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Objective:To investigate the effect of Buyang Huanwutang （BHT） on proliferation and differentiation in neural stem cells （NSCs） after oxygen-glucose deprivation/reoxygenation （OGD/R） injury. Method:NSCs isolated from the hippocampus of SD rats were cultured and randomly divided into a normoxia group， a model group， a BHT group， a rapamycin （Rapa） group， and a combination group ［autophagy inhibitor 3-methyladenine （3-MA） combined with BHT］. The 20% blank serum was used in the normoxia group， and 20% BHT-medicated serum in the BHT group. The doses of Rapa and 3-MA were 1 μmol·L<sup>-1</sup> and 5 mmol·L<sup>-1</sup>， respectively. The cells were subjected to OGD/R except those in the normoxia group. The cell morphology was observed under a light microscope. NSCs were confirmed by immunofluorescence detection of nestin expression. The viability and proliferation of NSCs were assessed by cell counting kit-8 （CCK-8） assay and 5-ethynyl-2-deoxyuridine （EdU） labeling， respectively. Furthermore， Ad-mCherry-GFP-LC3B fluorescence assay was performed to investigate autophagy. The effect of BHT on autophagy-related protein expression was detected by western blot assay. Brain derived neurotrophic factor （BDNF）， <italic>β</italic>-tubulin Ⅲ， and glial fibrillary acidic protein （GFAP） were evaluated by immunofluorescence assay. Result:OGD/R significantly reduced the cell viability of rat NSCs as compared with the normoxia group. Compared with the model group， the BHT group exhibited significantly improved viability of rat NSCs （<italic>P</italic><0.01）. BHT induced the production of autophagosomes in NSCs after OGD. The BHT group showed increased expression of microtuble-associated protein 1 light chain 3Ⅱ (LC3Ⅱ） and Beclin-1 （<italic>P</italic><0.05，<italic>P</italic><0.01） and slightly changed p62 compared with the normoxia group， and significantly up-regulated LC3Ⅱ and Beclin-1 （<italic>P</italic><0.05，<italic>P</italic><0.01） and down-regulated expression of p62 （<italic>P</italic><0.01） compared with the model group. The Rapa group had similar effect as the BHT group （<italic>P</italic><0.05，<italic>P</italic><0.01）， while the combination group inhibited the activity of autophagy （<italic>P</italic><0.01）. As indicated by the results of ad-mCherry-GFP-LC3B， compared with the normoxia group， the model group showed increased fluorescence intensity （<italic>P</italic><0.01）， and the BHT and Rapa groups could further increased the fluorescence intensity of autophagy （<italic>P</italic><0.01）， while the combination group inhibited autophagy activity （<italic>P</italic><0.01）. Immunofluorescence results revealed that compared with the normoxia group， the model group displayed significantly reduced positive cells of EdU， <italic>β</italic>-tubulin Ⅲ， GFAP， and BDNF （<italic>P</italic><0.01）， and the BHT and Rapa groups exerted similar protective and promoting effects （<italic>P</italic><0.05，<italic>P</italic><0.01）， while the combination group partially blocked the neuroprotection and differentiation ability of BHT （<italic>P</italic><0.05）. Conclusion:BHT pretreatment can effectively protect rat NSCs against OGD-induced injury and promoted proliferation and differentiation by up-regulating autophagy.
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The paper uses the methods of bibliometrics and mathematical statistics to carry out statistical analysis on the scientific pa pers in Neural Stem Cells (NSCs) in 2007-2016 recorded by the SCI database from the aspects of distribution of years,countries,institutions,journals,funds and high-frequency keywords,and generally evaluates the development hotspots and application prospect of NSCs in the recent 10 years.
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Aim Toclarifytheeffectofacteosideon proliferation of neural stem cells (NSCs ) from adult mice,as well as the involved signaling pathway.Meth-ods NSCswereisolatedfromthesubventricularzone (SVZ)of adult C57BL/6 mice,then identified by im-munofluorescence staining with Nestin,the marker of NSCs.NSCs were exposed to acteoside (5,10,20,40μmol·L-1 )in absence of mitogen(EGF/bFGF)for 24 h.We employed CCK8 assay to detect NSCs viability and BrdU staining to identify NSCs proliferation.We performed Western blot to quantify the expression level ofp-AktinducedbyacteosideonNSCs.Results With-out mitogen,acteoside increased NSCs proliferation by activating p-Akt,which can be blocked by LY294002, the inhibitor of PI3K/AKT signaling pathway.Conclu-sion ActeosidepromotestheproliferationofNSCsfrom adult mice by activating PI3K/AKT pathway.
