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Induction of airway smooth muscle proliferation and airway hyperreactivity after exposure to airborne particles

暴露于空气颗粒后诱导气道平滑肌增殖和气道高反应性

基本信息

批准号：
18590866
负责人：
TAMAOKI Jun
金额：
$ 2.51万
依托单位：
Tokyo Women's Medical University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18590866/
关键词：
Airway smooth muscle Remodeling Cell proliferation Epidermal growth factor Asthma Air pollution Ultrafine particle Protein synthesis 蛋白合成

项目摘要

In this in vitro and in vivo experiments, exposure to ambient ultrafine particles induced airway inflammatory reactions and tissue remodelling, as well as airway hyperresponsiveness. To determine whether ultrafine carbon black (ufCB) affects proliferation of airway smooth muscle cells and, if so, what the mechanism of action is, we studied human primary bronchial smooth muscle cell cultures. Incubation of cells in the serum-free medium with ufCB increased incorporations of [3H]-thymidine and [3H]-leucine into cells in a time- and dose dependent manner. This effect was attenuated by a free radical scavenger and a NADH/NADPH oxidase inhibitor, and completely inhibited by pretreatment with the epidermal growth factor receptor (EGF-R) tyrosine kinase inhibitors AG1478 and BIBX1382, and the mitogen-activated protein kinase (MEK) inhibitor PD98059. Transfection of a dominant negative mutant of H-Ras likewise abolished the effect ufCB. Stimulation with ufCB also induced processing of membrane anchored pro-heparin-binding (HB) -EGF, release of soluble HB-EGF into the medium, association of phosphorylated EGF-R and Shc with glutathione-S-transferase-Grb2 fusion protein, and phosphorylation of extracellular signal-regulated kinase (ERK) . Pretreatment with each AG1478, [Glu^52]Diphtheria toxin, a specific inhibitor of HB-EGF, and neutralizing HB-EGF antibody inhibited ufCB-induced ERK activation. Furthermore, exposure of mice to ufCB induced an increased responsiveness to inhaled methacholine, and effect that was likewise inhibited by EGFR inhibitors. Thus, ufCB causes oxidative stress-mediated proliferation of airway smooth muscle and a resultant airway hyperreactivity, presumably through the processing of HB-EGF and EGF-R and ERK cascade activation.

在体外和体内实验中，暴露于环境超细颗粒诱导气道炎症反应和组织重塑，以及气道高反应性。为了确定超细炭黑（ufCB）是否会影响气道平滑肌细胞的增殖，如果是这样，其作用机制是什么，我们研究了人原代支气管平滑肌细胞培养。在含有ufCB的无血清培养基中孵育细胞，以时间和剂量依赖性方式增加了[3 H]-胸苷和[3 H]-亮氨酸向细胞中的转化。这种作用被自由基清除剂和NADH/NADPH氧化酶抑制剂减弱，并被表皮生长因子受体（EGF-R）酪氨酸激酶抑制剂AG 1478和BIBX 1382以及促分裂原活化蛋白激酶（MEK）抑制剂PD 98059预处理完全抑制。H-Ras的显性负突变体的转染同样消除了ufCB的作用。用ufCB刺激还诱导膜锚定的前肝素结合（HB）-EGF的加工，可溶性HB-EGF释放到培养基中，磷酸化EGF-R和Shc与谷胱甘肽-S-转移酶-Grb 2融合蛋白的缔合，以及细胞外信号调节激酶（ERK）的磷酸化。用AG 1478、[Glu^52]白喉毒素（HB-EGF的特异性抑制剂）和中和HB-EGF抗体预处理可抑制ufCB诱导的ERK激活。此外，小鼠暴露于ufCB诱导对吸入乙酰甲胆碱的反应性增加，并且这种作用同样被EGFR抑制剂抑制。因此，ufCB引起氧化应激介导的气道平滑肌增殖和由此产生的气道高反应性，推测是通过HB-EGF和EGF-R和ERK级联激活的处理。