Ozone ALTERS Airway Smooth Muscle FUNCTION IN ASTHMA

描述(由申请人提供)：尽管臭氧(03)可导致哮喘加重、乙酰甲胆碱气道高反应性(AHR)和中性粒细胞气道炎症，并伴随着TNFpha和IL-1β水平的增加，但在哮喘和过敏原诱导的AHR中介导臭氧效应的机制仍不清楚。初步数据显示，臭氧和变应原联合使用显著增加了中性粒细胞、基础气道张力、AHR和KCl诱导的气道平滑肌(ASM)力的产生，这在一定程度上表明了ASM兴奋-收缩偶联的直接作用。我们推测，臭氧通过促进中性粒细胞介导的气道炎症和直接增加激动剂诱导的ASM力量的产生来增强过敏原诱导的小鼠AHR。为了验证这些假设，在目标1中，将使用耗尽或增加中性粒细胞和/或细胞因子的策略来确定中性粒细胞、肿瘤坏死因子和IL-1β在介导臭氧增强型过敏性AHR和呼吸道炎症中的作用。肺阻力和PENH将测量AHR，并将通过表征细胞图谱和细胞因子和趋化因子的水平来评估支气管肺泡灌洗中的呼吸道炎症。在目标2中，使用人类ASM细胞和来自过敏原和臭氧处理的野生型和敲除小鼠的小鼠气管环，将确定臭氧对关键的ASM兴奋-收缩偶联通路的影响。这些基本途径包括臭氧对激动剂诱导的钙反应、肌醇磷脂代谢、RhoA/RhoA激酶激活和肌球蛋白轻链磷酸酶抑制的钙敏化过程的影响。每个激动剂诱导的信号事件在调节臭氧和过敏原对ASM细胞功能或对气管环力产生的影响方面的充分性和必要性将被确定。这些研究将确定臭氧增强过敏原诱导的AHR的机制，并为预防哮喘恶化和降低哮喘发病率和死亡率的新治疗目标提供洞察力。

DESCRIPTION (provided by applicant): Although ozone (O3) induces asthma exacerbations, methacholine airway hyperresponsiveness (AHR) and neutrophilic airway inflammation associated with increases in levels of TNFalpha and IL-1beta, the mechanisms that mediate ozone effects in asthma and in allergen-induced AHR remain unknown. Preliminary data shows that ozone and allergen in combination markedly increased airway neutrophils, baseline airway tone, AHR and KCl-induced airway smooth muscle (ASM) force generation, suggesting in part a direct effect on ASM excitation-contraction coupling. We postulate that O3 enhances allergen-induced AHR in mice by promoting neutrophil-mediated airway inflammation and by directly increasing agonist-induced ASM force generation. To test these hypotheses, in Aim 1, the role of neutrophils, TNFalpha and IL-1beta in mediating ozone-enhanced allergic AHR and airway inflammation will be determined using strategies to deplete or augment airway neutrophils and/or cytokines. Lung resistance and Penh will measure AHR and airway inflammation will be assessed in bronchoalveolar lavage by characterizing cell profiles and levels of cytokines and chemokines. In Aim 2, using human ASM cells and murine tracheal rings from allergen-and ozone-treated wild type and knockout mice, the effects of ozone on critical ASM excitation-contraction coupling pathways will be defined. These essential pathways include ozone effects on agonist-induced: calcium responses, phosphoinositide metabolism and calcium sensitization processes of RhoA/RhoA kinase activation and of myosin light chain phosphatase inhibition. The sufficiency and necessity of each agonist-induced signaling event in mediating ozone and allergen effects on ASM cell function or on tracheal ring force generation will be determined. These studies will identify the mechanisms by which ozone enhances allergen-induced AHR and provide insight into new therapeutic targets to prevent asthma exacerbations and decrease asthma morbidity and mortality.