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目的 应用微阵列基因芯片比较三氧化二砷 (As2 O3)对HL60细胞作用后细胞周期素相关基因表达谱的变化。方法 As2 O3作用于HL60细胞 ,TRIzol试剂一步法提取作用前后HL60细胞的mRNA ,经逆转录反应 ,用Cy3标记的脱氧三磷酸尿苷 (Cy3 dUTP)和Cy5 dUTP两种不同的荧光染料将处理前后的HL60细胞mRNA分别标记成两种DNA探针 ,并与载有一组靶基因的表达谱芯片进行杂交 ,通过扫描分析筛选As2 O3作用前后HL60细胞表达有差异的基因 ;用电镜和原位凋亡检测法、流式细胞术及DNA凝胶电泳等方法检测细胞凋亡。结果 筛选出HL60细胞在As2 O3作用前后表达有差异的基因共 82条 ,主要涉及细胞周期调控类、凋亡相关蛋白及应激反应蛋白类、信号传导类等基因 ,其中表达上调的有 34条 ,表达下调的有 48条。流式细胞仪检测发现终浓度 1 5μmol/L的As2 O3对HL60细胞诱导凋亡的作用很强。对照组的细胞凋亡率为 1 7% ,而实验组细胞的凋亡率为 2 6 1 %。结论 细胞周期素B1、增殖细胞核抗原、类胰岛素生长因子结合蛋白等可能参与As2 O3诱导HL60细胞凋亡的发生机制
Objective To compare the expression changes of cyclin-related genes in HL60 cells treated with As 2 O 3 by microarray microarray. Methods The effect of As2 O3 on HL60 cells was detected by TRIzol reagent. The mRNA of HL60 cells was detected by reverse transcription reaction before and after treatment with Cy3-labeled deoxyuridine triphosphate (Cy3 dUTP) and Cy5 dUTP HL60 cells were labeled with two kinds of DNA probes respectively and hybridized with a set of expression microarray containing target genes. The differentially expressed genes of HL60 cells were screened by scanning analysis. Detection, flow cytometry and DNA gel electrophoresis and other methods to detect apoptosis. Results The results showed that there were 82 genes differentially expressed in HL60 cells before and after As2 O3 treatment, which mainly involved in cell cycle regulation, apoptosis related proteins, stress response proteins and signal transduction genes. Among them, 34 , Down regulation of 48. Flow cytometry showed that As 2 O 3 at a final concentration of 15 μmol / L had a strong effect on inducing apoptosis in HL60 cells. The apoptosis rate of the control group was 17%, while the apoptosis rate of the experimental group was 26%. Conclusion Cyclin B1, proliferating cell nuclear antigen, insulin-like growth factor-binding protein may be involved in the mechanism of As2 O3-induced HL60 cell apoptosis