CAJ | 학술논문

目的研究5-氟尿嘧啶对肝癌细胞HepG2增殖的影响并探讨其可能的作用机制.方法利用实时PCR方法分析microRNA-373(miR-373)在肝细胞癌组织及肝癌细胞中的表达水平,并检测经5-氟尿嘧啶处理的HepG2细胞miR-373的表达.利用蛋白印迹方法检测HepG2细胞转染miR-373模拟物或经5-氟尿嘧啶处理后,靶基因PPP6C的表达变化.设计拯救实验(Rescue assay)并利用CCK-8方法检测细胞增殖情况.结果 miR-373在肝细胞癌组织及细胞系中表达上调.与正常肝细胞对照比较,HepG2细胞miR-373上调2.94倍(P＜0.01).5-氟尿嘧啶可以有效抑制HepG2细胞内源性miR-373的表达水平(约50％,P＜0.01,48 h),并上调PPP6C的表达水平(约2.1倍,48 h).过表达miR-373可以阻遏5-氟尿嘧啶对HepG2细胞增殖的影响,CCK-8细胞增殖分析显示,细胞增殖分别恢复81％ (P ＜0.05,24 h)、84％ (P ＜0.01,48 h).结论 5-氟尿嘧啶可通过下调HepG2细胞内源性miR-373的表达,促进靶基因PPP6C蛋白表达,抑制细胞增殖.
목적 연구5-불뇨밀정대간암세포HepG2증식적영향병탐토기가능적작용궤제.방법 이용실시PCR방법분석microRNA-373(miR-373)재간세포암조직급간암세포중적표체수평,병검측경5-불뇨밀정처리적HepG2세포miR-373적표체.이용단백인적방법검측HepG2세포전염miR-373모의물혹경5-불뇨밀정처리후,파기인PPP6C적표체변화.설계증구실험(Rescue assay)병이용CCK-8방법검측세포증식정황.결과 miR-373재간세포암조직급세포계중표체상조.여정상간세포대조비교,HepG2세포miR-373상조2.94배(P＜0.01).5-불뇨밀정가이유효억제HepG2세포내원성miR-373적표체수평(약50％,P＜0.01,48 h),병상조PPP6C적표체수평(약2.1배,48 h).과표체miR-373가이조알5-불뇨밀정대HepG2세포증식적영향,CCK-8세포증식분석현시,세포증식분별회복81％ (P ＜0.05,24 h)、84％ (P ＜0.01,48 h).결론 5-불뇨밀정가통과하조HepG2세포내원성miR-373적표체,촉진파기인PPP6C단백표체,억제세포증식.
Objective To investigate the inhibitory effect of 5-fluorouracil (5-FU) on hepatocellular carcinoma (HCC) cell growth and to elucidate its potential molecular mechanism.Methods Real-time PCR analysis was conducted to determine the expression of miR-373 in HCC tissue specimens and HCC cell lines.The expression of miR-373 was also evaluated in HepG2 cells after 5-FU treatment.Western blot analysis was performed to detect the protein levels of PPP6C,a verified target of miR-373,with transfection of miR-373 mimics or 5-FU treatment.A rescue assay was conducted to investigate the cell growth in HepG2 cells by using CCK-8.Results miR-373 expression was up-regulated in both HCC tissues and cell lines.miR-373 expression depicted about 2.94-fold augment in HepG2 cells as compared to normal liver cells control (P ＜0.01).5-FU treatment led to a significant decrease of miR-373 levels (approximately 50％,P ＜0.01,48 h) and resulted in a marked increase of PPP6C protein (approximately 2.1-fold,48 h) in HepG2 cells.The overexpression of miR-373 could prevent the impact of 5-FU treatment on cell growth in HepG2 cells and CCK-8 assay showed that HepG2 cell growth was rescued approximately 81％ and 84％ at 24 h (P ＜ 0.05) and 48 h (P ＜ 0.01),respectively.Conclusion 5-FU can repress endogenous miR-373 level,which activates the expression of downstream targeted gene PPP6C,thereby exerting an inhibitory effect on HepG2 cells.