The role of epidermal growth factor receptor (EGFR) in mucin MUC5AC gene expression in airways

我们在体内和体外研究了EGFR在哮喘发病机制中的作用。用卵清蛋白(OVA)致敏(0d、7d)和激发(14d、15d、16d)。分别于第14天(OVA激发前)、第17、24、31天和第38天进行形态学分析(Alcian Blue/PAS染色、MUC5AC和EGFR免疫组织化学染色)、气道高反应性测定(Penh)和细胞分析(BAL)。第17天，OVA致敏和激发大鼠呼吸道上皮细胞的MUC5AC和EGFR免疫反应均显著增强。在没有额外的OVA刺激的情况下，在第38天观察到炎症细胞浸润和杯状细胞增生的自发消退与EGFR表达的减少。相比之下，AHR持续了三周。气管内滴注选择性EGFR酪氨酸激酶抑制剂AG1478可显著降低所有已建立的哮喘表型，并伴随着呼吸道上皮细胞的凋亡。在体外实验中，转化生长因子α与NCI-H292细胞孵育24小时后，Bc l-2、MUC5AC和粘液糖偶联物的产生增加，这一作用可被AG1478的预处理所抑制。AG1478也可诱导粘液产生细胞的凋亡，但MEK抑制剂PD98059对其无影响。半胱氨酸氨基转移酶抑制剂Z-VAD-fmk可抑制AG1478诱导的细胞凋亡。根据这些结果，我们认为EGFR的表达和激活可能在哮喘表型的发展中起关键作用，EGFR的酪氨酸激酶抑制剂可能是一种有用的治疗候选药物，可用于哮喘患者气道重塑的恢复。

We determined the role of EGFR in pathogenesis of asthma both in vivo and in vitro. Brown Norway rats were sensitized (day 0, day 7) and challenged (days 14, 15 and 16) with ovalbumin (OVA). Morphological analysis (Alcian blue/PAS staining, immunohistochemistry for MUC5AC and EGFR), measurement of airway hyperreactivity (penH) and cell analysis (BAL) were performed on days 14 (before OVA challenge), 17, 24, 31 and day 38. On day 17, OVA-sensitized and challenged rats showed a marked increase in both MUC5AC and EGFR immunoreactivity in airway epithelium. Without additional OVA challenge, spontaneous resolution of both inflammatory cell infiltration and goblet cell hyperplasia were observed simultaneously with the reduction of EGFR expression on day 38. In contrast, AHR persisted for three weeks. Intratracheal instillation of the selective EGFR tyrosine kinase inhibitor AG1478 induced a significant reduction in all established asthma phenotypes, which was accompanied by apoptosis of airway epithelial cells. In in vitro studies, incubation of NCI-H292 cells with TGFα for 24h increased Bcl-2, MUC5AC and mucous glycoconjugates production, effects that were inhibited by pretreatment of AG1478. AG1478 also induced apoptosis in mucous producing cells, but a MEK inhibitor PD98059 had no effect on apoptosis. Z-VAD-fmk, a caspase inhibitor reduced AG1478-induced apoptosis. From these results, we concluded that the expression and activation of EGFR may play a crucial role in the development of asthma phenotypes and that a tyrosine kinase inhibitor of EGFR may be a useful therapeutic candidate for the restoration of airway remodeling in patients with asthma.