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目的:转录因子STAT5在慢性粒细胞白血病(chronic myeloid leukemia,CML)中组成性激活,并与CML细胞恶性表型密切相关,本研究用cDNA芯片检测方法探讨诱骗核苷酸(decoy ODNs)抑制STAT5对白血病K562细胞株凋亡相关基因的影响。方法:分别提取decoy ODNs处理前后的K562细胞总RNA,逆转录合成Cy3、Cy5标记的cDNA探针,与人14K基因表达谱cDNA芯片(V2.0)杂交,扫描获得数据后,用Genespring软件分析对照组和实验组的差异表达基因,半定量RT- PCR验证cDNA芯片结果。结果:2张cDNA芯片检测重复性良好(R=0.9799)。在13824个标记的基因中,检出413个上调基因,其中包括:TIAF1、GAB1、GYPA、DAPK3、TNFRSF1B、IRF1和PML等在内的18个凋亡相关基因;在检出的332个下调基因中,发现PIM1、CCND2、CCND1、MYC和BCL2L1等在内的11个凋亡相关基因;部分基因经半定量RT-PCR证实与cDNA芯片结果一致。结论:Decoy ODNs抑制STAT5信号通路后可引起K562细胞多种凋亡相关基因的变化,本实验为进一步研究STAT5信号通路所调控的凋亡相关基因提供了依据。
AIM: The transcription factor STAT5 is constitutively activated in chronic myeloid leukemia (CML) and is closely related to the malignant phenotype of CML. In this study, we investigated the effects of decoy ODNs on STAT5 On K562 leukemia cell apoptosis related genes. Methods: Total RNA was extracted from K562 cells before and after treatment with decoy ODNs. Cy3 and Cy5 labeled cDNA probes were reverse transcribed and hybridized with human 14K gene cDNA microarray (V2.0). After the data were obtained by scanning, the data were analyzed by Genespring software The differentially expressed genes in the control and experimental groups were verified by semi-quantitative RT-PCR. Results: Two cDNA chips showed good reproducibility (R = 0.9799). Of the 13824 labeled genes, 413 genes were up-regulated, including 18 apoptosis-related genes including TIAF1, GAB1, GYPA, DAPK3, TNFRSF1B, IRF1 and PML. 11 apoptosis-related genes including PIM1, CCND2, CCND1, MYC and BCL2L1 were found. Some of the genes were confirmed by semi-quantitative RT-PCR to be consistent with that of cDNA microarray. CONCLUSION: Decoy ODNs can inhibit the expression of many apoptosis-related genes in K562 cells after inhibiting the STAT5 signaling pathway. This study provides a basis for further study of apoptosis-related genes regulated by STAT5 signaling pathway.