| Objectives To investigate the effects and mechanism of SDF-1?secreted by hUC-MSCs on the nerve function restoration of HIBD.Methods 1.The cell-surface antigens of hUC-MSCs were detected by flow cytometry;inducing differentiations and proliferation ability of the hUC-MSCs were identificated.2.After HIBD damage for 5 days using Rice method,the hUC-MSCs were transplanted into intracerebroventricular of rats,and the equal volume of PBS was injected as its control group.One day after cell transplantation,the injured brain tissue were collected for HE staining;the h SDF-1? release in the injured hippocampus was detected by ELISA.The learning-memory function and novel object exploration capacity were respectively detected by Morris water maze test and Object-in-place task following the h UC-MSCs transplantation for 4 weeks.The levels of SDF-1?,CXCR4,Nestin,PI3K/AKT protein expressions in the hippocampus were detected by Western Blotting.Hippocampalneurogenesis were observed using Nestin and Brdu immunofluorescence double staining.3.Primary neural stem cells of rats were cultured and identified by Nestin staining.The primary neural stem cells damaged by OGD were separately cocultured with the hUC-MSCs.Meanwhile,the neural stem cells with OGD treatment were served as its control group(OGD group),and hSDF-1 antibody was added to neutralize hSDF-1? in the coculturing medium.The levels of hSDF-1? secretion were tested in the medium of the three groups by ELISA.The proliferation changes of the injured neural stem cells were detected by a CCK-8 kit.The levels of PI3K/AKT protein expressions were detected in the OGD neural stem cells by Western Blotting.Results 1.More than 95% of hUC-MSCs exhibited CD44+,CD105+,HLA-ABC+,HLA-DR-and CD45-.The hUC-MSCs can be differatiated into osteogenic,adipogenic and chondrogenic.2.Following the hUC-MSCs transplantation,the cells in the hippocampus were more closely arranged,the cell swelling,nuclear condensation and vacuoles were significantly improved compared with those of the PBS group.During the 2-5 day's hidden platform training,the spending time to find the platform was less in the h UC-MSCs group than that of the PBS group(P<0.001).On the probe trial tests(Day 6),the times of crossing the platform quadrant were increased in the hUC-MSCs group compared with that of the PBS group(P=0.0023).And the rats transplanted by hUC-MSCs spend much moretime to explore new things than that of the PBS rats(P=0.0016).Furthermore,with the hUC-MSCs transplantation,the expression level of Nestin protein in hippocampus was higher than that of the PBS group(P<0.0001)and more newborn neural stem cells were found in the hippocampus(P=0.0229).The level of hSDF-1 ? secretion in the hippocampus was increased(P=0.0096),and the expression levels of SDF-1? and CXCR4 were significantly increased in the h UC-MSCs rats(P=0.0022;P=0.0287).Further study found that the hUC-MSCs transplantation activated the expression levels of PI3 K,AKT and p-AKT in the hippocampus of HIBD rats(P=0.0491;P=0.0040;P=0.0228).3.After separately cocultured with the hUC-MSCs,the level of hSDF-1? release in the coculture medium was statistically increased compared with that of OGD alone group(P=0.0021),and significantly decreas ed by the cocultured with hSDF-1? antibody treatment(P=0.036).The hUC-MSCs coculture induced the proliferation of the OGD-injured neural stem cells(P=0.0011),which was reduced by the hSDF-1 antibody treatment compared with the hUC-MSCs coculture group(P<0.0001).Morevoer,the levels of PI3 K and p-AKT expressions in the OGD-injured neural stem cells were obviously increased following separately cocultured with the hUC-MSCs(P=0.0133;P=0.0044),and were decreased by the hSDF-1antibody treatemtn in the hUC-MSCs coculture group(P=0.0109,P=0.002).Conclusion The hUC-MSCs transplantation enhances hippocampal hSDF-1? secretion to increase number of new born neural stem cell in DG of hippocampus,which mechanism maybe upregulate the levels of SDF-1?and CXCR4 protein expression to activate the PI3K/AKT signaling pathway and induce neural stem cells regeneration. |
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