Transplanted Bone Marrow Stromal Cells Labeled With Feridex In Rat Parkinson's Disease Models

Parkinson's disease (PD) is a progressive neurodegenerative disorder, which is primarily characterized by degeneration of the dopaminergic neurons of the nigrostriatal pathway. Although levodopa is still considered as the gold standard of antiparkinsonian drug therapy, chronic levodopa treatment is associated with the development of adverse events in the majority of patients, such as motor fluctuations, dyskinesias, and neuropsychiatric problems. Nowadays, most of the PD research has been focused on cell transplantation, which could be a potential therapeutics.Bone Marrow Stromal Cells (BMSCs) are stem cells with the characteristics of self-renewing, multi-potency and easily expanding in vitro. Therefore, BMSCs are ideal target cells for cell transplantation. Under specific conditions, BMSCs are capable to differentiate into other cell types, such as neural stem cells, neural cells and glial cells. A study has shown that there was a functional recovery with BMSCs transplantation in a PD animal model. BMSCs are able to differentiate into neural stem cells, neural cells and glial cells and dopamine cells when they are transplanted into caudate putamen of PD rats. Our research was focused on: (1) Cultivation,differentiation and identification of BMSCs from SD rats.(2) Establishing 6-HODA PD model with simple method. (3) Transplanting BMSCs labeled with Feridex and bromodeoxyuridine (BrdU) to treat Parkinson's disease in SD rats. (4) To examine the effect of treatment with BMSCs in Parkinson's disease rat model, and explore the methods of labeling bone marrow stromal cells with Feridex and to monitor the labeled cells after transplantation into the PD rats with MRI scanning.Part I Cultivation and Differentiation of BMSCs in VitroObjective:To explore the cultivation,differentiation and identification of BMSCs from SD rats.Methods:BMSCs were isolated from SD rats by anchoring culture.The proliferative characteristics were observed in primary and passage cultures. High purified BMSCs were selected and induced into neurons through the neurotrophic factors.The number of different immunoreactive cells was detected by nestin,NSE,GFAP and tyrosine hydroxylase (TH) immunocytochemistry.Results:The fourth generation cells were highly purified BMSCs. EGF, bFGF, BDNF and TRA might induce BMSCs into nestin,NSE and GFAP-positive cells, but no TH-positive cell was found. Conclusion:BMSCs can be amplified and purified in short time.Combination of cytokines may have synergic effects on proliferation and differentiation of BMSCs.PartⅡStudy on Injection 6-OHDA into Medial Fore Brain Bundles and Establish the Rat Model with Parkinson's DiseaseObjective:To establish a simple and effective rat model in Parkinson`s disease. Methods:80 SD rats were selected. 6-OHDA was injected into two points of medial forebrain bundles on the right sides. One burr hole was made, two injection points are–1.8mm antero-posterio to and–2mm lateral to the bregma, -8.5mm ventral to the dura mater and–1.8mm antero-posterio to and–2mm lateral to the bregma, -7.5mm ventral to the dura mater. In 6 weeks, apomorphine-induced rotation test was performed to examine the disease progress in this rat model.Further examined the lesions of substantia nigra and striatum of PD rats in vivo under MRI. Results:54 rats were induced to show obviously rolling behavior at the fourth week after lesion of substantia nigra (>7 rolls/min).The Success ratio of PD model was 75%.The lesioned substantia nigra of PD rats showed obviously low MRI signal.The low signal regions shrank in correspondence with the time extended, and disappeared at the end of the third week. Conclusions:The PD rat model gotten from bi-point lesions of medial forebrain bundles by small dose of 6-OHDA is effective with a high success rate. It's a perfect model to study PD.MRI scanning can be used to observe the lesions of PD rat model in vivo continually, which is an available means to evaluate and examine the PD rat model objectively. Part III Transplanted Bone Marrow Stromal Cells labeled with Feridex In Rat Parkinson's Disease Models by MRI measurementObjective: To examine the effect of treatment with BMSCs in Parkinson's disease rat model, and explore the methods of labeling bone marrow stromal cells with Feridex and to monitor the labeled cells after transplantation into the PD rats with MRI scanning. Methods: 45 PD rats were randomly divided into three groups: intravenous group,intra-arterial group and intracerebral group.They were transplanted with BMSCs labeled with Feridex and bromodeoxyuridine (BrdU). Another 5 PD rats were untreated group. Functional outcome measurements were performed using the apomorphine induced rotation test at 2,4,6 and 8 weeks after treatment. At the same time MRI scanning was performed to monitor the transplanted cells. After MR imaging,two rats of each group were killed and Prussian blue staining and immunofluorescent staining of the histological sections were performed. Results: A decrease of per minute numbers of apomorphine induced rotation test was noted in rats of intravenous group,intra-arterial group and intracerebral group.The decrease was more significant in intracerebral group. BMSCs colabeled with Feridex and BrdU could migrate into the lesions and differentiate after transplanted into PD rats. MRI scanning is a useful and noninvasive tool to track the location and distribution of the labeled BMSCs. Conclusions: Treatment with BMSCs may improve neurological outcome in PD rat model. Fefidex can be used to label BMSCs in vitro and MRI opens up the possibility of in vivo tracking of Fefidex-labeled BMSCs after transplantation.