DHX9调控白血病细胞增殖及凋亡的作用机制研究  

Mechanism of DHX9 in Regulating Proliferation and Apoptosis of Leukemia Cells

作　　者：黄楠芳 宋扬[1] 郭娟 张征[1] 李晓[1] 许峰 HUANG Nanfang;SONG Yang;GUO Juan(Department of Hematology,Shanghai Sixth People′s Hospital Affiliated to Shanghai Jiao Tong University School of Medicine,Shanghai 200233,China)

机构地区：[1]上海交通大学医学院附属第六人民医院血液内科,200233

出　　处：《医学研究杂志》2023年第9期29-34,共6页Journal of Medical Research

基　　金：国家自然科学基金资助项目(81770122)。

摘　　要：目的研究DHX9在白血病患者中的表达水平,以及对白血病细胞增殖及凋亡的影响和作用机制。方法分析GEO数据库中白血病及对照样本DHX9基因表达情况,探索其表达差异。通过慢病毒感染人白血病细胞系K562,构建DHX9敲低细胞系。采用CCK-8法检测细胞增殖,Annexin V-PI法检测细胞凋亡,基因表达谱筛查DHX9潜在靶向通路,通过实时荧光定量聚合酶链反应(real-time quantitative polymerase chain reaction,RT-qPCR)和Western blot法验证DHX9调控的机制。结果分析GEO数据库中的MILE研究(GSE13159),该研究纳入AML、MDS、CML及正常对照的基因表达谱分析,结果显示,AML组DHX9基因表达水平较对照组明显增高(P=0.007),较MDS组及CML组也明显增高(P均<0.001);而CML组DHX9表达明显低于AML组、MDS组及对照组(P均<0.001)。本研究构建了DHX9敲低的白血病细胞株K562(DHX9相对表达量为0.48±0.06,P<0.01)。与对照组比较,DHX9敲低的K562细胞呈现明显增高的自身凋亡率(P<0.001),对抗肿瘤药物阿扎胞苷(5-azacitidine,AZA)的凋亡敏感度也明显增加(P<0.05)。CCK-8增殖实验显示,DHX9敲低细胞增殖能力减弱(P<0.001)。通过基因表达谱分析获得了2264个差异基因,生物信息学分析显示,差异表达基因主要富集在癌症通路、凋亡和p53通路(P均<0.001)。Western blot法检测结果显示,DHX9敲低增加凋亡相关蛋白caspase-9的表达,降低Bcl-2表达。结论从正常对照到MDS再到AML,DHX9表达逐步增加预示较高的肿瘤增殖度和白血病转化风险,DHX9可能通过抑制p53介导的细胞凋亡促进白血病细胞增殖。Objective To investigate the expression level of DHX9 in patients with leukemia,and the effect and mechanism on the proliferation and apoptosis of leukemia cells.Methods The expression of DHX9gene between leukemia and control samples in GEO database was analyzed to explore the difference in expression.Human leukemia cell line K562 was infected with lentivirus to construct DHX9 knockdown cell line.CCK-8 assay was used to detect cell proliferation and Annexin V-PI flow cytometry was used to detect cell apoptosis.Gene expression profiles were used to screen potential DHX9 targeting pathways.Real-time quantitative polymerase chain reaction(RT-qPCR)and Western blot were used to verify the mechanism of DHX9 regulation.Results The MILE study(GSE13159)in GEO database was analyzed.The gene expression profiles of AML,MDS,CML and normal controls were included in this study,and the results showed that the expression level of DHX9gene in AML patients was significantly higher than that in the control group(P=0.007).In addition,the expression level of DHX9 in AML patients was significantly higher than that in MDS group and CML group(all P<0.001).The expression level of DHX9 in CML group was significantly lower than that in AML group,MDS group and control group(all P<0.001).This study constructed leukemia cell line K562 with DHX9 knockdown(the relative expression of DHX9 was 0.48±0.06,P<0.01).Compared with the control cells,K562 cells with DHX9 knockdown showed significantly higher self-apoptosis rate(P<0.001),and the apoptosis sensitivity of antitumor drug 5-azacytidine(AZA)was also significantly increased(P<0.05).CCK-8 proliferation assay showed that the proliferation ability of DHX9 knockdown cells was reduced(P<0.001).2264differentially expressed genes were obtained by gene expression profiles analysis.Bioinformatics analysis showed that differentially expressed genes were mainly enriched in the cancer pathway,apoptosis and p53 pathway(all P<0.001).Western blot analysis showed that DHX9 knockdown increased the expression of

关 键 词：DHX9 急性髓系白血病 P53 细胞凋亡 

分 类 号：R552[医药卫生—血液循环系统疾病]