Objective: To investigate the effects of the epidermal growth factor receptor(EGFR) inhibitor Gefitinib on airway inflammation and airway remodelling in asthmatic C57BL/6 mice, and to analyze its possible mechanisms. Methods: Male C57BL/6 mice, aged 6-8 weeks, were randomly assigned into five groups: Group A (control group), Group B (asthma group), Group C (asthma+20 mg/kg gefitinib group), Group D (asthma+40 mg/kg gefitinib group), and Group E (40 mg/kg gefitinib group), with seven mice per group. Mice were sensitized by intraperitoneal injection of a mixture of 0.2 ml solution containing OVA and Al(OH)3 [20 μg OVA+2 mg Al(OH)3 dissolved in 0.2 ml of physiological saline] at Day 0 and 14. Starting from Day 25 to 31, Group B, C, and D were challenged with nebulization of 1% OVA solution (8 ml) to induce asthma, once a day for approximately 40 minutes, with continuous aerosolization for 7 days. Group C and D were given 0.2 ml of Gefitinib dissolved in 0.5% carboxymethylcellulose sodium (CMCNa) by gavage half an hour before challenging, and Group E was simultaneously given with 0.2 ml of Gefitinib dissolved in 0.5% CMCNa only. Group A and B were given an equivalent volume of 0.5% CMCNa by gavage. After 24 h of final challenge, the bronchoalveolar lavage fluid (BALF) was prepared for the determination of total cell count and eosinophil count. The levels of total immune globulin E (IgE) in serum and interleukin (IL)-4, IL-5 and IL-13 in BALF and lung tissue homogenates were measured by ELISA. The mRNA expression levels of IL-4, IL-5, IL-13 in lung were measured. Immunohistochemistry and Western blot experiments were used to detect the expression levels of EGFR in lung tissues. Results: In Group B, the level of total IgE in serum, total cell count, eosinophil count, the levels of IL-4, IL-5, IL-13 in BALF and the phosphorylation of EGFR and its downstream activation in lung were higher than those in Group A (all P<0.05). The levels of total IgE in serum [(261.32±44.38) ng/ml, (194.09±52.39) ng/ml vs (1 023.70±105.51) ng/ml], total cell count [(23.70±4.08)×105/ml, (14.92±4.06)×105/ml vs (35.36±6.30)×105/ml], eosinophil count [(108.00±13.69)×104/ml, (67.00±17.28)×104/ml vs (147.86±20.06)×104/ml], IL-4 [(36.42±4.48) pg/ml, (30.45±8.12) pg/ml vs (58.72±7.17) pg/ml], IL-5 [(16.20±4.62) pg/ml, (13.38±5.14) pg/ml vs (23.46±5.38) pg/ml], IL-13 [(18.45±7.28) pg/ml, (14.33±7.70) pg/ml vs (104.12±24.66) pg/ml] in BALF of Group C and D were lower than those in Group B (all P<0.05). The levels of IL-4, IL-5, and IL-13 as well as their mRNA levels in the lung tissue of Group C and D were lower than those in Group B (all P<0.05). In Group C and D, the positive expression rate of phosphorylated epidermal growth factor receptor (p-EGFR) in lung tissue [(40.53±6.80)%, (23.60±4.42)% vs (70.78±5.36)%], p-EGFR/EGFR (61.68±7.48, 51.13±5.19 vs 105.90±11.66), phosphorylated extracellular regulated protein kinase (p-Erk)/extracellular regulated protein kinase (Erk) (75.28±7.11, 47.54±4.83 vs 98.76±4.71), and phosphorylated protein kinase B (p-Akt)/protein kinase B (Akt) (96.24±5.40, 68.52±2.73 vs 103.30±4.52) was lower than those of Group B (all P<0.05). There was no statistically significant difference in the relevant indicators between Group A and E (all P>0.05). Conclusion: Gefitinib may alleviate airway inflammation and airway remodeling in asthmatic mice by inhibiting EGFR phosphorylation and affecting the activation of downstream Erk and Akt.
目的： 探讨表皮生长因子受体（EGFR）抑制剂吉非替尼对哮喘C57BL/6小鼠气道炎症和气道重塑的作用，并分析其作用机制。 方法： 用随机数字法将雄性6~8周龄C57BL/6小鼠分为A组（对照组）、B组（哮喘组）、C组（哮喘+20 mg/kg吉非替尼组）、D组（哮喘+40 mg/kg吉非替尼组）和E组（40 mg/kg吉非替尼组），每组7只。分别于第0、14天腹腔内注射卵清蛋白（OVA）及Al（OH）3［20 μg OVA+2 mg Al（OH）3溶于0.2 ml生理盐水］混合液0.2 ml致敏；B、C、D组分别于第25至31天开始雾化吸入1% OVA溶液8 ml激发哮喘，1次/d，每次约40 min，连续雾化7 d。C、D组每次雾化激发前半小时灌胃给予0.5%羧甲基纤维素钠（CMCNa）溶解的吉非替尼0.2 ml，E组仅同时给予0.5% CMCNa溶解的吉非替尼0.2 ml。A组和B组灌胃给予等量的0.5% CMCNa。末次激发24 h后，收集小鼠支气管肺泡灌洗液（BALF）进行总细胞计数及嗜酸性粒细胞计数，ELISA检测各组小鼠血清总IgE、BALF及肺组织匀浆中白细胞介素（IL）-4、IL-5和IL-13水平；检测肺组织中IL-4、IL-5和IL-13的mRNA表达水平；免疫组化、Western印迹实验检测肺组织中EGFR等指标的表达水平。 结果： B组小鼠血清总IgE水平，BALF中总细胞计数、嗜酸性粒细胞计数、IL-4、IL-5、IL-13水平，肺组织中EGFR的磷酸化及下游激活水平均高于A组（均P<0.05）。C、D组小鼠血清总IgE水平［（261.32±44.38）、（194.09±52.39）比（1 023.70±105.51）ng/ml］，BALF中总细胞计数［（23.70±4.08）×105个/ml、（14.92±4.06）×105个/ml比（35.36±6.30）×105个/ml］、嗜酸性粒细胞计数［（108.00±13.69）×104个/ml、（67.00±17.28）×104个/ml比（147.86±20.06）×104个/ml］，IL-4［（36.42±4.48）、（30.45±8.12）比（58.72±7.17）pg/ml］、IL-5［（16.20±4.62）、（13.38±5.14）比（23.46±5.38）pg/ml］、IL-13［（18.45±7.28）、（14.33±7.70）比（104.12±24.66）pg/ml］水平均低于B组（均P<0.05）；肺组织中IL-4、IL-5、IL-13水平及其mRNA水平均低于B组（均P<0.05）。C、D组小鼠肺组织中磷酸化表皮生长因子受体（p-EGFR）阳性表达率［（40.53±6.80）%、（23.60±4.42）%比（70.78±5.36）%］、p-EGFR/EGFR（61.68±7.48、51.13±5.19比105.90±11.66）、磷酸化细胞外调节蛋白激酶（p-Erk）/细胞外调节蛋白激酶（Erk）（75.28±7.11、47.54±4.83比98.76±4.71）和磷酸化蛋白激酶B（p-Akt）/蛋白激酶B（Akt）（96.24±5.40、68.52±2.73比103.30±4.52）比值均低于B组（均P<0.05）。A、E组相关指标差异均无统计学意义（均P>0.05）。 结论： 吉非替尼可能通过抑制EGFR磷酸化，影响下游Erk和Akt的活化，进而缓解哮喘小鼠气道炎症和气道重塑。.