四氢紫堇萨明对Aβ_(25-35)诱导AD细胞模型凋亡的影响及其机制研究
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摘要
探讨四氢紫堇萨明(SQZJSM)对阿尔茨海默病(AD)细胞模型凋亡的影响及作用机制。采用Aβ_(25-35)诱导神经细胞PC-12损伤建立AD细胞模型,通过流式细胞仪检测凋亡率,高通量高内涵分析系统观察细胞核形态并检测线粒体膜电位(MMP)变化,Western blot法检测凋亡相关蛋白的表达。结果显示,SQZJSM可改善模型细胞受损的细胞核形态,同时显著减少模型细胞凋亡率和胞质Cyt C含量,上调MMP,降低Bax、Cleaved-Caspase 3、Cleaved-Caspase 9蛋白表达,增加Bcl-2、p-Akt/T-Akt表达(P <0. 05); PI3K/AKT抑制剂LY294002可阻断SQZJSM对模型细胞的上述改善作用。以上结果表明四氢紫堇萨明可显著改善Aβ_(25-35)诱导的AD细胞模型凋亡,其机制可能与激活PI3K/Akt信号通路调控内源性线粒体凋亡途径相关。
To explore the effect of tetrahydrocorysamine on the apoptosis of Alzheimer's disease( AD) cell model and its possible mechanism. Alzheimer's diseasecell model was established by Aβ_(25-35) inducing PC-12 cells. Flow cytometry was used to measure the apoptosis rate,high-throughput and high-content analysis system was used to observe the nuclear morphological changes and detect mitochondrial membrane potential( MMP) changes,the expression of apoptosis-related protein was detected by Western blot. The results showed that SQZJSM can improve the damaged nuclear morphology of model cells,and significantly reduce the apoptosis rate and Cyt C level in the cytoplasm,up-regulate the level of MMP,decrease the expression of Bax,Cleaved-Caspase3,Cleaved-Caspase 9 protein,and increase the level of Bcl-2,p-Akt/T-Akt( P < 0. 05); PI3K/AKT inhibitor LY294002 can block the improvement of SQZJSM on model cells. The above results indicate that tetrahydrocorysamine can significantly reduce the apoptosis rate of AD cell model induced by Aβ_(25-35),which may be related to the activation of PI3K/Akt signaling pathway to regulate endogenous mitochondrial apoptosis.
To explore the effect of tetrahydrocorysamine on the apoptosis of Alzheimer's disease( AD) cell model and its possible mechanism. Alzheimer's diseasecell model was established by Aβ_(25-35) inducing PC-12 cells. Flow cytometry was used to measure the apoptosis rate,high-throughput and high-content analysis system was used to observe the nuclear morphological changes and detect mitochondrial membrane potential( MMP) changes,the expression of apoptosis-related protein was detected by Western blot. The results showed that SQZJSM can improve the damaged nuclear morphology of model cells,and significantly reduce the apoptosis rate and Cyt C level in the cytoplasm,up-regulate the level of MMP,decrease the expression of Bax,Cleaved-Caspase3,Cleaved-Caspase 9 protein,and increase the level of Bcl-2,p-Akt/T-Akt( P < 0. 05); PI3K/AKT inhibitor LY294002 can block the improvement of SQZJSM on model cells. The above results indicate that tetrahydrocorysamine can significantly reduce the apoptosis rate of AD cell model induced by Aβ_(25-35),which may be related to the activation of PI3K/Akt signaling pathway to regulate endogenous mitochondrial apoptosis.
引文
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2 Wang SS,Zhang Z,Zhang X,et al.Advances in immunotherapy of Alzheimer’s disease with traditional Chinese medicine[J].Acta Pharm Sin(药学学报),2018,53:1023-1029.
3 Shoji M.Drug therapy for Alzheimer’s disease[J].Brain Nerve,2010,62:777-785.
4 Zhang HB,Sun Y.Effect of huperzine A on the expression of amyloidβ-protein in transgenic mice brain of Alzheimer's disease[J].Nat Prod Res Dev(天然产物研究与开发),2017,29:290-293.
5 Asai M,Iwata N,Yoshikawa A.Berberine alters the processing of Alzheimer’s amyloid precursor protein to decrease Abeta secretion[J].Biochem Biophys Res Commun,2007,352:498-502.
6 Wang H,Bi FJ,Lin T,et al.RP-HPLC fingerprint of corydalis yanhusuo and content determination of nine alkaloids[J].J Chin Med Materls(中药材),2017,40:624-629.
7 Bai X,Yang J,Liu F,et al.The anti-aging effect of corydalis yanhusuo W.T.Wang on aging mice induced by D-galactose[J].Guizhou Med J(贵州医药),2008,32:399-401.
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9 Maccioni RB,Munoz JP,Barbeito L,et al.The molecular bases of Alzheimer’s disease and other neurodegenerative disorders[J].Arch Med Res,2001,32:367-381.
10 D Puzzo,W Gulisano,O Arancio.The keystone of Alzheimer pathogenesis might be sought in Aβphysiology[J].Neuroscience,2015,307:26-36.
11 Yuan JY,Yankner BA.Apoptosis in the nervous system[J].Nature,2009,40:802.
12 Barage SH,Sonawane KD.Amyloid cascade hypothesis:pathogenesis and therapeutic strategies in Alzheimer’s disease[J].Neuropeptides,2015,52:1-18.
13 Yu Y,He W.Effects of Imperatorin on the learning and memory impairment of Aβ1-42-induced model of Alzheimer’s disease in mice[J].Nat Prod Res Dev(天然产物研究与开发),2017,29:1030-1034.
14 Orike N,Middleton G,Borthweick E,et al.Role of PI 3-kinase,Akt and Bcl-2-related proteins in sustaining the survival of neurotrophic factor-in-dependent adult sympathetic neurons[J].J Cell Biol,2001,154:995-1005.
15 Chen Q,Ye HY,Chen YQ,et al.Ferulic acid protects against apoptosis of PC12 cells induced by kainic acid[J].Chin JPathophysiol(中国病理生理杂志),2013,29:1175-1180.
16 Spencer SL,Sorger PK.Measuring and modeling apoptosis in single cells[J].Cell,2011,144:926-939.
17 Tait SW,Green DR.Mitochondrial regulation of cell death[J].Csh Perspect Biol,2013,5(9):64-74.
18 Yin G.Effects of overexpressed AKT on induced apoptosis[D].Wuhan:Huazhong University of Science and Technology(华中科技大学),2011.
19 Wang CH,Chen TT,Li GX,et al.Simvastatin preventsβ-amyloid 25-35-impaired neurogenesis in hippocampal dentate gyrus throughα7nAChR-dependent cascading PI3K-Akt and increasing BDNF via reduction of farnesyl pyrophosphate[J].Neuropharmacology,2015,97:122-132.