重组腺病毒转导的MSC导向的CNTF基因靶向性治疗MS对髓鞘和轴突的保护作用

目的:探讨重组腺病毒转导的骨髓间充质细胞导向(MSC)的睫状神经营养因子(CNTF)基因靶向性治疗多发性硬化对髓鞘和轴突的保护机制.方法:先构建、扩增、纯化Ad-CNTF-IBES-GFP,在体外培养MSC细胞.将1×108Ad-CNTF-IRES-GFP转染MSCS1×106,测定上清液中CNTF的浓度.再用MOG 35-55建立C57BL/6小鼠EAE模型,将Ad-CNTF-IRES-GFP转染的MSC移植治疗EAE小鼠,观察其病情评分;免疫荧光观察其病变部位的外源性MSO数量;免疫组化观察少突胶质细胞前体细胞(OPC)-NG2( )细胞在病变部位的数量,Western blot和免疫组化法观察神经生长因子CNTF表达,免疫组化观察促凋亡蛋白Caspase-3表达;病理观察髓鞘、轴突损伤情况;电镜观察髓鞘和轴突以及ODC损伤情况;所有的数据用(x)±s表示.采用SPSS 13.0进行统计分析.结果用多因素方差分析,P＜0.05差异有显著性.结果:MSC-Ad-CNTF-IRES-GFP治疗的EAE小鼠病情显著减轻.不仅平均发病时间缩短,发病率下降,病情减轻,而且病情严重程度也明显减轻.外源性MSC出现在EAE的脊髓病变部位.治疗后的EAE小鼠脊髓病变部位的NG2( )表达明显增加,促凋亡蛋白Caspaae-3的表达明显降低.治疗后的EAE小鼠病变部位的髓鞘和轴突变性明显减轻.结论:MSC-AD-CNTF-GFP可有效治疗EAE动物.其机制可能是:Ad-CNTF-IRES-GFP转染的MsC细胞定向去病变部位,升高其部位CNTF水平和NG2( )细胞数量,减轻其部位髓鞘、轴突及ODC损伤,减少促凋亡蛋白Caspase-3表达.

MSCs transfected by rAd-CNTF-IRES-GFP protect myelin and axon in multiple sclerosis model

Objective:To investigate the efficiency and mechanism for MSCs transfected by rAd-CNTF-IRES-GFP to protect myelin and axon in the model of multiple sclerosis:experimental allergic encephalomyelits(EAE)in C57BL/6 mice.Methods:Ad-CNTF-IRES-GFP vector was constructed with PDC316-CNTF-IRES-GFP and PBHG plasmids,and transfected into 293-LP cells and then MSCs by liposome pathway to amplify the recombinant Ad.The expression of CNTF was confirmed in vitro by immunofluorescence.MSCs transfected by Ad-CNTF-IRES-GFP ...