首页|基于DKK3调控探讨藏红花素对Aβ25-35诱导神经元损伤的保护作用

基于DKK3调控探讨藏红花素对Aβ25-35诱导神经元损伤的保护作用

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目的:探究藏红花素通过调控dickkopf WNT 信号通路抑制因子 3(Dickkopf3,DKK3)对Aβ25-35 诱导神经细胞损伤的保护作用及机制.方法:Aβ25-35 诱导小鼠海马神经细胞系HT22 细胞模拟细胞损伤,CCK8 增殖实验检测藏红花素最佳干预浓度.qPCR 检测 DKK3 沉默质粒(DKK3 siRNA)以及DKK3 过表达质粒(DKK3-OE)的DKK3 mRNA表达,随后进行细胞转染.细胞分为正常组、Aβ25-35组、Aβ25-35 +藏红花素组(1.0 μmol/L)、Aβ25-35 + DKK3 siRNA 组、Aβ25-35 + siRNA 对照组、DKK3-OE组、空白质粒对照组、DKK3-OE +藏红花素组(1.0 μmol/L)组.采用流式细胞数检测细胞凋亡,Tubulin β染色观察细胞神经突触形态,Western blot检测细胞Aβ、DKK3、β-catenin、GSK-3β、p-GSK-3β、Caspase-3、Bax、Bcl-2 蛋白表达.结果:0.5~2.0 μmol/L藏红花素可显著上调HT22 细胞活力(P<0.05).与正常组相比,Aβ25-35 组和DKK3-OE组细胞凋亡显著增加,突触长度、分支数显著减少,Aβ、DKK3、p-GSK-3β/GSK-3β、Caspase-3、Bax蛋白表达显著升高,β-catenin、Bcl-2 蛋白蛋白表达显著降低(P<0.01).与DKK3-OE组相比,藏红花素干预则能逆转以上结果.与Aβ25-35 组相比,Aβ25-35 +藏红花素组和Aβ25-35 + DKK3 siRNA组细胞的凋亡显著降低,突触长度、分支数显著增加,Aβ、DKK3、p-GSK-3β/GSK-3β、Caspase-3、Bax蛋白表达显著降低,β-catenin、Bcl-2 蛋白蛋白表达显著升高(P<0.05).结论:藏红花素对Aβ25-35 诱导神经细胞损伤具有保护作用,其作用与通过抑制DKK3 调控GSK-3β/β-catenin信号通路表达有关.
Neuroprotective Effect of Crocin on Neuron Damage Induced by Aβ 25-35 by Mediating DKK3
Objective:To study the neuroprotective effect of Crocin mediated Dickkopf3(DKK3)on neuron damage induced by Aβ25-35 and its mechanism.Methods:Mouse hippocampal neuron HT-22 cells were in-duced by Aβ25-35,then CCK-8 test was used to determine optimal intervention concentration of Crocin.q-PCR was used to detect the DKK3 mRNA expression of DKK3 silencing plasmid(DKK3 siRNA)and DKK3 overexpression plasmid(DKK3-OE),then plasmids were transfected into cells.Cells we divided into normal group,Aβ25-35 group,Aβ25-35 + Crocin group(1.0 μmol/L),Aβ25-35 + DKK3 siRNA group,Aβ25-35 +siRNA control group,DKK3-OE group,plasmid control group,DKK3-OE + Crocin group(1.0 μmol/L).Flow cytometry assay was used to detect cell apoptosis,staining to observe cell synaptic morphology,Wes-tern blot was used to detect protein expressions of Aβ,DKK3,β-catenin,GSK-3β,p-GSK-3β,Caspase-3,Bax,Bcl-2.Results:0.5~2.0 μmol/L Crocin significantly could increase cell activity(P<0.01).Compared to normal group,apoptosis in Aβ25-35 group and DKK3-OE group were significantly increased,synaptic length and number of branches were significantly reduced,and protein expressions of Aβ,DKK3,p-GSK-3 β/GSK-3 β,Caspase-3 and Bax were significantly increased,the protein expressions of β-catenin and Bcl-2 were significantly decreased(P<0.01).Compared to DKK3-OE group,Crocin could reverse the above results.Compared to Aβ25-35 group,Aβ25-35 + Crocin group and Aβ25-35 + DKK3 siRNA group cell apoptosis were significantly decreased,synaptic length and number of branches were significantly increased,pro-tein expressions of Aβ,DKK3,p-GSK-3 β/GSK-3β,Caspase-3 and Bax were significantly decreased,while the protein expressions of β-catenin and Bcl-2 were significantly increased(P<0.05).Conclusion:Crocin has a protective effect on injury of nerve cell in-duced by Aβ25-35,it related to regulate expression of GSK-3 β/β-catenin signaling pathway by mediating DKK3.

Alzheimer's diseaseCrocinDKK3neuronGSK-3β/β-cateninapoptosisin vitro

杨晓佳、吴敏、江萌、刘立权

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杭州市中医院·浙江 杭州 310007

阿尔兹海默症 藏红花素 DKK3 神经元 GSK-3β/β-catenin信号通路 细胞凋亡 体外实验

浙江省中医药科技计划

2022ZB236

2024

中国中医药科技
中华中医药学会

中国中医药科技

影响因子:1.156
ISSN:1005-7072
年,卷(期):2024.31(1)
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