目的：探讨冬凌草甲素（Ori）逆转人黑色素瘤细胞顺铂（DDP）耐药的作用及其机制。方法：分别将黑色素瘤DDP耐药细胞A375/DDP 和M14/DDP 分为对照组、2 μmol/L Ori 组、4 mg/L DDP组和2 μmol/L Ori+4 mg/L DDP组。CCK-8法、Transwell实验、Annexin Ⅴ-FITC/PI 染色流式细胞术分别检测各组细胞的增殖活力、侵袭和迁移能力及凋亡水平，透射电子显微镜观察自噬小体，免疫荧光染色法观察微管相关蛋白轻链3（LC3）点状结构，WB 法检测A375/DDP 细胞自噬相关蛋白Beclin-1、p62、LC3Ⅱ和LC3Ⅰ的表达。结果: 与4 mg/L DDP组相比，2 μmol/L Ori+4 mg/L DDP组细胞增殖活力、迁移和侵袭能力均显著下降（均P<0.01），凋亡水平显著升高（P<0.01）。4 mg/L DDP组细胞中可见大量自噬小体以及LC3点状染色，但2 μmol/L Ori+4 mg/L DDP组仅可见少量。与对照组相比，4 mg/L DDP 组细胞中Beclin-1 和LC3Ⅱ/LC3Ⅰ的蛋白表达水平均显著升高（均P<0.01），p62 的蛋白表达水平显著降低（P<0.05 或 P<0.01）；与 4 mg/L DDP 组相比，2 μmol/L Ori+4 mg/L DDP 组细胞中Beclin-1 和LC3Ⅱ/LC3Ⅰ的表达水平均显著降低（均P<0.01），p62 的表达水平显著升高（P<0.05）。结论: Ori 可增加耐药黑色素瘤细胞对DDP的敏感性，此作用可能与其抑制DDP引起的细胞自噬有关。

Objective: To investigate the effect of oridonin (Ori) in reversing cisplatin (DDP) resistance in human melanoma cells and its related mechanism. Methods: Melanoma cisplatin-resistant cell lines A375/DDP and M14/DDP were divided into control group,2 μmol/L Ori group, 4 mg/L DDP group and 2 μmol/L Ori+4 mg/L DDP group. Cell viability, invasion and migration, and apoptosis were detected by CCK-8, Annexin Ⅴ-FITC/PI staining flow cytometry and Transwell assay, respectively. The autophagosomes were observed by transmission electron microscopy, microtubule associated protein1 light chain 3 (LC3)-point structure was observed by immunofluorescence staining, and the expressions of autophagy related proteins Beclin-1, p62, LC3Ⅱ and LC3Ⅰ of A375/DDP cells were detected by WB.Results: Compared with 4 mg/LDDP group, cell proliferation viability, migration and invasion ability of 2 μmol/L Ori+4 mg/L DDP group were significantly decreased (all P<0.01), and apoptosis level was significantly increased (P<0.01). A large number of autophagosomes and LC3-point structures were observed in 4 mg/LDDPgroup; however, only a fewwere observed in 2 μmol/L Ori+4 mg/L DDP group. Compared with control group, the protein expression levels of Beclin-1 and LC3Ⅱ/LC3Ⅰ in 4 mg/L DDP group were significantly increased (all P<0.01), but the protein expression level of p62 was significantly decreased (P<0.05 or P<0.01); compared with 4 mg/L DDP group, the protein expression levels of Beclin-1 and LC3Ⅱ/LC3Ⅰin 2 μmol/L Ori+4 mg/L DDP group were significantly decreased (all P<0.01), but the protein expression level of p62 was significantly increased (P<0.05). Conclusion: Ori can increase the sensitivity of drug-resistant melanoma cells to DDP, which may be related to the inhibition of DDP-induced autophagy.

河南省科学技术厅基金（No. 212102310684）