松胞素B对白血病K562细胞基因表达谱的影响

Analysis of Gene Expression Patterns of Leukemia K562 Cells after Cytochalasin B Treatment

背景与目的:近年来兴起的DNA芯片技术能在一次杂交中同时监测数以千计的基因表达,能大大加速肿瘤药物作用机制的研究和药物治疗靶点的发现。本研究的目的是利用基因表达谱芯片研究松胞素B(cytochalasinB,CB,旧称细胞松弛素B)对白血病K562细胞基因表达谱的影响。方法:利用限制性显示RCR(restrictiondisplayPCR,RD-PCR)技术扩增K562细胞cDNA片段,取其中277个cDNA片段按设计的矩阵打印在玻片上,制成基因芯片;用CB处理K562细胞24h,分别提取对照组和处理组细胞总RNA,将两组等量的细胞总RNA纯化为mRNA,反转录为cDNA,利用RD-PCR技术进行扩增标记;与芯片杂交后扫描分析处理前后表达差异的基因。结果:在所收集的探针中,对照组和处理组细胞间存在差异表达的基因。共筛选到CB处理后表达下调的基因18条。结论:CB诱导后的K562细胞部份基因表达呈下调趋势,其中大部分基因与细胞增殖、信号转导、转录调节相关。

BACKGROUND &OBJECTIVE: The advanced technique of DNA microarray makes it possible to monitor the expression of thousands of genes simultaneously in one hybridization experiment. This technique accelerates demonstration of anti tumor drug mechanisms and discovery of new drug targets. This study was designed to investigate the differential gene expression of K562 cells after cytochalasin B treatment using cDNA microarray. METHODS: Restriction display polymerase chain reaction (RD PCR) products of 277 human genes were spotted on a glass slide in microarray. K562 cells grew in RPMI 1640 medium with 10 μg/ml cytochalasin B. After 24 hours, the total RNA was isolated from K562 cells, and mRNA was purified. Both mRNA from the treated K562 cells and the controlled K562 cells were reversely transcribed into cDNA and labeled with two different fluorescence dyes: Cy5 or Cy3, using a method of restriction digestion and PCR labeling (RD PCR). The probes were hybridized to the cDNA microarrays. After high stringent washing,the cDNA microarray was scanned for the fluorescent signals and showed difference between the two cells. RESULTS: Among the 277 target genes, 18 down regulated genes were identified after cytochalasin B treatment. CONCLUSION: There is a consistent tendency toward lower expressed genes in partial K562 cells after cytochalasin B treatment. Most down regulated genes were correlated with cell proliferation, signal transduction, and transcription factor.