Role of macrophages in organophosphorus pesticide-induced airway hyperreactivity

描述（由申请人提供）：我们已经证明，在豚鼠模型中，有机磷农药（OPs）引起气道高反应性，这是剂量相关的，并与自身抑制毒蕈碱M2受体的功能丧失有关，该受体通常限制支配气道平滑肌的副交感神经释放乙酰胆碱。我们最近报道了抗原致敏改变了致敏豚鼠而非非致敏豚鼠中op诱导的气道高反应性的机制，涉及il -5依赖机制。在非致敏动物中，OPs如何引起神经元M2功能障碍尚不清楚，但我们的初步数据表明，这种作用不是通过胆碱酯酶抑制或与神经元M2受体的直接拮抗相互作用介导的。相反，OPs似乎通过对巨噬细胞的影响间接影响神经元M2受体功能，因为使用脂质体包裹的氯膦酸盐消耗巨噬细胞可以防止op诱导的气道高反应性。我们的假设是，OPs激活巨噬细胞上调炎症细胞因子的表达和释放，这些炎症细胞因子在各种气道高反应性模型中被证明会导致M2受体功能障碍。我们提出了四个目标来检验这一假设。在Aim 1中，我们将使用体内生理测量来证实巨噬细胞介导OPs引起的气道高反应性，并确定它们的作用是否随着时间的推移而改变，正如在臭氧诱导的气道高反应性中观察到的嗜酸性粒细胞一样。目的2将利用从经op处理的豚鼠和未经op处理的豚鼠的支气管肺泡灌洗液中分离的巨噬细胞来检测OPs对巨噬细胞表达和气道高反应性炎症细胞因子释放的影响。在Aim 3中，我们将使用原代神经细胞培养来确定OP诱导的巨噬细胞介质是否直接与神经相互作用以改变M2受体的表达或功能或神经的结构可塑性。目的4将证实目的2和3中鉴定的op诱导巨噬细胞介质的体内生理相关性。机制研究对于制定OP诱导的气道高反应性的预防和治疗方法至关重要，敏感性个体（过敏）和非敏感性个体之间可能存在差异，并且对于确定OP暴露对人类健康构成的风险至关重要。鉴于哮喘患病率日益增加、人类特别是儿童广泛接触有机磷农药以及恐怖分子使用有机磷农药和神经毒剂的可信威胁，这些研究对公共卫生的影响具有重要意义。

DESCRIPTION (provided by applicant): We have demonstrated that in a guinea pig model, organophosphorus pesticides (OPs) cause airway hyperreactivity that is dose-related and associated with the functional loss of autoinhibitory muscarinic M2 receptors that normally limit acetylcholine release from parasympathetic nerves that innervate airway smooth muscle. We recently reported that sensitization to antigen alters the mechanisms underlying OP-induced airway hyperreactivity to involve IL-5-dependent mechanisms in the sensitized but not the non-sensitized guinea pig. How OPs cause neuronal M2 dysfunction in the non-sensitized animal is not known but our preliminary data indicate that this effect is not mediated by cholinesterase inhibition or direct antagonistic interactions with neuronal M2 receptors. Rather, OPs appear to influence neuronal M2 receptor function indirectly via effects on macrophages since depletion of macrophages using liposome-encapsulated clodronate protects against OP-induced airway hyperreactivity. It is our hypothesis that OPs activate macrophages to upregulate expression and release of inflammatory cytokines previously shown to cause M2 receptor dysfunction in various models of airway hyperreactivity. We propose four Aims to test this hypothesis. In Aim 1, we will use in vivo physiological measurements to confirm that macrophages mediate airway hyperreactivity caused by OPs and determine whether their role changes over time, as has been observed for eosinophils in ozone-induced airway hyperreactivity. Aim 2 will utilize macrophages isolated from bronchoalveolar lavage collected from OP-treated versus untreated guinea pigs guinea pigs to examine the effect of OPs on macrophage expression and release of inflammatory cytokines implicated in airway hyperreactivity. In Aim 3, we will use primary nerve cell cultures to determine whether OP- induced macrophage mediators interact with nerves directly to alter M2 receptor expression or function or the structural plasticity of nerves. Aim 4 will confirm the in vivo physiological relevance of OP-induced macrophage mediators identified in aims 2 and 3. Mechanistic studies are critical to developing preventive and therapeutic approaches for OP-induced airway hyperreactivity, which are likely to differ between sensitized (allergic) and non-sensitized individuals, and for determining the risks to human health posed by OP exposures. The public health implications of these studies are significant in light of the increasing prevalence of asthma, the wide spread exposure of humans and especially children to OPs and the credible threat of terrorist use of OP pesticides and nerve agents.