| Objective:To investigate whether the level of VA in vivo can regulate the Wnt/?-catenin signaling pathway through RA signaling pathway,and then affect the proliferation of neural stem cells in the hippocampus after HIBD,thereby affecting the prognosis of nerve tissue function.Methods:The model of VA intervention was divided into four groups:VAS,SVAS and VAT.The experimental rats of four groups began a continuous Morris water maze experiment about 5 days from 28d to 32d after damage.Hippocampal tissues of four groups were collected with EDU proliferation staining and Nestin antibody markers in 3d and 7d after damage.PCR and Western blot were used to detect the mRNA and protein level of RAR?,Akt,p-Akt,GSK3?,p-GSK3?,?-catenin,and CyclingD1 in3d and 7d after damage.The neural stem cells were isolated from hippocampus of fetal rat and were identified by cytochemical staining with Nestin antibody.Neural stem cells were interfered with RA concentrations and oxygen glucose deprivation injury(oxygen glucose deprivation,OGD).CCK8 cell proliferation was used to detect the proliferation rate at different time after injury.PCR and Western blot were used to detect the mRNA and protein level of RAR?,Akt,GSK3?,?-catenin,and CyclingD1 of primary neural stem after injury.Results:The results of Morris water maze show that the latency of finding platform in group SVAS is shorter than that of the other three groups,while the average latency time of finding platform in group VAD and group VAT is longer than that of group VAS and group SVAS,and the number of crossing platform in group SVAS is more than that of group VAD(P<0.05).The results of EDU proliferation detection in 3d and 7d after injury indicated that SVAS group and VAS group was higher than group VAD and group VAT.The results of gene expression for RAR?showed that group VAS and group SVAS was higher than group VAD and VAT group(P<0.05)in the hippocampus in 3d and 7d after HIBD.After HIBD in 3d the mRNA level of?-catenin,Akt and GSK3?for group VAS and group SVAS was higher than group VAD and VAT group(P<0.05),and the mRNA levels of?-catenin and GSK3?showed that group SVAS was higher than those in the other three groups.CyclinD1 after HIBD injury,the level of mRNA in hippocampus in 3d and 7d for group SVAS was higher than those in the other 3 groups(P<0.05).After ODG,the proliferation of 12h,24h,48h and 72h after hypoxia was the lowest after the intervention of 50umol/LRA(P<0.05).And 5umol/L has the highest value added(P<0.05).The proliferation of 1umol/LRA and 5umol/LRA intervention group was higher than that in simple OGD group(P<0.05).The results of PCR gene detection showed that the expression of RAR?under the intervention of 1umol/LRA,5umol/LRA and 10umol/LRA was significantly higher than that of group OGD(P<0.05).The expression of?-catenin was higher in 5umol/LRA after OGD than in group OGD(P<0.05).Cyclin D1 under the intervention of 5umol/LRA and10umol/LRA was significantly higher than group OGD(P<0.05).The mRNA level of under the intervention of 1umol/LRA,5umol/LRA and10umol/LRA after OGD was significantly higher than group OGD(P<0.05).The protein level of RAR?,CyclinD1,GSK3?and?-catenin under the intervention of 1umol/L,5umol/L and 10umol/LRA were higher than those in OGD group.Conclusions:Suitable VA can promote the proliferation of neural stem cells.It is preliminarily revealed that VA level regulates Wnt/beta-catenin signaling pathway through RA signaling pathway,and then affects the proliferation of neural stem cells in HIBD inferior hippocampus,thereby affecting the prognosis of nerve tissue function. |
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