Survival And Differentiation Of Neural Stem Cells From Rat Hippocampus After Transplantation Into The Adult Rat Brain

Alzheimer's disease is one of the major neurodegenerative disease that cause progressive cognitive and behavior deterioration. Pathological changes in the brain of AD patients are senile plagues, neurofibrillary tangles and the region-specific degeneration of neuronal population, particularly in the areas connected to the hippocampus and pallium. Drug therapy is widely used in clinic but without long-term effects. Neural transplantation might be an essential therapeutic method for curing AD. However, because of the lack of autografts donor tissues and the immune responses by allograft donor tissues which destroy allograft donor cells, this way is not satisfied either. Neural stem cells(NSCs) is a kind of stem cells with the potential to self-renew and to generate astrocytes, oligodendrocytes and neurons. These characters make it a perfect donor cells for transplantation. In central neural system of adult mammals, NSCs mainly exist in the subventricular zone(SVZ) of lateral ventricle and the dentate gyrus(DG) of hippocampus. About neural stem cells from the dentate gyrus(DG) of hippocampus, the outcomes only focus on mechanisms of development and differentiation in vitro. It is meaningful task to study the differentiation of it in vivo, especially in the region of hippocampus which is a important pathological region of AD.This study choose the NSCs from dentate gyrus of the hippocampus in newborn rats as the donor cells, which were grafted into CAi region of hippocampus in AD model rats. Hoechst33258-label technique was applied to detect the survival and migration ofNSCs in vivo for three different times(2,4,6 weeks after NSCs grafting). Then double-label technique(Hoechst33258-label technique and immunofluorescence technique) was applied to detect the differentiation of NSCs into astrocytes and neurons, in vivo, especially into these neurons expressing ChAT⁺ or Glu⁺. The differentiation ratios of those cells were also observed. Meanwhile, behaviral tests were applied to examine the curative effect of grafting NSCsMaterials and MethodNscs isolated from the dentate gyrus of the hippocampus were cultured and labeled Hoechst33258 at the second passage and then transplanted into the hippocampus of AD model rats, which is established by injection of A β_1-40.behavioral test of AD models rat: Health male SD rats were divided into 5 groups by random: normal group, AD group, 2-week-transplantation group, 4-week-transplantation group and 6-week-transplantation group. Y-maze was used to evaluate the learning and memory ability of 5 group rats.Morphologic test of grafted cells: The brains of 2, 4, 6-week-transplantation-group rats were made freezing slice, which were observed straightly to make sure the survival and migration of the grafted NSCs with Hoechst33258. Then the section stained by Hoechst33258 would be carried immunofluorescence procedure with antibody against NF-200, GFAP, ChAT, as well as Glu to identify the differentiation of neural stem cells in the host brain.Results1. Behaviral testThe numbers of learning in normal group, AD group, 2,4,6-week-transplantation group were 34.83 + 7.59, 98.17±1.83, 39.50±3.62, 38.33+5.87, 37.17±5.91; the number of memory in these groups were 18.33 + 4.18, 77.17 + 8.00, 21.50±3.49, 20.50+4.62, 20.33 + 4.91. Result showed that, as comparing with normal rats, the ability of learning and memory in AD rats were decreased remarkably(P<0.05) while comparing with AD rats, this ability of that in 2,4,6-week-transplantation groups enhanced obviously(P<0.05). But, there were no significant difference betweentransplantation groups and normal groups (i>>0.05) and there were no significant difference among three transplantation groups either (P>0.05) .2. Survival and migrate of NSCs in the host brainThe grafted cells distributed in niddle-path, the point of injection and move to opposite directions along hippocampal gyms. And migration distance was increased with the time of transplantation (the fastest distance of migration was 4, 6, 7mm respectively after transplantation for 2, 4, 6 weeks). The most grafted cells survived well.3. Differentiation of NSCs in the host brainWhile 2 weeks after transplantation, grafted NSCs did not expressed GFAP antigen, NF-200 antibody and so on. After transplantation for 4, 6 week, in the graft, many cells expressed GFAP antigen, a few cells expressed NF-200 and few cells express ChAT, Glu antigen, suggesting grafted Nscs differentiate.Conclusion1. The grafted Nscs can be not only alive in the brains of AD model rats but also differentiate into neurons, especially these neurons: cholinergic neurons and glutamate neurons.2. Transplanting NSCs can canameliorate the spatial memory of AD rats.