积雪草酸通过抑制耐药相关蛋白表达增强U87MG胶质瘤细胞对紫杉醇的敏感性

目的： 探索积雪草酸（asiatic acid，AA）对紫杉醇（paclitaxel, PTX）耐药性胶质瘤细胞的抑制作用及其可能的作用机制。 方法： CCK-8实验、实时荧光定量PCR、Western blotting检测AA对成胶质细胞瘤U87MG细胞的增殖、凋亡的影响。浓度递增法构建PTX耐药性细胞株PR-U87MG，以U87MG细胞为对照，CCK-8实验验证PR-U87MG细胞对PTX的耐药性，实时荧光定量PCR、Western blotting检测PR-U87MG细胞中MDR1、LRP mRNA及蛋白的表达水平。AA和PTX单独或联合处理PR-U87MG细胞，CCK-8实验、实时荧光定量PCR、Western blotting检测各组细胞增殖活力及凋亡的变化。 结果： 成功构建PTX耐药性细胞株PR-U87MG。AA可以剂量依赖方式抑制U87MG细胞和PR-U87MG细胞的增殖活力（P<0.01），并明显促进其凋亡（P<0.01）。与AA或PTX单独处理组相比，联合处理组中PARP1的蛋白水平显著减少（P<0.01），caspase 3的裂解量显著增加（P<0.01），耐药相关蛋白P-糖蛋白1 （P-dycoprotein 1, Pgp-1）和LRP表达水平显著减少（P<0.01）。 结论： AA可有效增强U87MG胶质瘤细胞株对PTX的敏感性，其机制可能与AA抑制具有药物排出功能的耐药蛋白Pgp-1和LRP表达有关。

Asiatic acid enhances the chemosensitivity of U87MG glioma cells to paclitaxelthrough inhibiting the expression of drug resistance related proteins

Objective: To explore the inhibitive effect of asiatic acid (AA) on paclitaxel (PTX)-resistant glioma cells and its possiblemechanism. Methods: The effects of AA on the proliferation and apoptosis of glioblastoma U87MG cells were detected by CCK-8 as-say, Real-time quantitative polymerase chain reaction (qPCR) and Western blotting. The drug-resistant glioma cell line PR-U87MG wasestablished by culturing the cells in concentration-increasing PTX. With U87MG cells as control, the PTX-resistance of PR-U87MGcells was confirmed using CCK-8 assay, and the mRNA and protein levels of MDR1 and LRP were measured with qPCR and westernblotting. PR-U87MG cells were treated with AA, PTX or AA+PTX, and then the cell viability and apoptosis of each group were mea-sured with CCK-8 assay, qPCR and Western blotting. Results: PTX-resistant PR-U87MG cell line was successfully established. AA in-hibited the viability of U87MG and PR-U87MG cells in a dose-dependent manner (P<0.01) and significantly promoted their apoptosis(P<0.01). Compared with the group treated with AA or PTX alone, the group treated with the combination of AA and PTX had signifi-cantly decreased protein levels of PARP1 (P<0.01), drug-resistant related proteins (Pgp-1 and LRP [lung resistance protein], all P<0.01), and markedly increased caspase 3 (P<0.01). Conclusion: AA could effectively enhance the sensitivity of U87MG cells to PTX,and the mechanism may be related to the suppressed expression of drug efflux-associated proteins Pgp-1 and LRP.