摘要
目的探讨粉尘螨变应原Derf1mRNA疫苗对哮喘小鼠的特异性免疫治疗效果。方法 50只BALB/c小鼠随机分为5组(10只/组),分别为PBS组、Derf1变应原致敏组、Derf1变应原免疫治疗组、β-actin mRNA免疫治疗组和Derf1 mRNA免疫治疗组。分别于第0、第7和第14天,PBS组小鼠腹腔注射PBS,其余4组小鼠则腹腔注射10μg Der f 1进行致敏,建立小鼠哮喘模型。自第21天起,除PBS组小鼠雾化吸入PBS,其他4组小鼠均雾化吸入100μg/ml Der f 1变应原,30 min/次,1次/d,连续7 d,观察并记录小鼠哮喘发作情况。5组小鼠于最后1次雾化吸入致敏2周后,背部皮下分别注射100μl PBS、1μg Der f1(维持致敏)、10μg Derf1(免疫治疗)、2μgβ-actin mRNA和2μg Derf1mRNA,1次/周,连续3周。最后1次皮下注射后2周处死小鼠。收集各组小鼠的支气管肺泡灌洗液(BALF),ELISA法检测γ干扰素(IFN-γ)和白介素13(IL-13)水平,并计数嗜酸粒细胞(EOS);收集各组小鼠脾组织,分离脾细胞,除PBS组外,其他4组均加入10μg/ml Der f 1培养72 h,ELISA法检测脾细胞培养上清液中IFN-γ和IL-13水平;取各组小鼠眼球血,ELISA法检测血清中总IgE以及变应原特异性IgE(sIgE)、IgG1(sIgG1)和IgG2a(sIgG2a)抗体水平。HE染色观察各组小鼠肺组织切片。结果除PBS组外,其他4组小鼠雾化吸入致敏后,均出现急性哮喘发作症状。小鼠BALF中,Der f 1 mRNA免疫治疗组和Der f 1免疫治疗组的IFN-γ水平分别为(897.56±105.73)和(864.48±70.62)pg/ml,均明显高于Derf1变应原致敏组[(209.05±52.28)pg/ml]和β-actin mRNA免疫治疗组[(219.47±64.72)pg/ml](P<0.01);两者IL-13水平分别为(241.64±31.41)和(321.94±41.07)pg/ml,则明显低于Derf1变应原致敏组[(520.62±43.77)pg/ml]和β-actin mRNA免疫治疗组[(507.22±42.26)pg/ml](P<0.01);两者EOS数量分别为(1.33±0.44)×105和(1.48±0.39)×105个/ml,均明显低于Derf1变应原致敏组[(3.54±0.52)×105个/ml]和β-actin mRNA免疫治疗组[(2.98±0.53)×105个/ml](P<0
Objective To investigate the effect of Der f 1 mRNA molecules for specific immunotherapy on murine model of asthma. Methods Fifty BALB/c mice were randomly divided into 5 groups： PBS group, Der f 1 sensitization group, Der f 1 specific immunotherapy（SIT） group, β-actin mRNA SIT group, and Der f 1 mRNA SIT group. On days0, 7 and 14, mice in PBS group received PBS injection; mice in the other groups were intraperitoneally injected with10 μg Der f 1. At day 21, the mice in the 4 experimental groups were challenged with a 30-min inhaled dose of Der f 1（100 μg/ml） for 7 successive days. Two weeks after the final sensitization, the mice in the above five groups were immunized by intradermal injection with PBS, 1 μg Der f 1, 10 μg Der f 1, 2 μg β-actin mRNA, and 2 μg Der f 1 mRNA,respectively for 3 times at one-week intervals. Two weeks after the last intradermal injection, all mice were sacrificed and bronchoalveolar lavage fluid（BALF） was collected. ELISA was performed to detect the levels of IFN-γ and IL-13 in BALF, the number of eosinophils in the BALF was recorded. Splenocytes were prepared, and cultured with Der f 1 allergen（10 μg/ml） for 72 h. Splenocytes of PBS group was cultured without Der f 1 allergen. The levels of IFN-γ and IL-13 in splenocyte culture supernatant were measured by ELISA, as well as serum antibody levels of total IgE, allergenspecific IgE（sIgE）, sIgG1, and sIgG2 a. Lung sections were stained in hematoxylin and eosin, and observed under the microsope. Results Except for PBS group, mice in the other 4 group showed symptoms of acute asthma attack. Compared with Der f 1 sensitization group [（897.56±105.73） pg/ml] and β-actin mRNA SIT group [（219.47±64.72） pg/ml],the level of IFN-γ in BALF from Der f 1 mRNA SIT group [（897.56±105.73） pg/ml] and Der f 1 SIT group [（864.48±70.62）pg/ml] significantly increased（P 0.01）. However, the level of IL-13 in BALF from Der f 1 mRNA SIT group[（241.64±31.41） pg/ml] and Der f 1 SIT group [（321.94 �
出处
《中国寄生虫学与寄生虫病杂志》
CAS
CSCD
北大核心
2014年第4期268-273,共6页
Chinese Journal of Parasitology and Parasitic Diseases
基金
国家自然科学基金(No.81172790
81270091)
国家级大学生创新创业训练项目(No.201310368022
201310368031)~~
关键词
粉尘螨
变应原
mRNA疫苗
特异性免疫治疗
Dermatophagoides farinae
Allergen
mRNA vaccine
Specific immunotherapy
