Role of macrophages in organophosphorus pesticide-induced airway hyperreactivity

描述（由申请方提供）：我们已经证明，在豚鼠模型中，有机磷农药（OP）引起气道高反应性，这与剂量相关，并与自身抑制性毒蕈碱M2受体的功能丧失相关，该受体通常限制乙酰胆碱从支配气道平滑肌的副交感神经释放。我们最近报道，抗原致敏改变了OP诱导的气道高反应性的机制，涉及致敏豚鼠而非非致敏豚鼠的IL-5依赖性机制。OP如何在非致敏动物中引起神经元M2功能障碍尚不清楚，但我们的初步数据表明，这种作用不是由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，OP似乎通过对巨噬细胞的作用间接影响神经元M2受体功能，因为使用脂质体包封的氯膦酸盐消除巨噬细胞可防止OP诱导的气道高反应性。我们的假设是，OP激活巨噬细胞以上调先前显示在各种气道高反应性模型中引起M2受体功能障碍的炎性细胞因子的表达和释放。我们提出了四个目标来检验这一假设。在目标1中，我们将使用体内生理测量来确认巨噬细胞介导OP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变化，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OP处理与未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OP对巨噬细胞表达和与气道高反应性有关的炎性细胞因子释放的影响。在目的3中，我们将使用原代神经细胞培养物来确定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诱导的巨噬细胞介质的体内生理相关性。机制研究对于开发OP诱导的气道高反应性的预防和治疗方法至关重要，这可能在致敏（过敏）和非致敏个体之间存在差异，并确定OP暴露对人类健康造成的风险。鉴于哮喘日益流行，人类特别是儿童广泛接触有机磷农药，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具有重大意义。

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.