Secretory Phospholipase A2s in Airway Pathophysiology

描述(申请人提供)：呼吸道病理生理学中的分泌型磷脂酶A2。二十烷基类化合物合成失调在哮喘的免疫生物学中起着核心作用。新的证据表明，上皮细胞可以通过控制花生四烯酸(AA)的释放来调节二十烷类化合物的产生，花生四烯酸(AA)被白细胞用来产生促炎性二十烷酸。磷脂酶A2(PLA2)S是一组酶，催化膜磷脂释放AA的限速步骤，启动白三烯(LT)S和前列腺素(PG)S的产生。最近，9个分泌性PLA2(SPLA2)S被发现与著名的Iva胞浆PLA2(CPLA2a)协同作用，释放AA并优先启动促炎症二十烷类化合物的合成，如半胱氨酰白三烯(CysLT)S。我们的总体假设是，上皮细胞分泌的PLA2上调介导了促炎二十烷类物质的产生。哮喘的一种表型是运动诱导的支气管收缩(EIB)，其中二十烷类化合物合成失调起关键作用。具有这种表型的哮喘患者在EIB期间，其呼吸道中CysLTs的基础水平增加，CysLTs和其他二十烷基类化合物如PGD2的释放维持了支气管收缩。在初步研究中，我们发现sPLA2组X(sPLA2-X)在EIB哮喘患者的诱导痰中较正常对照组升高，在运动刺激后的呼吸道中升高，并在呼吸道上皮细胞中特异表达。在哮喘小鼠模型中，上皮细胞是sPLA2-X的主要来源，sPLA2-X的遗传缺陷显著抑制了哮喘的发展、气道炎症、支气管高反应性和重塑。在特定的目标1中，我们将进行一项基线的支气管内活检研究，以确定患有EIB的哮喘患者是否有呼吸道上皮中sPLA2的上调。将比较患有EIB的哮喘患者、无EIB的哮喘患者和正常对照组在AA释放的关键调控点以及CysLTs和其他二十烷类化合物合成途径方面的差异。在特定的目标2中，我们将在这三组中进行一项运动挑战研究，以确定在EIB期间，上皮中sPLA2的激活是否启动了CysLts和其他二十烷类化合物的持续释放。将检查上皮内AA释放的主要控制点、sPLA2释放到呼吸道中以及驻留在呼吸道内的白细胞中。在特定的目标3中，我们将对每组单独分离的原代支气管上皮细胞进行体外研究，并与外周血嗜酸粒细胞共同培养，以检测上皮sPLA2在哮喘上皮细胞释放AA和上皮激活白细胞CysLT合成中的作用。这些研究集中在一种特定疾病表型中二十烷类化合物产生的免疫生物学，旨在提供重要的信息，导致哮喘的新疗法。与公共卫生相关。在发达国家，哮喘是年轻人最常见的慢性病，成人患病率为7.2%，儿童患病率为10.8%。2002年，哮喘导致484,000人住院，190万人到急诊科就诊，1390万人到门诊就诊。全国每年因哮喘造成的经济负担超过127亿美元。目前迫切需要开发新的哮喘治疗方法，因为长期的哮喘预防性治疗对大约三分之一的哮喘患者无效，而且目前的治疗方法都没有改变这种疾病的自然病史。这些数据突显了开发新疗法的必要性，这些疗法将提高哮喘治疗的有效性。在这项应用中，我们将重点放在导致哮喘患者促炎性二十烷类化合物产生增加的机制上。初步数据强烈暗示分泌型磷脂酶A2组X是促炎性二十烷类化合物产生的关键调节因子。我们的目标是确定sPLA2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，以提高哮喘治疗的有效性，改变哮喘的自然病程。

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.