R-脊椎蛋白3对小鼠神经干细胞增殖及Wnt/β-catenin通路的影响
The effect of R-spondin3 on proliferation of neural stem cells and Wnt/β-catenin signals in mice
查看参考文献24篇
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文摘
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目的观察R-脊椎蛋白3(Rspo3)对小鼠神经干细胞(NSCs)增殖的影响及其机制.方法分离、悬浮培养孕14~15 d的CD1胎鼠大脑侧脑室下区(SVZ)NSCs并通过免疫荧光法鉴定.取第3代处于对数生长期的NSCs分为实验组和对照组,体积各为1 mL.实验组添加储存浓度为50 μg/mL的Rspo3蛋白0.8 μL(终浓度40 ng/mL),对照组添加等体积的NSCs培养液.干预后6 h通过BrdU细胞渗入实验法检测2组NSCs增殖情况.干预后4 h和8 h采用Western blotting实验检测β-catenin蛋白的表达.结果免疫荧光染色结果显示90%以上细胞呈巢蛋白和SOX2阳性.经过诱导分化后,部分细胞呈神经元核抗原或胶质纤维酸性蛋白阳性,证明实验中提取分离的细胞为NSCs.BrdU细胞渗入实验法显示实验组细胞增殖率为1.56±0.03,明显高于对照组(1.04±0.04),差异有统计学意义(P<0.05).Western blotting实验显示4 h和8 h时实验组β-catenin蛋白相对表达量分别为1.09±0.10、1.20±0.13,对照组分别为0.56±0.05、0.83±0.04,实验组明显高于对照组,差异有统计学意义(P<0.05).结论Rspo3蛋白可在体外促进小鼠NSCs增殖,其机制可能与Wnt/β-catenin信号通路的激活有关. |
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其他语种文摘
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Objective To explore the effect of R-spondin3 on the proliferation of neural stem cells (NSCs) and its mechanism in mice.Methods The mouse NSCs derived from the subventricular zone of E14-15d CD1 mice were confirmed by immunofluorescence assay. The NSCs after 3 passages of culture were chosen and randomly divided into 2 groups (V=1 mL). In the experimental group, 0.8 µL of R-spondin3 with an initial concentration of 50 µg/mL was added (final concentration: 40 ng/mL) while in the control group an equal amount of culture fluid was added. The proliferation of the cells in the 2 groups was detected by 5-Bromo-2-deoxy Uridine (BrdU) kits after the cells were treated by R-spondin3 for 6 hours. The protein expression of β-catenin was measured by western blotting after the cells were treated by R-spondin3 for 4 and 8 hours.Results Under optical microscopy, the round and bright cells grew in culture medium and easily accumulated to become neurospheres. Immunofluorescence assay showed that over 90% of the cells expressed Nestin and SOX2 and that some of them expressed NeuN or GFAP after induced differentiation. Brdu proliferation test showed that the proliferation rate of Brdu~+/DAPI~+ for the experimental group (1.56±0.03) was significantly higher than that for the control group (1.04±0.04) (P<0.05). Western blotting showed that the expression levels of β-catenin were increased at both 4h and 8h after treatment for the experimental group (1.09±0.10 and 1.20±0.13), significantly higher than those for the control group (0.56 ± 0.05 and 0.83 ± 0.04) (P<0.05).Conclusions R-spondin3 can promote in vitro proliferation of NSCs in mice, which may be associated with activated Wnt/β-catenin signal pathways. |
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来源
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中华神经医学杂志
,2018,17(4):344-348 【扩展库】
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DOI
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10.3760/cma.j.issn.1671-8925.2018.04.004
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关键词
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R-spondin3
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神经干细胞
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Wnt/β-catenin信号通路
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地址
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山西医科大学第二临床医学院, 太原, 030002
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合肥市第一人民医院神经外科, 230032
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陆军总医院附属八一脑科医院, 北京, 100700
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语种
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中文 |
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文献类型
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研究性论文 |
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ISSN
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1671-8925 |
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学科
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细胞生物学 |
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基金
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国家自然科学基金
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文献收藏号
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CSCD:6230690
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