超顺磁氧化铁标记骨髓间充质干细胞移植治疗帕金森病大鼠模型的组织学研究

Study of SPIO Labeled or BrdU Labeled Bone Marrow Mesenchymal Stem Cells in Parkinson's Rat Model following Transplantation

目的:研究应用超顺磁氧化铁(SPIO)和BrdU标记骨髓间充质干细胞(MSCs)移植治疗帕金森病(PD)大鼠模型,观察移植的MSCs在PD模型鼠脑内的迁移及分化,探讨MSCs移植治疗PD的可能性。方法:分离和培养大鼠骨髓间充质干细胞,一组MSCs用脂质体转染法进行SPIO标记,另一组MSCs用BrdU标记;制作PD大鼠模型,将不同标记的MSCs分别移植到PD大鼠右侧纹状体区,利用阿朴吗啡观察PD鼠的行为学变化。应用透射电镜和免疫荧光双标法观察移植后的MSCs增殖、迁徙和分化。结果:电镜显示,SPIO标记的MSCs呈类神经元形态。免疫酶标显示移植后4周和12周BrdU-MSCs组和SPIO-MSCs组的注射侧黑质区TH免疫反应阳性细胞较对照组注射侧黑质区有增多趋势,但无统计学差异。免疫荧光组化结果表明,MSCs移植后4周,在移植区可见少量BrdU和Nestin双标为阳性的神经前体细胞,12周时可见少量双标的神经细胞,未发现明显的BrdU和TH双标为阳性的细胞。移植组与对照组相比行为学(旋转次数)有明显差异(P<0.05)。结论:MSCs移植治疗能改善PD模型大鼠的行为学。MSCs进入PD模型大鼠脑内可存活并处于增殖状态,有广泛迁移并在局部微环境的作用下能向神经前体细胞和神经细胞转化,但数目较少。故其改善PD大鼠神经功能的机理值得进一步探讨。

Objective.-To investigate curative effects after the transplantation of superparamagnetie iron oxide （SPIO） and Br dU labeled MSCs in the treatment of PD models. Methods：MSCs from adult bone marrow of SD rats were cuhured, isolated and purified. Labeled MSCs was achieved with the magnetic resonance contrast agent SPIO. and Lipofectamine was used as transfection agent to achieve high transfer efficiencies. MSCs were also labed by BrdU. SPIO -labeled MSCs or BrdU labeled MSCs were transplanted to the right striatal area of 6-OHDA PD rat models. Rotational behavior was assessed in each group at 1 , 2, 4, 6 weeks following transplantation. Brain frozen sections were carried out immunofluorescene double staining. Electron micro scope was used to detect the ultra structure of MSCs. Results： Histochemical staining of brain tissue shows that BrdU labeled MSCs were moved from the transplanted points to the nearby areas. Neuron like cells were detected by Electron microscope. Some BrdU/Nestin positive cells could be found in the implanted region of BrdU-labebed MSCs at 4 week after transplanta tion, but disappeared at 12 week. At this time some BrdU/MAP 2 positive cells appeared except BrdU/TH positive culls. Conclusion：SPIO particles could label MSCs effectively, and MR1 detection of SP10 labeled cells was a promising method and novel approach to analyse the MSCs following transplantation to PD models.