实验性自身免疫性神经炎相关细胞的免疫机制

Associated cell-mediated immune mechanisms of experimental autoimmune neuritis

目的建立 P2或 P0多肽诱导的实验性自身免疫性神经炎(EAN)大鼠模型,确定诱导EAN 的优选抗原和剂量,探讨 EAN 相关细胞免疫机制。方法实验组用 P2_(57-81)或 P0_(180-199)多肽加完全弗氏佐剂(FCA)免疫 Lewis 大鼠,对照组单用 FCA 免疫,致敏后每日对大鼠进行临床评分,比较高峰期最大评分,致敏第14天进行淋巴细胞增殖试验,测定 CD_4^+T/淋巴结单个核细胞(MNC)和CD_4^+CD_(25)^+T/CD_4^+T 细胞百分比,并进行坐骨神经病理检查。结果实验大鼠瘫痪高峰期最大评分,P2_(57-81)200μg组显著高于 P2_(57-81)100μg组和 P0_(180-199)200μg组(均P<0.01),P2_(57-81)100μg组与P0_(180-199)200μg组差异元统计学意义;P2_(57-81)100μg组和 P2_(57-81)200μg组对 P2_(57-81)多肽刺激反应性显著高于对照组(均 P<0.01),P0_(180-199)200μg组对 P0_(180-199)多肽刺激反应性显著高于对照组(P<0.01),P2_(57-81)100μg组和 P0_(180-199)200μg组对相应致敏多肽刺激反应性显著低于 P2_(57-81)200μg组(均P<0.05);CD_4^+T/淋巴结 MNC 百分比在各组间差异无统计学意义;CD_4^+CD_(25)^+T/CD_4^+T 细胞百分比,P2_(57-81)200μg组显著低于 P2_(57_81)100μg组、P0_(180-199)200μg组和对照组(均 P<0.01),P2_(57-81)100μg组和P0_(180-199)200μg组显著低于对照组(均P<0.01),P0_(180-199)200μg组显著低于 P2_(57-81)100μg组(P<0.01);EAN 急性期坐骨神经以炎性细胞浸润为主,P2_(57-81)200μg组重于其他实验组(均 P<0.01),P2_(57-81)200μg组慢性期无炎性细胞浸润,而表现多发性局灶性脱髓鞘和神经纤维崩解未恢复。结论200μg P2_(57-81)多肽是诱导 EAN 的优选抗原,EAN 致病与 CD_4^+T 细胞数量无关,而与致病性 T 细胞活性增强及CD_4^+CD_(25)^+T 细胞减少有关。

Objective To establish P2 or P0 peptide-induced experimental autoimmune neuritis （EAN） in the Lewis rats and to explore the optimal type and doses of antigen inoculated to induce EAN and the associated cell-mediated immune mechanisms. Methods Lewis rats were classified into EAN and control groups. The EAN rats were immunized by injection into both hind footpads of inoculums containing 100 or 200 μg of P2 57-81 peptide or 200 μg of P0 180-199 peptide and Freund＇ s complete adjuvant （ FCA）, and the control rats were immunized with FCA only. Clinical scores were compared when they were at peak time of paralysis. Lymphocyte proliferation assay, the ratio of CD4^＋ T cells to lymphatic monocytes and percentage of CD4^＋ CD25^＋T cells to CD4^＋ T cells obtained on day 14 post-immunization were examined. Histopathological assessment of sciatic nerves was made. Results Peak clinical scores of P2 57-81 200 μg group were dramatically higher than those in P2 57-81 100 μg group and P0 180-199 200 μg group （both P 〈0. 01 ） , and no differences were found between P2 57-81 100 μg group and P0 180-199 200 μg group. Proliferating responses of lymphatic monocytes to P2 57-81 peptide in P2 57-81 100 μg group and P2 57-81 200 μg group were higher than the Control group （both P 〈0. 01 ）, the responses to P0 180-199 peptide in P0 180-199 200 μg group was higher than the control group （ P 〈 0. 01 ）, and the responses to the inoculated peptide in P2 57-81 100 μg group andP0 180-199 200 μg group were lower than that in P2 57-81 200 μg group （ both P 〈 0. 05 ）. There were no differences of ratio of CD4^＋ T cells to lymphatic moncyes between groups. The ratio of CD4^＋ CD25^＋ T to CD4^＋ T cells in P2 57-81 200 μg group was dramatically lower than those in P2 57-81 100 μg group （P 〈0. 01 ）, P0 180-199 200μg group （P 〈0. 01 ） and control group （P 〈0. 01 ）, and those in P2 57-81 100 μg group and P0 180-199 200 μg group were lower than control group （both P 〈0.01 ）, and that in P0 180-199 200 μg group were lower than that in P2 57-81 100μg group （ P 〈0. 01 ）. Sciatic nerve in P2 57-81 200 μg group histologically revealed more severe inflammation than other EAN groups, and during the acute period of disease it was characterized by infiltration of inflammatory cells, whereas during the chronic period it was mainly presented by remnant multiple localized demyelination or collapse of nerve fibers without infiltration of inflammatory cells. Condusion P2 57-81 peptide is an optimal antigen at a dose of 200 μg to induce experimental autoimmune neuritis in the Lewis rats, whose severity and course are correlated with the enhanced activity of pathogenic T cells and the decreased number of CD4^＋ CD25^＋T cells, other than the number of CD4^＋ T cells.