Page 55 - 《中国药房》2020年第24期

人参总皂苷对 D-半乳糖致 PC12 细胞衰老的改善作用及其机制

研究 Δ

＊
乔巨慧 ，赵大庆，刘美辰 ，睢博文，刘 颖，邢 欣（长春中医药大学吉林省人参科学研究院，长春 130117）
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中图分类号 R285 文献标志码 A 文章编号 1001-0408（2020）24-2993-07
DOI 10.6039/j.issn.1001-0408.2020.24.08

摘 要 目的：研究人参总皂苷对D-半乳糖致PC12细胞衰老的改善作用及其机制。方法：将大鼠嗜铬细胞瘤（PC12）细胞用D-半
乳糖处理以建立细胞衰老模型。采用CCK-8法筛选D-半乳糖的建模浓度和人参总皂苷的给药浓度。试验设正常对照组、模型组
和人参总皂苷低、高浓度组，检测各组细胞衰老情况、细胞凋亡率、细胞周期和细胞中线粒体膜电位（MMP）、三磷酸腺苷（ATP）、活
性氧（ROS）水平以及凋亡相关蛋白[B淋巴细胞瘤2（Bcl-2）及其相关蛋X蛋白（Bax）、细胞色素C（Cyt-C）]和氧化损伤相关蛋白[核
因子 2 相关因子 2（Nrf2）、血红素氧合酶 1（HO-1）]表达水平，另设阳性药物组[5 mmol/L N-乙酰-L-半胱氨酸（NAC）]和阳性对照
组[D-半乳糖+5 mmol/L NAC]比较氧化损伤相关蛋白水平。结果：D-半乳糖能显著抑制PC12细胞的存活率，临界浓度为20 mg/mL；
人参总皂苷可显著增加D-半乳糖诱导衰老细胞的存活率，半数效应浓度为65 μg/mL，故将后续试验人参总皂苷的低、高浓度设置
为55、65 μg/mL。与正常对照组比较，模型组衰老细胞数明显增加，细胞凋亡率和G1期细胞比例均显著增加，S期细胞比例、细胞
中MMP、ATP含量和Bcl-2蛋白以及线粒体中Cyt-C蛋白表达水平均显著降低，ROS含量和Bax、Nrf2蛋白及胞浆内Cyt-C蛋白表
达均显著升高（P＜0.05或P＜0.01）。与模型组比较，人参总皂苷低、高浓度组衰老细胞数明显减少，细胞凋亡率和G1期细胞比例
均显著降低，S期细胞比例、细胞中MMP、ATP（除低浓度组外）含量和Bcl-2、Nrf2、HO-1蛋白以及线粒体中Cyt-C蛋白表达水平均
显著升高，ROS（除低浓度组外）含量和Bax蛋白及胞浆内Cyt-C蛋白表达水平均显著降低；阳性对照组细胞内Nrf2、HO-1蛋白表
达水平亦显著升高（P＜0.05或P＜0.01），但均低于人参总皂苷组。结论：人参总皂苷可改善D-半乳糖诱导的P12细胞衰老，其机
制可能与激活Nrf2抗氧化信号途径拮抗D-半乳糖诱导的氧化应激、缓解其所致的线粒体功能障碍有关。
关键词 人参总皂苷；D-半乳糖；PC12细胞；衰老；线粒体；凋亡相关蛋白；氧化损伤相关蛋白

Study on Improvement Effects of Total Ginsenosides on D-galactose Induced PC12 Cell Senescence and Its
Mechanism
QIAO Juhui，ZHAO Daqing，LIU Meichen，SUI Bowen，LIU Ying，XING Xin（Institute of Ginseng Research，
Changchun University of TCM，Changchun 130117，China）

ABSTRACT OBJECTIVE：To study the improvement effects of total ginsenosides on the senescence of PC12 cells induced by
D-galactose and its mechanism. METHODS：Rat pheochromocytoma（PC12）cells were treated with D-galactose to establish cell
senescence model. CCK-8 method was used to screen the D-galactose modeling concentration and total ginsenosides concentration.
Normal control group，model group，total ginsenosides low and high concentration groups were set up. Cell senescence，cell
apoptosis rate，apoptotic cycle and mitochondrial membrane potential（MMP），cell adenosine triphosphate（ATP） and reactive
oxygen species（ROS）levels in each group were detected. The expression of apoptosis related proteins [B lymphoma 2（Bcl-2）and
its related egg X protein（Bax），cytochrome C（Cyt-C）] and oxidative damage related proteins [nuclear factor 2 related factor 2
（Nrf2），heme oxygenase 1（HO-1）] were detected. In addition，positive drug group [5 mmol/L N-acetyl-L-cysteine（NAC）] and
positive control group [D-galactose + 5 mmol/L NAC] were set up to compare the levels of oxidative damage related proteins.
RESULTS：D-galactose could significantly inhibit the survival rate of PC12 cells，with a critical concentration of 20 mg/mL. The
total ginsenosides could significantly increase the survival rate of D-galactose induced senescent cells with a median effective
concentration（EC50 ）of 65 μg/mL，and then the low and high concentrations of total ginsenosides were set at 55 and 65 μg/mL.
Compared with normal control group，the number of aging cells increased，the apoptotic rate and percentage of G1 phase were
significantly increased in model group. the percentage of S phase，MMP and ATP contents，the protein expression of Bcl-2 and
Cyt-C in mitochondria were decreased significantly， while
Δ 基金项目：国家自然科学基金资助项目（No.81703663）；中央本
ROS content，the protein expression of Bax，Nrf2 and Cyt-C
级重大增减支项目（No.2060302）；吉林省教育厅“十三五”科学技术项
protein in endochylema were increased significantly（P＜0.05
目（No.JJKH20190478KJ）
or P＜0.01）. Compared with model group，the number of
＊硕士研究生。研究方向：中药生物技术。电话：0431-86177401。
E-mail：sunqiao150509@163.com aging cells reduced，the apoptosis rates and percentage of G1
# 通信作者：副研究员，博士。研究方向：中药有效成分及其传统 phase were significantly decreased in total ginsenosides low
功效。电话：0431-86177401。E-mail：liumc0367@163.com and high concentration groups，the percentage of S phase，the

中国药房 2020年第31卷第24期 China Pharmacy 2020 Vol. 31 No. 24 ·2993 ·