Molecular basis of age-dependent changes in airway smooth muscle functions

项目概要/摘要 临床研究表明，随着年龄的增长，呼吸功能会显着丧失。这导致减少 生活质量、其他疾病的倾向增加以及治疗以下疾病的气雾剂药物输送无效 阻塞性呼吸道疾病。然而，年龄依赖性丧失的细胞和分子基础 呼吸尚未建立。气道平滑肌（ASM）在调节气道 通过影响支气管的机械（收缩和松弛）特性来影响呼吸。 在此，我们建议建立由于衰老而在 ASM 中发生的分子变化，这些变化是导致 ASM 丧失的原因。 老年人的呼吸功能。初步研究表明收缩和松弛减弱 与“年轻”大鼠相比，老年（“老年”）大鼠气道和 ASM 细胞的反应。附加数据表明 衰老促进 ASM 中的“表型可塑性”或“转换”，这是在某些疾病中观察到的现象 平滑肌收缩表型转变为增殖/合成表型的过程。 肌球蛋白重链和平滑肌α-肌动蛋白（收缩表型标记蛋白）的表达较低 与年轻大鼠相比，从老年大鼠获得的 ASM 中。此外，全局基因表达谱分析 ASM 细胞中肌生长抑制素的表达降低，肌生长抑制素是 TGF-b 家族的成员，已知可抑制 肌细胞的生长和增殖。最后，ASM 细胞的初步数据表明细胞内 随着年龄的增长，向收缩剂和松弛剂发出信号，激活 G 蛋白偶联受体 (GPCR) 关于 ASM。基于这些研究，我们假设 ASM 随着年龄的增长而经历表型调节 导致收缩和舒张反应能力下降； GPCR 信号传导能力的丧失是造成这种情况的原因 反应性丧失，部分是由于随年龄增长而发生的 TGF-b 通路激活丧失所致。在 具体目标 1 我们建议使用大鼠和人类建立 ASM 表型的年龄依赖性变化 通过采用肌动描记器、光磁扭转细胞计数等新型工具来检测气道和 ASM 细胞 牵引显微镜。在具体目标 2 中，我们建议为表型转换建立机制基础 通过详细描述第二信使的表达和活动的变化，随着年龄的增长而发生的 ASM， 调节 ASM 功能的 GPCR 信号传导的效应器和调节器。在具体目标 3 中，我们提出了研究 表征肌生长抑制素和 TGF-b 对气道和 ASM 表型的影响，并确定 TGF-β信号传导逆转与年龄相关的合成/增殖表型的能力。总的来说， 这些研究旨在确定 ASM 功能中与年龄相关的分子变化，这些变化有助于年龄 老年人群呼吸功能下降相关。我们的发现可能有助于开发工具 改善呼吸功能，修改阻塞性肺疾病的诊断和治疗方案 老年人的疾病。

Project Summary/Abstract Clinical studies have demonstrated a significant loss of respiratory function with age. This results in reduced quality of life, increased propensity for other diseases, and ineffective aerosol drug delivery for the treatment of obstructive respiratory diseases. However, the cellular and molecular basis for age-dependent loss of respiration has not been established. Airway smooth muscle (ASM) plays a significant role in the regulation of respiration by influencing the bronchial tone due to its mechanical (contraction and relaxation) properties. Herein we propose to establish molecular changes that occur in the ASM due to aging that account for loss of respiratory function in the elderly. Preliminary studies demonstrate diminished contractile and relaxation responses of airways and ASM cells in aged ("Old") rats compared to "Young" rats. Additional data suggest aging promotes "phenotype plasticity" or "switching" in ASM, a phenomenon observed in certain disease processes in which the smooth muscle contractile phenotype is changed to a proliferative/synthetic phenotype. Expression of myosin heavy chain and smooth muscle a-actin (contractile phenotype marker proteins) is lower in ASM obtained from Old rats compared to Young rats. Furthermore, global gene expression profile analysis in ASM cells revealed decreased expression of myostatin, a member of the TGF-b family, known to inhibit the growth and proliferation of myocytes. Lastly, preliminary data from ASM cells suggest diminished intracellular signaling with age to both contractile and relaxant agents that activate G protein-coupled receptors (GPCRs) on ASM. Based on these studies we hypothesize that ASM undergoes phenotype modulation with age that results in decreased contractile and relaxant responsiveness; a loss of GPCR signaling capacity underlies this loss of responsiveness, mediated in part by a loss of TGF-b pathway activation that occurs with age. In Specific Aim 1 we propose to establish age-dependent changes in ASM phenotype using rat and human airways and ASM cells by employing novel tools such as myograph, optical magnetic twisting cytometry and traction microscopy. In Specific Aim 2, we propose to establish the mechanistic basis for a phenotype switch in the ASM that occurs with age by detailing changes in the expression and activity of second messengers, effectors and regulators of GPCR signaling that regulate ASM function. In Specific Aim 3, we propose studies to characterize the effects of myostatin and TGF- b on airways and ASM phenotype, and determine the capacity of TGF- b signaling to reverse the synthetic/proliferative phenotype associated with age. Collectively, these studies seek to identify age-dependent molecular changes in ASM function that contribute to the age- associated decline in respiratory function in the elderly population. Our findings may help develop tools to improve respiratory functions, and modify diagnostic and treatment regimens for obstructive pulmonary diseases in the elderly.