SD 大鼠食物过敏动物模型建立及其消化系统超微结构变化

摘要

Objective To investigate the methodology of establishing food allergy animal model in SD rats and the ultrastructure change of the digestive system.Method 30 SD rats were randomly divided into ovalbumin group(20 rats)and normal saline group(10 rats).Intraperitoneal injection of low dose of sensitization and enhanced sensitization with OVA were performed in OVA group rats on day 1 and day 15 respectively.High-doses intragastric OVA challenge was performed in OVA group rats every two days since day 20 to day 49. Intraperitoneal injection and intragastric administration with same dose normal saline was performed in control group.Serum OVA-IgE antibodies were detected by using ELISA method at week 1,2,3,4,5,6, 7 respectively.Diarrhea was observed among the rats.Rats were sacrificed at week 6,5 cm long duodenum was clipped from ligament of Treitz to distal end.Toluidine blue and HE staining were performed for the mast cell integrity and pathological changes assessment of intestine respectively after fixation,embedding and sectioning.The ultrastructure of intestinal mucosa was observed under transmission electron microscope. Results The OVA-IgE level of OVA group increased gradually from week 4 to 6,but declined at week 7, compared to the normal control group,the difference was statistically significant.20 rats were all attacked by diarrhea at weeks 6.Jejunal mucosal eosinophil density [(16.48 ±2.32)cells/HPF vs.(1.70 ±0.42) cells/HPF]and mast cell number [(9.02 ±1.55)cells/HPF vs.(1.84 ±0.87)cells/HPF]were higher, but the mast cell integrity percentage [(36.50% ±12.54%)vs.(92.20% ±6.87%)]was lower than control group,The difference was statistically significant (P <0.01).The rarefaction and shedding of villi, obscure cell junction,extensive mitochondrial swelling,dilatation and degranulation endoplasmic reticulum was observed in small intestine in OVA group under the light and electron microscope.Conclusions The food allergy SD rats model was established successfully based on low-dose intraperitoneal injection and high-dose intragastric challenge in 6 weeks.Small intestinal mucosal injury obscure cell junction can be observed in OVA sensitized rats,which may play an important role in the pathogenesis of food allergy.%目的：探讨 SD 大鼠食物过敏动物模型建立的适宜条件及该模型消化系统超微结构的改变。方法30只 SD 大鼠随机分为食物过敏组（20只），生理盐水组（10只）。食物过敏组第1天及第15天以卵清蛋白（ovalbumin，OVA）低剂量腹腔注射进行基础致敏及强化致敏；第20天起以 OVA 进行高剂量隔日灌胃激发至第49天；对照组分别以等量生理盐水进行腹腔注射及灌胃。分别于1、2、3、4、5、6和7周采用酶联免疫吸附试验检测血清 OVA-IgE 抗体水平，同时观察大鼠腹泻情况。第6周后处死动物，取其十二指肠 Treitz 韧带以下5 cm，经固定、包埋、切片，甲苯胺兰染色行肥大细胞完整性评估，HE 染色下观察肠道病理变化，电镜观察肠黏膜超微结构。结果食物过敏组 OVA-IgE 水平在第4～6周逐渐升高，第7周下降，与对照组相比差异有统计学意义（P ＜0．01）。第6周食物过敏组大鼠均出现腹泻。食物过敏组与对照组肠黏膜嗜酸粒细胞密度[（16．48±2．32）个/HPF vs．（1．70±0．42）个/HPF]、肥大细胞数总数[（9．02±1．55）个/HPF vs．（1．84±0．87）个/HPF]和完整性百分率[（36．50％±12．54％）vs．（92．20％±6．87％）]比较，差异有统计学意义（均 P ＜0．01）。电镜和光镜下观察，食物过敏组动物小肠绒毛稀疏、脱落，细胞连接模糊，线粒体广泛肿胀，内质网扩张，内质网脱颗粒。结论采用 SD 大鼠通过低剂量腹腔注射与高剂量灌胃激发至6周可成功建立理想食物过敏动物模型，卵清蛋白致敏大鼠小肠肠黏膜损伤明显，细胞连接模糊，可能在食物过敏发病机制中起重要作用。