siRNA沉默HOTAIR表达对人胶质瘤细胞增殖和侵袭的影响  

The effect of siRNA-mediated silencing of the HOTAIR gene on cell proliferation and invasion in human glioma cell

作　　者：郭志钢[1] 王涵 王冠[1] 李妍哲 田宇[1] 李朝晖[1] GUO Zhi-gang;WANG Han;WANG Guan(Department of Neurosurgery,China-Japan Union Hospital of Jilin University,Changchun 130033,China;Department of Laboratory,Affiliated Hospital of Changchun Chinese Medical University,Changchun 130010,China)

机构地区：[1]吉林大学中日联谊医院神经外二科,吉林长春130033 [2]长春中医药大学附属医院检验科,吉林长春130010

出　　处：《中国实验诊断学》2018年第10期1830-1833,共4页Chinese Journal of Laboratory Diagnosis

基　　金：教育部"新世纪优秀人才支持计划"资助项目(NCET-06-0306);吉林省科技发展计划国际科技合作项目(20130413028GH);吉林省卫生计生青年科研课题(2015Q005)

摘　　要：目的探讨小干扰RNA(small interference RNA,siRNA)沉默HOX转录反义RNA(HOX transcript antisense RNA,HOTAIR)基因对人胶质瘤细胞增殖和侵袭的影响。方法首先利用荧光定量逆转录聚合酶链反应(quantitative reverse transcriptase-polymerase chain reaction,qRT-PCR)检测HOTAIR在人胶质瘤细胞A172和正常星形胶质细胞HA1800中的表达。然后合成si-HOTAIR,用Lipofectamine 2000转染至A172细胞,同时设立阴性对照(negative control,NC)和空白对照(Blank),qRT-PCR检测转染效率。分别采用噻唑蓝(MTT)法、流式细胞术和Transwell侵袭实验检测下调HOTAIR表达对人胶质瘤细胞A172增殖、周期及侵袭的影响。结果 qRT-PCR结果显示,HOTAIR mRNA在A172和HA1800中的相对表达量分别为1.12±0.08和2.67±0.03(P<0.01)。转染siHOTAIR 24h、48h及72h后,A172细胞中HOTAIR表达水平分别下降(38.27±3.52)%、(70.16±0.57)%和(81.54±0.41)%(P<0.01)。MTT结果显示转染si-HOTAIR 48h和72h后,A172细胞的增殖率分别下降(17.38±0.39)%(P<0.05)和(30.54±1.22)%(P<0.01)。流式细胞术显示下调HOTAIR表达能够阻滞A172细胞从G0/G1期向S期转换。Transwell实验结果显示si-HOTAIR组A172细胞穿膜细胞数为36.1±1.5,显著低于NC组的76.8±2.4(P<0.01),降低HOTAIR表达水平能够降低A172细胞的侵袭能力。结论下调HOTAIR表达能够抑制胶质瘤细胞增殖和侵袭能力,HOTAIR有望成为胶质瘤基因治疗的潜在靶点。Objective To determine the in vitro effect of HOTAIR gene silencing by siRNA（small interference RNA）on human glioma cell proliferation and invasion.Methods Expression level of HOTAIR was detected in human glioma cells line A172 and human astrocytes HA1800 by quantitative reverse transcriptase-polymerase chain reaction（qRT-PCR）.Then,HOTAIR siRNA（si-HOTAIR）and a negative control siRNA（NC）were designed and transfected into glioma cell line A172 using Lipofectamine 2000.Transfection efficiency was detected by qRT-PCR.Finally,cell proliferation,cell cycle and cell invasion ability were measured using MTT,flow cytometry and transwell assay respectively.Results The expression level of HOTAIR mRNA was 2.67±0.03 in A172 cells and 1.12±0.08 in HA1800.The HOTAIR mRNA expression level was significantly increased in A172 cells compared with HA1800 cells（P〈0.01）.Si-HOTAIR transfection was observed to significantly reduce the expression level of HOTAIR mRNA in A172 cell.24,48 and 72 hafter si-HOTAIR transfection,the expression level of HOTAIR mRNA decreased by（38.27±3.52）%,（70.16±0.57）% and（81.54±0.41）%（P〈0.01）,respectively.MTT results showed that the viability of A172 cell was significantly inhibited by transfection with si-HOTAIR.The cell proliferation rate in si-HOTAIR group decreased by（17.38±0.39）% 48 hafter transfection（P〈0.05）,and（30.54±1.22）% 72 hafter transfection（P〈0.01）.Flow cytometry results showed that the cell population in the G1 phase was increased but the S phase population was decreased after HOTAIR gene silencing compared with the results observed for the NC cells.The in vitro transwell assay revealed that the number of invasive cells in si-HOTAIR group was significantly decreased than in the control group（P〈0.01）.The invasiveness of A172 cells was significantly suppressed after siHOTAIR transfection.Conclusion The HOTAIR gene was demonstrated to play a key regulatory role in the proliferation and invasion of human glioma cells.HOTAIR co

关 键 词：SIRNA HOTAIR 胶质瘤 增殖 侵袭 

分 类 号：R739.4[医药卫生—肿瘤]