CAJ | 학술논문

目的：重组过表达EZH2基因3'非翻译区的MCF-7细胞株中筛选靶向miRNAs，并进行细胞和组织定量分析。方法重组细胞株感染慢病毒文库，利用细胞毒性药物筛选后抽提细胞基因组，以此为模板PCR扩增出miRNA前体，测序确定miRNAs名称，并对其进行PCR定量分析。结果在重组细胞株中共筛选出7种miRNAs，依次为miR-15b、miR-16-2、miR-181b2、miR-217、miR-224、miR-329-1、miR-487b，这些新型miRNAs用生物信息学软件PicTar和TargetScan无法预测。Real-time PCR发现，与乳腺癌细胞MCF-7相比，正常乳腺细胞株HBL-100中miR-217、miR-329-1、miR-487b高表达；乳腺癌组织中仅miR-15b及miR-16-2表达增加，与癌旁组织相比差异有统计学意义（P<0.05）。结论在重组过表达EZH23'-UTR的乳腺癌MCF-7细胞株中结合慢病毒文库可筛选出用生物信息学软件未预测的新型靶向miRNAs，这些新型miRNA在正常乳腺细胞与肿瘤细胞及组织中存在差异表达。
목적：중조과표체EZH2기인3'비번역구적MCF-7세포주중사선파향miRNAs，병진행세포화조직정량분석。방법중조세포주감염만병독문고，이용세포독성약물사선후추제세포기인조，이차위모판PCR확증출miRNA전체，측서학정miRNAs명칭，병대기진행PCR정량분석。결과재중조세포주중공사선출7충miRNAs，의차위miR-15b、miR-16-2、miR-181b2、miR-217、miR-224、miR-329-1、miR-487b，저사신형miRNAs용생물신식학연건PicTar화TargetScan무법예측。Real-time PCR발현，여유선암세포MCF-7상비，정상유선세포주HBL-100중miR-217、miR-329-1、miR-487b고표체；유선암조직중부miR-15b급miR-16-2표체증가，여암방조직상비차이유통계학의의（P<0.05）。결론재중조과표체EZH23'-UTR적유선암MCF-7세포주중결합만병독문고가사선출용생물신식학연건미예측적신형파향miRNAs，저사신형miRNA재정상유선세포여종류세포급조직중존재차이표체。
Objective To screen novel miRNAs targeting EZH2 3' untranslated region (UTR) in recombinational MCF-7 breast cancer cells over-expressing EZH2 3' UTR and quantitative analyze the expressions of the screened miRNA in breast cancer cells and tissues. Methods A lentiviral library was transfected into the recombinant cell line MCF-7. The cells were screened with cytotoxic agents before extraction of the genome for amplification of the miRNA precursors using PCR. The screened miRNAs were identified with sequence analysis and their expressions were analyzed quantitatively with real-time PCR in breast cancer cells and tissues. Results Seven miRNAs were screened from the recombinant MCF-7 cells, namely miR-15b, miR-16-2, miR-181b2, miR-217, miR-224, miR-329-1, and miR-487b, all of which failed to be predicted by bioinformatics software. Real-time PCR showed that miR-217, miR-329-1, and miR-487b were over-expressed in MCF-7 cells, and the expression of miR-15b and miR-16-2 was significantly increased in cancer tissues compared with the adjacent tissues (P<0.05). Conclusions Novel targeted miRNAs that can not be predicted by bioinformatics software were successfully screened from MCF-7 breast cancer cells over-expressing EZH2 3' UTR. These miRNAs are expressed differentially between normal breast cells and breast cancer tissues.