摘要
目的:探讨lncRNAASB16-AS1在结直肠癌(CRC)中的表达及其对CRC细胞增殖的影响。方法:采用逆转录定量PCR(qRT-PCR)检测26对CRC及其配对癌旁组织、4株CRC细胞(HT-29、SW-480、SW-620及LOVO)中lncRNAASB16-AS1的表达,选择2株lncRNAASB16-AS1表达最高的CRC转染ASB16-AS1干扰片段作为干扰组,实验设置对照组(不转染)及阴性对照组(转染阴性对照干扰片段),采用CCK-8法检测转染0、24、48、72及96h时细胞的增殖情况,qRT-PCR法检测转染48h时细胞miR-185-5p表达;将含有ASB16-AS1野生型序列或突变序列的质粒与miR-185-5pmimic或阴性对照mimic共转染至293T细胞,转染24h后采用双荧光素酶法检测各组细胞的荧光强度,观察lncRNAASB16-AS1对miR-185-5的调控作用。结果:qRT-PCR结果显示,lncRNAASB16-AS1在CRC组织中的表达水平显著高于癌旁组织(P<0.01),CRC细胞HT-29及LOVO中lncRNAASB16-AS1的表达水平显著高于SW-480及SW-620(P<0.01);与阴性对照组比较,转染ASB16-AS1干扰片段的HT-29和LOVO细胞中lncRNAASB16-AS1的表达水平显著降低(P<0.01);转染48h开始,干扰组HT-29和LOVO细胞的增殖水平显著低于阴性对照组(P<0.05);转染48h时,下调ASB16-AS1后细胞中miR-185-5p表达水平显著升高(P<0.05);双荧光素酶结果显示,野生型ASB16-AS1和miR-185-5pmimic共转染,显著降低293T细胞中双荧光素酶的活性(P<0.05)。结论:CRC的发生发展的机制与lncRNAASB16-AS1上调有关。
Objective:To evaluate the expression of lncRNA ASB16-AS1 in colorectal cancer(CRC)tissues and investigate the effect of lncRNA ASB16-AS1 on the proliferation of CRC cells.Method:The expression of lncRNA ASB16-AS1 in 26 pairs of CRC and paraplastic tissues were determined by qRT-PCR.The expression of lncRNA ASB16-AS1 in 4 CRC cell lines was also detected.The two lncRNA ASB16-AS1-high-expressed cells were transfected with lncRNA ASB16-AS1 siRNA or NC siRNA,respectively.Cell proliferation activities in 0 h,24 h,48 h,72 h or 96 h were examined by CCK-8 assay.Dual-Luciferase reporter assay was also used to screen microRNA response elements of miR-185-5p in lncRNA ASB16-AS1.The expression of miR-185-5p was also detected.Result:The level of lncRNA ASB16-AS1 was significantly increased in CRC tissues compared with paraplastic tissues(P<0.01).HT-29 and LOVO cells showed the highest level of lncRNA ASB16-AS1 in the 4 CRC cell lines.Transfect with lncRNA ASB16-AS1 siRNA significantly inhibited cell proliferation in the 2 CRC cells when compared with the NC siRNA transfected cells.The expression of miR-185-5p was significantly up-regulated by the inhibition of lncRNA ASB16-AS1(P<0.01).Moreover,dual-Luciferase reporter assay showed that co-transfection with wild type lncRNA ASB16-AS1 and miR-185-5p mimic significantly reduced the relative luciferase activity when compared with the other groups.Conclusion:lncRNA ASB16-AS1 is closely related to the occurrence and development of CRC,lncRNA ASB16-AS1 may be a novel target in the prevention and treatment of CRC.
作者
石峰
唐青
魏晓为
SHI Feng;TANG Qing;WEI Xiaowei(Funing People's Hospital,Yancheng 224000,Jiangsu,China;Department of Oncology,Nanjing First Affiliated Hospital of Nanjing Medical University,Nanjing 210006,Jiangsu,China)
出处
《贵州医科大学学报》
CAS
2019年第10期1167-1172,共6页
Journal of Guizhou Medical University
基金
国家自然科学基金面上项目(81773240)
江苏省自然科学基金面上项目(BK20181118)
南京市卫生青年人才第二层次
