The Improvement Of Intracerebral Transplantation Of Neural Stem Cells Combination With Microglial Cells In Parkinson Disease Model

Objective:1. Neural stem cells (NSCs) were combined with microglial cells in the cell-transplant therapy in order to investigation the therapeutic effect of locomotor function in PD2. RT-PCR and fluorescence immunoassay results were analyzed for the mechanism of molecular biology experiment improvement.Methods:1. PD-model-rats were established by right intracerebral stereotaxic injection of 6-OHDA into rat medial forebrain bundle (MFB) and caudate putamen nucleus (Cpn) region. Apomorphine induced rats that can rotate more than 7rpm were regarded as prosperity models.2. The NSCs were come from the embryo that confirmd 14 days of gestation. The brain is removed, and then brain stem were pinched off from the body.3. Microglial cells were collected from SD rats of neonatal 1～2d. The microgial cells were cultured mixed for 7-9d, then the cells were separation and purification.4. NSCs were cultured in vitro with microglial cells and they were prepare to be translation.PD-model-rats were randomly devided into NSCs combination with microglial cells and contral group.5. Adjusting steps test and vibrissae-elicited forelimb placing test were used to excessive bracing reactions and PD-model-rats’ control.6. And the expression of TH was analyzed by immunofluorescence histochemistry and RT-PCR.Results:1. There were 216-OHDA stereotactic injection PD rats established successfully. The success rate of models were 70%.20 of them were selected randomly and divided into experimental group and control group, each of them were 10 PD model rats,6 ones were used for behavioral tests, and the others were prepaired for molecular biological detection.2. NSCs became to neural spheres after 7 days’cultivation in vitro. And the results of dififerentiation cells immunofluorescence showd positive in GFAP and TH.3. Microglia were determinated by CD11b fluorescence immunoassay and cellular purification of microglia was more than 95%.4. Microglia promoted the NSCs differentiation into TH positive cells.5. Adjusting steps test and vibrissae-elicited forelimb placing test shows the improvement of motion integration in the NSCs combination with microglial cells group (P<0.05). Apomorphine induced rotation were reduced by the cells transplantation therapy (P<0.05)6. PD-model-rats’ TH expression were up regulation to the control group.Conclusion:1. NSCs can differentiate into neurons and glial cells. NSCs cultured with microglia cells can promote the differentiation of NSCs to DA neurons in vitro.2.6-OHDA model rats transplanted of NSCs and microglia can improve the motor function of PD rats.3. Transplantation of neural stem cells combination with microglial cells improved PD models’motion function and the proofs could be confirmd by immunofluorescence histochemistry and RT-PCR.