Piezo2在EC细胞中的定位和机械激活。AIM 2的实验将确定Piezo2的激活如何与单个EC细胞和肠道器官中5-羟色胺的释放相耦合，以及Piezo2的激活如何与胃肠动力相耦合。这些目的得到了强有力的初步数据的支持，这些数据表明Piezo2是人和小鼠结肠EC细胞特有的，机械刺激激活了原代EC细胞中的Piezo2，导致5-羟色胺的释放，而Piezo2在结肠上皮中的阻断降低了运动能力。为了验证中心假设，我们将使用免疫组织化学和超分辨率成像、电生理学和光遗传学以及单细胞和肠道器官的机械刺激、电化学和新技术的5-羟色胺测量、野生型和转基因动物新的体外和体内结肠运动记录。这项工作将 在梅奥诊所的学术培养环境中进行，并在胃肠病和肝病科的全力支持下进行。候选人将得到若干顾问的支持，这些顾问是候选人确定为必要培训的领域的专家。他将由一个强大的指导委员会(Gianrico Farrugia博士和Nicholas LaRusso博士)指导。两人都是国际权威--胃肠动力领域的离子通道专家法鲁贾博士和霍普金斯博士。 拉鲁索在上皮生物学和机械感觉方面。作为这项工作的结果，候选人将极大地促进我们对人体生理学中EC细胞机械敏感性机制的理解，目标是为治疗策略提供一个新的平台，并为R01的应用获得数据。