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目的检测雌激素在小鼠实验性自身免疫性脑脊髓炎(EAE)中的预防性治疗作用并初步探讨其体内作用机制。方法用髓鞘少突胶质细胞糖蛋白(MOG_(35-55))和完全弗氏佐剂(CFA)免疫C57BL/6J小鼠制备EAE模型;雌激素于建模始每日皮下注射50μmol/L进行治疗,通过发病指数评分、脊髓病理分析、炎症因子水平检测,并应用流式细胞仪(FACS)及实时荧光定量PCR(q PCR)评价治疗效果;进一步通过分析小鼠脾细胞及脊髓相关功能分子CD4/PD-1、CD19/PD-L1、CD4/CD25/Foxp3、CD19/CD21/CD23及CD19/CD5/CD1d~(high)的表达变化,探讨雌激素可能的作用机制。结果雌激素预防性治疗可有效延缓EAE小鼠的发病,减轻脱髓鞘病变。病理结果显示,雌激素治疗能减少小鼠脊髓组织炎性细胞的浸润,同时下调外周炎性因子TNF-a及IL-17A的分泌表达(P<0.01);与模型组相比,雌激素治疗下调T淋巴细胞表面活化分子CD69的表达(P>0.05)并可显著上调CD4~+T细胞表面PD-1的表达(P<0.05);同时,雌激素治疗后PD-1的配体CD19+B细胞表面PD-L1的表达明显上调(P<0.01),脾细胞中CD19~+CD21~(high)CD23~(low)(P<0.01)及CD19~+CD5~+CD1d~(high)(P<0.01)的比例也有不同程度的升高,但对脾细胞中CD4~+CD25~+Foxp3~+的表达无明显作用。结论雌激素预防性治疗可延缓EAE小鼠的发病进程,其作用机制可能与上调PD-1/PD-L1通路并上调调节性B细胞水平有关。
Objective To investigate the preventive effect of estrogen in mice with experimental autoimmune encephalomyelitis (EAE) and to explore its mechanism in vivo. Methods C57BL / 6J mice were immunized with myelin oligodendrocyte glycoprotein (MOG_ (35-55)) and complete Freund’s adjuvant (CFA) to prepare EAE model. Estrogen was injected daily subcutaneously at a dose of 50μmol / L, and the therapeutic effect was evaluated by the score of onset index, the pathological analysis of the spinal cord and the level of inflammatory cytokines, and the therapeutic effect was evaluated by flow cytometry (FACS) and real-time quantitative PCR (qPCR). Furthermore, To explore the possible mechanism of action of estrogen by examining the expression of functional molecules CD4 / PD-1, CD19 / PD-L1, CD4 / CD25 / Foxp3, CD19 / CD21 / CD23 and CD19 / CD5 / CD1d high. Results Estrogen preventive treatment can effectively delay the onset of EAE mice and reduce the demyelinating lesions. The results of pathology showed that estrogen treatment can reduce inflammatory cell infiltration in spinal cord tissue of mice and down-regulate secretion of TNF-a and IL-17A (P <0.01). Compared with model group, estrogen treatment (P <0.05), and down-regulated the expression of PD-1 on the surface of CD4 ~ + T lymphocytes (P <0.05). Meanwhile, the expression of CD19 + The expression of PD-L1 on B cells was significantly upregulated (P <0.01). The levels of CD19 ~ + CD21 ~ (high) CD23 ~ (low) and CD19 ~ + CD5 ~ + CD1 ~ P <0.01), but there was no significant effect on the expression of CD4 ~ + CD25 ~ + Foxp3 ~ + in spleen cells. Conclusion Estrogen prophylactic treatment can delay the onset of EAE in mice. The mechanism may be related to up-regulation of PD-1 / PD-L1 pathway and up-regulation of regulatory B cells.