CDK抑制剂NSC649890联合顺铂对人骨肉瘤细胞U2-OS的抑制作用

CDK inhibitor NSC 649890 combined with or without cisplatin to inhibit human osteosarcoma U2-OS cell. An experimental study

[目的]探讨CDK抑制剂NSC 649890联合顺铂对人骨肉瘤U2-OS细胞抑制作用及可能的机制。[方法]MTT法检测分别采用50、100、200、400、800 nmol/L的NSC 649890 2、4、6、8、10μg/ml顺铂、50 nmol的NSC649890联合2μg/ml顺铂、300nmol的NSC 649890联合6μg/ml顺铂培养48 h后细胞增殖情况,金氏q值评价联合用药效应;流式细胞仪、Western blotting检测分别采用300 nmol/L的NSC 649890、6μg/ml顺铂及二者联合培养48 h后细胞凋亡及凋亡通路相关蛋白表达情况,以上实验均以单纯U2-OS细胞作为对照组。[结果]U2-OS细胞经不同浓度顺铂、NSC 649890单独处理48 h后,细胞抑制率随两种药物浓度增加而显著升高,比较差异均有统计学意义(P<0.05)。在联合用药组中,顺铂与NSC 649890联合浓度(2μg/ml+50 nmol/L)组两药为协同作用,(6μg/ml+300nmol/L)组两药为相加作用。流式细胞仪检测显示,顺铂组、NSC 649890组及联合组细胞凋亡率显著高于对照组(P<0.05)。Western blotting检测结果显示,与对照组相比,顺铂和NSC 649890单独作用后均可下调Bcl-2蛋白、procaspase-3蛋白的表达水平及上调Bax蛋白的表达水平,差异均有统计学意义(P<0.05),而两者联合作用时效果更加显著(P<0.05)。[结论]NSC 649890能增强顺铂对人骨肉瘤U2-OS细胞的抑制作用。

[Objective] To investigate the effect of CDK inhibitor NSC 649890 combined with cisplatin on human osteo- sarcoma cell line U2-OS and its possible mechanism. [Methods] The cytostatic effect of NSC 649890 （50, 100, 200, 400, and 800 nmol/L） , cisplatin （2, 4, 6, 8, and 10 μg/ml） , and NSC 649890 combined with cisplatin （50 nmol＋2 Wg/ml, 300 nmol＋6 wg/ml） on U2-OS ceils were evaluated by MTr assay at 48 hours after culture. The drug combination effect was evalu- ated using Jin＇ s Q formula. The influence of NSC 649890 （300 nmol/L） , cisplatin （6 μg/ml） , and NSC 649890 （300 nmol/L） combined with cisplatin （6 μg/ml） on the apoptosis of U2-OS cells were determined by flow cytometry at 48 hours after culture.The expression of apoptosis signaling pathway-related proteins in U2-OS cells treated with NSC 649890 （300 nmol/L） , cisplatin （6 I.Lg/ml） , and NSC 649890 （300 nmol/L） combined with cisplatin （6 μg/ml） were measured by Western blotting at 48 hours after culture. The untreated U2-OS cells were involved as control group. [Results] The separate treatment of U2-OS with different concentrations of cisplatin and NSC 649890 for 48 h showed that the cell growth inhibition rate was dose dependent （P〈0.05） . In the drug combination group, a synergistic effect was presented in the 2μg/ml cisplatin combined with 50 nmol/L NSC 649890 group, while an additive effect was presented in the 6μg/ml cisplatin combined with 300 nmol/L NSC 649890 group. The results of flow cytometry showed that the cisplatin group,NSC 649890 group, and combination group had significantly higher apoptosis rates than the control group （P〈0.05, respectively） . The results of Western blot- ting indicated that compared with the control group, cisplatin or NSC 649890 alone could down-regulate the expression of Bcl-2 protein and procaspase-3 protein and up-regulate the expression of Bax protein （P〈0.05） , however, a combina- tion of the two had a more significant effect （P〈0.05） .[Conclusions] NSC 649890 can enhance the inhibitory effect of cisplatin on human osteosarcoma U2-OS cells.