The Experimental Research On Transplantation Of NSCs Into Brain Of The Rats With Cerebral Hemorrhage And The Expression Of VEGF, FN And LN After Transplantation

Objective: To realize the biological characteristics about growth, proliferation and differentiation of neural stem cells(NSCs) from rats cultured in vitro into. By transplanting neural stem cells cultured in vitro into brain of the rats with cerebral hemorrhage, to observe the refection about impaired nervous function of rats, to investigate pathological changes at focal brain tissue, and survivorship, migration and proliferation of transplanted cells, to realize the expression of vascular endothelial growth factor (VEGF), fibronection(FN) and laminin(LN), then to research the reparation about transplantation of neural stem cells towards nervous function after cerebral hemorrhage and the effect of changes about VEGF, FN and LN on neural circuit reestablishment.Methods: The animal model of rats with cerebral hemorrhage was duplicated by injecting collagenase into globus pallidus(GP). The neural stem cells were isolated from brain of rats, then were cultured in vitro, transfered of culture and identified by immunocytochemistry. Bromodeoxyuridine(BrdU) was incorporated into DNA to mark neural stem cells. Experimental rats were divide into model group(group A), transplantation group(group B), sham operation group(group C) and normal control group(group D). In group B, neural stem cells with marker were transplanted into brain of the rats with cerebral hemorrhage. At the 1st day, the 7th day, the 14th day and the 30th day after operation, nervous function defection and refection of rats was observed in each group by Longa EZ score, brain tissue of each group were extracted, pathological changes of each group's brain tissue were observed after hematoxylin-eosin(HE) staining, immunohistochemistry staining was used to detect neurocytes in brain tissue of each group, to determine the expression of VEGF, FN and LN of each group on hematoma (or impairment and transplantation) peripheral zone and normal tissue zone and to observe the expression and distribution of BrdU in transplantation group.Results: (1) Neural stem cells of neonatal rats could keep on propagating and integrate into nerval sphere in the defined medium containing bFGF and EGF, and could express positive reaction of nestin by immunohistochemistry staining. (2) After injecting collagenase into globus pallidus, the rats appeared some degrees of nervous function defection but could recuperate gradually. In sham operation group, only a few rats appeared slight nervous function and recuperated rapidly. In transplantation group, the nervous function defection of rats also recuperated rapidly after transplantation, and the Longa EZ score were better than model group at the 7th day and the 14th day(P<0.05). (3) In model group and transplantation group, typical haematoma and relative pathological changes such as hydrops, apomorphosis, necrosis cellular proliferation and could be observed from brain tissue. In sham operation group, pathological lesion were slight. The morphous and distribution of GFAP+ cells and NF-L+ cells changed because of haematoma, lesion and transplantation. At the 1st day, nestin+ cells around hematoma zone and transplantation zone slightly augmented in transplantation group, but nestin+ cells were normal in model group and transplantation group at the the 7th day, the 14th day and the 30th day. (4) Comparing with model group, sham operation group and normal control group, the expression of VEGF,FN at the 7th day, the 14th day, the 30th day and the expression of LN at the 7th day, the 14th day in transplantation group were higher (P<0.01). There were BrdU+ cells around hematoma zone and transplantation zone in transplantation group.Conclusion: Transplantation of neural stem cells cultured in vitro into brain of the rats with cerebral hemorrhage could enhance the refection about impaired nervous function of rats. The transplanted neural stem cells could survival, proliferate, migrate and differentiate locally. The expression of VEGF, FN and LN increased after transplantation of neural stem cells(NSCs) into brain of the rats with cerebral hemorrhage, which perhaps provide condition for reparation of impairment, proliferation and migration of extrinsic source neural stem cells and reestablishment neural circuit.