Repair of Airway Epithelium Following Chlorine Lung Injury

描述（由申请人提供）：急性高水平暴露于损害呼吸道的化学品可导致危及生命的肺损伤。氯气是一种剧毒的呼吸道刺激物，被认为是一种化学威胁剂，因为它可能在工业事故或恐怖袭击中释放。像许多肺部毒物一样，氯会对呼吸道上皮细胞造成损伤。呼吸道上皮细胞屏障的丧失导致在急性肺损伤中观察到的其他后遗症，包括肺水肿、气道阻塞、炎症和呼吸道感染。有效修复呼吸道上皮和恢复屏障功能是解决氯诱导肺损伤的关键。呼吸道上皮在损伤后通过干细胞的增殖和分化进行修复。参与气道上皮修复的一种类型的细胞是基底细胞。低亲和力神经生长因子受体（NGFR）已被确定为基底细胞以及其他组织中上皮干细胞的标志物，但这种蛋白质在干细胞功能中的作用尚不清楚。NGFR结合多种神经营养因子配体，包括神经生长因子（NGF）、脑源性神经营养因子和神经营养因子3。在所提出的研究中要检验的假设是NGFR控制氯诱导的肺损伤后肺上皮干细胞的增殖、存活和分化。在特定目标1中，将从正常小鼠和缺乏功能性NGFR的突变小鼠的气管中纯化基底细胞。将在存在或不存在NGFR配体的情况下测量培养细胞存活、增殖和分化成其他类型上皮细胞（纤毛细胞和Clara细胞）的能力。在特定目标2中，根据主要研究者实验室制定的方案，将野生型、NGFR敲除和NGF转基因小鼠暴露于氯气。将通过鼻内递送配体或通过使用先前在主要研究者实验室开发的NGF过表达转基因小鼠来测试神经营养因子的作用。将在氯暴露后的不同时间测量气管上皮修复的程度。在具体目标3中，将研究神经营养素对家兔氯暴露后上皮修复的影响。这些研究将使用家兔，因为该种属的气道中基底细胞分布更广，与人类更相似。拟议的实验不仅将研究上皮修复的机制，还将促进治疗剂的开发，这些治疗剂可用于刺激吸入有毒化学物质后肺损伤的消退。 公共卫生相关性：本申请是为了回应PAR-10-019，转化研究中的CounterACT探索/开发项目（R21）而提交的。拟议的实验旨在了解肺在吸入有毒气体后如何自我修复，目的是开发基于刺激肺细胞加速修复的肺损伤治疗方法。这种类型的研究是通过寻求资助机会公告，因为担心美国。S.在恐怖袭击中有意释放的剧毒化学品或在工业事故或自然灾害中无意释放的剧毒化学品可能对平民造成不利影响。

DESCRIPTION (provided by applicant): Acute high-level exposures to chemicals that damage the respiratory tract can cause life-threatening lung injury. Chlorine gas is a highly toxic respiratory irritant that is considered a chemical threat agent because of the possibility that it could be released in industrial accidents or terrorist attacks. Chlorine, like many pulmonary toxicants, produces injury to epithelial cells lining the respiratory tract. Loss of the respiratory epithelial cell barrier contributes to other sequelae observed in acute lung injury, including pulmonary edema, airway obstruction, inflammation, and respiratory infections. Efficient repair of respiratory epithelium and restoration of barrier function are essential for resolution of chlorine-induced lung injury. The epithelium of the respiratory tract is repaired following injury through the proliferation and differentiation of stem cells. One type of cell involved in the repair of airway epithelium is the basal cell. The low-affinity nerve growth factor receptor (NGFR) has been identified as a marker for basal cells, as well as for epithelial stem cells in other tissues, but the role of this protein in stem cell function is unknown. NGFR binds multiple neurotrophin ligands, including nerve growth factor (NGF), brain-derived neurotrophic factor, and neurotrophin 3. The hypothesis to be tested in the proposed studies is that NGFR controls the proliferation, survival, and differentiation of lung epithelial stem cells following chlorine-induced lung injury. In Specific Aim 1, basal cells will be purified from the trachea of normal mice and from mutant mice lacking functional NGFR. The ability of the cultured cells to survive, proliferate, and differentiate into other types of epithelial cells (ciliated cells and Clara cells) will be measured in the presence or absence of NGFR ligands. In Specific Aim 2, wild-type, NGFR knockout, and NGF transgenic mice will be exposed to chlorine gas according to a protocol established in the Principal Investigator's laboratory. Effects of neurotrophins will be tested by delivering ligands intranasally or by using NGF-overexpressing transgenic mice previously developed in the Principal Investigator's laboratory. The extent of tracheal epithelial repair will be measured at various times after chlorine exposure. In Specific Aim 3, the effect of neurotrophins on epithelial repair following chlorine exposure in rabbits will be investigated. Rabbits will be used for these studies because of the wider distribution of basal cells in the airways in this species which is more similar to that in humans. The proposed experiments will not only investigate mechanisms of epithelial repair, but will also foster the development of therapeutic agents that could be used to stimulate resolution of lung injury following inhalation of toxic chemicals. PUBLIC HEALTH RELEVANCE: This application is submitted in response to PAR-10-019, CounterACT Exploratory/Developmental Projects in Translational Research (R21). The proposed experiments are designed to understand how the lung repairs itself after inhalation of a toxic gas with the goal of developing therapy for lung injury based on stimulating lungs cells to accelerate repair. This type of research is sought through the Funding Opportunity Announcement because of concerns that U. S. civilians could be adversely affected by highly toxic chemicals released intentionally in terrorist attacks or unintentionally in industrial accidents or natural disasters.