CELLULAR BASIS OF AIRWAY SECRETION CLEARANCE COUPLING

气道分泌间隙耦合的细胞基础

• 批准号: 2029614
• 负责人: Richard John Bookman
• 金额: $ 20.65万
• 依托单位: UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-01-01 至 1999-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2029614
• 关键词: G protein; acetylcholine; calcium flux; cell motility; muscarinic receptor; receptor coupling; respiratory airway clearance; respiratory epithelium; respiratory function; sheep; video microscopy

Tracheal epithelial cells play a vital role in pulmonary host defense by clearing foreign materials from the airways through the process of mucociliary clearance. Despite this critical role, our understanding of the cellular and molecular mechanisms responsible for the regulation of mucociliary clearance is far from complete. The broad, long-term hypothesis of this project is that effective clearance depends on cooperative actions and coordinated responses of ciliated cells and mucus-secreting cells. We refer to this as 'secretion-clearance coupling'. Parasympathetic cholinergic input to the airway, acting via muscarinic receptors, plays an important role in regulating mucociliary clearance and these receptors are found on both ciliated cells and submucosal glands. In this proposal, we seek to understand the mechanisms responsible for the regulation of ciliary beating frequency (CBF) by acetylcholine (ACh). We hypothesize that ACh, acting through m3 muscarinic receptors on ciliated cells, has dual, opposing actions on CBF and, further, that these actions are mediated by changes in cytoplasmic free Ca2+ concentration [Ca2+]i resulting from ACh activation of this G-protein coupled receptor. The net result of these opposing actions serves to give the ciliated cell an extraordinary degree of fine control in regulating CBF responses. In order to test this hypothesis, we will examine, using high resolution digital video microscopy, mechanisms by which ACh can raise and lower [Ca2+]i, measure the tightness of coupling between Ca2+ and CBF, and begin to identify signaling molecules that mediate such coupling. The specific aims are: I) To define the mechanisms responsible for regulating [Ca2+]i in ciliated cells in response to ACh. II) To characterize the kinetics of coupling between [Ca2+]i and CBF. Ill) To characterize the molecular site of Ca2+ action. An increased understanding of the normal mechanisms that regulate ciliary beating will help to define the basis for defective mucociliary clearance in such common diseases as asthma and chronic bronchitis. Present day therapy for mucociliary dysfunction is unsatisfactory. The proposed experiments will help to identify molecular targets that might serve as the basis for novel treatment strategies for such airway diseases.

气管上皮细胞在肺宿主防御中发挥重要作用 通过以下过程清除气道中的异物 粘膜纤毛间隙。尽管发挥着至关重要的作用，但我们的理解 负责调节的细胞和分子机制 粘液纤毛的清除还远未完成。广泛、长期 该项目的假设是有效清除取决于 纤毛细胞的合作行动和协调反应 分泌粘液的细胞。我们将此称为“分泌清除” 耦合'。副交感神经胆碱能输入气道，通过 毒蕈碱受体，在调节粘膜纤毛中起重要作用 清除和这些受体被发现在纤毛细胞和 粘膜下腺。在本提案中，我们试图了解 负责调节纤毛跳动频率的机制 (CBF) 由乙酰胆碱 (ACh) 产生。 我们假设 ACh 通过 m3 毒蕈碱受体作用于 纤毛细胞，对 CBF 有双重相反的作用，而且， 这些作用是由细胞质游离 Ca2+ 的变化介导的 该 G 蛋白的 ACh 激活导致 [Ca2+]i 浓度 偶联受体。这些相反行动的最终结果是 赋予纤毛细胞非凡的精细控制能力 调节 CBF 反应。为了检验这个假设，我们将 使用高分辨率数字视频显微镜检查机制 ACh 可以升高和降低 [Ca2+]i，测量 Ca2+ 和 CBF 之间的耦合，并开始识别信号传导 介导这种偶联的分子。具体目标是： I) 定义负责调节 [Ca2+]i 的机制 纤毛细胞对 ACh 的反应。 II) 表征[Ca2+]i 和CBF 之间的耦合动力学。 III) 表征 Ca2+ 作用的分子位点。 加深对正常调节机制的了解 纤毛跳动将有助于确定有缺陷的粘膜纤毛的基础 清除哮喘和慢性支气管炎等常见疾病。 目前对粘液纤毛功能障碍的治疗并不令人满意。这 拟议的实验将有助于识别可能的分子目标 作为此类气道新治疗策略的基础 疾病。