胰岛素联合骨髓基质干细胞治疗帕金森病模型鼠

The curative effectiveness of insulin in combination with bone marrow stromal cells for parkinson＇ s disease rat models

目的 观察胰岛素联合骨髓基质干细胞（BMSCs）治疗帕金森病（PD）模型鼠的疗效.方法 采用贴壁法培养、纯化BMSCs,流式细胞仪检测第3代BMSCs细胞的表面标志表达情况;将6-OHDA单侧损毁成功的PD模型大鼠21只随机分为3组,每组各7只,分别移植BMSCs 12 μl（细胞密度为1×10^5个/μl）、同数量的BMSCs＋胰岛素50 nmol/L和生理盐水12μl于模型鼠右侧纹状体.在干预后4周分别观察模型大鼠阿朴吗啡诱导的旋转行为的变化,运用免疫荧光观察每组2只模型大鼠脑内BMSCs细胞存活情况以及高效液相HPLC法检测每组5只模型人鼠脑内双侧黑质-纹状体区DA含量的变化.结果 （1）第3代的BMSCs表面标志物CD29、CD34、CD44和CD45表达分别为97.1%、0.0%、99.0%和0.5%;（2）细胞移植治疗4周后联合组旋转次数较BMSCs组（P〈0.05）和生理盐水组（P〈0.01）明显减少;（3）免疫荧光观察模型鼠脑内BMSCs细胞存活情况：BrdU阳性细胞数量胰岛素＋BMSCs组干预4周后BrdU阳性细胞数比BMSCs组明显增多（P〈0.01）;（4）HPLC结果示：胰岛素联合组模型鼠脑内黑质.纹状体区DA含量虽较其他两组增加,但差异无统计学意义（P〉O.05）.结论 胰岛素联合BMSCs移植治疗PD模型的疗效总体好于BMSCs组和生理盐水组.

Objective To observe the curative effectiveness of insulin in combination with bone marrow stromal cells （BMSCs） for parkinson＇ s disease （PD） rat models. Methods （1） From BMSCs cultivating pints of view, Adherent method was used to raise and repeatedly purify BMSCs. In addition flow cytometry was exploited to detect expression of CD29, CD34, CD44 and CD45, which all the markers exis ted on the cell＇ s surface of the BMSCs in the third generation; （2） The succeed PD rat models （n = 21） were randomizefy divided to three groups,which were employed as injected BMSCs （1.2×10^6 cells/each model） ,insulin （50 nmol/L） associates BMSCs and normal saline group （12 μl）, respectively. Four weeks later,the rotating frequency was calculated and the cell＇ s survival was tested by the immunofluores cent method. The DA level in the brain （substantia nigra and striatum） was detected by HPLC. Results （1） CD29, CD34, CD44 and CD45 BMSCs in the third generation were expressed 97. 1% ,0. 0% , 99.0% ,0.5%. （2） The rotating frequencies in the insulin group combined BMSCs was much more de creased in comparson with the BMSCs （P 〈0. 05） and the normal saline group （P 〈 0. 01）. （3） The cell＇ survival was to observe. BrdU positive cells in the group interfered by insulin combined BMSCs were much more than the BMSCs group （P 〈0. 01）. （4） DA concentration in the group of insulin combined BMSCs were increased compared to the other 2 groups, however it did not have the statistical significance （P 〉0.05）. Conclusion The curative effect of the insulin combined BMSCs interfered the rat model was better than BMSCs treated group alone. It is essencial to provide further evidence in vivo on insulin protect do paminergic neurons.