Treatment Of Traumatic Brain Injury In Rats With Intravenous Administration Of Human Umbilical Cord Blood Mesenchymal Stem Cell

Object: It is observed that transplantation of HUCBMSCs(Human umbilical cord blood mesenchymal stem cell)for treatment of traumatic brain injury(Traumatic brain injury)in rats, through the Neurological Severity Score Points,the difference expression of NGF(Never growth factor),BDNF (Brain-derived neurotrophic factor) and apoptotic cell number, so that influence injury partial NGF, BDNF expression.Finally, it affect the local micro-damage environment and a reduction in apoptosis, and promote the recovery of neurological function.Methods:Taking the forth generation HUCBMSCs, which are marked by Brdu(5-bromo-2-deoxyuridine) ahead 72 hours before implantation.The marked cells is detected the cell marker rate by immunocytochemical, is tested the cell activity using trypan blue exclusion experiment. 90 healthy male Sprague-Dawley rats were divided randomly into sham-operated group(30), Injury control group(30),treated group(30).The three groups were divided randomly into post inject 3d,7d,14d,21d,28d groups,every group had 6 rats. It is the improved methods of production of free-falling Feeney that is adopted for the model of TBI.Successful model for 24 hours, the mice of treated group were injected 3×106个HUCBMSCs which are solved in 1ml PBS by mouse tail vein,and the mice of sham injury group and the control group were injected the equal volume of PBS solution.All mice were not using immunosuppressive agents.Using HE staining, immunohistochemistry, in situ hybridization on the injury in rats at different time points after injection,in order to observe the morphological changes, Brdu, BDNF and NGF factors, as well as the apoptosis.In addition, the rats are evaluated by Neurological Severity Score Points before killed at every time points. The HUCBMSCs from Chinese Academy of Medical SciencesInstitute of Hematology are the second generation. The adherent cells were spindle to form large flat fibroblast morphology, and cell plasma boundaries clearly, the nuclear center, there is 1 ~ 2 nucleoli, 80 % cell fusion, was arranged swirl. Taking the forth genertation cells ,which are incorporated Brdu,it is showed that 80 ~ 90% cell express Brdu positive by immunohistochemical staining.So it indicate 80 ~ 90% HUCBMSCs were marked.Testing the remaining cell lines after transplantation by trypan blue exclusion, it is showed that more than 90% cell survival.By Neurological Severity Score Points,the neuroal improvement of the rats in contral group is superior to the treatment group from the 7 day after adminstration of HUCBMSCs.The sham injury group structure of rat brain cells is in normal;Injury damage to the cortex of rats in control group shows that the organizational structure of damage, loss, neuronal degeneration and cell body shrinkage, there is glial cells and inflammatory cell infiltration at the trauma surrounding. The treatment group is less change than the the control group.The cells labeled Brdu which gathered obviously damage zone and its surrounding areas and survival can be seen in the transplantation group. However, with time, the cells labeled Brdu gradually reduced.Only a very small number of nerve cells were TUNEL positive in the sham injury group. Substantial TUNEL staining positive cells were seen in the surrounding brain damage area following traumatic brain injury and there is significant difference comparing with sham injury group(P<0.05) .TUNEL-positive cells significantly decreased in transplantation group in surrounding brain injury areas, but it is significantly higher than the injury control group(P<0.05) .Over time, the number of TUNEL-positive cells showed a downward trend in the injury control group and the transplantation group.Only a very small number of nerve cells were BDNF,NGF positive in the sham injury group. Substantial BDNF,NGF positive cells were seen in the surrounding brain damage area following traumatic brain injury, about 14 days after injected,followed by a gradual decline ,and there is significant difference comparing with sham injury group(P<0.05) . BDNF,NGF-positive cells significantly increased in treated group in surrounding brain injury areas, but it is significantly higher than the control group(P<0.05) ,about 14 days after injected, followed by a gradual decline in 21,28days,but it is still higher than the control groupConclusion:1. HUCBMSCs can migrate to damaged areas by intravenous.2. Unused immunosuppressants, HUCBMSCs can survive for at least four weeks in the healthy immune function in xenogeneic (rat) brain, and rats did not happen transplant-related deaths.3. The transplantation of HUCBMSCs improve the neuron which in the surrounding brain damage area,increase the expression of BDNF,NGFand BDNFmRNA, as well as ,decrease the apoptosis of neuron.