芪仙汤对苯并芘加剧哮喘小鼠气道炎症及黏液高分泌的影响及机制研究

Study on the Effect and Mechanism of Qixian Decoction on Airway Inflammation and Mucus Hypersecretion in Mice with Benzopyrene-Exacerbated Asthma

目的观察芪仙汤(黄芪、仙灵脾、巴戟天等)对暴露于大气污染物苯并芘(BaP)的哮喘小鼠气道炎症及黏液分泌的影响,并探讨其分子机制。方法将雌性BALB/c小鼠随机分为空白组、哮喘组[卵白蛋白(OVA)致敏]、BaP组(OVA+BaP)和芪仙汤组(OVA+BaP+芪仙汤37.5 g·kg^(-1)),每组10只。采用OVA致敏法复制小鼠哮喘模型,并以BaP滴鼻法模拟大气污染暴露。采用HE、AB-PAS染色法分别检测小鼠肺组织炎症及黏液分泌水平;qRT-PCR法检测肺组织白细胞介素4(IL-4)、白细胞介素13(IL-13)mRNA表达;免疫组化法检测肺组织黏蛋白5AC(MUC5AC)及磷酸化ERK(p-ERK)蛋白表达;DCFH-DA探针法检测肺组织活性氧(ROS)水平。结果与空白组比较,哮喘组小鼠的气道、血管周围炎症细胞明显增多,气道黏膜出现中断、不连续,炎症评分显著升高(P<0.01);小鼠肺组织IL-4、IL-13 mRNA表达明显上调(P<0.05);小鼠的气道黏液分泌显著增多(P<0.01);小鼠气道MUC5AC、p-ERK蛋白表达显著上调(P<0.01);小鼠肺组织ROS水平明显升高(P<0.05)。与哮喘组比较,BaP组小鼠的炎症细胞浸润进一步加剧,肺泡间隔断裂明显,炎症评分显著升高(P<0.01);小鼠肺组织IL-4、IL-13 mRNA表达显著上调(P<0.01);小鼠的气道黏液显著增多(P<0.01);小鼠气道MUC5AC、p-ERK蛋白表达显著上调(P<0.05,P<0.01);小鼠肺组织ROS水平显著升高(P<0.01)。与BaP组比较,芪仙汤组小鼠的炎症细胞浸润、气道黏膜连续性均明显改善,炎症评分显著降低(P<0.01);小鼠肺组织IL-4、IL-13 mRNA表达显著下调(P<0.01);小鼠的气道黏液分泌显著减少(P<0.01);小鼠气道MUC5AC、p-ERK蛋白表达显著下调(P<0.05,P<0.01);小鼠肺组织ROS水平显著降低(P<0.01)。结论芪仙汤可改善大气污染物BaP加剧的哮喘小鼠气道炎症和黏液分泌,具有抗肺损伤作用,可能与其对ROS和ERK通路的抑制有关。

Objective To observe the effects of Qixian Decoction(composed of Astragali Radix,Herba Epimedii,Morindae Officinalis Radix,etc.) on airway inflammation and mucus secretion in asthmatic mice exposed to the atmospheric pollutant benzopyrene(BaP),and to explore the molecular mechanisms.Methods Female BALB/c mice were randomly divided into blank group,asthma group[ovalbumin(OVA)sensitization],BaP group(OVA+BaP)and Qixian Decoction group(OVA+BaP+Qixian Decoction 37.5 g·kg^(-1)),with 10 mice in each group.The mouse asthma model was replicated using the OVA sensitization method,and exposed to atmospheric pollution was simulated by BaP nasal drip method.The levels of lung inflammation and mucus secretion were detected by HE and AB-PAS staining;the mRNA expressions of interleukin 4(IL-4)and interleukin 13(IL-13)in lung tissue were detected by qRT-PCR,and the protein expressions of mucin 5AC(MUC5AC)and phosphorylated ERK(p-ERK)in lung tissue were detected by immunohistochemistry;the lung tissue reactive oxygen species(ROS)levels were measured by the DCFH-DA probe method.Results Compared with the blank group,the inflammatory cells around airway and blood vessels in asthma group were significantly increased;the airway mucosa was interrupted and discontinuous,and the inflammation score was significantly increased(P<0.01);the mRNA expressions of IL-4 and IL-13 in mice lung tissue were significantly up-regulated(P<0.05);the airway mucus secretion of mice was significantly increased(P<0.01);the protein expressions of MUC5AC and p-ERK in airway of mice were significantly up-regulated(P<0.01);the ROS level in mouse lung tissue was significantly increased(P<0.05).Compared with the asthma group,the inflammatory cell infiltration in BaP group was further aggravated,the alveolar septum was significantly broken,and the inflammatory score was significantly increased(P<0.01);the mRNA expressions of IL-4 and IL-13 in mouse lung tissue were significantly up-regulated(P<0.01);the airway mucus in mice was significantly increased(P<0.01);the protein expressions of MUC5AC and p-ERK in mice airway were significantly up-regulated(P<0.05,P<0.01);and the ROS level in mice lung tissue was significantly increased(P<0.01).Compared with the BaP group,the inflammatory cell infiltration and airway mucosal continuity in the mice in the Qixian Decoction group were significantly improved,and the inflammation score was significantly reduced(P<0.01);the mRNA expressions of IL-4 and IL-13 in mice lung tissue were significantly down-regulated(P<0.01);the airway mucus secretion in mice was significantly reduced(P<0.01);the protein expressions of MUC5AC and p-ERK in mice airway were significantly down-regulated(P<0.05,P<0.01);the ROS level in mice lung tissue was significantly reduced(P<0.01).Conclusion Qixian Decoction has effects in relieving airway inflammation and mucus secretion in mice with asthma exacerbated by atmospheric pollutants BaP and the anti-lung damage effects were possibly related to its inhibition of ROS and ERK pathways.