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目的探讨抑制Akt信号通路活性能否提高视网膜母细胞瘤(RB)SO-Rb50细胞对卡铂的敏感性。设计实验研究。研究对象SO-Rb50细胞株。方法 CCK-8法检测卡铂作用于细胞的IC50值;蛋白印迹法检测Akt、p-Akt、caspase 3、bax、p21蛋白在药物作用前后表达量的改变;流式细胞仪法检测药物作用前后细胞周期的改变。主要指标IC50值,蛋白表达量,细胞周期中各期细胞百分比。结果卡铂作用于SO-Rb50细胞48小时的IC50值为(30.1±5.9)mg/L;分别用5μM和10μM的LY294002抑制细胞Akt信号通路的活性后,卡铂作用于SO-Rb50细胞的IC50值分别为(16.3±0.83)mg/L和(8.64±0.11)mg/L。用卡铂(30mg/L)、LY294002(5μM和10μM)单独或联合作用于SO-Rb50细胞48 h后,细胞Akt活性随LY294002浓度的增加而降低,caspase-3蛋白生成增加,bax蛋白生成增加,p21蛋白在卡铂作用下生成显著增加;LY294002对细胞周期的进程无明显影响,卡铂使G0/G1期细胞百分比从(57.89±2.16)%下降至(12.11±2.87)%,G2/M期细胞从(0.34±0.34)%增加至(43.62±11.01)%,S期细胞无明显变化;两种药物联合作用并未改变卡铂对细胞周期的影响。结论抑制Akt信号通路可提高SO-Rb50细胞对卡铂的敏感性,抑制Akt信号通路可能是提高RB化疗疗效的一个靶点。
Objective To investigate whether inhibiting the activity of Akt signaling pathway can improve the sensitivity of carboplatin to retinoblastoma (RB) SO-Rb50 cells. Design experiment research. Research object SO-Rb50 cell line. Methods The IC50 values of carboplatin on cells were detected by CCK-8 assay. The expression of Akt, p-Akt, caspase 3, bax, p21 protein before and after drug treatment was detected by Western blotting. Cell cycle changes. The main indicators of IC50 value, protein expression, cell cycle percentage of cells in each phase. Results IC50 value of carboplatin in SO-Rb50 cells for 48 hours was (30.1 ± 5.9) mg / L. After IC50 of 5μM and 10μM LY294002 were respectively used to inhibit the Akt signaling pathway, the IC50 of carboplatin in SO-Rb50 cells Values were (16.3 ± 0.83) mg / L and (8.64 ± 0.11) mg / L, respectively. After treated with either carboplatin (30mg / L), LY294002 (5μM and 10μM) alone or in combination with SO-Rb50 cells for 48 h, the Akt activity decreased with the increase of LY294002 concentration and the production of caspase-3 protein and bax protein increased , while the expression of p21 protein was significantly increased under the action of carboplatin. LY294002 had no significant effect on the cell cycle progression. The percentage of cells in G0 / G1 phase decreased from (57.89 ± 2.16)% to (12.11 ± 2.87)%, and the percentage of G2 / M The cell number increased from (0.34 ± 0.34)% to (43.62 ± 11.01)%, and there was no significant change in S phase cells. The combination of the two drugs did not change the effect of carboplatin on cell cycle. Conclusion Inhibition of Akt signaling pathway can enhance the sensitivity of SO-Rb50 cells to carboplatin, and inhibition of Akt signaling pathway may be a target for improving the efficacy of RB chemotherapy.