曲格列酮对前列腺癌PC-3细胞转移潜能影响分子机制探讨

Metastatic potential effect of troglitazone on human prostate cancer PC-3 cells and its molecular mechanism

目的:探讨曲格列酮对人前列腺癌PC-3细胞增殖、侵袭和迁移的影响。方法:PC-3细胞经不同浓度曲格列酮(0、1×10-6、1×10-5和1×10-4 mol/L)处理后,采用MTT法检测各组细胞增殖水平;细胞划痕实验和细胞侵袭试验检测侵袭及迁移能力的改变;蛋白质印迹法检测细胞Id1、MMP-2和MMP-9蛋白的表达水平。结果:MTT检测显示,不同浓度曲格列酮对PC-3细胞抑制作用明显,并呈浓度和时间依赖性,P<0.001。细胞划痕实验显示,PC-3细胞经不同浓度曲格列酮作用后,1×10-6 mol/L组细胞迁移率为(70.67±10.42)%,1×10-5 mol/L组为(49.00±6.00)%,1×10-4 mol/L组为(20.33±5.05)%,明显低于对照组的(80.5±6.86)%,差异有统计学意义,P<0.01,且呈显著剂量依赖关系,F=78.57,P<0.001。细胞体外侵袭实验显示,1×10-6 mol/L组穿越Transwell滤膜的PC-3细胞数为85.17±15.28,1×10-5 mol/L组为70.33±7.63,1×10-4 mol/L组为52.50±8.87,明显低于对照组的137.67±15.09,差异有统计学意义,P<0.001,且呈显著剂量依赖关系,F=53.99,P<0.001。细胞体外迁移实验显示,1×10-6 mol/L组迁移至Transwell下室的细胞数为88.83±17.02,1×10-5 mol/L组为71.67±9.44,1×10-4 mol/L组为54.50±9.35,明显低于对照组的142.00±17.40,差异有统计学意义,P<0.001,且呈显著剂量依赖关系,F=44.72,P<0.001。蛋白质印迹法检测结果显示,对照组Id1蛋白相对表达量为12.67±7.23,1×10-6 mol/L组为24.67±6.51,1×10-5 mol/L组为36.67±6.11,1×10-4 mol/L组为49.33±4.93;对照组MMP-2蛋白相对表达量为9.00±4.36,1×10-6 mol/L组为25.33±8.62,1×10-5 mol/L组为42.00±7.55,1×10-4 mol/L组为58.67±12.06;对照组MMP-9蛋白相对表达量为13.00±3.61,1×10-6 mol/L组为25.67±6.43,1×10-5 mol/L组为38.33±8.96,1×10-4 mol/L组为53.67±6.43。曲格列酮显著下调PC-3细胞Id1、MMP-2和MMP-9蛋白表达水平,P值均<0.05。结论:曲格列酮能有效抑制PC-3细胞增殖与侵袭迁移,同时下调Id1、MMP-2和MMP-9蛋白表达水平。

OBJECTIVE： To investigate the inhibitory effect of troglitazone on the proliferation,migration and inva- siveness of prostate cancer PC-3 cells. METHODS： PC-3 cells were cultured with different concentration of troglitazone（0, 1 × 10-6 , 1 × 10-s and 1 ×10-4 mol/L）. Cell proliferation was measured by MTT methods ; cell migration and invasiveness were measured by invasion and migration transwell system;protein expression of Idl,MMP-2 and MMP-9 were measured by Western blot. RESULTS： MTT assay showed that troglitazone, in a dose- and time- dependent manner, inhibited the growth of PC-3 cells（P〈0. 001）. Cell scratch assay showed that the migration rates of guoup 1 ×10-6 , 1 × 10-5 and 1 × 10-4 mol/L were （70.67 ± 10. 42） %, （49.00± 6.00） % and （20.33 ± 5.05） %, which were significantly lower than those in the control groups（P〈0.01）. They show a typical dose-effect relationship（F= 78.57, P〈0. 001）. Invasiveness assay showed that the numbers of invading of guoup 1× 10-6 , 1 × 10-5 and 1 ×10-4 mol/L were 85.17± 15.28,70.33 ± 7. 63 and 52.50 ±8. 87 respectively, which were significantly lower than those in the control groups （137.67 ±15.09, P〈 0. 001）. They showed a typical dose effect relationship（F= 53.99 ,P〈0. 001）. Migration assay showed that the numbers of migrating of guoup 1 × 10-6 , 1 × 10-5 and 1 × 10-4 mol/L were 88.83± 17.02,71.67 ± 9.44 and 54.50 ± 9.35 respec- tively, which were significantly lower than those in the control groups（142.00±17.40, P〈0. 001）. They showed a typical dose effect relationship（F=44.72,P〈0. 001）. Western blot showed that the protein expressions of Idl in different gu- oups was 12.67±7.23 of control, 24. 67± 6.51 of group 1 ×10-6 mol/L,36. 67±6. 11 of group 1 ×10-5 mol/L and 49.33±4.93 of group 1 × 10-4 mol/L. The protein expression of MMP-2 in different guoups were 9.00±4.36 of control, 25.33±8.62 of group 1 × 10-6 mol/L,42. 00±7. 55 of group 1 × 10-6 mol/L and 58.67±12.06 of group 1 × 10-4 mol/L. The protein expression of MMP-9 in different guoups were 13.00±3.61 of control, 25.67 ±6.43 of group 1× 10-6 mol/L, 38. 33±8.96 of group 1 × 10-5 mol/L and 53.67± 6.43 of group 1× 10-4 mol/L. The protein expression of Idl, MMP-2 and MMP-9 were all decreased significantly after treatment compared with the control cells（P〈0.05）. CONCLUSIONS： Troglitazone could inhibit the proliferation,migration and invasiveness of prostate cancer PC-3 cells. At the same time,it could down-regulates the expression of Idl, MMP-2 and MMP-9.