Regulation of Proinflammatory Signaling Pathways by CFTR

描述(由申请人提供)：本提案的主要目标是 确定在肺中介导的细胞内信号通路 与囊性纤维化相关的慢性炎症和感染过程 (Cf)。初步数据显示，呼吸道上皮细胞系来源于 Cf气管分泌大量促炎因子IL8。始终如一地，基恩 CFTR治疗或CPX药物治疗可抑制IL8的基线水平 分泌，同时仍允许生理性IL8分泌以响应 细菌的存在。在正常的上皮细胞中，IL8的分泌是由 NFkB途径的激活和药物基因组学研究表明 NFkB途径基因亚群与IL-8的分泌平行 Cftr、cpx和铜绿假单胞菌暴露。因此，我们假设 CFTR抑制CF上皮细胞分泌IL-8的机制 是通过直接作用于促炎症的细胞内信号通路。我们 希望确定和研究这一途径，并提出以下具体建议 目标。目标1：确定cftr和cpx的信号通路 抑制CF上皮细胞分泌IL8。我们将使用基因芯片 以确定其表达被CFTR改变的基因，或因存在 CPX。目标2：确定CFTR或CPX允许的分子机制 Cf上皮细胞对假单胞菌的生理反应 铜绿假单胞菌。我们将确定IL8启动子中的顺式作用元件 介导CFTR、CPX和铜绿假单胞菌的激活或抑制。目标3：达到 确定CFTR中抑制CF上皮细胞分泌IL8的亚区 细胞。我们将准备表达cftr的病毒载体。 删除了不同的子域，并确定CFTR域(S) 抑制IL-8分泌所必需的。预期、创新和影响： 我们期望这项研究将产生有价值的新知识。 这将为开发治疗CF的新疗法提供帮助。建议数 假设驱动的研究是创新的，因为生物化学和 假说驱动的药物基因组学以前从未以这种方式被用于 研究单基因疾病，如囊性纤维化。这件事的影响 研究将确定CFTR影响能力的新途径 肺上皮细胞对细菌攻击做出反应。

DESCRIPTION (provided by applicant): The principal goal of this proposal is to identify the intracellular signaling pathways in the lung that mediate the chronic processes of inflammation and infection associated with cystic fibrosis (CF). Preliminary data indicate that airway epithelial cell lines derived from CF trachea secrete massive levels of pro-inflammatory IL8. Consistently, gene therapy with CFTR, or drug therapy with CPX, suppresses baseline levels of IL8 secretion, while still allowing physiological IL8 secretion in response to the presence of bacteria. In normal epithelial cells, IL8 secretion is mediated by activation of the NFkB pathway, and pharmacogenomic studies indicate that a subset of NFkB pathway genes parallel the secretion of IL8 as a function of CFTR, CPX and exposure to P. aeruginosa. We have therefore hypothesized that the mechanism by which CFTR suppresses IL8 secretion from CF epithelial cells is by direct action on a pro-inflammatory intracellular signaling pathway. We wish to identify and study this pathway, and propose the following Specific Aims. Aim #1: To determine the signaling pathways by which CFTR and CPX suppress IL8 secretion from CF epithelial cells. We will use cDNA microarrays to identify genes whose expression is altered by CFTR, or by the presence of CPX. Aim #2: To identify the molecular mechanisms by which CFTR or CPX permit CF epithelial cells to respond physiologically to the presence of Pseudomonas aeruginosa. We will identify cis-acting elements within the IL8 promoter which mediate activation or suppression by CFTR, CPX and P. aeruginosa. Aim #3: To identify subdomains in CFTR that suppress IL8 secretion from CF epithelial cells. We will prepare viral vector constructs expressing CFTR from which different subdomains have been deleted, and identify the CFTR domain(s) required for suppression of IL8 secretion. Expectations, Innovation and Impact: Our expectations are that this research will generate valuable new knowledge that will be useful for the development of novel therapies for CF. The proposed hypothesis-driven research is innovative because biochemistry and hypothesis-driven pharmacogenomics have not previously been used in this way to investigate a single gene disease such as cystic fibrosis. The impact of this research will be to identify novel pathways by which CFTR affects the ability of lung epithelial cells to respond to bacterial assault.