黄花蒿花粉过敏小鼠鼻炎模型的优化与稳定性研究

Study on optimization and stability of Artemisia annua pollen-induced allergic rhinitis model in mice

目的探索黄花蒿花粉变应性小鼠鼻炎模型维持稳定的激发条件。方法以BALB/c小鼠为实验动物,选用黄花蒿花粉变应原提取物为致敏蛋白质,于1 d、4 d、7 d对每只小鼠皮下注射0.1 ml的以Art a1计浓度为20μg/ml的黄花蒿花粉变应原提取液。致敏结束后停息1周,用以Art a1计浓度为500μg/ml的黄花蒿花粉变应原提取液对小鼠进行首次滴鼻激发,每鼻孔10μl。将模型组分为3组分别连续激发7 d、10 d、14 d,每隔4周再激发7 d,探索首次激发时间。空白对照组小鼠以生理盐水进行免疫和滴鼻激发。造模后观察各组小鼠行为学和鼻部病理学变化,检测致敏小鼠体液免疫和细胞免疫应答变化。确定最佳首次滴鼻激发周期后,进一步比较不同激发频次对致敏效果的影响。结果抗原首次激发后,10 d组小鼠过敏症状明显高于7 d和14 d组,且10 d和14 d组小鼠血清特异性IgE抗体水平明显高于7 d组。与空白对照组比较,二次激发后3个模型组小鼠组仍维持明显过敏症状;鼻部均有明显的上皮细胞增生、鼻甲肿大、炎细胞增多等病理变化;血清特异性IgE抗体水平均显著增高;抗原特异性IL-4和IL-6淋巴细胞增殖明显,其中10 d和14 d组高于7 d组。不同激发频次对过敏模型的稳定性有较大影响,每隔2周激发一次的致敏小鼠过敏症状以及血清中抗原特异性抗体水平均显著高于每隔4周激发的小鼠。结论本研究通过对比不同首次抗原激发时间以及不同激发频次对黄花蒿致敏小鼠模型的影响,最终优化了过敏小鼠评价模型建立的参数,为脱敏治疗药物疗效评价体系的建立提供参考。

Objective To optimize the challenge scheme for establishing a stable mouse model of Artemisia annua pollen-induced allergic rhinitis.Methods BALB/c mice were subcutaneously injected with 0.1 ml allergen extract containing 20μg/ml Art a1 from Artemisia pollen on 1 d,4 d and 7 d.One week after the sensitization,these mice were divided into three groups and intranasally challenged with Artemisia annua pollen allergen extract containing 500μg/ml Art a1 for 7(7 d group),10(10 d group)and 14(14 d group)consecutive days,respectively.The first challenge was followed by another 7 days of challenge every four weeks.Blank control group was set up through sensitizing and challenging BALB/c mice with normal saline.Behavioral changes and nasal pathological changes were observed.The changes in humoral and cellular responses were also detected.After the first challenge cycle was decided,the challenge frequency was further optimized.Results After the first challenge,the allergic symptoms of mice in 10 d group were significantly severe than those in 7 d and 14 d groups,and the levels of serum specific IgE antibody in 10 d and 14 d groups were significantly higher than that in 7 d group.After the second challenge,the mice in the three model groups still had obvious allergic symptoms as compared with the blank control group.There were obvious pathological changes in the nose,including epithelial cell proliferation,turbinate enlargement and inflammatory cell increase.Moreover,the level of serum specific IgE antibody increased significantly and the proliferation of antigen-specific IL-4 and IL-6 lymphocytes was significantly up-regulated,especially in 10 d and 14 d groups.The frequency of challenge had a great impact on the stability of the allergic model.The allergic symptoms of sensitized mice challenged every two weeks were significantly severe than those of mice challenged every four weeks and the level of serum antigen-specific antibody was also higher.Conclusions This study optimized the first challenge cycle and challenge frequency for establishing a mouse model of Artemisia annua pollen-induced allergic rhinitis,which provided reference for the establishment of drug efficacy evaluation system for desensitization therapy.