氯离子在气道上皮应激中的早期信号分子作用研究

申请者观察到人支气管上皮细胞（BEC）等多种细胞在不同应激刺激下出现胞内氯离子浓度增加及其动力学方式的差异，推测氯离子可能作为气道上皮应激的早期信号分子。氯离子流动的通道门控机制差异导致氯离子浓度动力学方式差异，进而激活不同的氯离子敏感蛋白及其信号通路，产生不同的应激反应定式，包括增殖修复、炎症和免疫激活定式。为确定氯离子动力学方式-氯离子信号通路-细胞应激功能定式的对应关系，拟研究：①应激刺激下BEC内氯离子浓度增加的动力学方式，以及相对应的通道机制；②胞内氯离子浓度增加伴随的细胞功能改变，包括细胞增殖和修复速率、抗原摄取和提呈、炎性细胞粘附激活等，建立氯离子动力学方式与免疫或炎症定式的对应关系；③筛选并验证具有氯离子结合域且参与上述细胞功能的蛋白质。该研究将确定氯离子作为新的气道上皮应激早期信号分子，阐明气道上皮应激反应的早期启动机制，为解释气道上皮应激后的不同反应定式及转归提供依据。

In the previous study, we observed a decrease of chloride ions flux and Cl-1 accumulation in various kind of cells including bronchial epithelial cells (BEC), insulted by different stress stimuli including temperature stress, acid-base stress, antigen insulting and oxidant, etc. However, different dynamics of Cl-1 accumulation could be induced by different kind of stimulus. Therefore, a hypothesis can be speculated that chloridion is possibly the early signal molecules during airway epithelium stress. The differences among mechanisms of channel gating for chloride ions flux results in the differences of dynamics of Cl-1 accumulation, which activate those Cl-1 sensed-proteins and its signal pathway, consequently induce different stress responses on airway epithelial cells including proliferation and wound repair, immune response, and inflammatory adhesive activation, etc. To certificate the hypothesis, this study aim at to definite Cl-1 as the early signal molecules during airway epithelium stress, and make sure the relation of the dynamics of Cl-1 accumulation—Cl-1 signal pathway—stress response type of cells. The main research content includes: ①the dynamics of Cl-1 accumulation in BEC stimulated by different stress, foe example, either transient accumulation or persistent accumulation, and different gating mechanism of channels including CFTR and CACC; ②the change of cellular function with Cl-1 accumulation in BEC, observing cells proliferation and speed of wound repair, uptake and presentation of antigen by BEC, adhesion with inflammatory cells such as PMB,EOS and monocytes, secretion of inflammatory cytokines, and definite the relationship between dynamics of Cl-1 accumulation and immune type or inflammation type; ③combine whole wide genome siRNA data bank and protein data bank (PDB), selecting those proteins which both involve in above stress response and has the domain to bind Cl-1 ,and for further verification. This research will discover the early mechanism in airway epithelium stress, and provide reasons to explain the different response type and end results of airway epithelium after being stressed.