B细胞连接蛋白的表达抑制小鼠实验性自身免疫性脑脊髓炎

B-cell linker protein inhibits experimental autoimmune encephalomyelitis

目的研究B细胞连接蛋白BLNK(B-cell linker protein)的表达对小鼠实验性自身免疫性脑脊髓炎(experimentalautoimmune encephalomyelitis,EAE)的影响并初步探讨其可能的机制。方法 MOG35-55多肽免疫BLNK缺陷(BLNK-deficient,BLNK-/-)鼠及C57BL/6鼠制备EAE小鼠模型,观察实验动物的临床症状及中枢神经系统的病理学变化;应用ELISA方法检测血清中的MOG特异性IgG和IgM抗体水平;采用流式细胞术检测脾脏中调节性T细胞的变化;分选的脾脏B淋巴细胞采用RT-PCR技术评价B细胞所分泌的细胞因子的变化。结果 BLNK-/-鼠临床症状评分明显高于C57BL/6鼠(P<0.01);HE染色及LFB染色结果显示,BLNK-/-鼠与C57BL/C鼠比较炎症感染灶及脱髓鞘病灶明显增多。BLNK-/-鼠血清中的MOG特异性IgG和IgM抗体水平显著低于C57BL/C鼠(P<0.005)。BLNK-/-鼠与C57BL/6鼠相比,脾脏中调节性T细胞百分比及B细胞所分泌的IL-10的表达明显减少,而IL-2的表达显著增高(P<0.05);IFN-γ的表达水平在2组小鼠之间无统计学差异(P>0.05)。结论 BLNK的表达抑制EAE,其机制可能是通过对B细胞分泌的Th1/Th2细胞因子的调节来实现。

This study was performed to evaluate the role of B-cell linker protein （BLNK） on the development of experimental autoimmune encephalomyelitis （EAE） and its possible mechanism. BLNK-deficient （BLNK^-/-） mice and C57BL/6 mice were immunized with MOG peptide in CFA to construct EAE model, and the EAE clinical score and pathological severity were observed. Serum IgM and IgG anti-MOG Ab levels were measured by ELISA; the percentage of regulatory T cells from spleen was analyzed by/low cytometry. PT-PCR was used to determine the production of cytokines in B cells from spleen. Data showed that BLNK^-/- mice have augmented disease course of EAE compared with wild-type mice （P〈0.01）. HE and Luxol Fast Blue-Periodic acid （LFB） staining showed that areas of inflammation and demyelination were more widespread in BLNK-/- mice when compared with wild-type mice at day 16. However, BLNK^-/- mice showed lower serum IgM and IgG anti-MOG Ab levels compared with wild-type mice （P〈0.005）. Furthermore, the percentage of regulatory T cell was significantly reduced in BLNK^-/- mice compared to wild-type mice （P〈0.05）. In addition, the IL-10 mRNA expression of Th2 cytokine in BLNK^-/- mice was decreased compared with wild-type mice （P〈0.05）. By contrast, Thl cytokine expression, IL-2, was increased in BLNK^-/- mice compared with wild-type mice （P〈0.05）. While the mRNA expression of IFN-γ was slightly but not significantly elevated in BLNK^-/- mice （P〉0.05）. Thus, BLNK expression in B cells is critical for termination of EAE responses, possibly through modulating the Th1/Th2 cytokine balance in B cells.