异体骨髓间充质干细胞对EAE大鼠神经再生保护作用的研究

目的观察异体骨髓间充质干细胞(BMSCs)移植后在EAE大鼠体内存活、增殖、分化,探讨BMSCs对EAE的神经再生的修复作用及迁徙分化情况。方法 SD雌性大鼠45只,随机分为3组,其中EAE模型组20只,正常对照组5只,EAE模型BMSCs移植组20只。重组免疫蛋白MBP68-86诱导建立EAE的动物模型,在免疫后早期第5天静脉输注Wistar大鼠的BMSCs(4×106/只,0.4mL)。观察输注后大鼠神经功能变化和组织学变化,利用免疫组化方法检测不同时间脑脊髓神经元特异性烯醇化酶(NSE)、胶质纤维酸性蛋白(GFAP)、2’,3’-环腺苷酸-3’-磷酸二酯酶(CNPase)表达,观察BMSCs移植后EAE大鼠体内神经元、星形胶质细胞和少突胶质细胞数量。结果 EAE大鼠免疫后早期移植BMSCs,神经功能评分无论是高峰期评分还是平均评分显著低于EAE模型组。BMSCs移植组在第7天NSE阳性细胞数较EAE模型组表达水平增多,但并无显著性差异,而第15天、第21天和第30天NSE阳性细胞数明显高于EAE模型组的表达水平(P0.05)。BMSCs移植组各时间点CNPase染色阳性细胞数明显高于EAE模型组的表达水平(P0.05),随着时间迁移,BMSCs移植组CNPase染色阳性细胞数表达逐渐增加。BMSCs移植组各个时间点GFAP阳性细胞数与EAE模型组比较无显著性差异。病理检查显示EAE大鼠免疫后早期移植BMSCs可使脑组织炎症反应减轻,脱髓鞘病灶的数目也明显少于EAE模型组。结论预防性静脉输注BMSCs可能通过免疫调节,抑制免疫反应所致的神经细胞损伤,减少神经元和少突胶质细胞的凋亡,从而发挥神经保护作用,但不能排除仍有部分BMSCs在局部微环境信号的诱导下向脱髓鞘区迁徙并分化为髓鞘形成细胞的可能。     

Relative susceptibility of SJL/J and B10.S mice to experimental allergic encephalomyelitis (EAE) is determined by the ability of prethymic cells in bone marrow to develop into EAE effector T cells

SJL/J mice are highly susceptible to actively induced experimental allergic encephalomyelitis (EAE), whereas B10.S mice are resistant. However, both strains share the H-2^s haplotype. We have previously shown that the relative susceptibility of SJL/J and B10.S mice to acute EAE correlates, respectively, with high and low responsiveness to myelin basic protein (MBP), as determined by cloning and limiting dilution analysis of in vitro T cell proliferation. Here, we have investigated the ability of SJL/J and B10.S mice to generate EAE-effector T cells in vivo. We have developed a new mouse strain, B10.S Thy 1.1, that differs at the Thy 1 locus from SJL/J and B10.S mice (both Thy 1.2) but has the same MHC and resistance pattern to EAE as do B10.S mice. Using radiation bone marrow chimeras formed between SJL/J and B10.S Thy 1.1 mice, we have shown that a population of radiosensitive prethymic cells in SJL/J bone marrow has an intrinsic potential to generate EAE-effector T cells, whereas that in B10.S Thy 1.1 bone marrow does not. This lack of detectable EAE effector cells in B10.S Thy 1.1 mice does not appear to be due to the generation of suppressor T cells or to a detect in antigen-presenting cells. Moreover, the potential of SJL/J bone marrow to generate EAE-effector T cells is not inhibited by the concomitant presence of B10.S Thy 1.1 bone marrow cells, thymocytes or dendritic cells in mixed chimeras. Hence, the relative susceptibility of SJL/J and B10.S mice to EAE appears to be directly related to the respective responder status of their T cells to MBP, as evidenced by their ability (or inability) to generate EAE-effector T cells. This high and low responder status appears in turn to be linked to non-MHC background genes, although this has not been established formally.     

回药扎里奴思方联合BMSCs移植对脑缺血再灌注大鼠BBB紧密连接蛋白的影响

目的观察扎里奴思方联合BMSCs移植对MCAO模型大鼠BBB上Occludin和Claudin mRNA表达的影响。方法 250只SD大鼠随机分为假手术组、模型组、扎方组、移植组和联合组,除假手术组10只外,其余各组15只;线栓法制备MCAO模型,体外全骨髓贴壁法培养及扩增BMSCs;大鼠灌胃给药[14.6 g/(kg·d)],BMSCs悬浮液经颈内动脉移植入脑(2×106/200μl);移植后1 d、3 d、7 d、14 d取材,干湿重法检测脑含水量,Real timePCR技术检测Occludin和Claudin mRNA表达。结果模型大鼠脑含水量较假手术组增加(P0.01),Occludin和Claudin mRNA表达降低(P0.01);与模型组比较,扎方、移植及联合各3 d、7 d、14 d组脑含水量降低(P0.01),各组各时间点Occludin和Claudin mRNA表达增高(P0.01);与移植组比较,扎方1 d组脑含水量降低(P0.05),3 d组Occludin mRNA表达增高(P0.01),7 d组表达降低(P0.01),扎方1 d组Claudin mRNA表达增高(P0.05),7 d、14 d组表达降低(P0.01),联合各组脑含水量均降低,以1 d、14 d明显(P0.01),各组Occludin和Claudin mRNA表达均增高(P0.01);扎方与联合组比较,联合各组脑含水量降低,以7 d、14 d组明显(P0.01,P0.05),Occludin和Claudin mRNA表达增高(P0.01);同组间比较,均以7 d组变化显著,脑含水量呈先增后减趋势,Occludin和Claudin mRNA表达呈先减后增趋势,14 d有明显改善(P0.01)。结论脑缺血再灌注损伤后BBB受损,通透性改变,脑水肿形成,以7 d最为明显;扎里奴思方和BMSCs移植均可不同程度改善脑缺血后脑水肿程度,以二者联合应用作用显著,其机制可能与干预Occludin和Claudin mRNA动态表达有关。