全反式维甲酸增强顺铂对A549 细胞化疗敏感性及对Survivin mRNA 和COX-2 mRNA表达的影响

All-trans Retinoic Acid Can Enhance Chemosensitivity of Human Non-Small Cell Lung Cancer Cell Line A549 to Cisplatin and Suppress Expression of mRNA and Cox- 2 mRNA in vitro

  • 摘要: 目的:体外观察全反式维甲酸(all-trans retinoic acid ,ATRA)增强顺铂(cisplatin,DDP )对人类非小细胞肺癌细胞株A549 细胞的增殖抑制及对凋亡抑制蛋白Survivin mRNA和环氧化酶-2(cyclooxygenase- 2,COX-2)mRNA 表达的影响。方法:应用不同浓度组DDP(0.5、5、50mg/L)、ATRA(0.1、1、10μ mol/L)以及联合用药组(ATRA 1 μ mol/L,DDP 5mg/L),处理肺腺癌细胞株A549 细胞,采用四甲基偶氮唑盐(MTT)比色法观察不同浓度DDP 组、不同浓度ATRA 组及联合用药组对A549 细胞生长的影响;应用实时荧光定量聚合酶链反应(real-time polymerase chain reaction ,RT-PCR)检测DDP 组、ATRA 组及联合用药组处理前后A549 细胞中Survivin mRNA和COX-2 mRNA 表达变化;应用流式细胞术观察DDP 组、ATRA 组及联合用药组处理前后细胞凋亡率。结果:与空白对照组相比,单独应用DDP 、ATRA 处理A549 细胞均诱导细胞凋亡,且呈浓度依赖性。与单独应用DDP 的作用相比,联合用药组可更显著抑制A549 的增殖,增加细胞的凋亡率(P<0.05),并增强对A549 细胞Survivin mRNA和COX-2 mRNA表达的抑制作用(P<0.05);并且,流式细胞术测定结果显示联合用药组的早期凋亡率(7.37± 3.83)% 、中晚期凋亡率(34.37±2.08)% 、继发性坏死率(7.44± 0.46)% 均较单独应用DDP 组高(3.55± 0.75)% 、(6.62± 0.33)% 、(3.03± 0.05)% ,P 均<0.05。结论:全反式维甲酸能够明显提高非小细胞肺癌对顺铂的敏感性,其机制可能与抑制非小细胞肺癌细胞Survivin mRNA和COX-2 mRNA 的表达有关。

     

    Abstract: Objective:To investigate the influence of cisplatin (DDP) combined with all-trans retinoic acid (ATRA) on the growth and apoptosis of human non-small cell lung cancer A549 cell line and on the expression of Survivin mRNA and COX-2 mRNA in this cell line in vitro. Methods:Different concentrations of DDP (0.5mg/L, 5mg/L, and 50mg/L), different concentrations of ATRA (0.1 μ mol /L, 1 μ mol /L, and 10μ mol /L) and DDP combined wi th ATRA (DDP 5mg/L and ATRA1 μ mol /L) were used in cultures for the pulmonary carcinoma A 549 cell line in different groups. The influence on A 549 growth induced by different concentrations of DDP, different concentrations of ATRA, and DDP combined with ATRA were analyzed by MTT assay. Expression of Survivin mRNA and COX- 2 mRNA levels in the A549 cells were detected by R-T PCR before and after treatment. Flow cytometry (Annexin V-FITC/PI Staining Methods) was used to determine the apoptosis index of the A 549 cells before and after treatment. Results: DDP alone or ATRA alone was found to inhibit the proliferation of the A549 cells ( P<0.05), and the apoptotic effect was increased in a concentration-dependent manner (P<0.05). Compared with DDP alone, DDP combined with ATRA significantly inhibited the proliferation of the A 549 cells ( P<0.05), increased the apotosis rate of the cells and suppressed Survivin mRNA and COX-2 mRNA expression in the cells ( P<0.05). Flow cytometry results showed that the early apoptosis rate, middle and late apoptosis rate and secondary necrosis rate in the DDP combined with ATRA group were (7.37± 3.83)%, (34.37± 2.08)%, and (7.44± 0.46)%, respectively, higher than those of the DDP group ( 3.55± 0.75)%, (6.62± 0.33)%, and (3.03± 0.05)%, P<0.05. Conclusion:ATRA can significantly increase the chemosensitivity of human non-small cell lung cancer cells to DDP. The mechanism may be related to inhibition of Survivin mRNA and COX-2 mRNA expression.

     

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