骨髓间充质干细胞干预治疗实验性自身免疫性脑脊髓炎大鼠的研究

Study of intervention treatment of bone marrow mesenchymal stem cells for experimental autoimmune encephalomyelitis in rats

目的 探讨骨髓间充质干细胞（BMSCs）干预治疗实验性自身免疫性脑脊髓炎（EAE）大鼠的效果及其机制,以及最佳的治疗时机.方法 36只健康雌性SD大鼠随机分为正常对照组、EAE模型组和免疫后当天、第5d、第10 d、第15 d注入BMSCs干预治疗组（d0、d5、d10、d15 BMSCs组）.用MBP68-86作免疫原制作EAE大鼠模型,分别在免疫后相应时间点静脉注入Wistar大鼠的BMSCs或生理盐水.观察大鼠的发病状况、神经功能缺损评分（NDS）和脑、脊髓组织病理改变.用流式细胞仪检测周围血T细胞亚群,ELISA法检测血干扰素-γ （IFN-γ）、肿瘤坏死因子-α（TNF-α）、转化生长因子-β1（TGF-β1）、白介素（IL）-4和IL-10水平.结果 正常对照组大鼠均正常.与EAE模型组比较,d0 BMSCs组和d5 BMSCs组大鼠的发病数减少,发病时间延迟,高峰期和平均NDS降低,外周血CD4＋T细胞明显降低,Treg细胞明显升高,IFN-γ和TNF-α水平明显降低,IL-10水平明显升高（P ＜0.05～0.01）,脑、脊髓组织的病变明显减轻;d10 BMSCs组CD4 ＋/CD8＋比值明显降低（P＜0.05）.结论 预先用BMSCs干预治疗EAE大鼠有显著的效果,其机制可能为BMSCs通过上调IL-10表达来调节T细胞的增殖活化及功能,抑制异常免疫反应.而在发病后给予BMSCs则无明显效果.

Objective To explore the effects and its mechanism,and the optimal time of intervention treatment of bone marrow mesenchymal stem cells （BMSCs） for experimental autoimmune encephalomyelitis （EAE） in rats.Methods Thirty-six SD female rats were randomly divided into normal control group,EAE model group and BMSCs treatment groups with BMSCs injectted at right soon,5 d,10 d,15 d of immune （d0,dS,d10 and d15 BMSCs groups）.For induction of EAE,SD rats were immunized with MBP68-86,and BMSCs of Wistar rats or water were administered intravenously in post-immunization.Throughout the study,rats were evaluated for neurological defect score （NDS） of active disease and histopathologic examination.The expressions of T cells in peripheral blood were analyzed by flow cytometric analysis.The concentrations of interferon-γ（IFN-γ）,tumor necrosis factor-α （TNF-α）,transforming growth factor-β1 （TGF-β1）,interleukin （IL）-4 and IL-10 in peripheral blood were determined by ELISA.Results Rats in normal control group were all normal.Compared with EAE model group,the onset rats were reduced,the onset time was delayed,the peak and mean NDSs were significantly decreased,the expression of CD4 ＋ T cells in peripheral blood was significantly decreased,the expression of Treg cells was significantly increased,the levels of IFN-γand TNF-α were significantly decreased and the level of IL-10 was significantly increased in d0 BMSCs group and d5 BMSCs group （P ＜ 0.05-0.01）,and their brain,spinal cord histologic lesions were obviously reduced ; while the ratio of CD4 ＋/CD8 ＋ was significantly decreased in d10 BMSCs group （P ＜ 0.05）.Conclusions Prophylactical using of BMSCs can be effective in treating EAE.The mechanism is that the BMSCs may increase the expression of IL-10 and then regulate the activation,proliferation and function of T cells,and inhibit the abnormal immune responses.BMSCs couldn＇t be effective after onset of EAE.