Calcium Dependent Signaling Pathways in Epithelial Cells

上皮细胞中钙依赖性信号通路

• 批准号: 6993585
• 负责人: Alice S Prince
• 金额: $ 39.3万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-12-10 至 2008-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6993585
• 关键词: Pseudomonas aeruginosa; Staphylococcus aureus; apical membrane; bacteria infection mechanism; cAMP response element binding protein; calcium flux; calmodulin dependent protein kinase; caveolins; chloride channels; cystic fibrosis; gene expression; genetically modified animals; glycolipids; host organism interaction; inositol phosphates; interleukin 8; laboratory mouse; mucosal immunity; nuclear factor kappa beta; phosphatidylinositol 3 kinase; phospholipase C; protein localization; receptor expression; respiratory epithelium; respiratory infections; tissue /cell culture; toll like receptor; transcription factor

DESCRIPTION (provided by applicant): Airway epithelial cells are active participants in the immune defense of the lung. As part of the mucosal immune system, it seems likely that airway cells should share major signaling pathways with "professional" immune cells. T and B cells generate Ca2+ fluxes in response to antigen as a major mechanism to activate gene expression. Mucosal epithelial cells also respond to bacterial ligands with the activation of this common intracellular messenger. In airway epithelial cells Ca2+ dependent signaling pathways are activated by Gram positive and Gram negative bacteria to stimulate expression of cytokines and particularly the chemokine IL- 8. Immediately upon contact by either S. aureus or P.aeruginosa, epithelial cells release Ca2+ from intracellular stores. These Ca2+ fluxes are sufficient to induce downstream signaling, NF-kappaB activation and IL-8 transcription. We postulate that a spatially organized receptor complex, which includes glycolipids and toll like receptors is mobilized to the apical surface of airway cells in response to specific bacterial ligands. In the experiments proposed, we will identify the components of this lipid raft signaling complex that are important in epithelial activation. The generation of Ca2+ fluxes appears to be a common central theme in the activation of immune signaling. We will establish how epithelial cells generate Ca2+ fluxes, the involvement of PI3K and PLCgamma in response to bacterial ligands, as well as the participation of Src, Ras, Btk, and CaMK's. Bacterial signaling is mediated by toll like receptors and particularly TLR2 in airway cells. We will determine how the activation of TLR2 generates Ca2+ signaling, which kinases are involved, and how they are co-localized at the apical pole of airway cells. This will be accomplished by studying signaling in both TLR2 null and caveolin-1 null mice and in epithelial cell lines derived from these animals. We will delineate the major components of this signaling cascade, identifying which Ca2+-dependent kinases, phosphatases and transcription factors including NFAT and CREB as well as NF-kappaB are involved. Such Ca2+ activated responses are likely to be important in airway inflammation induced by chronic bacterial infection, as in airway diseases such as cystic fibrosis. Exactly how CFTR dysfunction affects Ca2+ signaling will be explored. CFTR mutations may influence basal Ca2+ levels in the cell. Repeated apical exposure to bacteria may cause receptor clustering and increase the efficiency of epithelial signaling. By identifying the components of these pathways it should be possible to target therapy to ameliorate airway inflammation in diseases such as cystic fibrosis.

描述（由申请人提供）：气道上皮细胞是对肺部免疫防御的积极参与者。作为粘膜免疫系统的一部分，气道细胞似乎应该与“专业”免疫细胞共享主要的信号通路。 T和B细胞响应抗原作为激活基因表达的主要机制而产生Ca2+通量。 粘膜上皮细胞还会通过这种常见的细胞内信使的激活对细菌配体反应。在气道上皮细胞中，CA2+依赖性信号传导途径通过革兰氏阳性和革兰氏阴性细菌激活，以刺激细胞因子的表达，尤其是趋化因子IL-8。金黄色葡萄球菌或P.Aeruginosa接触后，立即从细胞内存储接触上皮细胞释放Ca2+。这些CA2+通量足以诱导下游信号传导，NF-kappab激活和IL-8转录。我们假设一个空间组织的受体复合物（包括糖脂和类似受体的收费）被动员到气道细胞的顶部表面，以响应特定的细菌配体。在提出的实验中，我们将确定在上皮激活中很重要的脂质筏信号传导复合物的成分。 Ca2+通量的产生似乎是免疫信号传导激活的常见中心主题。我们将建立上皮细胞如何产生Ca2+通量，PI3K和PLCGAMMA响应细菌配体的参与以及SRC，RAS，BTK和CAMK的参与。细菌信号传导是由TOLL像受体一样，尤其是气道细胞中TLR2介导的。我们将确定TLR2的激活如何生成Ca2+信号传导，激酶涉及哪些激酶，以及它们如何在气道细胞的顶端共定位。这将通过研究TLR2 NULL和Caveolin-1 Null小鼠以及从这些动物衍生的上皮细胞系中的信号传导来实现。我们将描述该信号级联的主要组成部分，以确定涉及哪些Ca2+依赖性激酶，磷酸酶和转录因子以及NF-kappab。这种CA2+活化反应可能在慢性细菌感染引起的气道炎症中很重要，例如在气道疾病（例如囊性纤维化）中。 CFTR功能障碍将如何影响CA2+信号传导。 CFTR突变可能影响细胞中的基础Ca2+水平。反复接触细菌可能会导致受体聚类并提高上皮信号的效率。 通过鉴定这些途径的成分，应该可以靶向治疗以减轻诸如囊性纤维化等疾病的气道炎症。