实验性变态反应性脑脊髓炎小鼠胸腺中MOG特异性胸腺细胞分化机制探讨

The study on the differentiation mechanism of MOG-specific thymocytes in the thymus of experimental autoimmune encephalomyelitis mice

为观察实验性变态反应性脑脊髓炎(experimental autoimmune encephalomyelitis,EAE)小鼠胸腺细胞的变化,探讨其相关分化机制,选择C57BL/6小鼠构建EAE模型,并根据EAE病程分为4组(EAE诱导后第0、5、10和15天);使用计数板记录4组小鼠的胸腺细胞数,并记录临床评分分别为0、1、3.5的EAE小鼠和PBS对照小鼠的胸腺细胞数;应用FACS分析EAE诱导后第0、5、10和15天的胸腺细胞亚群——双阴性(double negative,DN)细胞、双阳性(double positive,DP)细胞、CD4单阳性(CD4 single positive,CD4SP)细胞、CD8单阳性(CD8 single positive,CD8SP)细胞的比例,并对第10天小鼠胸腺细胞亚群进行凋亡分析。结果显示:与第0天比较,胸腺细胞数在EAE诱导后第5天升高(P<0.01),之后第10天和第15天逐渐降低(P<0.05);EAE小鼠的临床评分越高胸腺细胞数越低(P<0.05);EAE诱导后第10天胸腺细胞凋亡比例最高,胸腺细胞亚群中DP细胞的凋亡比例最高;4组小鼠DN、CD4SP、CD8SP细胞比例逐渐升高,而DP细胞比例逐渐降低。这提示EAE诱导后胸腺细胞增殖分化进程加快,可能机制为在疾病发生之前髓鞘少突胶质细胞糖蛋白(myelin oligodendrocyte glycoprotein,MOG)抗原对于潜在分化成MOG特异性T细胞的DP细胞的阳性选择有加速作用,这些细胞最终迁移至致病部位,使患者表现出临床症状。

In order to study the changes of thymocytes in experimental autoimmune encephalomyelitis(EAE)mice and explore its related differentiation mechanism,we constructed EAE model in C57 BL/6 mice and divided them into four groups(0,5,10 and 15 days after EAE induction)according to the course of EAE;The thymocytes of four groups were recorded by blood counting board,and the thymocytes of EAE mice with clinical score 0,1,3.5 and PBS control mice were also recorded;FACS was used to analyze the proportion of thymocyte subsets:double negative(DN),double positive(DP),CD4 single positive(CD4 SP),CD8 single positive(CD8 SP)on day 0,5,10 and 15.The apoptosis of thymocyte subsets in mice on day 10 was analysed.The results showed that,compared with on day 0,the number of thymocytes increased on days 5(P<0.01),and then decreased gradually on days 10 and 15 after EAE induction(P<0.05).The higher the clinical score,the lower the thymocyte count(P<0.05);The apoptotic proportions of overall thymocytes and the DP subset were the highest on day 10;The proportions of DN,CD4 SP,CD8 SP cells increased gradually,while the proportion of DP cells decreased gradually in the four groups.These data suggest that the process of proliferation and differentiation of thymocytes after EAE induction is accelerated,and the possible mechanism is that myelin oligodendrocyte glycoprotein(MOG)antigen accelerates the positive selection of DP cells that potentially differentiate into MOG-specific T cells before the onset of the disease,and these cells eventualy lmigrate to the pathogenic site and cause clinical symptoms.