Secretory Phospholipase A2s in Airway Pathophysiology

描述（由申请人提供）：气道病理生理学中的分泌型磷脂酶A2。类花生酸合成失调在哮喘的免疫生物学中起着重要作用。新出现的证据表明，上皮细胞可以通过控制白细胞用于产生促炎性类花生酸的花生四烯酸（AA）的释放来调节类花生酸的产生。磷脂酶A2（PLA 2）是一组催化AA从膜磷脂释放的限速步骤的酶，其引发白三烯（LT）和洋地黄素（PG）的产生。最近，已经鉴定了9种分泌型PLA 2（sPLA 2），其与充分描述的IVa组细胞溶质PLA 2（cPLA 2a）协同作用以释放AA并优先启动促炎性类二十烷酸如半胱氨酰白三烯（CysLT）的合成。我们的总体假设是，上皮中分泌型PLA 2的上调介导哮喘中促炎性类花生酸的产生增加。其中类花生酸合成失调起关键作用的哮喘表型是运动诱导的支气管收缩（EIB）。具有这种表型的哮喘患者气道中CysLT的基础水平增加，并且CysLT和其他类花生酸如PGD 2的释放在EIB期间维持支气管收缩。在初步研究中，我们发现sPLA 2 X组（sPLA 2-X）在EIB哮喘患者诱导痰中相对于正常对照升高，在运动激发后气道中增加，并且特异性表达于气道上皮。在小鼠哮喘模型中，上皮细胞是sPLA 2-X的主要来源，并且sPLA 2-X的遗传缺陷显著抑制气道炎症、支气管高反应性和重塑的发展。在具体目标1中，我们将进行基线支气管内活检研究，以确定EIB哮喘患者气道上皮中sPLA 2是否上调。在AA释放的关键调节点的差异，以及CysLTs和其他类花生酸的合成途径将与EIB哮喘，哮喘无EIB，正常对照组之间进行比较。在具体目标2中，我们将在这三组中进行运动挑战研究，以确定上皮中sPLA 2的激活是否会在EIB期间启动CysLT和其他类花生酸的持续释放。将检查上皮内、从释放到气道中的sPLA 2和驻留在气道内的白细胞中的AA释放的主要控制点。在具体目标3中，我们将使用从各组单独分离并与外周血嗜酸性粒细胞共培养的原代支气管上皮细胞进行体外研究，以检查上皮sPLA 2对哮喘上皮释放AA和上皮激活白细胞中CysLT合成的贡献。这些研究集中在特定疾病表型中类花生酸产生的免疫生物学上，旨在为哮喘的新疗法提供重要信息。公共卫生相关性。哮喘是发达国家年轻人中最常见的慢性疾病，成人患病率为7.2%，儿童为10.8%。2002年，哮喘导致484 000人住院，190万人急诊，1 390万人门诊。全国每年哮喘的经济负担超过127亿美元。目前迫切需要开发新的哮喘治疗方法，因为哮喘的长期预防性治疗对大约三分之一的哮喘患者无效，并且目前的治疗方法都不能改变疾病的自然史。这些数据强调了开发新疗法的必要性，这将提高哮喘治疗的有效性。在本申请中，我们专注于导致哮喘中促炎性类二十烷酸产生增加的机制。初步数据强烈暗示分泌型磷脂酶A2组X作为促炎性类花生酸产生的关键调节剂。我们的目标是确定sPLA 2在哮喘中的功能，以便开发新的治疗方法并在临床试验中进行测试，从而提高哮喘治疗的有效性并改变哮喘的自然史。

DESCRIPTION (provided by applicant): Secretory Phospholipase A2s in Airway Pathophysiology. Dysregulated eicosanoid synthesis plays a central role in the immunobiology of asthma. Emerging evidence indicates that the epithelium can regulate eicosanoid production by controlling the release of arachidonic acid (AA) used by leukocytes to generate pro-inflammatory eicosanoids. The phospholipase A2s (PLA2)s are a group of enzymes that catalyze the rate-limiting step of AA release from membrane phospholipids initiating the production of leukotrienes (LT)s and prostaglandins (PG)s. Recently, 9 secretory PLA2s (sPLA2)s have been identified that act in concert with the well-described group IVa cytosolic PLA2 (cPLA2a) to release AA and preferentially initiate synthesis of pro-inflammatory eicosanoids such as cysteinyl leukotrienes (CysLT)s. Our overall hypothesis is that upregulation of secretory PLA2s in the epithelium mediates increased production of pro-inflammatory eicosanoids in asthma. A phenotype of asthma where dysregulated eicosanoid synthesis plays a critical role is in exercise-induced bronchoconstriction (EIB). Asthmatics with this phenotype have increased basal levels of CysLTs in their airways, and the release of CysLTs and other eicosanoids such as PGD2 sustain bronchoconstriction during EIB. In preliminary studies, we found that sPLA2 group X (sPLA2-X) is elevated in induced sputum of asthmatics with EIB relative to normal controls, increases in the airways following exercise challenge, and is specifically expressed in airway epithelium. In the murine model of asthma, the epithelium is a major source of sPLA2-X, and genetic deficiency of sPLA2-X markedly inhibits the development airway inflammation, bronchial hyperresponsiveness and remodeling. In Specific Aim 1, we will conduct a baseline endobronchial biopsy study to determine if asthmatics with EIB have upregulation of sPLA2s in the airway epithelium. Differences in the key regulatory points for AA release, and the synthetic pathways for CysLTs and other eicosanoids will be compared between asthmatics with EIB, asthmatics without EIB, and normal controls. In Specific Aim 2 we will conduct an exercise challenge study in these three groups to determine if activation of sPLA2s in the epithelium initiates the sustained release of CysLTs and other eicosanoids during EIB. The major control points for AA release within the epithelium, from sPLA2s released into the airways, and in leukocytes residing within the airways will be examined. In Specific Aim 3, we will conduct in vitro studies with primary bronchial epithelial cells isolated from each of the groups alone and co-cultured with peripheral blood eosinophils to examine the contribution of epithelial sPLA2s to the release of AA from the asthmatic epithelium and the epithelial activation of CysLT synthesis in leukocytes. These studies focusing on the immunobiology of eicosanoid production in a specific disease phenotype are designed to provide important information leading to new therapies for asthma. PUBLIC HEALTH RELEVANCE. Asthma is the most prevalent chronic disease of young adults in the developed world with a prevalence of 7.2% in adults, and 10.8% in children. Asthma led to 484,000 hospitalizations, 1.9 million emergency department visits, and 13.9 million outpatient visits in 2002. The national annual economic burden of asthma is over 12.7 billion dollars. There is a pressing need to develop new therapies for asthma because long-term preventative treatments for asthma are ineffective in about a third of patients with asthma, and none of the current therapies alter the natural history of the disease. These data highlight the need to develop new therapies that will improve the effectiveness of treatments for asthma. In this application we focus on the mechanism leading to increased production of pro-inflammatory eicosanoid in asthma. Preliminary data strongly implicate secretory phospholipase A2 group X as a key regulator of pro-inflammatory eicosanoid production. Our goal is to determine the function of the sPLA2s in asthma so that novel therapies can be developed and tested in clinical trials that will improve the effectiveness of asthma treatments and alter the natural history of asthma.