Transcellular Communication in Airway Inflammation and Airway Hyperresponsiveness

描述（由申请人提供）：研究旨在研究上皮细胞（EPI）、巨噬细胞（MO）和气道平滑肌细胞（ASMC）被激活的机制，以传递粒细胞a)粘附/迁移信号，b) LTC4的产生，以及c)正常反应性气道转化为高反应性气道。核心假设是，受刺激的EPI、MO或ASMC分泌的内源性14 kDa组VPLA2 （gVPLA2）作为一种细胞间信使蛋白，在哮喘的人气道中高度表达，并在小鼠EPI、MO和ASMC的免疫致敏过程中被诱导。假设gVPLA2水解嗜酸性粒细胞富含磷脂酰胆碱的外质膜，通过一种独立于cPLA2激活的新机制引起随后的LTC4分泌。在Aim 1中，研究人员提出了一种假设，即[32-整合素介导的嗜酸性粒细胞粘附是由gVPLA2通过一种独立于mapk介导的cPLA2磷酸化的新机制调节的。研究表明，气道驻留细胞分泌的gVPLA2传递信号，诱导嗜酸性粒细胞（32-整合素）粘附。研究人员建议进一步研究gVPLA2在生理条件下诱导粘附的潜在途径。在Aim 2中，研究人员提出研究内源性gVPLA2在细胞间通讯中介导LTC4分泌增强的细胞进入和细胞内作用。进一步的研究建议通过使用针对gVPLA2或TAT-dn-cPLA2的中和单抗来测试gVPLA2的特异性。在Aim 3中，我们提出了研究gVPLA2和cPLA2在急性和慢性哮喘小鼠模型中介导气道炎症和气道高反应性（AHR）的机制。在初步数据的基础上，我们提出研究来验证内源性分泌gVPLA2介导免疫致敏小鼠AHR的假设。我们将评估MCL-3G1（一种抗gVPLA2的单抗）在oa攻击前/后小鼠中阻断乙酰胆碱或gVPLA2攻击的AHR的作用。我们将使用免疫致敏的gvpla2敲除和cpla2敲除小鼠进行进一步研究，以确定这些酶在AHR和炎症细胞体内迁移中的具体作用。来自这些研究的数据应该阐明一种新的内源性跨细胞通讯机制，该机制启动气道炎症和AHR。

DESCRIPTION (provided by applicant): Studies are proposed to examine the mechanisms by which epithelial cells (EPI), macrophages (MO) and airway smooth muscle cells (ASMC) are activated to communicate signals for granulocyte a) adhesion /migration, b) generation of LTC4, and c) conversion of normally reactive airways into hyperresponsive airways. The central hypothesis is that the endogenous 14 kDa group V PLA2 (gVPLA2) secreted from stimulated EPI, MO or ASMC serves as an intercellular messenger protein that is highly expressed in asthmatic human airways and is inducible during immune sensitization in mouse EPI, MO and ASMC. It is hypothesized that gVPLA2 hydrolyzes the phosphatidylcholine-rich outer plasma membrane of eosinophils to cause subsequent LTC4 secretion by a novel mechanism that is independent of cPLA2 activation. In Aim 1, studies are proposed to examine the hypothesis that [32-integrin-mediated eosinophil adhesion is regulated by gVPLA2 by a novel mechanism independent of MAPK-mediated cPLA2 phosphorylation. Studies are proposed to demonstrate that gVPLA2 secreted from airway resident cells transmits the signal for induction of (32-integrin adhesion in eosinophils. Further studies are proposed to elucidate the potential pathway by which adhesion is induced under physiological conditions by gVPLA2. In Aim 2, studies are proposed to examine the cellular entry and intracellular action of endogenous gVPLA2 in mediating augmented secretion of LTC4 during transcellular communication. Further studies are proposed to test the specificity of gVPLA2 by using a neutralizing mAb against gVPLA2 or TAT-dn-cPLA2. In Aim 3, studies are proposed to examine the mechanisms of gVPLA2 and cPLA2 in the mediation of airway inflammation and airway hyperresponsiveness (AHR) in acute and chronic murine models of asthma. Based upon preliminary data, studies are proposed to test the hypothesis that endogenous secretion of gVPLA2 mediates AHR in immune sensitized mice. The effect of MCL-3G1, a mAb against gVPLA2, in blocking AHR to methacholine or gVPLA2 challenge in pre/post OA-challenge mice will be assessed. Further studies using immune-sensitized gVPLA2-knockout and cPLA2-knockout mice will be generated to establish the specific role of these enzymes in AHR and inflammatory cell migration in vivo. Data derived from these studies should elucidate a new endogenous mechanism of transcellular communication that initiates airway inflammation and AHR.