局部致敏激发诱导变应性鼻炎模型的方法及机制分析.docx

局部致敏激发诱导变应性鼻炎模型的方法及机制研究摘要【目的】本实验主要探索局部滴鼻致敏及激发诱导实验性变应性鼻炎模型相矢的免疫学 指标及组织形态学特点•研究用最贴近自然的方式建立变应性鼻炎的模型，并对其 机 制进行初步研究方法】6周龄BALB/c雌性小鼠分为6组：AR腹腔注射模型组于第1、8、15天给 予0.4mg/ml OVA生理盐水辅以佐剂AI(OH)3腹腔致敏第29-33天和35-39天鼻腔滴 入25mg/ml OVA生理盐水进行激发AR大剂量(25mg/ml OVA)滴鼻模型组于第1—15 天给予鼻腔滴入25mg/ml OVA生理盐水致敏•第29-33天连续5天，间隔两天•在 34—39天连续5天间隔两天在41—45天连续滴鼻5天;AR中剂量(2.5mg/ml OVA) 滴鼻模型及AR小剂量(0.25mg/ml OVA)滴鼻模型组造模方法同大剂量组；麻黄素组前 期滴2周1%麻黄素后期同中剂量组；对照组以生理盐水代替另外各组小鼠会在实 验过程的22天、40天及46天尾静脉采血离心取血清，并均于末次激发后24h内取脾 脏单个核细胞培养3天后取细胞上清标本，保存于・80C结果】(1 )血清抗体：各实验组血清中OVA・specific IgE均较对照组及小剂量组显 著升高，模型组OVA・specific IgE较其他组显著升高，且小剂量、中剂量及大剂量组 的IgE呈递增趋势，麻黄素组较中剂量组显著升高；模型组、大剂量、中剂量、小剂 量及麻黄素组较对照组OVA・specific lgG1和OVA-specific lgG2a均显著升高。

在实验 实施过程中，小鼠体内抗体的动态变化发现小鼠体内IgE浓度在模型组、大剂量组、 中剂量组、小剂量及麻黄素组隨给药时间延长浓度逐渐升高，而且发现在后期麻黄素组 浓度高于大剂量组，总体结果小剂量、中剂量、大剂量的抗体浓度呈递增趋势，麻黄素 浓度早期低于大剂量，中期和大剂量无统计学差异，后期高于大剂量组各实验组小鼠 血清中lgG1均高于对照组•在实验早期阶段抗体浓度逐渐升高，而在实验第40天之 后抗体浓度均有下降趋势；lgG2a抗体浓度变化趋势和lgG1相仿•各组均显著高于 对照组•且在40天时各组抗体浓度均有明显下降趋势总体结果这三种抗体 也是随滴药剂量增加抗体浓度隨之增加（IgE 抗体浓度在大剂量和小剂量、中剂量和 麻黄素组、大剂量和模型组及麻黄素与模型组其组间比较PvO.05） 2 ）脾细胞培养 上清结果发现IL-4及IL-4/IFN-y比值在3种剂量组中隨给药剂量增加其OD值也逐渐 增加，该趋势与抗体结果一致•其中IL-4/IFN-y比值大剂量、中剂量和模型组明显升 高，IFN*在对照组及大剂量组明显高于其他实验组，有统计学意义3）HE染色结 果显示模型组大量嗜酸性粒细胞浸润，大剂量组、麻黄素组有明显嗜酸性粒细胞浸润， 中剂量组有嗜酸性粒细胞细胞浸润，而对照组和小剂量组未见嗜酸性粒细胞。

结论】：大剂量、中剂量、麻黄素组及模型组均造模成功，但免疫状态各有不同特点• IgE作为反应过敏性疾病的重要指标•其在麻黄素组中较中剂量组明显升高，小剂量、 中剂量和大剂量的免疫反应逐渐增强各组IgE隨刺激时间延长浓度逐渐升高・而 lgG1和lgG2a在后期出现下降趋势，且发现在后期麻黄素组IgE较大剂量组明显升 高，我们认为麻黄素破坏鼻黏膜后增加了小鼠过敏的易感性所以我们可以用贴近自然 的方式通过增加局部致敏变应原的剂量，延长激发时间以及前期对鼻黏膜的破坏干预建 立起接近常规模型组过敏程度的变应性鼻炎模型尖键词】动物模型；麻黄素；IgE; IgG 亚类；卵清蛋白；剂量AbstractObjective : This experiment mainly explore the local intranasal sensitization and challenge induced experime ntal allergic rhinitis model and their related immuno logical in dicators and morphological characteristics.Our research is in a mimic natural way to build the model of allergic rhinitis and a preliminary study on its mechanism.Method : 6 weeks of female BALB/c mice were divided into 6 groups: intraperitoneal injection of AR model group given 0.4 mg/ml OVA physiological saline with adjuvant Al (OH) 3 in 1,7,14 days,28-37 days for 10 consecutive days nasal drops with 25 mg/ml OVA physiological sali ne.AR in large dose (25 mg/ml OVA) intra nasal model group give n nasal drops with 25 mg/ml OVA physiological saline in 仁14 days,28-32 days for five consecutive days.interval of two days, in 34-38 days for five consecutive days, interval of two days,in the 40-44 days for 5 consecutive days.AR in middle dose (2.5 mg/ml OVA) intranasal model and AR in small dose (0.25 mg/ml OVA) intranasal model building method the same with large dose group.Ephedrine group dropping 2 weeks later with 1% ephedrine dose group,then in the same way as middle dose group.The control group accept physiological sali ne in stead.Caudal vein blood of all groups of mice in the experimental process of 22 days,40 days and 46 days is collected and we must take splenic mononuclear cell to culture 3 days after the last excitation in 24 , then collecting cultural supernatants to・ 80 C.Results : (1) Serum antibodies OVA - specific IgE in serum of the experimental group were significantly higher than the control group and small dose group and OVA ・ specific IgE of the model group is more sign ifica ntly in creased tha n the other groups.And IgE of the small dose group,medium dose group and high dose group showed a trend of increasing gradually.We see a more significant rise in ephedrine group than the model group.OVA - specific lgG1 and OVA ・ specific lgG2a of the high dose group, middle dose group, small dose group and ephedrine group were more significantly increased than the control group」n the experime nt,the dyn amic ch a nge of antibody in all experime ntal group found that the IgE co nee ntratio n in the model group, high-dose group,middle dose group,small dose group and ephedri ne group in creased as the exte nded time.The IgE concentratio n of the ephedrine group was higher than the large dose group in the late phase.The overall result is antibody concen tration in creased gradually as the does increased.The IgE concentration of the ephedrine group is lower than the high dose in early phase,the same as middle dose group in middle phasejate higher in the late phase」gG1 of all group in serum were higher than the control group and the antibody concentration in creased in the early stages of the experime nt and in the 40th day had a down ward trend.Similarly we found the same trend in the ch a nge of lgG2a concentration .Overall these three antibody in creased with the increase of dropping dose (IgE antibody concentration in large dose and small dose group,medium dose and ephedrine group,high dose and model group, ephedrine group and model group,comparing with each other, P < 0.05).(2) Spleen cell culture The OD value of IL-4 and IL・4/IFN・y in three dose groups also increased gradually with the dose increase as the trend in IgE antibody」L-4/IFN-yin the large dose group,middle dose group and the model g。