髓鞘碱性蛋白片段69-85诱导建立大鼠自身免疫性脑脊髓炎模型

Establishment of a Myelin Basic Protein Fragment-Induced Wistar Rat Model of Autoimmune Encephalomyelitis

目的探索以大鼠髓鞘碱性蛋白片段69-85（MBP69-85）为单一抗原,建立Wistar大鼠实验性自身免疫性脑脊髓炎（EAE）模型,并观察其病理改变。方法MBP69-85以生理盐水溶解,与完全福氏佐剂（CFA）充分混匀制备免疫乳剂;雌性Wistar大鼠70只,留取10只作为正常对照组,余者根据免疫乳剂组分的不同随机分为A、B、C组（n=20）。A组：MBP69-85每只50μg＋CFA（含卡介苗6 mg）;B组：MBP69-85每只25μg＋CFA（含卡介苗6 mg）;C组：MBP69-85每只25μg＋CFA（含卡介苗12 mg）。脑和脊髓组织切片进行HE染色,MBP及神经微丝（neurofilament,NF）免疫组化检测,观察神经组织的病理改变。结果A组内部分大鼠在免疫后第12-16天发病,表现为尾部及四肢无力或麻痹、斜颈等,平均临床症状评分为2.38±1.89;B、C组均未见发病;HE染色可见神经组织内炎细胞浸润,血管＂袖套＂样病灶形成;MBP及NF免疫组化染色显示病变组织内白质脱髓鞘及轴突损伤明显。结论EAE模型建立与免疫抗原的剂量有一定的依赖关系;佐剂中的卡介苗含量不是诱导大鼠发病的主要原因;本模型具有多发性硬化的典型临床表现及病理改变,是研究多发性硬化发病机制及治疗的可靠的动物模型。

Objective To establish an experimental autoimmune encephalomyelitis（EAE） model in Wistar rats with myelin basic protein fragment（MBP69-85） and observe its pathological changes.Methods MBP69-85 dissolved in normal saline was mixed with complete Freund＇s adjuvant（CFA）（including 6 mg bacille Calmette Guerin） to prepare the encephalitogenic emulsion.Ten out of 70 Wistar rats were chosen as a control group,the others were divided equally into group A,B,C according to the difference of the encephalitogenic emulsion.Rats in group A were immunized with 50 μg MBP69-85 ＋CFA（including 6 mg BCG）.Rats in group B were immunized with 25 μg MBP69-85＋CFA（including 6 mg BCG）.Rats in group C were immunized with 25 μg MBP69-85＋CFA（including 12 mg BCG）.The pathological changes of brain and spinal cord tissues were examined by light microscopy after HE staining and immunohistochemistry of MBP and NF.Results Some of the Wistar rats immunized with 50 μg MBP69-85 showed disorder at 12～16 days after immunization.The clinical symptoms included tail acratia or paralysis of tail and limbs,head tilt,etc.and the mean score was 2.38±1.89.There were infiltration of inflammatory cells inside nervous tissue and perivascular cuffings in HE stained sections.The immunohistochemistry of MBP and NF showed demyelination in the white matter and axon injury.Conclusion To some extent,the establishment of EAE depends on the dose of the immunizing antigen.The BCG in CFA was not the major cause of morbility of the rats.The EAE model induced with MBP69-85 in Wistar rats,showing typical clinical symptoms and pathological changes of multiple sclerosis,is a reliable animal model for the study of pathogenesis and treatment of multiple sclerosis.