促红细胞生成素对β-淀粉样蛋白诱导细胞凋亡的保护作用

Effects of erythropoietin on cell apoptosis induced by β-amyloid peptide

目的的探讨促红细胞生成素（Epo）对凝聚态β-淀粉样蛋白25～35片断（Aβ（25-35）诱导细胞凋亡的影响。方法人神经母细胞瘤细胞株（SH．SY5Y）体外传代培养，细胞进入对数生长期后进行实验。MTT比色法检测不同浓度Aβ（25-35）用24h后细胞活力变化；Hoechst33258核染色和Western blotting法分别观察和检测20μmol／LAβ（25-35）用24h后，细胞形态及细胞caspase-3、cleaved caspase-3水平的变化。运用同样方法检测不同浓度Epo预处理3h，对20μmol／LAβ（25-35）用24h所致细胞活力、细胞形态以及caspase-3和cleaved caspase-3水平改变的影响。结果结果显示不同浓度的Aβ（25-35）用24h后，SH—SY5Y细胞活力均明显下降，并呈剂量依赖性（P〈0．05）；10U Epo预处理可明显抑制Aβ（25-35）导的细胞活力下降（P〈0．05）。Hoechst33258核染色发现10UEpo预处理可明显减轻Aβ（25-35）导的细胞凋亡现象（P〈0．05）；Western blotting检测表明，10UEpo可显著抑制Aβ（25-35）导的cleaved caspase-3表达增高（P〈0．05）。结论Epo通过抑制Aβ（25-35）起的cleaved caspase-3表达增高而对其诱导的细胞凋亡发挥保护作用。本实验为研究阿尔茨海默病的发病机制及探索新的有效治疗药物提供了重要的理论基础。

Objective To investigate the effects of erythropoietin （Epo） on cell apoptosis induced by Aβ（25-35）. Methods SH-SY5Y cells were performed serial subcultivation, and entered into experiment during the exponential phase of growth. The viability of SH-SY5Y cells 24 h after induction by Aβ（25-35） with different concentrations were detected by MTT assay, and the morphology and levels of cleaved caspase-3 and caspase-3 24 h after induction by 20 p.mol/L Aβ（25-35） were observed and examined by Hoechst 33258 nucleus staining and Western blotting, respectively. After pretreatment with Epo of different concentrations for 3 h, the cell viability, morphology, and levels of cleaved caspase-3 and caspase-3 24 h after induction by 20 p, mol/L Aβ（25-35） were detected with the corresponding methods. Results It was revealed by MTT assay that 24 h after treatment with Aβ（25-35） of different concentrations, the viability of SH-SY5Y cells was significantly decreased in a dose-dependent manner（ P 〈0.05） , and various concentration of Epo could significantly prevent the cell viability induced by Aβ（25-35） （P 〈0.05）. Pretreatment with 10 U Epo could significantly inhibit the viability decrease induced by Aβ（25-35） （P 〈 0.05）. It was indicated by Hoechst 33258 nucleus staining that pretreatment with 10 U Epo could significantly reduce the cell apoptosis induced by Aβ（25-35） （P 〈0.05）. Results of Western blotting showed that pretreatment with 10 U Epo could inhibit the increased expression of cleaved caspase-3 induced by Aβ（25-35） （P 〈0.05）. Conclusion Epo may have protective effect on cell apoptosis induced by Aβ（25-35） via inhibiting the expression of cleaved caspase-3. Thus, it provides an important theoretic basis for the study of pathogenesis and management of Alzheimer＇s disease.