EOSINOPHIL APOPTOSIS IN ASTHMA

嗜酸性粒细胞在发病机制中是重要的效应细胞。 哮喘的症状。细胞因子白介素3(IL-3)、白介素5和粒细胞- 巨噬细胞集落刺激因子(GM-CSF)在 嗜酸性粒细胞的发育和激活。如果没有这些因素的影响 细胞因子、嗜酸性粒细胞迅速发生凋亡。我们已经确认 先前观察到嗜酸性粒细胞招募到呼吸道 节段性抗原挑战(SAC)显示人类存活时间延长 活体，与外周血嗜酸性粒细胞比较。此外，我们 最近确定延长的生存时间似乎是 主要是由于呼吸道分泌GM-CSF所致 体外嗜酸性粒细胞。分子机制的研究 在过去的几年里，参与细胞凋亡的人呈爆炸式增长。Ced- 3/ICE半胱氨酸蛋白酶基因家族是重要的促凋亡基因 而Bcl2和相关蛋白提供了重要的反转录因子。 调节作用和抑制细胞凋亡。我们最近确认了 ICE的四种新亚型(β、伽马、β-β)以及 新的Ced-3/ICE同系物，CPP32，Mch2和Mch3。值得注意的是，我们有 还在BAL嗜酸性粒细胞中鉴定了其中的几种蛋白质 哮喘患者。在这项调查中，我们建议研究 嗜酸性粒细胞程序性死亡中的半胱氨酸蛋白酶。我们会 首次鉴定已知的半胱氨酸蛋白酶基因及其产物 在BAL和外周血中的嗜酸性粒细胞，以及嗜酸性粒细胞可诱导 细胞系AML14。第二，我们将确定 糖皮质激素和嗜酸性粒细胞活性细胞因子对半胱氨酸蛋白酶的影响 基因表达，并系统地研究了 嗜酸性粒细胞活性细胞因子可能拮抗糖皮质激素调节 细胞凋亡。第三，交联型嗜酸性粒细胞表面APO- 1/Fas抗原对半胱氨酸蛋白酶基因表达的影响 以及确定凋亡抑制因子bcl2和p35在细胞周期中的作用 糖皮质激素和Fas介导的细胞凋亡。最后，我们提出了一种 新的凋亡半胱氨酸蛋白酶的鉴定和克隆策略 来自嗜酸性粒细胞和急性髓系白血病细胞系。机制的阐明 在体内，嗜酸性粒细胞的存活是通过什么来调节的 炎症是我们理解哮喘发病机制的关键。 此外，对这些通路的操纵将是 哮喘的治疗性干预。

Eosinophils are known to be important effector cells in the pathogenesis of asthma. The cytokines interleukin-3 (IL-3), IL-5, and granulocyte- macrophage colony-stimulating factor (GM-CSF) are important in eosinophil development and activation. Without the influence of these cytokines, eosinophils rapidly undergo apoptosis. We have confirmed previous observations that eosinophils recruited to the airways of humans by segmental antigen challenge (SAC) show a prolonged survival ex vivo, in comparison with peripheral blood eosinophils. Moreover, we have recently determined that the prolonged survival appears to be primarily due to the autocoid secretion of GM-CSF by the airway eosinophils ex vivo. Investigations into the molecular mechanisms involved in apoptosis have exploded in the last several years. The Ced- 3/ICE cysteine protease gene family are important pro-apoptotic mediators, while Bcl-2 and related proteins provide important counter- regulatory effects and inhibit apoptosis. We have recently identified four novel isoforms of ICE (beta, gamma, delta epsilon) as well as the novel Ced-3/ICE homologs, CPP32, Mch2 and Mch3. Significantly, we have also identified several of these proteins in BAL eosinophils from asthmatics. In this investigation we propose to study the role of cysteine proteases in programmed cell death in eosinophils. We will first characterize the known cysteine protease genes and their products in BAL and peripheral blood eosinophils, and the eosinophil inducible cell line AML14. Second, we will determine the effects of glucocorticoids and the eosinophil-active cytokines on cysteine protease gene expression, and also systematically investigate mechanisms by which the eosinophil-active cytokines might antagonize glucocorticoid mediated apoptosis. Third, the effect of crosslinking eosinophil surface APO- 1/FAS-antigen on cysteine protease gene expression will be examined, as well as determining the role of apoptotic inhibitors BcL-2 and p35 on glucocorticoid- and FAS-mediated apoptosis. Finally, we propose a strategy for identifying and cloning novel apoptotic cysteine proteases from eosinophils and the AML cell line. The elucidation of mechanisms by which eosinophil survival is regulated in vivo at sites on inflammation is critical to our understanding of asthma pathogenesis. Furthermore, manipulation of these pathways will be a key area for therapeutic intervention in asthma.