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Objective To study the effects of embryonic neural stem cells transplantion on trauma of red nucleus neu-rons of the rats with spinal cord injury.Methods NSCs in logarithmic phage were labeled with BrdU,a Sprague Dawley rat mode of spinal cord injury (SCI) was developed with electrocircuit control spinal cord injuring device.Thirty SD rats were randomly divided into three groups: sham group,SCI group and NSC group.The NSCs were trans-planted into injured site three days after SCI.Then NSCs labeled with Brdu were detected by immunohistochemisty,rubrospinal tract (RST) neurons were labeled by retrograde transport of the horseradish peroxidase (HRP) from the lesion site,which were taken by damaged axons and remained in the neurons,then the labeled red nucleus (RN) neurons were counted.Hind limb function of experimental rats was evaluated by a blinder observer using BBB open field locomotion rating score.Results BrdU positive NSCs were detected in the spinal cord after transplantation,the number of RST neurons labeled by HRP in NSC group was more than that in SCI group (P <0.01),the BBB score of NSC group was higher than SCI group (P <0.01).Conclusion The transplanted NSCs can survive in the injured site of spinal cord and protect RN,then promote more remarkably functional recovery after SCI.
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0.05).The transfected NSCs were confirmed to have the latent ability of multi-directional differentiation.Conclusion:NSCs can express HIF-1? and GFP steadily after transfected with the recombinant adenovirus and the bionomics is normal.
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@#ObjectiveTo investigate the effects of fluid percussion injury(FPI) on survival and differentiation of transplanted human embryonic neural stem cells (HNSCs) in rats. MethodsThe HNSCs were separated from the cerebral cortex of the 8-week-old fetal and were cultured in DMEM/F12 combinated with EGF, bFGF and LIF. The rat models of FPI were made with fluid percussion system. The HNSCs labeled with BrdU were transplanted into the injured zone 24 hours after brain injury, then the rats were killed at the 1st and 4th week post-transplanted stages, and the brain slices were stained with immunocytochemistry. The GFAP, MAP-2, and BrdU positive cells were investigated.ResultsThe transplanted HNSCs migrated to the whole brain, and differentiated into GFAP and MAP-2 positive cells. MAP-2 positive cells were observed at 1 week post-transplanted stage, on the contrary, more GFAP positive cells were discovered 4 weeks after transplantation. Part of the HNSCs migrated to the choroids plexus of the lateral ventricle and microvessels. ConclusionThe transplanted HNSCs survive in the injured zone, and differentiate into astrocytes gradually during the recovery. The host devours part of the HNSCs.
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@#ObjectiveTo investigate an effective method to isolate neural stem cells(NSCs).MethodsNSCs were dissociated by digestion with trypsin, EDTA and different doses of Dispase,and serum-free culture techniques and immunohistochemistry techniques were used to verifying the dissociated.ResultsA lot of single neural stem cells were obtained by using Dispase to digest neurosphere, and the cells could keep its structure and morphology.ConclusionIt is an ideal method by using Dispase to digest neurosphere for isolating NSCs.
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@#ObjectiveTo study the mechanism of differentiation of mesenchymal stem cells(MSCs) into neuron-like cells in vitro.MethodsMSCs of Wistar rats were separated and cultured, and then induced with DMSO and BHA in vitro. The specific marking proteins of neurons, glia and neural stem cells were detected before preinduction, at 24h after preinduction, at 6h, 24h, and 48h after neuronal induction.ResultsAfter the inducement, many MSCs turned into bipolar,multipolar and taper,and then intersected as network structure. Nestin was strong positive at 6h after neuronal induction, and decreased at 24h, 48h after the induction. NeuN was present at 6 h after neuronal induction, and increased at 24h, 48h after the induction.ConclusionMSCs can be induced into neural stem cells(NSCs) at first, and then differentiate into neuron-like cells in vitro.
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Objective:To explore the effects of IL-1 beta on the differentiation of NSCs into dopaminergic neurons.Methods: To isolate and culture mesencephalic neural stem cells in vitro.we used immunocytochemistry to detect TH-positive neurons induced by 10%FBS and interleukin-1 bate for 7days respectively.Results: The positive ratio of TH expression in induced cells by 10%FBS,IL-1? 10pg/ml,IL-1? 120pg/ml,IL-1? 150pg/ml,IL-1? 200pg/ml were 0.45%,0.6%,7.2%,12.5%and 8.75% respectively.The positive ratios of TH expression in induced cells by IL-1? 150pg/ml(12.5%) were much higher than that by 10%FBS(control 0.45%)(P