CFTR在高血压脑血管重构过程中平滑肌细胞表型转换的作用及机制研究

Phenotype switching of cerebrovascular smooth muscle cells is one of the key pathological change in hypertensive cerebrovascular remodeling. Our preliminary study have shown that down-regulation of CFTR protein expression in the process of cerebrovascular remodeling in 2 kidney 2 clamp hypertensive rats (2K2C), which is positive correlation with blood pressure change. In basilar artery smooth muscle cells (BASMCs), we found that angiotensin II (Ang II) increase cell proliferation and time-dependently reduced CFTR protein expression, which could be attenuated by AT1 receptor inhibitor. Our preliminary data indicated that CFTR is involved in cerebrovascular remodeling in hypertension and Ang II-stimulated BASMCs phenotype switching. However, it is not reported yet. Based on our preliminary data, we will employ Ang II-stimulated BASMCs phenotype switching cell model and hypertensive animal models (including 2K2C, spontaneously hypertensive rat（SHR） and Ang II infusion CFTR knockout mouse), through Patch clamp, Co-IP and other molecular biotechnology, to investigate the role of CFTR in hypertensive cerebrovascular remodeling and BASMCs phenotype switching. Furthermore, we will investigate the protein interaction between CFTR and AT1 receptor, to find out whether the protein interaction could mediated the downstream signal pathway change. The novelty of this proposal is to identify CFTR chloride channel as a novel molecular mechanism for BASMCs phenotype switching and thus hypertensive cerebrovascular remodeling , and will help in the search of new drug target for hypertensive cerebrovascular remodeling.

脑血管平滑肌细胞表型转换是高血压脑血管重构的关键病变。我们预实验发现高血压脑血管重构过程中CFTR蛋白表达下调，体外实验发现Ang II诱导脑血管平滑肌细胞表型转换时，呈时间依赖降低CFTR蛋白表达，该效应可被AT1拮抗剂所抑制，提示CFTR参与Ang II诱导脑血管平滑肌细胞表型转换和高血压脑血管重构过程，这尚未有研究报道。本项目拟在原有工作基础上，采用Ang II刺激脑血管平滑肌细胞表型转换模型和双肾双夹高血压大鼠、自发性高血压大鼠和CFTR基因敲除小鼠微量灌注Ang II等动物模型上，通过膜片钳、免疫共沉淀以及其他分子生物学技术，明确CFTR在高血压脑血管重构血管平滑肌细胞表型转换过程中的作用，揭示CFTR与AT1受体是否存在相互作用而介导下游信号通路改变。本项目从CFTR氯离子通道这一新的角度，揭示高血压脑血管重构平滑肌细胞表型转换的新靶点，为临床转化提供实验室依据。

脑血管平滑肌细胞表型转换是高血压脑血管重构的关键病变。囊性纤维化跨膜传导因子（CFTR）在高血压血管平滑肌细胞表型转换中的作用尚未清楚。本项目主要研究CFTR对Ang II诱导脑基底动脉血管平滑肌细胞表型转换的作用及其信号传导机制，并且在动物水平研究CFTR在高血压动脉血管的表达与高血压血管重构的关系，明确CFTR对高血压血管平滑肌细胞表型转换和血管重构的作用。我们研究发现Ang II诱导脑血管平滑肌细胞表型转换过程中，呈时间和浓度依赖地减少CFTR蛋白表达；过表达或者沉默CFTR蛋白表达显著抑制或增强Ang II诱导血管平滑肌细胞表型转换作用。进而发现CFTR与AT1受体通过直接相互作用，调控下游MAPK和Rho信号传导通路而参与血管平滑肌细胞表型转换。在不同类型高血压动物模型上我们发现高血压脑血管重构过程中CFTR蛋白表达下调，与血压和血管重构呈显著负相关关系。我们还发现在高血压患者腹主动脉血管平滑肌细胞CFTR表达显著降低。CFTR基因敲除显著增强Ang II诱导血压升高、血管收缩反应和血管重构等效应。本项目研究明确了CFTR在高血压脑血管重构血管平滑肌细胞表型转换过程中的作用，首次发现CFTR与AT1受体结合，调控AT1及其下游信号传导通路，为干预高血压RAS系统诱导高血压脑血管重构平滑肌细胞表型转换提供新的药物干预靶点。