Leukotrienes and Lung Inflammation

描述(申请人提供)：肥大细胞、巨噬细胞和多形核白细胞(PMN)通过合成和释放生物活性脂类白三烯(LT)B4和LTC4来启动和放大肺部的炎症。从细胞中释放出来后，LTC4被转化为LTD4。这些二十烷基类化合物通过调节白细胞募集、内皮通透性、成纤维细胞增殖和平滑肌收缩，帮助“塑造”肺部炎症反应。LTC4、LTD4和LTB4在哮喘和特发性肺纤维化(IFF)的发病机制中起中心作用。它们对宿主防御感染也是至关重要的。因此，细胞如何调节这些二十烷基类化合物的合成是这些疾病发病机制的核心。LTC4的形成需要核膜上至少四种蛋白质的功能相互作用。它们是胞浆磷脂酶A2(CPLA2)、5-脂氧合酶(5-LO)、5-脂氧合酶激活蛋白(FLAP)和LTC4合成酶；LTA被LTA4水解酶代谢产生LTB4。我们使用生物化学和分子成像相结合的方法来展示5-LO、FLOW和LTC4合成酶在核膜上的新型、激活依赖的大分子复合体中的相互作用，这些复合体包括额外的蛋白质。我们还鉴定了一种10 kDa的蛋白质(Association Protein 10 kDa，AP-10)，它在细胞激活后与复合体形成同时从翻盖上解离。这一建议的假设是，这些复合体是细胞激活和LT合成之间的纽带。这项提议的广泛的长期目标是了解这些复合体的组装、降解和分隔是如何在分子、细胞生物学和体内水平上调节LT的形成的。提出了三个具体目标。在具体目标1中，我们将确定5-LO膜复合体是如何组装和组织的。在特定目标2中，我们将鉴定AP-10蛋白。在特定的目标3中，我们将检验这样的假设，即LTC4和LTB4的合成在胞浆和细胞核之间划分，并且新发现的内膜5-LO复合体调节LTB4的合成。这些目标的完成将使我们能够确定炎症性肺部疾病的新机制，并可能找到治疗方法。

DESCRIPTION (provided by applicant): Mast cells, macrophages, and polymorphonuclear leukocytes (PMN), initiate and amplify inflammation in the lung by synthesizing and releasing the bioactive lipids leukotriene (LT)B4, and LTC4. After its release from cells, LTC4 is converted to LTD4. These eicosanoids help "shape" the pulmonary inflammatory response by regulating leukocyte recruitment, endothelial permeability, fibroblast proliferation, and smooth muscle contraction. LTC4, LTD4 and LTB4 are central to the pathogenesis of asthma and idiopathic pulmonary fibrosis (IFF). They are also critical for host defense to infection. How cells modulate the synthesis of these eicosanoids is therefore central to the pathogenesis of these diseases. The formation of LTC4 requires the functional interaction of at least four proteins on the nuclear envelope. These are cytosolic phospholipase A2 (cPLA2), 5-lipoxygenase (5-LO), the 5-lipoxygenase-activating protein (FLAP), and LTC4 synthase; the metabolism of LTA by LTA4 hydrolase yields LTB4. We have used a combination of biochemistry and molecular imaging to demonstrate the interaction of 5-LO, FLAP, and LTC4 synthase in novel, activation- dependent, macromolecular complexes on the nuclear envelope that include additional proteins. We have also identified a 10 kDa protein (Associated Protein 10 kDa, AP-10) that dissociates from FLAP after cell activation concurrent with complex formation. The hypothesis of this proposal is that these complexes are the link between cell activation and LT synthesis. The broad long-term objective of this proposal is to understand how the assembly, degradation, and compartmentalization of these complexes regulate the formation of LTs on molecular, cell biological, and in vivo levels. Three Specific Aims are proposed. In Specific Aim 1 we will determine how 5-LO membrane complexes are assembled and organized. In Specific Aim 2 we will identify the AP-10 protein. In Specific Aim 3 we will test the hypothesis that the synthesis of LTC4 and LTB4 is compartmentalized between the cytosol and nucleus and that newly identified inner membrane 5-LO complexes regulate the synthesis of LTB4. The completion of these aims will allow the identification of novel mechanisms and, potentially, therapeutic approaches, to inflammatory pulmonary diseases.