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PLK1基因沉默对K562/A02细胞周期、增殖和耐药性的影响
引用本文:刘林,张敏,邹萍,田蕾,刘芳. PLK1基因沉默对K562/A02细胞周期、增殖和耐药性的影响[J]. 中国实验血液学杂志, 2006, 14(2): 241-246
作者姓名:刘林  张敏  邹萍  田蕾  刘芳
作者单位:华中科技大学同济医学院附属协和医院血液病研究所,武汉,430022
摘    要:为了探讨小干扰RNA(siRNA)对K562/A02细胞Polo样激酶1(PLK1)基因的抑制作用及其对细胞周期、细胞增殖和耐药性的影响,将构建的针对PLK1mRNA的siRNA真核质粒,导入K562/A02细胞,用RT-PCR和Western-blot方法分析PLK1基因在转染前后的表达差异;台盼蓝拒染试验检测细胞存活率;流式细胞术(FACS)检测细胞周期和细胞内阿霉素(ADM)的积累量;MTT试验检测细胞对ADM的药物敏感性。结果显示,与对照组相比,转染24和48小时后,siRNA-PLK1质粒转染组的PLK1mRNA下降(34.7±2.1)%和(56.6±1.5)%,蛋白水平下降(49.9±3.2)%和(62.1±1.7)%;转染24和48小时后,细胞存活率下降了30%和59%;转染48小时后,G2/M期细胞比例提高了2.77倍,同时细胞内ADM积累量显著提高;对ADM药物敏感性的相对率为73.8%。结论:PLK1基因沉默能有效抑制K562/A02细胞生长,使细胞周期停滞在G2/M期,同时使得细胞内ADM积累量提高,对ADM敏感性增强。

关 键 词:PLK1 基因沉默  siRNA 质粒  K562 细胞/A02细胞  细胞周期  耐药性
文章编号:1009-2137(2006)02-0241-06
收稿时间:2005-04-18
修稿时间:2006-01-23

Effect of PLK1 Gene Silence on Cell Cycle, Proliferation and Drug Resistance in K562/A02 Cells
LIU Lin,ZHANG Min,ZOU Ping,TIAN Lei,LIU Fang. Effect of PLK1 Gene Silence on Cell Cycle, Proliferation and Drug Resistance in K562/A02 Cells[J]. Journal of experimental hematology, 2006, 14(2): 241-246
Authors:LIU Lin  ZHANG Min  ZOU Ping  TIAN Lei  LIU Fang
Affiliation:Department of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China.
Abstract:The study was purposed to investigate the effect of small interference RNA (siRNA) targeting Polo-like kinase 1 (PLK1) gene on cell cycle progression, proliferation and drug resistance in K562/A02 cells. siRNA plasmid vector specifically targeting PLK1 gene with enhanced green fluorescence protein (EGFP) was transfected into K562/A02 cells. Expressions of PLK1 mRNA and protein were assayed by RT-PCR and Western-blot; cell proliferation was evaluated by direct cell counting after trypan blue staining. Cell cycle and intracellular adriamycin (ADM) accumulation was determined by flow cytometry; 50% inhibition concentration (IC50) of ADM on K562/A02 cells was determined by MTT method. The results showed that, as compared with control cells, siRNA plasmid reduced PLK1 mRNA expression by (34.7 +/- 2.1)% for 24 hours and by (56.6 +/- 1.5)% for 48 hours, PLK1 protein significantly decreased simultaneously by (49.9 +/- 3.2)% and by (62.1 +/- 1.7)%. After being transfected for 24 and 48 hours, the rate of survival cells decreased by 30% and 59% respectively. Forty-eight hours after transfection, the ratio of K562/A02 cells at G2/M increased by 2.77-fold, at the same time, intracellular ADM accumulation increased and the relative efficiency of K562/A02 cells to ADM was 73.8%. It is concluded that PLK1 gene silence can inhibit K562/A02 cell proliferation, induce cell cycle arrest at G2/M, and increase intracellular ADM accumulation, so that enhance cell sensitivity to ADM.
Keywords:PLK1 gene silence   siRNA plasmid   K562 cell/A02 cell   cell cycle   drug resistance
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