Role of MEK1 in T cell function in asthma

描述(申请人提供)：哮喘是一个主要的公共卫生问题。T细胞介导的呼吸道炎症在慢性哮喘中起重要作用。虽然人们对炎症的发生机制知之甚多，但对炎症持续的机制却知之甚少。急性炎症通常由于负的体内平衡调节而消退。在初步研究中，我们观察到哮喘患者T细胞增殖增加。这种增殖对无能诱导和负性调节因子--转化生长因子--β和白介素10的作用不起作用。在探索这种难治性的机制时，我们观察到哮喘患者的T细胞表达增加的信号激酶MEK1，它刺激IL-2的产生。有趣的是，IL-2直接诱导MEK1。我们的假设是，IL-2驱动的MEK1的表达建立了一种正反馈机制，驱动T细胞的高增殖，并推翻了哮喘的负调控。我们还假设，MEK1除了激活胞浆中的ERK1/2外，还抑制细胞核中的基因抑制物SMRT，后者启动T细胞中的炎症基因转录。我们提出了三个具体目标。在特定目标1下，我们将研究哮喘T细胞MEK1表达增加的机制。我们将研究各种细胞因子和共刺激分子的作用。我们还将研究导致MEK1表达的信号传递过程。我们将对人类血液T细胞以及正常和转基因小鼠T细胞进行研究。在特定目的2下，我们将研究核MEK1在T细胞功能中的作用，检测其对基因抑制子SMRT的抑制作用，剖析核MEK1和ERK1/2在c-Fos基因启动子调控SMRT和ETS转录因子中的独立作用，最后探讨SMRT在哮喘患者T细胞功能中的作用。这些研究将涉及基因操作、染色质免疫沉淀和T细胞功能研究。在特定目标3下，我们将研究MEK1和IL-2在哮喘小鼠模型中对呼吸道炎症持续存在的贡献。我们专门开发了一种慢性哮喘的小鼠模型，它在许多方面模仿人类哮喘。我们将检测转基因小鼠品系和/或T细胞，以及抗IL2受体(抗CD25)抗体在这一慢性哮喘模型中对气道高反应性、炎症、粘液产生和气道重塑的影响。这项建议很重要，因为它解决了哮喘患者呼吸道炎症持续存在的机制。这项研究的结果将导致开发新的哮喘治疗方法。

DESCRIPTION (provided by applicant): Asthma is a major public health problem. T cell-mediated airway inflammation plays a major role in chronic asthma. Although much is known about the mechanism of initiation of inflammation, less is known about the mechanism of its persistence. Acute inflammation usually resolves due to negative homeostatic regulation. In preliminary studies we observed increased T cell proliferation in asthma. This proliferation is refractory to anergy induction and to the action of the negative regulators-TGF-beta and IL-10. In exploring the mechanism of this refractoriness we observed that T cells from asthmatic patients express increased levels of the signaling kinase MEK1, which stimulates IL-2 production. Interestingly, IL-2 directly induces MEK1. Our hypothesis is that the IL-2 driven MEK1 expression establishes a positive feedback mechanism that drives heightened T cell proliferation and overrides the negative regulation in asthma. We also hypothesize that MEK1, in addition to activating ERK1/2 in the cytosol, inhibits the gene repressor SMRT in the nucleus, which primes inflammatory gene transcription in T cells. We propose 3 specific aims. Under specific aim 1 we will study the mechanism of increased MEK1 expression in T cells from asthma. We will examine the contribution of various cytokines and co-stimulatory molecules. We will also examine the signaling processes that lead to MEK1 expression. We will perform studies with human blood T cells as well as normal and genetically modified mouse T cells. Under specific aim 2 we will study the role of nuclear MEK1 in T cell function, examine its inhibitory effect on the gene repressor SMRT, dissect the independent role of nuclear MEK1 and ERK1/2 in regulating SMRT and Ets transcription factors at the c-Fos gene promoter, and finally, investigate the role of SMRT in T cell function from asthmatic patients. These studies will involve gene manipulations, chromatin immunoprecipitation, and T cell functional studies. Under specific aim 3 we will examine the contribution of MEK1 and IL-2 to the persistence of airway inflammation in a mouse model of asthma. We have specifically developed a mouse model of chronic asthma, which mimics human asthma in many respects. We will examine genetically modified mouse strains and/or T cells, and an anti-IL2 receptor (anti-CD25) antibody for their effect on airway hyperreactivity, inflammation, mucus production and airway remodeling in this chronic asthma model. The proposal is important because it addresses the mechanism of persistence of airway inflammation in asthma. The results of this study will lead to the development of novel therapeutic modalities for asthma.