TRPV4通道介导的气道上皮功能失调在屋尘螨诱导哮喘中的作用研究

House dust mites （HDM）has been well-recognized as the leading allergen for asthma. Recent findings demonstrated HDM stimulation resulted in elevation of intracellular calcium（[Ca2+]i）, disruption of epithelial barrier function as well as exacerbation of airway hyper-responsiveness (AHR). We have previously shown TRPs, serving as non-selective cation channels, were critically involved in smooth muscle cell migration, proliferation, remodeling and contraction via regulating intracellular Ca2+ hemostasis. Here, we aim to evaluate the role of TRPV4 mediated epithelial dysfunction in HDM induced asthma by addressing these important issues: a, the effect of HDM treatment to the expression and function of epithelial TRPV4 channel, confirming the significance of TRPV4-derived Ca2+ influx in HDM-elevated epithelial [Ca2+]i; b, the contribution of TRPV4 to HDM caused alterations of epithelial electric resistance, apical junction complex and airway inflammation in vitro; c, the in vivo involvement of TRPV4 in HDM triggered AHR, airway epithelial barrier dysfunction, airway inflammation as well as airway remodeling. Hence, this project would identify the pivotal role of TRPV4 in HDM induced asthma, which might shed new insights for the prevention and treatment of asthma.

屋尘螨（HDM）是哮喘发病最常见的过敏原，可以增高气道上皮内钙离子浓度（[Ca2+]i）、破坏上皮屏障功能，加重气道高反应性。我们的前期研究证实非选择性阳离子通道TRP通过调节[Ca2+]i，参与调控平滑肌细胞迁移、增殖、重塑及收缩。本课题进一步探讨TRPV4 通道介导的气道上皮功能失调在HDM哮喘中的作用，研究拟从如下几方面开展：（1）探索HDM对气道上皮TRPV4表达的影响及通道功能的改变，验证TRPV4介导的钙内流是HDM致上皮[Ca2+]i升高的主要途径；（2）明确TRPV4对HDM所致上皮膜电阻和连接蛋白改变、炎症因子分泌的调控作用；（3）构建HDM哮喘模型，在整体水平确认TRPV4在HDM哮喘上皮通透性改变、气道炎症、气道高反应性和气道重塑中的地位。本项目将阐明气道上皮TRPV4通道在HDM所致哮喘中扮演的关键角色，为有效防治哮喘提供新思路，为开发哮喘治疗新药提供有力依据。

支气管哮喘是最常见的慢性气道疾病之一，屋尘螨（HDM）是最常见的气道过敏原，但HDM所致哮喘的具体机制尚不明确。瞬时受体电位（TRP）是可通透钙离子的非选择性阳离子通道超家族，可以被多种理化因素所激活。前期的研究显示TRPV4可能与哮喘密切相关，参与介导气道上皮钙内流，在吸入性化学损伤所致气道炎症及气道上皮通透性改变中发挥核心作用。本研究以人原代气道上皮细胞和气道上皮细胞系为研究对象，采用钙图、上皮膜电阻测定、ELASA、siRNA干扰等技术，从生理学、分子生物学及免疫性角度综合评价TRPV4通道在哮喘上皮细胞一系列病理生理学改变中的作用。研究发现，哮喘患者气道上皮TRPV4通道表达在转录水平和蛋白水平均显著高于非哮喘患者。HDM刺激正常气道上皮细胞可以上调TRPV4的表达。功能学研究发现：HDM上调气道上皮细胞胞内钙浓度，钙离子内流是胞内钙增加的来源，TRPV4参与介导这一过程；TRPV4通道在HDM刺激下活性增加；哮喘气道上皮TRPV4活性高于正常气道上皮；特异性阻滞剂HC-067047或者干扰TRPV4可以降低上皮膜电阻。HDM可以上调TRPV4内源性激动剂EET的分泌，阻断EET途径可以抑制HDM诱导的TRPV4表达上调。综上所述，TRPV4通过调节上皮细胞内钙浓度而调控上皮通透性参与哮喘发病。