Epithelial Acetylcholine Oral Biology and Pathology

上皮乙酰胆碱口腔生物学和病理学

基本信息

批准号：
7514038
负责人：
SERGEI A GRANDO
金额：
$ 10.06万
依托单位：
UNIVERSITY OF CALIFORNIA-IRVINE
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-04-01 至 2008-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7514038
关键词：
Acetylcholine Acetylcholinesterase Adhesions Agonist Apoptosis Binding Biological Biological Assay Birth Cell Cycle Cell Cycle Progression Cell Line Cell physiology Cells Cellular biology Cessation of life Chemicals Chewing Gum Choline O-Acetyltransferase Cholinergic Agents Cholinergic Receptors Chronic Class Code Coupled Data Depth Development Drosophila acetylcholine receptor alpha-subunit Environmental Tobacco Smoke Enzyme-Linked Immunosorbent Assay Enzymes Epithelial Epithelial Cells Epithelium Equilibrium Esophageal Esophageal mucous membrane Event Exhibits Exposure to GTP-Binding Proteins Gastrointestinal tract structure Gated Ion Channel Gingiva Glycoproteins Goals Health Human Human Cloning Immunohistochemistry In Vitro Inhalators Knockout Mice Long-Term Effects Mediating Mediator of activation protein Medical Messenger RNA Metabolic Methodology Molecular Morbidity - disease rate Mucous Membrane Mus Muscarinic Acetylcholine Receptor Muscarinics Neurons Nicotine Northern Blotting Nose Numbers Oligonucleotides Oral Pathology Pathway interactions Pharmaceutical Preparations Physiological Play Polymerase Chain Reaction Process Purpose Regulation Replacement Therapy Reverse Transcriptase Polymerase Chain Reaction Role Signal Pathway Signal Transduction Smokeless Tobacco Solid Staging Structure System Time Tobacco Tobacco Use Cessation Tobacco smoke Transcript Western Blotting Yin-Yang absorption aqueous cholinergic keratinocyte keratinocyte differentiation migration nicotine replacement novel oral biology oral cavity epithelium receptor research study stem

项目摘要

DESCRIPTION (provided by applicant): PURPOSE: To identify the medical and biological significance of the novel non-neuronal cholinergic system of the upper digestive tract, and elucidate the effects of pure nicotine (Nic) vs. tobacco products on nicotinic and muscarinic acetylcholine receptors (nAChR and mAChR) expressed by oral and esophageal keratinocytes (KC). BACKGROUND: ACh regulates vital functions of KC including proliferation, adhesion, migration, differentiation and apoptosis. The alpha3, alpha5, alpha7, alpha9, beta2, and beta4 nAChR subunits, the m2, m3, m4 and m5 mAChR subtypes, as well as the synthesizing enzyme choline acetyltransferase and the degrading enzyme acetylcholinesterase are expressed and function in human oral KC, and may target KC for direct effects of Nic. PRELIMINARY OBSERVATIONS: Concomitant emergence of different ACh receptors in the course of KC differentiation allows ACh to exhibit diverse and temporally regulation of the developing mucosal epithelium. A stimulatory effect of ACh on Ca2+ influx, mediated by nAChR, balances an inhibitory effect, mediated by mAChR, and simultaneous activation of both receptors produces a kind of a yin yang regulatory balance in KC. HYPOTHESES: 1) Each ACh receptor expressed by oral KC regulates uniquely the cell cycle progression so that the differentiation-determined changes in the repertoire of ACh receptors in a single keratinocyte can diversify the biological effects of ACh. 2) Chronic exposure to Nic disturbs the dynamic equilibrium between the nicotinic and the muscarinic pathways of keratinocyte control by ACh, leading to an aberrant cell cycle. SPECIFIC AIMS: 1) Identify contribution of each ACh receptor to the cell cycle progression using subtype-selective agonists and antagonists, antisence oligonucleotides, and knockout mice. 2) Identify long-term effects of Nic vs. aqueous extract of smokeless tobacco vs. environmental tobacco smoke on expression/function of keratinocyte ACh receptors and the Ca2+ pathways subserving ACh signaling in KC. METHODOLOGY: A combination of molecular biological (RT-PCR, real-time TaqMan PCR, Northern blots), immunological (immunohistochemistry, Western blots, ELISA), pharmacological and biological assays will be employed to quantitate ACh receptor-mediated changes of the cell cycle progression (Aim #1) as well as effects of Nic and tobacco products on the structure and function of keratinocyte ACh receptors (Aim #2). SIGNIFICANCE: The role of non-neuronal ACh in oral cell biology and pathology will be elucidated, which will help explain some of the mechanisms of deleterious effects of tobacco products in the upper digestive tract.

描述（由申请人提供）：目的：确定医疗和新型非神经元胆碱能系统的生物学意义上消化道，并阐明纯尼古丁 (Nic) 与尼古丁 (Nic) 的影响烟草制品对烟碱和毒蕈碱乙酰胆碱受体（nAChR 和 mAChR）由口腔和食管角质形成细胞（KC）表达。背景：乙酰胆碱调节 KC 的重要功能，包括增殖、粘附、迁移、分化和凋亡。 α3、α5、α7、α9、β2、和 beta4 nAChR 亚基，m2、m3、m4 和 m5 mAChR 亚型，以及合成酶胆碱乙酰转移酶和降解酶乙酰胆碱酯酶在人口腔 KC 中表达并发挥作用，并且可能 Nic 直接影响的目标 KC。初步观察：伴随 KC 分化过程中不同 ACh 受体的出现使得 ACh 对发育中的粘膜表现出多样化和暂时的调节作用上皮。 ACh 对 Ca2+ 内流的刺激作用，由 nAChR 介导，平衡 mAChR 介导的抑制作用和同时激活两种受体的相互作用在 KC 中产生一种阴阳调节平衡。假设：1) 口服 KC 表达的每种 ACh 受体都独特地调节细胞周期进展，从而使分化决定的变化单个角质形成细胞中的 ACh 受体库可以使角质形成细胞多样化 ACh 的生物学效应。 2) 长期接触尼克会扰乱动态角质形成细胞的烟碱和毒蕈碱途径之间的平衡受 ACh 控制，导致细胞周期异常。具体目标：1) 确定每个 ACh 受体对细胞周期进程的贡献亚型选择性激动剂和拮抗剂、反义寡核苷酸，以及基因敲除小鼠。 2) 确定 Nic 与水提取物的长期影响无烟烟草与环境烟草烟雾对表达/功能的影响角质形成细胞 ACh 受体和 Ca2+ 通路促进 ACh 信号传导 KC。方法学：结合分子生物学（RT-PCR、实时荧光定量PCR） TaqMan PCR、Northern 印迹）、免疫学（免疫组织化学、Western blot）印迹法、ELISA）、药理学和生物学测定将用于定量 ACh 受体介导的细胞周期进展变化（目标 #1）以及 Nic 和烟草制品对结构和功能的影响角质形成细胞 ACh 受体（目标#2）。意义：非神经元 ACh 的作用口腔细胞生物学和病理学方面的知识将得到阐明，这将有助于解释烟草制品对上层社会产生有害影响的一些机制消化道。