胎肝特异性癌胚RNA——AK003710的筛选及功能研究

目的 筛选胎肝特异性癌胚RNA,探讨其对肝癌细胞增殖、侵袭能力的影响.方法 对小鼠肝发育和肝再生的长链非编码RNA(IncRNA)芯片数据进行生物信息学分析,筛选得到在胎肝和再生肝脏中均高表达的候选IncRNA,利用实时定量PCR在小鼠肝癌组织中对候选IncRNA进行表达水平检测,综合芯片分析及实时定量PCR结果,选择表达差异最明显的IncRNA行进一步研究.首先在胎肝和肝再生模型中验证该IncRNA的表达,然后利用siRNA技术干扰该IncRNA,再通过EdU标记技术和Transwell实验分别检测细胞的增殖能力和侵袭能力.结果 生物信息学分析筛选得到在胎肝和再生肝脏中均高表达的7条IncRNA,其中3条在小鼠肝癌组织中较正常肝组织高表达,3条表达无差异,1条无表达;高表达的3条IncRNA中,IncRNA-AK003710在小鼠肝发育和肝再生芯片中的差异表达倍数最高,可作为进一步研究对象.进一步验证显示IncRNA-AK003710在胎肝及多种肝损伤修复模型中表达上调;功能实验表明对IncRNA-AK003710靶向干扰后能降低小鼠肝癌细胞Hepa1-6的增殖和侵袭能力.结论 IncRNA-A003710在胎肝、再生肝组织及肝癌组织中均高表达,是胎肝特异性癌胚RNA,可调控肝癌细胞的增殖及侵袭过程,有望成为肝癌治疗的潜在靶点.

Screen and preliminary function study of fetal liver specific oncofetal RNA AK003710

Objective To screen for a fetal liver specific oncofetal RNA and to study its effect on the proliferation and mobility of hepatocellular carcinoma (HCC) cells. Methods By overlapping the microarray results of mouse fetal liver and regenerated liver following partial hepatectomy (PH), we obtained several candidate lncRNAs which were highly expressed in both fetal liver and regenerated liver tissues, and then the expression of these candidate lncRNAs in HCC tissue were also detected. The most differentially expressed lncRNA in the mouse HCC tissues as detected by real-time PCR were chosen for further research. By real-time PCR,the expression of those lncRNA in fetal liver and regenerated liver tissue was verified. For the function study, EdU labeling system and Transwell experiment were carried out to determine the proliferation ability and mobility after knocking down the lncRNA by siRNA transfection in HCC cell line Hepa1-6. Results Seven candidate lncRNAs which were highly expressed in both fetal liver and regenerated liver tissues were obtained; 3 of them were overexpressed in HCC tissues compared with normal liver tissues, 3 had no differential expression, and 1 was undetectable. Among the 3 overexpressed lncRNAs, lncRNA-AK003710 was the most differentially expressed lncRNA according to microarray and real-time PCR results. And then we detected that lncRNA-AK003710 was overexpressed in the fetal liver and 3 kinds of injured liver tissues. Furthermore, the proliferation ability and mobility of Hepa1-6 were impaired after knocking down lncRNA-AK003710 in HCC cell line Hepa1-6. Conclusion lncRNA-AK003710 is overexpressed in the fetal liver, regenerated liver tissues, and HCC tissues; therefore it is an oncofetal RNA that can regulate the proliferation and invasion of HCC cells and may be a potential therapeutic target for HCC.