Redox-based Fas signaling in allergic airway disease

描述(申请人提供)：死亡受体Fas在肺细胞上表达，在那里它可以传递信号，调节不同的结果。我们最近发现，氧化剂过氧化氢(H202)促进小鼠肺上皮细胞中Fas的聚集，并且H202使用Fas向c-Jun-N末端激酶(JNK)发出信号，这是一个关键的信号模块，允许细胞感知和响应应激。Fas信号转导的结果不是严格意义上的细胞凋亡，还包括形态改变、炎症等。在过敏性呼吸道疾病小鼠的呼吸道中，促生存因子NF-kB被长期激活，这可以抵消Fas激活的促凋亡作用。在过敏性呼吸道炎症期间，肺的氧化还原环境改变为促氧化状态，H202存在于过敏性呼吸道疾病小鼠的肺中。因此，这项建议中要检验的假设是，H202介导的过敏性呼吸道疾病小鼠肺上皮中JNK的激活通过Fas的氧化激活促进了气道高反应(AHR)和重塑。在具体目标1中，我们将确定H202是否通过使用过氧化氢酶过表达的小鼠和使用降低H202的药物来促进过敏性呼吸道疾病小鼠的JNK激活、AHR和重塑。我们还将直接管理H202的来源，以确定H202是否足以激活JNK，并诱导AHR，以及这是否需要功能Fas。在特定的目标2中，我们将确定Fas在过敏性肺病小鼠中是否被氧化和寡聚，这种情况是否发生在呼吸道上皮细胞中，以及对H202的需求。在具体目标3中，我们将利用系统缺乏Fas的LPR小鼠，或通过建立表达CC10-dN-Fas的小鼠，来确定过敏性呼吸道疾病小鼠呼吸道上皮细胞Fas在JNK活化、AHR和气道重塑中的需求。最后，在特定的目标4中，我们将通过四环素诱导的方式，通过在传导呼吸道细胞中表达显性阴性版本的JNK1转基因小鼠，来确定呼吸道上皮细胞JNK激活在AHR中的作用。这些方法在确定关键的氧化剂靶点和上皮信号事件方面至关重要，这些信号事件对过敏性呼吸道疾病的病理生理学至关重要。

DESCRIPTION (provided by applicant): The death receptor Fas is expressed on lung cells where it can transduce signals that mediate diverse outcomes. We recently showed that the oxidant, hydrogen peroxide (H202) promotes clustering of Fas in mouse lung epithelial cells, and that H202 uses Fas to signal to c-Jun-N-terminal kinase, (JNK), a critical signaling module that allows cells to sense and respond to stress. The outcome of Fas signaling is not strictly apoptosis, but also including shape changes, inflammation etc. The pro-survival factor, NF-kB is activated in a prolonged fashion in airways of mice with allergic airway disease, which can counteract the pro-apoptotic effect of Fas activation. During allergic airways inflammation, the redox environment of the lung is changed towards a pro-oxidant state, and H202 is present in lungs of mice with allergic airways disease. Thus, the hypothesis to be tested in this proposal is that H202- mediated activation of JNK in lung epithelium of mice with allergic airways disease contributes airways hyper-responsiveness (AHR) and remodeling, through oxidative activation of Fas. In Specific Aim 1, we will determine whether H202 promotes JNK activation, AHR, and remodeling in mice with allergic airways disease via the use of catalase over-expressing mice, and the use of pharmacologic agents that lower H202. We will also directly administer a source of H202, to determine whether H202 is sufficient to activate JNK, and elicit AHR, and whether this requires functional Fas. In Specific Aim 2, we will determine whether Fas is oxidized and oligomerized in mice with allergic lung disease, and whether this occurs in airway epithelium, and the requirement for H202 herein. In Specific Aim 3, we will determine the requirement of airway epithelial Fas in JNK activation, AHR, and remodeling in the airways of mice with allergic airway disease by using LPR mice, which lack Fas systemically, or via the creation a CC 10-DN-Fas expressing mouse. Lastly, in Specific Aim 4 we will determine the contribution of airway epithelial JNK activation in AHR through the generation of a transgenic mouse expressing a dominant negative version of JNK1 in cells of the conducting airways, in a tetracycline inducible manner. These approaches are crucial in defining critical oxidant targets and epithelial signaling events crucial to the patho-physiology of allergic airways disease.