Stereotaxic Transplantation Of Bone Marrow Mesenchymal Stem Cells In Treatment Of Cerebral Ischemia Reperfusion Injury In Rats

Objective:To observe the effects of bone marrow mesenchymal stem cells (BMSCs) transplantation for treatment of cerebral ischemia reperfusion injury in rats and investigate its possible mechanism.Methods:BMSCs were isolated using density gradient centrifugation combined with adherent screening method and labeled by10mg/L of BrdU.The male SD rats were grouped randomly into transplantation, PBS and sham transplant groups after MCAO model was formed by suture occluded method.7days after injury, rats in transplantation group were given BMSCs stereotactic transplantation into the striatum on the damaged side. Rats in PBS group were given the same volume of PBS and nothing was given though stereotactic transplantation to sham transplant groups.Evaluate their moving ability by mNSS scoring3d and lw(before transplantation)after injury, and lw,2w,3w,4w,2m after therapy. The learning and memory ability of rats were evaluated with Morris Water Maze2w after the model.Rats were sacrificed in2month after injury to the immunohistochemical staining including BrdU single-staining,Brdu+Nestin、Brdu+MAP-2、Brdu+GFAP、 Brdu+vWF double-staining,MPO and CD11single-staining. BMSCs for transplantation were observed the survival and differentiation in host brain by Immunohistochemical staining.Results:(1) By the way of density gradient centrifugation combine with adherence screening method to separate BMSCs. The separated cells was detected by flow cytometer, result display the cells express CD90、CD71, did not express CD45、CD3、 CD14which is the surface markers of hematopoietic lineage cells,according with the characteristics of BMSCs, such as the form and cell-surface marker.The cells possess favourable self-renewal ability and multi-directional differentiation potential, confirm the cells we separated is BMSCs.(2)The rats developed typical symptoms of neurological impairment after they were subjected to MCAO. TTC staining shows ischemic regions were pale,which proved the model is success.(3)The mNSS score of transplantation group was better than control(P<0.05).(4) The escape latency of hidden platform in the transplantation group were obviously decreased as compared with PBS and sham transplant groups group(P<0.05).The number of crossing the platform was increased obviously(P<0.05).(5)BrdU+cells and Immunohistochemical double staining positive cells(Brdu+Nestins、Brdu+MAP-2、Brdu+GFAP、Brdu+vWF) were seen in the injury center and surroundings of rats in transplantation group,there were no CD11and was MPO immunohistochemical single-staining positive cells.Conclusion:BMSCs could be separated and amplification in vitro. BMSCs stereotactic transplantation can survived and differentiated in the host brain. BMSCs transplantation is able to improve the recovery of the neural function of ischemic brain injury rats. The mechanism may be related to the transplanted cells differentiating into neuron-like and glial-like cells, secretion or promoting the secretion of neurotrophic factor of host cells and angiogenesis.In this study,no obvious immune rejection cells were found, it has high safety.