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目的观察并比较C57BL/6小鼠初次和加强免疫Ⅱ型胶原后的特异性细胞及体液免疫应答,探讨特异性免疫应答在胶原诱导关节炎发病中的意义。方法用鸡Ⅱ型胶原(CⅡ)在C57BL/6小鼠尾部皮内注射,诱导胶原性关节炎(CIA)。分别取初次免疫后19d及加强免疫后7、28dCIA小鼠脾脏淋巴细胞,CⅡ体外刺激扩增后,采用5-溴脱氧尿嘧啶(BrdU)掺入法和流式细胞术分别测定其扩增情况和表型;并通过流式细胞术测定外周血中细胞内细胞因子干扰素(IFN)-γ、白细胞介素(IL)-4的水平分析Th1和Th2亚群的变化;同时用酶联免疫吸附试验(ELISA)测定不同时期CIA模型血清中抗CⅡ抗体的表达。结果①免疫后不同时间,CIA模型组外周血中CD4+IFN-γ+细胞百分率明显高于对照组(P<0.01),但CIA模型组内不同时间CD4+IFN-γ+细胞百分率差异无统计学意义;②特异性增生实验显示,CIA模型组CD4+T细胞中BrdU+细胞率均高于对照组,但加强免疫后BrdU+细胞率明显低于初次免疫(72±6)%,并且有逐渐降低的趋势;初次免疫后CD4+IFN-γ+细胞百分率为(13±4)%,加强免疫后阳性细胞百分率也下降;③ELISA法测定不同时间血清中CIA模型组抗CⅡ抗体水平,28d吸光度(A)值达到最高,其后逐渐减低,到35d左右(即加强免疫后14d)出现较低的抗体水平。结论C57BL/6小鼠加强免疫反应中特异
Objective To observe and compare the specific cellular and humoral immune responses of C57BL / 6 mice after initial and intensive immunization of type Ⅱ collagen and explore the significance of specific immune responses in the pathogenesis of collagen-induced arthritis. Methods Collagen induced arthritis (CIA) was induced intradermally in the tail of C57BL / 6 mice with chicken type Ⅱ collagen (C Ⅱ). Spleen lymphocytes and CⅡ splenic lymphocytes were stimulated at 19d after immunization and at 7 and 28d after immunization, respectively. After amplification in vitro, the proliferation of CIA mice was detected by BrdU incorporation and flow cytometry And phenotype. The levels of cytokines IFN-γ and IL-4 in peripheral blood were measured by flow cytometry. Th1 and Th2 subsets were analyzed by enzyme-linked immunosorbent assay Adsorption assay (ELISA) was used to determine the expression of anti-CⅡ antibody in serum of CIA model in different periods. Results ① The percentage of CD4 + IFN-γ + cells in CIA model group was significantly higher than that in control group at different time points after immunization (P <0.01), but the percentage of CD4 + IFN-γ + cells in CIA model group was not statistically different (2) The proliferation of BrdU + cells in CD4 + T cells in CIA model group was higher than that in control group, but the BrdU + cell rate was significantly lower than that of the first immunization (72 ± 6% ; The percentage of CD4 + IFN-γ + cells after primary immunization was (13 ± 4)%, and the percentage of positive cells after booster immunization also decreased. ③The levels of anti-CⅡantibody in CIA model group ) Reached the highest value, then gradually reduced to about 35d (ie, 14d after boosting) lower antibody levels. Conclusion C57BL / 6 mice specific immune response