奥沙利铂对人肺腺癌A549细胞增殖凋亡及Survivin和Cyclin D1表达的影响

Effects of Oxaliplatin on cells growth and apoptosis and Survivin,Cyclin D1 expressions of lung adenocarcinoma cell line A549

目的研究奥沙利铂(oxaliplatin,L-OPH)对人肺腺癌A549细胞株的生长抑制作用和对Survivin、Cyclin D1基因表达的影响,探讨奥沙利铂抑制肺癌细胞增殖的机制。方法以不同浓度(0、7.81、15.63、31.25、62.50和125.00μg/mL)的奥沙利铂干预体外培养的肺腺癌细胞株A549,分别于24、48和72 h后,采用MTT方法检测细胞光密度值,分析A549细胞的增殖活性。采用流式细胞术检测其细胞周期变化。采用Hoechst33342染色方法在荧光显微镜下观察细胞形态变化。采用RT-PCR、Western-blotting方法检测Survivin和Cyclin D1的mRNA及蛋白的表达情况。结果①奥沙利铂能有效抑制A549细胞的增殖(P<0.05),呈时间-剂量依赖性。②奥沙利铂能改变A549细胞的周期分布并诱导其凋亡(P<0.05),呈时间-剂量依赖性。③奥沙利铂作用48 h后A549细胞Survivin、Cyclin D1基因及蛋白的表达明显低于对照组,差异有统计学意义(P<0.05),且该抑制效应呈浓度依赖性。结论奥沙利铂可以抑制A549细胞株的增殖、改变其周期分布、诱导其凋亡,且呈一定的剂量-时间依赖性;其机制可能与奥沙利铂下调Survivin和Cyclin D1基因的表达有关。

【Objectives】 To study the inhibition of Oxaliplatin on cells growth of A549 cell line and the effects on the expressions of Survivin,Cyclin D1,discuss the possible mechanism of inhibition of Oxaliplatin on the proliferation of lung cancer cells.【Methods】 MTT assay was used to measure the optical density（OD） of A549 cells to analyze the proliferation of A549 cells,when treated with different concentrations（0,7.81,15.63,31.25,62.50,125.00 μg/mL） of Oxaliplatin for 24,48 and 72 h in vitro.Cell-cycle kinetics were analyzed by flow cytometry（FCM）.Hoechst 33342 was used to observe the changes of cell by using the fluorescence microscope.The expressions of Survivin and CyclinD1 mRNA and relative proteins activity were analyzed by Reverse transcription-polymerase chain reaction（RT-PCR） and Western-blotting.【Results】 The proliferation of A549 cells were effectively inhibited（P 0.05） by Oxaliplatin in a time-dose dependent manner.Oxaliplatin can change the distribution of cell cycle,induce the apoptosis of A549 cells（P 0.05）,in a time-dose dependent manner.After treated by Oxaliplatin for 48 hours,the expressions of genes and proteins of Survivin and Cyclin D1 in A549 cells were obviously lower than those in normal A549 cells（P 0.05）,while the inhibitory effects presented in dose-dependent manner.And the differences were statistically significant.【Conclusions】 Oxaliplatin can inhibit the proliferation of A549 cell line,change the distribution of cell cycle and induce the apoptosis of A549 cells,in a time-dose dependent manner.The mechanism may be related to the down-regulation of Survivin and Cyclin D1 genes expressions treated by Oxaliplatin.