| The study and reaserch of culture and proliferation and differentiation on NSCs, as well as its occurrence mechanisms plays a significant role on ischemic cerebrovascular disease. Recently, the hippocampus is thought a important construction that is closely connected with study and retention and so on. Study discovers that rat's hippocamp contains undifferentiated neural precursor cells that can differentiate into new neurons and glials supplementing adult brain tissue under the specific condition ,and this can bring new pathway for research and cure of central lesion and degenerative. If regulating factors of NSCs'prolifiration and differentiation is knowed , it can be controlled to differentiate into one kind of cells , and greatly affluences source of neural transplantation.In this study, the hippocampal neural stem cells cultured neonate rat are researched, based on simulated cerebral ischemia and reperfusion in vitro. We investigated the acitivity, proliferation and differentiation of cultured hippocampal neural stem cells and Xue Suan Tong Injection with biochemical method and immunocytochemical technique.1 the culture and identification of Neural stem cells of neonate rats and the effect of Panax pseudo-ginseng on neural stem cellsWe selected hippocampus cells of neonate rats, and investigated the effect of Panax pseudo-ginseng on the cells proliferation by primary cell culture, passage, immunofluorescence and MTT colorimetry. The results showed that cells have Nestin expression and the basic attributes (properties) of neural stem cells which belong to the stem cells in the central nervous system. We adopted MTT colorimetry to measure the cell proliferation effects of Panax pseudo-ginseng different in concentrations at the same time point and those in the same concentration at different time points. The results showed that Panax pseudo-ginseng promotes cell proliferateion at 4.375～560μg/ml, the most at 17.5μg/ml. When the concentration is 1120～4480μg/ml, there is no significance between the treatment and the control group which possibly caused by the reason that the promoting and prohibiting effects are on the balance. Until the Panax pseudo-ginseng administrated for 24h, can it promote the neural stem cells to proliferate.2 Establishment of Ischemia reperfusion model of hippocampus neural stem cells in neonate ratsIn this study, Ischemia reperfusion model of hippocampus neural stem cells in neonate rats was erected by OGD method, and effect of OGD method on ippocampus neural stem cells cultured was investigated by MTT colorimetry.The results showed that cytoactive in different time steped up at first compared to control group after OGD,and then decreased ,and significant deviation in 4h and 6h.Cytoactive was in phase of heighten in 1h,and in stage of heighten in 2h.cytoactive was lower than control group in 4h that was obviously statistical significance in contrast to 2h(P<0.05). cytoactive was obviously lower than control group in 6h,and with extremely significant deviation ( P<0.01 ) ,which was obviously statistical significance in contrast to 4h.That showed cells were damaged by OGD and the damage of cells presented with certain regularities.3 Influence of RP on the influence of cultured hippocampal neural stem cells in vitro and effect of Panax pseudo-ginsengIn this study, we adopted the immunofluorescence double labeling and MTT colorimetry to investigate the effects of Panax pseudo-ginseng on the proliferation, differentiation and self-renew of the neural stem cells. The results shown that compared with the control group, the area density, optical density and numbers of the Nestin/Brdu,Nestin/Vimentin,Nestin/Tuj-1 positive cells in the Panax pseudo-ginseng group are much higher with significance.The MTT colorimetry also showed that compared with the control group,the OD value of model group was obviously descendent with significant difference,and compared with model group the value of model group obviously increased with significant difference.This is to say Panax pseudo-ginseng can promote the survival and proliferation of the hippocampus neural stem cells of the neonate rats in vitro, and also differentiate into neurons and glia,and also showed that Panax pseudo-ginseng protected the damaged hippocampus neural stem cells.4 Influence of RP on the influence of Ca2+ and membrane of cultured hippocampal neural stem cells in vitro and effect of Panax pseudo-ginsengIn this study,we adopted the NOVOstar to investigate the effects of Panax pseudo-ginseng on Ca2+ and membrane of cultured hippocampal neural stem cells in vitro and relation of Ca2+ and membrane and the proliferation and differentiation of the neural stem cells.The results showed ca2+ density of NSCs increased after is-4h. fore treatment of Panax pseudo-ginseng inhibited the increasing of density in NSCs.after RP,ca2+ density of NSCs had the tendency of rising,was climax in RP-2h,and began to low in RP-3h.in the same time point,the ca2+ density of NSCs in the treatment groups was all lower than the model groups,and among with significant difference(P<0.05).Membrance potential of NSCs decreased after is-4h.fore treatment of Panax pseudo-ginseng inhibited the increasing of membrance potential in NSCs. after RP, membrance potential of NSCs had the tendency of decreasing,was minmum point in RP-2h,and began to rise in RP-3h.in the same time point,the membrance potential of NSCs in the treatment groups was all higher than the model groups,and among with significant difference. The results also showed ca2+ density and Membrance potential of NSCs changed around a certain limit.This is to say Panax pseudo-ginseng achieved the protectiong role of cells by inhibiting ecto- ca2+ to inflow and endogenous ca2+ to release and changed membrane potential of cells by altering ion concentration in intra-cellular and ecto.the study also showed ca2+ density and Membrance potential of NSCs changed around a certain limit with the development and change of cell proliferation and differentiation.Bove of all, in this study we successfully isolate and culture neural stem cells and erect simulated cerebral ischemia and reperfusion in vitro;based on this, we reveal that the traditional Chinese medicine Panx pseudo-ginseng not only can promote proliferation and fifferentiation of normal cultured neural stem cells , but also protect hippocampal neural stem cells based on simulated cerebral ischemia and reperfusion in vitro, and investigate its mechanism , which provides favourable theory knowledge and significant experiment evidences to the prophylaxis and treatment of Chinese herbs for cerebrovascular diseases.
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