目的 观察不同浓度β-淀粉样蛋白（amyloid β-protein25-35，Aβ_25-35）和六味地黄丸含药血清（medicated serum of Liuwei Dihuang Pill，MSLDP）对PC12（pheochromocytoma cells）细胞及FoxO3a（Forkhead box protein O 3a）、p-FoxO3a蛋白表达的影响，研究六味地黄丸含药血清预处理对Aβ_25-35损伤AD（Alzheimer’s disease）细胞模型的保护作用。方法 配制Aβ_25-35母液和制备六味地黄丸含药血清，用含有不同浓度的Aβ_25-35（10 μmol·L^-1、20 μmol·L^-1、40 μmol·L^-1）和六味地黄丸含药血清（2.5%、5%、10%、20%、30%）的培养基培养PC12细胞，噻唑蓝（MTT）法检测细胞活力，同时观察细胞形态的变化，免疫印迹法检测细胞中FoxO3a及p-FoxO3a蛋白表达量的变化。取合适浓度的六味地黄丸含药血清预处理Aβ_25-35诱导的AD细胞模型，观察细胞形态、活力及FoxO3a、p-FoxO3a表达量的变化。结果 ①Aβ_25-35显著降低PC12细胞存活率（P<0.01），并造成细胞形态的改变，细胞内p-FoxO3a表达量降低，而FoxO3a的表达升高。②六味地黄丸含药血清显著提高PC12细胞存活率（P<0.01），并促使细胞增加分化趋势，细胞内FoxO3a表达降低，而p-FoxO3a的表达随着血清浓度的增加而增加。③六味地黄丸含药血清预处理能够明显降低Aβ_25-35对PC12细胞的损伤，与模型组相比，六味地黄丸含药血清干预组FoxO3a的蛋白表达降低（P<0.01），p-FoxO3a的蛋白表达显著升高（P<0.05）。结论 六味地黄丸含药血清能够提高AD细胞模型的细胞活性，对PC12细胞具有明显的保护作用，磷酸化FoxO3a蛋白表达水平的提高可能是其相关的作用机制。

Objective To observe the effects of different concentrations of Aβ_25-35 and medicated serum of Liuwei Dihuang Pills (MSLDP) on rat adrenal pheochromocytoma cells (PC12) and the expression of FoxO3a and p-FoxO3a proteins, and to study the protective effect of MSLDP pretreatment on Aβ_25-35 injury AD cell model.Methods We prepared Aβ_25-35 mother liquor and prepare medicated serum of Liuwei Dihuang Pills, cultured PC12 cells with different concentrations of Aβ_25-35 (10μmol·L^-1, 20μmol·L^-1, 40μmol·L^-1) and MSLDP (2.5%, 5%, 10%, 20%, 30%) medium, MTT method was used to detect cell viability and observe changes in cell morphology. Western Blot was used to detect changes in the expression of FoxO3a and p-FoxO3a proteins in the cells. We took the appropriate concentration of MSLDP to pretreat the AD cell model induced by Aβ_25-35, and observed the changes in cell morphology, viability and FoxO3a and p-FoxO3a expression.Results ①Aβ_25-35 reduced the survival rate of PC12 significantly (P<0.01), and caused changes in cell morphology. The expression of p-FoxO3a in PC12 cells decreased, but the expression of FoxO3a increased. ②MSLDP improved the survival rate of PC12 cells significantly (P<0.01), and promoted the cells to increase their differentiation tendency, the expression of FoxO3a in the cells decreased, and the expression of p-FoxO3a increased with the increase of concentration. ③MSLDP pretreatment can significantly reduce the damage of Aβ_25-35 to PC12 cells. Compared with the model group, the protein expression of FoxO3a in the MSLDP intervention group decreased (P<0.01), and the protein expression of p-FoxO3a increased significantly (P<0.05).Conclusion MSLDP can increase the activity of Aβ_25-35-induced AD cell models and have a protective effect on PC12 cells. The mechanism may be related to the increase of phosphorylated FoxO3a protein expression.

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河南省科技厅河南省科技攻关项目（212102311084）：六味地黄丸通过调控FoxO3a靶点防治早期AD的抗氧化功效研究，负责人：宋军营；河南省科技厅中原科技创新领军人才计划资助项目（204200510022）：六味地黄丸介导PPARγ/Nrf2/TXNIP信号通路防治早期老年痴呆的作用机制，负责人：张振强；河南省教育厅河南省高校科技创新团队支持计划项目（21IRTSTHN026）：中西医结合防治重大慢病，负责人：张振强。