多西紫杉醇联合二烯丙三硫协同诱导前列腺癌细胞凋亡的研究

Synergetic Cell Apoptotic Effect Induced by Docetaxel Combined with Diallyl Trisulfide in Prostate Cancer Cells

【目的】探讨多西紫杉醇联合二烯丙三硫(DATS)对前列腺癌PC-3细胞的协同杀伤效应及其机制。【方法】PC-3细胞接种96孔板,分别给予多西紫杉醇(5～60 nmol/L)和DATS(5～60μmol/L)单药处理24、48、72 h;联合用药采用15μmol/L DATS联合不同浓度多西紫杉醇(5～60 nmol/L)处理48 h。MTT法检测多西紫杉醇和(或)DATS对PC-3细胞生长的影响。DAPI染色、流式细胞仪检测多西紫杉醇、DATS及二者联合对PC-3细胞凋亡和细胞周期的影响;Western-blot测定Bcl-2、Bax、Bcl-xL/xS及细胞色素C、caspase-9、caspase-3的表达及活性变化。【结果】多西紫杉醇、DATS均能抑制PC-3细胞的生长,DATS可明显提高多西紫杉醇的杀伤效应,DATS(15μmol/L)联合多西紫杉醇(10 nmol/L)作用48 h的细胞生存率为(50.1±1.0)%,低于多西紫杉醇(67.4±1.0)%和DATS(69.2±1.8)%,二者具有协同抑瘤作用;联合组使细胞凋亡率及G2/M期细胞比率明显增高,并更能显著抑制Bcl-2、Bcl-xL的表达,而且联合组使细胞色素C的释放及caspase-9、caspase-3的裂解活化时间提前了8 h。【结论】多西紫杉醇及DATS共同诱导细胞凋亡是二者发挥协同效应抑制前列腺癌细胞生长的重要机制,其可能与细胞周期阻滞、Bcl-2、Bcl-xL的表达降低、细胞色素C的释放及caspase-9、caspase-3的活性变化有关。

[Objective] To study the killing effect of docetaxel combined with diallyl trisulfide （DATS） in prostate cancer cell line PC-3 and further to determine the mechanism. [ Methods ] PC-3 cells were treated with various concentrations of docetaxel or （and） DATS. Cell viability was estimated using MTr assay. Apoptotic cells were detected following staining with DAPI. The percentage of sub G1 and cell cycle were confirmed by flow cytometry. Protein levels of apoptosis regulating proteins were determined using Western bolt including Bcl-2, Bax, Bcl-xL/xS, cytochrome c, caspase-9, and caspase-3. [Results] Docetaxel or DATS alone could inhibit the proliferation of PC-3 cells. For combination group, the cell viability was decreased to （50.1±1.0）% to compare with （67.4 ± 1.0）% of docetaxel at 10 nmol/L and （69.2 ± 1.8）% of DATS at 15 μmol/L separately and there was a synergistic effect between these two drugs. Apoptotic rate and the cell number in S-phase and G-phase of the combination group were higher than other two drugs. The combination treatment could significantly reduce the Bcl-2, Bcl-xL expression and promote the release of cytochrome c and the cleavage of caspase-9 and caspase-3. [ Conclusion ] The combination of docetaxel with DATS can inhibit the proliferation of prostate cancer cells congenerously by inducing apoptosis, which may be correlated with S-and G2-phase arrest, the down-regulation of Bcl-2, Bcl-xL, the release cytochrome c and the activation of caspase-9 and caspase-3.