lncRNA HOTAIRM1通过miR-17-5p调控慢性粒细胞白血病细胞的生长和转移  

LncRNA HOTAIRM1 regulates growth and metastasis of chronic myeloid leukemia cells via miR-17-5p

作　　者：苗玉迪[1] 高瑛[1] 侯丽敏[1] 胡星星 张虹[1] 赵乔佳杰 魏旭仓[2] MIAO Yudi;GAO Ying;HOU Limin;HU Xingxing;ZHANG Hong;ZHAO Qiaojiajie;WEI Xucang(Department of Hematology,Shaanxi Provincial People’s Hospital,Xi’an 710061,China;Blood Disease Laboratory,Shaanxi Provincial People’s Hospital)

机构地区：[1]陕西省人民医院血液内科,西安710061 [2]陕西省人民医院血液病研究室

出　　处：《山西医科大学学报》2022年第12期1499-1510,共12页Journal of Shanxi Medical University

基　　金：陕西省自然科学基础研究计划项目(2021JM-548)。

摘　　要：目的探究长链非编码RNA(lncRNA)HOX反义基因间RNA髓系1(HOTAIRM1)调控慢性粒细胞白血病(CML)细胞生长和转移的可能分子机制。方法qRT-PCR法检测60例CML患者和60例健康体检者的血清HOTAIRM1和miR-17-5p水平。将CML细胞系(K562细胞)作如下分组:①对照组(未转染)、siRNA-NC组(转染siRNA-NC)、siRNA-HOTAIRM1组(转染siRNA-HOTAIRM1)、pcDNA-NC组(转染pcDNA-NC)和pcDNA-HOTAIRM1组(转染pcDNA-HOTAIRM1);②对照组(未转染)、NC-mimic组(转染NC-mimic)、miR-17-5p-mimic组(转染miR-17-5p-mimic)、NC-inhibitor组(转染NC-inhibitor)和miR-17-5p-inhibitor组(转染miR-17-5p-inhibitor);③siRNA-HOTAIRM1+NC-inhibitor组(共转染siRNA-HOTAIRM1和NC-inhibitor)和siRNA-HOTAIRM1+miR-17-5p-inhibitor组(共转染siRNA-HOTAIRM1和miR-17-5p-inhibitor)。细胞不同转染后,通过MTT实验检测细胞增殖,AnnexinⅤ/PI双染色实验检测细胞凋亡,Transwell实验检测细胞迁移和侵袭,qRT-PCR检测细胞中HOTAIRM1、miR-17-5p、WNT5A和β-catenin的mRNA水平,Western blot检测细胞中WNT5A和β-catenin的蛋白水平。通过双荧光素酶报告基因实验验证HOTAIRM1与miR-17-5p、miR-17-5p与WNT5A的靶向关系。结果与健康体检者相比,CML患者的血清HOTAIRM1水平升高(t=12.253,P<0.001),血清miR-17-5p水平降低(t=15.601,P<0.001)。与对照组或siRNA-NC组相比,siRNA-HOTAIRM1组HOTAIRM1相对表达量、WNT5A和β-catenin的mRNA和蛋白相对表达量、细胞活力、迁移和侵袭细胞数量均降低(P<0.05),而miR-17-5p相对表达量升高(P<0.05)。与对照组或pcDNA-NC组相比,pcDNA-HOTAIRM1组HOTAIRM1相对表达量、WNT5A和β-catenin的mRNA和蛋白相对表达量、细胞活力、迁移和侵袭细胞数量均升高(P<0.05),而miR-17-5p相对表达量降低(P<0.05)。HOTAIRM1靶向抑制miR-17-5p,miR-17-5p靶向抑制WNT5A(P<0.05)。与对照组或NC-mimic组相比,miR-17-5p-mimic组miR-17-5p相对表达量升高,WNT5A和β-catenin的mRNA和蛋白相对表达量降低,细胞活力降低,迁移和侵袭细�Objective To explore the possible molecular mechanism of long non-coding RNA(lncRNA)HOX transcript antisense intergenic RNA(HOTAIRM1)regulating the growth and metastasis of chronic myeloid leukemia(CML)cells.Methods Serum HOTAIRM1 and miR-17-5p levels in 60 CML patients and 60 healthy subjects were detected by qRT-PCR.The CML cell line K562 cells were divided into the following groups:①control group(no transfection),siRNA-NC group(transfected with siRNA-NC),siRNA-HOTAIRM1 group(transfected with siRNA-HOTAIRM1),pcDNA-NC group(transfected with pcDNA-NC)and pcDNA-HOTAIRM1 group(transfected with pcDNA-HOTAIRM1);②control group(no transfection),NC-mimic group(transfected with NC-mimic),miR-17-5p-mimic group(transfected with miR-17-5p-mimic),NC-inhibitor group(transfected with NC-inhibitor)and miR-17-5p-inhibitor group(transfected with miR-17-5p-inhibitor);③siRNA-HOTAIRM1+NC-inhibitor group(co-transfected with siRNA-HOTAIRM1 and NC-inhibitor)and siRNA-HOTAIRM1+miR-17-5p-inhibitor group(co-transfected with siRNA-HOTAIRM1 and miR-17-5p-inhibitor).After different transfection,the cell proliferation was detected by MTT assay,the cell apoptosis was detected by AnnexinⅤ/PI double staining assay,and the cell migration and invasion abilities were detected by Transwell assay.The levels of HOTAIRM1,miR-17-5p,WNT5A mRNA andβ-catenin mRNA in cells were detected by qRT-PCR,and the protein levels of WNT5A andβ-catenin in K562 cells were detected by Western blot.Results Compared with healthy subjects,the serum HOTAIRM1 level was increased in CML patients(t=12.253,P<0.001),while the serum miR-17-5p level was decreased(t=15.601,P<0.001).Compared with control group or siRNA-NC group,the relative expression of HOTAIRM1,the relative mRNA and protein expression of WNT5A andβ-catenin,the cell viability,the number of migrating and invasive cells were all decreased in siRNA-HOTAIRM1 group(P<0.05),while the relative expression of miR-17-5p was increased(P<0.05).Compared with control group or pcDNA-NC group,the relative expression of

关 键 词：慢性粒细胞白血病 生长 转移 长链非编码RNA HOTAIRM1 miR-17-5p WNT5A Β-CATENIN 

分 类 号：R733.7[医药卫生—肿瘤]