MTMR14基因缺失增强哮喘气道高反应及其机理研究

Asthma is a common respiratory disease, which is mainly characterized by enhanced contraction of airway smooth muscle---airway hyperresponsiveness (AHR). Since the mechanisms of AHR are very complicated, it is very difficult to cure it. Therefore, it is very important to identify its mechanism. With the support of our previous NSFC grant (No: 81170227), we found that MTMR14, a phosphatidylinositol phosphatase, is highly expressed in airway smooth muscle of both human and mice, however, its expression level decreased dramatically in either asthmatic patients or mice. Deletion of MTMR14 significantly enhanced the contraction of tracheal rings and increased intracellular calcium concentration in airway smooth muscle. As we all know, calcium is closely coupled to airway hyperresponsiveness. Therefore, we hypothesize that its deletion might enhance airway smooth muscle contraction and accelerate airway hyperresponsiveness by regulating the intracellular calcium level. In order to test this hypothesis, in this project, we will perform the following two studies by using WT and MTMR14 KO mice: 1. To clarify the relationship of MTMR14 and airway hyperresponsiveness; 2. To investigate the underlying mechanisms from the following several aspects: intracellular calcium level, RyRs channels and other ion channels by using electrophysiological, imaging, and molecular biological technologies, etc. This study is expected to find a new mechanism for airway hyperresponsiveness and a new molecular target for clinical treatment of asthma.

哮喘是一种常见的呼吸系统疾病，其主要特点是气道平滑肌收缩增强，即气道高反应（AHR）。AHR的形成机理复杂多样，导致哮喘的治疗十分困难，因而对其机理的研究尤为重要。在前一NSFC课题（81170227）支持下，我们发现磷酸酶MTMR14在人和小鼠的气管平滑肌中均表达，然而在哮喘患者或小鼠中其表达均显著降低；MTMR14缺失导致气管平滑肌收缩增强及细胞内钙显著升高。钙离子与气道高反应密切相关，因此我们推测MTMR14缺失可能通过调节气管平滑肌细胞内钙以促进气管平滑肌收缩增强进而引发气道高反应。本项目拟以野生型和MTMR14基因敲除小鼠为研究对象开展以下研究：1.阐明MTMR14与哮喘气道高反应之间的关系；2.利用电生理、成像及分子生物学等手段，从钙离子、调节钙的RyRs通道以及其它通道等方面揭示MTMR14缺失增强气道高反应的机理。有望发现气道高反应新机制并为哮喘治疗提供新的靶点。

MTMR14是一种新型磷酸酶，在睾丸、骨骼肌和心肌中高表达，其缺失会导致细胞内钙紊乱，引起众多肌肉疾病，如中央核肌病、骨骼肌衰老、肌无力和肌肉疲劳等。此外，该基因还能调控细胞的增殖、迁移以及动脉粥样硬化的形成。哮喘是一种以气道高反应性和慢性气道炎症为特征的异质性疾病，通常由支气管和微气管的慢性炎症、气道阻塞而引起呼吸紊乱。作为细胞内重要的第二信使，Ca2+在调控哮喘的作用中起到十分重要的作用。在骨骼肌中，我们已经报道MTMR14的缺失可以导致细胞内钙的紊乱从而影响骨骼肌的正常功能，而我们随后的研究发现MTMR14在气管平滑肌中也是高表达的。因而我们推测MTMR14的缺失可能影响气管平滑肌的收缩以及哮喘的形成。. 为了验证该假设，我们首先检测了MTMR14在正常小鼠及哮喘小鼠中的表达情况，我们发现在哮喘小鼠中MTMR14的表达是下调的，这提示MTMR14可能在哮喘形成中发挥一定的作用。接着我们用野生型和MTMR14敲除型小鼠建立哮喘模型，用以研究MTMR14的缺失对哮喘小鼠气道高反应性的影响及其机理。实验结果如下：MTMR14敲除型哮喘小鼠气道炎症细胞浸润、气道壁重塑、气道高反应性及呼吸系统阻力均显著增强；进一步研究发现MTMR14的缺失导致Th2细胞因子IL-4、IL-13表达上调、黏蛋白Muc5ac、TNF、NOX4的表达也显著上调，这与前面的表型发现是一致的。此外我们还发现MTMR14的缺失导致了气管平滑肌中细胞内钙增加、L型钙通道电流异常，这可能与MTMR14的缺失导致了L型钙通道家族中Cav1.2表达下调以及胞内环境紊乱有关。综上所述，我们的结果表明：MTMR14的缺失可通过影响气管平滑肌细胞内钙水平而影响气管平滑肌细胞L型钙通道电流进而导致气道高反应。