目的 研究人参皂苷Rh₂逆转HepG2细胞对5-氟尿嘧啶（5-Flu）耐药性的作用。方法 构建对5-Flu耐药的HepG2细胞系（HepG2-5-Flu），MTT法分别测定人参皂苷Rh₂和5-Flu（二者质量浓度均为15.63、31.25、62.50、125.00、250.00、500.00、1 000.00、2 000.00 μg/mL）、人参皂苷Rh₂和5-Flu不同比例（5∶1、3∶1、1∶1、1∶3、1∶5）的混合药物（总浓度同单给药）对HepG2-5-Flu细胞存活率的影响，利用协同作用系数（CI）获得人参皂苷Rh₂和5-Flu的最佳协同作用浓度；通过Annexin V/PI双染色流式细胞术检测5-Flu（375 μg/mL）、人参皂苷Rh₂＋5-Flu（人参皂苷Rh₂∶5-Flu=1∶3，浓度为125、375 μg/mL）对细胞凋亡影响；实时荧光定量PCR（qRT-PCR）法检测细胞P-糖蛋白（P-gp）和多药耐药相关蛋白（MRP）的mRNA表达水平。结果 人参皂苷Rh₂对HepG2-5-Flu细胞的半数抑制浓度（IC₅₀）为223.3 μg/mL，而5-Flu的IC₅₀为8.34 μg/mL。人参皂苷Rh₂与5-Flu浓度比为5∶1、3∶1、1∶1、1∶3、1∶5时，混合药物对细胞的IC₅₀分别为324.7、224.0、219.0、135.6、105.9 μg/mL。人参皂苷Rh₂和5-Flu比例为5∶1时，二者没有协同作用（CI>1）；当人参皂苷Rh₂和5-Flu比例为3∶1和1∶1时，药物浓度大于62.5 μg/mL时才有较小的协同作用（0.62和5-Flu比例为1∶3和1∶5时，大多数药物浓度下都有较好的协同作用（02＋5-Flu组细胞坏死、早期凋亡、晚期凋亡的比例均显著升高，活细胞比例显著下降（P<0.05、0.01、0.001），MDR1和P-gp 2个耐药相关蛋白的mRNA表达显著下调（P<0.01）。结论 人参皂苷Rh₂能够逆转HepG2细胞对5-Flu的耐药性，机制可能与下调MDR1和P-gp表达相关。

Objective To study anti-liver cancer efficacy and overcome the drug resistance of 5-fluorouracil (5-Flu) that synergistically enhanced by ginsenoside Rh₂. Methods HepG2 cell line (HepG2-5-Flu) resistant to 5-Flu was constructed, Ginsenoside Rh₂ and 5-Flu (15.63, 31.25, 62.50, 125.00, 250.00, 500.00, 1 000.00, 2 000.00 μg/mL) were determined by MTT method, different proportions (5:1, 3:1, 1:1, 1:3, 1:5) of ginsenoside Rh₂ and 5-Flu (the total concentration was the same as single administration) on the survival rate of HepG2-5-Flu cells, and the optimal synergistic concentration of Ginsenoside Rh₂ and 5-FLu was obtained by using the synergistic coefficient. The effects of 5-Flu (375 μg/mL) and ginsenoside Rh₂+ 5-Flu (Ginsenoside Rh₂:5-Flu=1:3, 125, 375 μg/mL) on apoptosis were detected by Annexin V/PI double staining flow cytometry. The mRNA expression levels of P-gp and MRP were detected by real-time quantitative PCR (qRT-PCR). Results The 50% inhibitory concentration (IC₅₀) of Ginsenoside Rh₂ against HepG2-5-Flu cells was 223.3 μg/mL, and that of 5-Flu was 8.34 μg/mL. The IC₅₀ of ginsenoside Rh₂ and 5-Flu were 324.7, 224.0, 219.0, 135.6 and 105.9 μg/mL when the concentration ratio of ginsenoside Rh₂ and 5-flu was 5:1, 3:1, 1:1, 1:3 and 1:5, respectively. When the ratio of ginsenosides Rh₂ and 5-Flu was 5:1, there was no synergistic effect (CI > 1). When the ratio of ginsenoside Rh₂ and 5-Flu was 3:1 and 1:1, the concentration of ginsenoside Rh₂and 5-Flu was greater than 62.5 μg/mL (0.6 < CI < 1). When the ratio of ginsenoside Rh₂ and 5-Flu was 1:3 and 1:5, most drugs had a good synergistic effect (0 2 + 5-Flu group were significantly increased, and the percentages of living cells were significantly decreased (P < 0.05, 0.01, 0.001), and the mRNA expressions of MDR1 and P-GP were significantly down-regulated (P < 0.01). Conclusion Ginsenoside Rh₂ can reverse the resistance of HepG2 cells to 5-Flu, and the mechanism may be related to the down-regulation of MDR1 and P-GP expression.

R285.5

陕西省中医药管理局中医药科研课题（LCMS016）