目的 探讨哈巴苷对β淀粉样蛋白(Aβ_25-35)诱导的PC12细胞毒性的影响。方法 PC12细胞以哈巴苷(20、40、80 μmol/L)预孵育3 h后以Aβ_25-35(30 μmol/L)损伤24 h;MTT法检测细胞增殖;检测细胞上清液中丙二醛(MDA)、谷胱甘肽(GSH)、超氧化物歧化酶(SOD)水平;采用Annexin-V-FITC双染法检测细胞凋亡率;检测活性氧(ROS)的生成;Western blotting法检测Akt、Bcl-2和Bax蛋白表达。结果 与对照组比较,Aβ_25-35使PC12细胞的存活率显著降低(P<0.01),细胞上清液中MDA水平显著升高(P<0.01)、GSH和SOD水平显著降低(P<0.01),细胞内ROS水平显著升高(P<0.01),细胞凋亡率显著增加(P<0.01),细胞内p-Akt、Bcl-2蛋白表达下调(P<0.01),Bax蛋白表达显著上调(P<0.01);哈巴苷预孵育PC12细胞3 h,使细胞上清液中的MDA水平降低,GSH和SOD水平显著升高(P<0.05、0.01);明显剂量依赖性抑制细胞凋亡;上调p-Akt和Bcl-2蛋白表达(P<0.05、0.01),并下调Bax蛋白表达(P<0.05、0.01)。结论 哈巴苷改善Aβ_25-35诱导的PC12细胞毒性,可能通过激活PI3K/Akt信号通路和上调细胞内SOD水平发挥作用。

Objective The main purpose of the present study is to investigate the influence of harpagide on amyloid-β_25-35 (Aβ_25-35)- induced cytotoxicity. Methods Oxidative stress was assessed by measuring MDA, glutathione (GSH), and superoxide dismutase (SOD) levels. Cell viability was assessed by MTT assay. Cell apoptosis detection was performed using an Annexin-V-FITC Apoptosis Detection Kit. The production of ROS was determined using a ROS Assay Kit. Western blotting detection was carried out to detect the protein expression. Results Our studies showed that pretreatment with harpagide could reduce Aβ_25-35-induced oxidative stress. Harpagide markedly inhibited cell apoptosis in a dose dependent manner. More importantly, harpagide increased the PI3K/AKt and Bcl-2 family protein ratios on pre-incubation with cells for 3 h. Conclusion One possible mechanism of harpagide to improve Aβ_25-35-induced PC12 cytotoxicity may be through the up-regulation of SOD and activation of the PI3K/AKt pathway.