双类似物鼻黏膜耐受对实验性自身免疫性重症肌无力幼鼠作用机制的研究

The mechanism of prophylactic effects of nasal tolerance with a dual analogue on experimental autoimmune myasthenia gravis in young mice

摘要:

目的：对实验性自身免疫性重症肌无力(EAMG)幼鼠选择双类似物（Lys262-Ala207）通过不同时间点进行鼻黏膜免疫耐受预防性给药后，观察其临床及免疫指标的变化，评价临床疗效及探讨鼻黏膜耐受对EAMG的预防作用机制。方法：用乙酰胆碱受体单克隆抗体建立C57BL/6小鼠EAMG模型，并在致敏前10 d(A组)及致敏当日(B组)经鼻黏膜滴入Lys262-Ala207诱导黏膜耐受，检测给药后A,B组及相应对照组CA组、CB组小鼠的临床症状及各项指标的改变情况。结果：①A，B两组临床症状明显轻于相应对照组,且A组临床症状轻于B组。②A，B组低频重复电刺激（RNS）出现衰减的阳性率低于各自对照组；③A，B两组小鼠血清乙酰胆碱受体抗体AchRAb(IgG)水平明显低于各自对照组，且A组轻于B组。④调节性CD4+CD25+T细胞含量A组（4.516±0.598）明显高于B组（3.671±0.300），且A，B组明显高于各自对照组。结论：Lys262-Ala207鼻黏膜预防耐受可有效地抑制重症肌无力的临床症状，为采用该途径治疗重症肌无力的可行性提供了理论依据。

Abstract:

OBJECTIVE: To study the prophylactic effects of nasal tolerance with a dual analogue, Lys262-Ala207, on the mouse model of experimental autoimmune myasthenia gravis (EAMG) and the underlying mechanism. METHODS: Mouse model of EAMG was induced by intraperitoneal injection of mAb35. Lys262-Ala207 or PBS was given nasally before 10 days (study group A and control group A) or on the day (study group B and control group B) of immunization for 10 days. Clinical syndromes were evaluated after immunization. Serum level of acetylcholine receptor antibody (AChR-Ab) IgG was detected using ELISA. The number of monouclear cells expressing CD4+ and CD4+ CD25+T from spleen was measured using flow cytometry. RESULTS: Compared with the corresponding control groups, the clinical syndromes were improved (P<0.01) in mice from the study groups A and B. The positive rate of the repetitive nerve stimulation (RNS) test in the study groups A and B was significantly lower than that in the corresponding control groups (P<0.01). The study group A showed lower positive rate of RNS than the study group B (P<0.05). The serum levels of AChR-Ab IgG in the study groups A and B (15.01±1.09 and 19.23±1.31 μg/mL) decreased compared with that in the corresponding control groups (28.12±1.28 and 29.35±1.28 μg/mL) (P<0.01). The study group A mice had lower serum AChR-Ab IgG levels than the study group B (P<0.01). The number of CD4+ CD25+T cells in the study groups A (4.516±0.598%) and B (3.671±0.300%) increased significantly compared with that in the corresponding control groups (2.661±0.411% and 2.412±0.500%) (P<0.01) and more CD4+ CD25+T cells were found in the study group A than in the study group B (P<0.01). CONCLUSIONS: Nasal administration with dual analogues may ameliorate clinical syndromes in EAMG rats, which may be associated with decreased serum AChR-Ab IgG levels and increased number of CD4+ CD25+T cells from spleen.