抑制Akt通路提高视网膜母细胞瘤对卡铂敏感性的实验研究  

Inhibition of Akt activity enhances the sensitivity of retinoblastoma SO-Rb50 cell line to carboplatin in vitro

作　　者：张浩[1] 王怡琛[2] 李彬[2] 杨素红[1] 宫月秋[1] 

机构地区：[1]首都儿科研究所附属儿童医院眼科,100020 [2]首都医科大学附属北京同仁医院北京同仁眼科中心北京市眼科研究所眼科学与视觉科学北京市重点实验室,100005

出　　处：《眼科》2015年第6期405-408,共4页Ophthalmology in China

摘　　要：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 discuss the correlation between Akt activity and the chemosensitivity of retinoblastoma SO-RbSO cells to carboplatin. Design Experimental study. Participants SO-RbS0 cell line. Methods Cytotoxicity of carboplatin was determined using a standard colorimetric cell counting kit-8 （CCK-8） assay. Western Blot was used to detect the changes of Akt, p-Akt, caspase 3, bax, p21 expression with drug treatments. Flow cytometer was used to examine the cell cycle distribution before and after drug treatments. Main Outcome Measures ICS0 values, expression of proteins, cell percentage of each phase of cell cycle. Results ICS0 value of car- boplatin on SO-RbSO cells was （30.1±5.9）mg/L. Preincubating cells with LY294002 to inhibit Akt activity, ICS0 values to carboplatin were lowered to （16.3±0.83）mg/L for 5μM and （8.64±0.11）mg/L for 10 μM LY294002. Treated with Carboplatin or LY294002 alone or their combinations for 48 hours, p-Akt expressions of cells were all reduced, caspase-3, bax expressions were all increased and p21 expressions were increased only with carboplatin treatment. The percentage of each cell phase did not change much due to the applica- tion of LY294002 alone, while the exposure to carboplatin caused a significant accumulation of cells in G2/M phase [from （0.34±0.34）% to （43.62±11.01）%] with a concomitant reduction in G0/G1 phase [from （57,89±2.16）% to （12.11±2.87）%], and the additive combination with LY294002 did not further alter the cell percentage of each cell phase. Conclusion Inhibition of Akt activity can increase the sen- sitivity of SO-Rb50 cells to carboplatin and Akt may be a target for reducing drug resistance that improves the treatment for retinoblas- toma.

关 键 词：视网膜母细胞瘤 药物疗法 卡铂 AKT信号通路 

分 类 号：R739.7[医药卫生—肿瘤]