Hyperthermia-augmented epithelial apoptosis and acute lung injury

描述（由申请人提供）： 肺上皮细胞对于正常的气体交换和维持物理和免疫屏障对抗环境危险至关重要。上皮细胞的急性丧失可能产生破坏性后果，包括气体交换、表面活性剂合成、气道阻塞和继发感染易感性增加。调节上皮细胞存活的过程对于肺上皮如何响应急性和慢性环境挑战至关重要，对肺功能和宿主存活具有重要影响。我们已经确定了新的途径，通过这些途径，高温，如发热、临床高温综合征和环境暴露，加速肺上皮细胞凋亡并加重肺损伤。本申请的总体目标是阐明发热范围内的高热改变肺上皮损伤和恢复的机制。我们的中心假设是，与死亡受体（例如TNF受体-1/TNFR1或fas）的活化同时的高温暴露加速了死亡诱导信号复合物的组装，增强了下游促凋亡途径，并抑制了抗凋亡因子。我们的假设的基本原理是基于我们最近的研究表明：（1）在HS（42 C）或发热中同时暴露于高温，（39.5C）范围在TNF处理或Fas活化后大大增加和加速小鼠MLE 15肺上皮细胞中的半胱天冬酶-3活化和凋亡，并且不依赖于NFB和主要的热活化转录因子HSF-1，和（2）在TNF处理后存在有限的时间窗，但Fas活化后不存在，此后细胞被保护免于HS增强的细胞凋亡（在初步数据部分中讨论）。我们将利用MLE-15小鼠肺上皮细胞、BEAS 2B人肺上皮细胞和原代培养的人小气道上皮细胞（SAECs），以及体内LPS诱导的小鼠ALI模型来实现以下具体目标：（1）确定高温如何改变TNF和Fas激活的死亡诱导信号复合物、它们的组成和功能以及对细胞存活与死亡的影响;（2）描述高温对JNK MAP激酶信号传导的影响以及TNF处理的细胞中细胞存活与死亡的后果;（3）分析高温促进人肺上皮细胞凋亡的机制;以及（4）分析加速上皮细胞凋亡对LPS诱导的肺损伤的贡献。我们期望这些研究的结果将确定在临床相关条件下导致肺损伤的新的病理生理学途径。目标4中的药理学研究将提供新的临床前数据，可能导致未来的ALI患者治疗。

DESCRIPTION (provided by applicant): Lung epithelium is critical to normal gas exchange and maintenance of a physical and immu- nologic barrier against environmental dangers. Acute loss of epithelium can have devastating consequences, including loss of gas exchange, surfactant synthesis, airway obstruction, and increased susceptibility to secondary infections. The processes that regulate epithelial cell sur- vival are critical to how lung epithelium responds to acute and chronic environmental challenges with important consequences for lung function and host survival. We have identified new path- ways by which hyperthermia, such as occurs in fever, clinical hyperthermia syndromes and en- vironmental exposures, accelerate apoptosis in lung epithelium and worsen lung injury. The overall objective of this application is to elucidate the mechanisms by which hyperthermia within the febrile range modifies lung epithelial injury and recovery. Our central hypothesis is that ex- posure to hyperthermia concurrent with activation of death receptors (e.g. TNF Receptor- 1/TNFR1 or fas) accelerates assembly of the death inducing signaling complexes, augments downstream pro-apoptotic pathways, and inhibits anti-apoptotic factors. The rationale for our hypothesis is based on our recent studies demonstrating that (1) co-exposure to hyperthermia in the HS (42C) or febrile (39.5C) range greatly increases and accelerates caspase-3 activation and apoptosis in mouse MLE15 lung epithelial cells after TNF treatment or fas activation and is independent of NFB and the major heat activated transcription factor, HSF-1, and (2) there is a limited time window following TNF treatment but not fas activation after which cells are pro- tected from HS-augmented apoptosis (discussed in the Preliminary Data section). We will utilize MLE-15 mouse lung epithelial cells, BEAS2B human lung epithelial cells, and primary cultured human small airway epithelial cells (SAECs), and an in vivo intratracheal LPS-induced mouse ALI model to pursue the following specific aims: (1) determine how hyperthermia alters TNF- and fas-activated death-inducing signaling complexes, their composition, and function and the consequences for cell survival vs. death; (2) delineate the effect of hyperthermia on JNK MAP kinase signaling and the consequences for cells survival vs. death in cells TNF-treated cells; (3) analyze the mechanisms of hyperthermia-enhanced apoptosis in human lung epithelial cells, and (4) analyze the contribution of accelerated epithelial apoptosis to LPS-induced lung injury. We expect that the results of the proposed studies will identify new pathophysiologic pathways that contribute to lung injury under clinically relevant conditions. The pharmacologic studies in aim 4 will provide new preclinical data that may lead to future therapies in patients with ALI.