PM_(2.5)对哮喘小鼠气道重塑的作用机制研究

Mechanism of PM_(2.5) on Airway Remodeling in Asthmatic Mice

目的 探讨PM_(2.5)对卵清蛋白(OVA)诱导的哮喘小鼠气道重塑的影响,并分析其潜在机制。方法 将36只SPF级雌性BALB/c小鼠随机分为对照组、哮喘组、哮喘+PM_(2.5)组,每组12只。哮喘组、哮喘+PM_(2.5)组于第1、7、14天腹腔注射OVA致敏,第21~27天连续进行1%OVA雾化激发,每次30 min,共7次。其中哮喘+PM_(2.5)组每次于OVA激发前30 min给予100μg PM_(2.5)混悬液滴鼻干预。对照组用生理盐水处理。3组均在末次激发24 h后进行气道阻力检测,处死动物后收集小鼠肺泡灌洗液(BALF)及肺组织标本。通过HE、PAS和Masson染色分别评估小鼠肺部炎症、杯状细胞增生和胶原纤维沉积情况,免疫组织化学法检测肺组织α-平滑肌肌动蛋白(α-SMA)的表达;通过ELISA法检测小鼠BALF中白细胞介素-4(IL-4)和IL-5水平;通过Western blot法检测小鼠肺组织p38丝裂原活化蛋白激酶(p38 MAPK)/核因子-κB(NF-κB)信号通路相关蛋白表达。结果 与哮喘组比较,哮喘+PM_(2.5)组气道阻力及炎症评分升高,杯状细胞增多,胶原纤维增生明显,BALF中IL-4、IL-5水平也显著增高(P<0.05)。与哮喘组比较,哮喘+PM_(2.5)组肺组织中转化生长因子-β_(1)(1.25±0.17 vs 0.87±0.25)、p-p38 MAPK/p38 MAPK(1.24±0.24 vs 0.74±0.19)、p-NF-κB p65/NF-κB p65(1.02±0.14 vs 0.70±0.10)蛋白表达水平升高(P<0.05)。结论 PM_(2.5)可通过激活p38 MAPK/NF-κB信号通路加重哮喘小鼠的气道重塑。

Objective To investigate the effect of PM_(2.5) on airway remodeling induced by ovalbumin(OVA) in asthmatic mice,and to analyze its underlying mechanism.Methods In total,36 SPF female BALB/c mice were randomly divided into control group,asthma group and asthma+PM_(2.5) group,with 12 mice in each group.Mice in the asthma group and the asthma+PM_(2.5) group were sensitized by intrabitoneal injection of OVA on days 1,7 and 14,and were stimulated by 1% OVA atomization on days 21-27(30 min each time for a total of 7 times).Mice in asthma+PM_(2.5) group were given 100 μg PM_(2.5) suspension nasal drops at 30 min before each OVA stimulation.The control group was treated with normal saline.Airway resistance was detected at 24 h after the last stimulation of mice in the three groups.Bronchoalveolar lavage fluid(BALF) and lung tissue samples were collected after the animals were sacrificed.Hematoxylin-eosin(HE) staining,PAS staining and Masson staining were used to evaluate the pulmonary inflammation,trophy cell hyperplasia and collagen fiber deposition.The expression of α-smooth muscle actin(α-SMA) in lung tissue was detected by immunohistochemistry,and the levels of interleukin-4(IL-4) and interleukin-5(IL-5) in BALF were determined by ELISA.The expression level of p38 mitogen-activated protein kinase(p38 MAPK)/nuclear factor-κB(NF-κB) signaling pathway related proteins in mouse lung tissues was detected by Western blot.Results Compared with the asthma group,the airway resistance and inflammation scores of asthma +PM_(2.5) group were increased,Goblet cells were increased,the collagen fiber hyperplasia was obvious,and the levels of IL-4 and IL-5 in BALF were also significantly increased(P<0.05).Compared with the asthma group,the protein expression levels of transforming growth factor-β(1.25±0.17 vs.0.87±0.25),p-p38 MAPK/p38 MAPK(1.24±0.24 vs.0.74±0.19),p-NF-κB p65/NF-κB p65(1.02±0.14 vs.0.70±0.10) in lung tissue of asthma +PM_(2.5) group were increased(P<0.05).Conclusion PM_(2.5) can aggravate airway remodeling in asthmatic mice by activating the p38 MAPK/NF-κB signaling pathway.