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目的探讨以改组尘螨Ⅱ类变应原基因Der f2和Der p2后表达的蛋白为变应原,对小鼠哮喘模型的特异性免疫治疗效果。方法随机将160只清洁级BALB/c小鼠分为PBS组(阴性对照组),Der f2和Der p2免疫治疗组(阳性对照组),哮喘组,改组变应原免疫治疗M01、M03、M08、M10蛋白治疗组。用尘螨提取液于0、7、14 d腹腔注射致敏激发BALB/c小鼠,第21天雾化吸入激发,连续7 d,其中各免疫治疗组于第25～27天雾化前30 min分别用Der f2、Der p2和改组变应原进行特异性免疫治疗,PBS组则用PBS进行腹腔注射和雾化吸入。最后1次雾化24 h后,引颈处死。分别观察肺组织病理变化、支气管肺泡灌洗液(BLAF)白细胞计数,酶联免疫吸附试验(ELISA)测定BLAF和脾细胞培养上清液细胞因子IL-4、IL-17和IFN-γ的含量及血清中特异性IgG1、IgE抗体水平变化。结果与哮喘组比较,改组变应原免疫治疗组和阳性治疗组肺部炎症显著减轻,BALF中的总细胞数及嗜酸性粒细胞数,血清中抗原特异性IgG1、IgE抗体均显著降低;免疫治疗组(包括阳性对照)的BALF和脾细胞培养上清液中的细胞因子IL-4、IL-17均明显降低,IFN-γ含量显著升高。结论通过基因改组获得的尘螨Ⅱ类变应原改组疫苗免疫治疗小鼠哮喘,可有效降低尘螨引起的小鼠肺部炎症。
OBJECTIVE: To investigate the immunotherapeutic effect of asthma model in mice by using the protein expressed after Der f2 and Der p2 of the mite class II allergens were reorganized. Methods 160 clean BALB / c mice were randomly divided into PBS group (negative control group), Der f2 and Der p2 immunotherapy group (positive control group), asthma group, shuffled allergen immunotherapy M01, M03, M08 , M10 protein treatment group. BALB / c mice were sensitized by intraperitoneal injection of dust mite extract on day 0, day 7, and day 14, and inhaled on the 21st day for 7 days. The immunization groups were immunized 30 days before the day 25 to 27 min respectively with Der f2, Der p2 and shuffled allergens for immunotherapy, while PBS group was injected intraperitoneally and nebulized with PBS. After the last atomization for 24 h, the neck was sacrificed. The pathological changes of lung tissue, the number of leukocyte in bronchoalveolar lavage fluid (BLAF) and the levels of cytokines IL-4, IL-17 and IFN-γ in the culture supernatant of BLAF and splenocytes were detected by enzyme linked immunosorbent assay (ELISA) And serum IgG1, IgE antibody level changes. Results Compared with the asthma group, the lung inflammation in the shuffled group and the untreated group were significantly reduced, and the total number of cells and eosinophils in BALF and the levels of IgG1 and IgE in serum were significantly decreased. The cytokines IL-4, IL-17 in the culture supernatant of BALF and spleen cells of the treatment group (including the positive control) were significantly decreased, and the content of IFN-γ was significantly increased. Conclusion Immune treatment of mice with asthma by shuffling mite allergen Ⅱ allergen induced by gene shuffling can effectively reduce lung inflammation in mice induced by dust mite.