The Experimental Study For The Therapy Of Engrafting Hypoxic Preconditioning HOM-MSCs Into PD Model Rats

OBJECTIVE: Through the experiment of transplanting human olfactory mucosa mesenchymal stem cells into the rats with Parkinson's disease to investigate the effects of rats' behavior and the survival,distribution and direction of hOM-MSCs after cells transplantation.Thus,in order to explore the effect of hOM-MSCs transplantation on PD rats and the effect of hypoxic preconditioning.METHODS:(1)All animals were anaesthetised via intraperitoneal(i.p.)injection of chloral hydrate on concentration of 10%and fixed on the stereotaxic instrument.The injection of 6-hydroxydopamine(6-OHDA)solution were performed by microinfusion pump with a continual speed.The solution was injected into the left medial forebrain bundle(MFB)using a 10?L Neuro Syringe(Shenzhen,China)with a 33-gauge needle.Rats,which turns to right over six times per minute,were selected and randomly divided into three groups: saline group(A),hOM-MSCs normoxic group(B),hOM-MSCs hypoxic preconditioning group(C)and each group contains 10 rats.(2)Cells were selected to transfect the green fluorescent protein lentivirus vector(GFP-lentiviral vector),to detect the transfection and expression of green fluorescent protein lentivirus vector with fluorescence microscope while cells have suitable density at fourth passage.(3)Rotation test were performed to define the successful PD model's rats that cycling over six times per minute,to count the total cycles in 10 minutes.(4)Nuero-Syringe was connected with a microinfusion pump to transplant /engraft hOM-MSCs that were pretreated into the contralateral corresponding position of the injured /lesioned area(model area)under the assistance of the rat skull stereotactic orientation.(5)Rats' behaviors were evaluated on the second day after to establish PD model,the first week after engraftment,and the fourth week after engraftment,and to count total cycles in 10 minute.(6)All rats were sacrificed for pathological examination or immunohistochemical examination after the last behavioral evaluation was completed 4 weeks after transplantation.RESULTS:(1)Green fluorescent protein lentivirus vector(GFP-lentiviral vector)can reliably transfect and stably express green fluorescent protein(GFP)in hOM-MSCs.(2)Green fluorescent protein lentivirus vector(GFP-lentiviral vector)labeled cells is a reliable tracer method.(3)HOM-MSCs still survived in the rat's brain after 4 weeks of transplantation and migrated to injured area.(4)Rat's behavior disorders have been enhanced after cells engraftment.(5)HOM-MSCs can differentiate into neuron in rat brain after transplantation.(6)Hypoxic preconditioning can promote the differentiation of hOM-MSCs into specific neurons after transplantation,significantly improved the behavior of PD rats after transplantation.CONCLUSIONS:(1)OM-MSCs are able to migrating,to survival and to differentiate into neurons after cells transplantation in PD rats.(2)Transplanting OM-MSCs is an effective therapeutic method for PD model rats,what's more,it will have more significantly effective effects if cells have been hypoxically predicted before engrafting.(3)Hypoxia preconditioning promotes the differentiation of hOM-MSCs into dopaminergic neurons after transplantation.