摘要
目的研究miR-144-3p靶向调控ATP结合转运蛋白G家族成员2(ATP-binding transporter G family member 2,ABCG2)对胃癌(gastriccancer,GC)HGC-27细胞侵袭和迁移能力的影响,并对其作用机制进行初步探讨.方法通过qRT-PCR检测人GCHGC-27细胞株和人胃黏膜上皮GES-1细胞株中miR-144-3p和ABCG2的表达情况;通过靶基因预测软件预测miR-144-3p和ABCG2靶向结合位点,通过双荧光素酶报告基因实验检测二者靶向结合关系;通过qRT-PCR检测miR-144-3p mimic或miR-144-3p inhibitor转染GC细胞后miR-144-3p及ABCG2的表达水平;通过明胶酶谱实验检测对转染ABCG2 siRNA的GC细胞中基质金属蛋白酶-2(matrix metalloproteinase-2, MMP-2)和基质金属蛋白酶-9(matrix metalloproteinase-9, MMP-9)活性的影响;通过Transwell侵袭和迁移实验检测转染miR-144-3p mimic、miR-144-3p inhibitor、ABCG2 siRNA对GC细胞侵袭和迁移能力的影响.结果与正常细胞组人胃黏膜上皮GES-1细胞株相比, GC细胞组人GCHGC-27细胞株中miR-144-3p的表达量明显降低, ABCG2的表达水平明显升高,差异具有统计学意义(P<0.05);靶基因预测miR-144-3p和ABCG2 3’UTR存在结合位点,双荧光素酶报告基因实验证实了miR-144-3p和ABCG2靶向结合关系;与对照组相比,转染miR-144-3p mimic组GC细胞中miR-144-3p的表达明显升高, ABCG2的表达明显降低,细胞侵袭和迁移能力显著降低,差异具有统计学意义(P<0.05),转染miR-144-3p inhibitor则表现出相反的作用;明胶酶谱实验检测结果表明ABCG2 siRNA转染可显著抑制GC细胞中MMP-2和MMP-9蛋白活性,抑制GC细胞侵袭和迁移能力,差异具有统计学意义(P<0.05).结论 MiR-144-3p能够抑制GC细胞侵袭和迁移能力,其作用机制与靶向调控ABCG2-MMP-2/9信号通路有关.
AIM To investigate the effect of targeted regulation of ATPbinding transporter G family member 2（ABCG2） signaling pathway by miR-144-3p on the invasion and migration of gastric cancer（GC） cells HGC-27, and to explore the underlying mechanism. METHODS The expression of miR-144-3p and ABCG2 in human GC cell line HGC-27 and human gastric mucosal epithelial cell line GES-1 was detected by qRT-PCR. The target gene prediction software was used to predict whether miR-144-3p binds to ABCG2, and the binding site was used to detect whether miR-144-3p targets ABCG2 by dual luciferase reporter gene assay. The expression of miR-144-3p and ABCG2 was detected by qRT-PCR after transfection of miR-144-3p mimic or miR-144-3p inhibitor into GC cells. The expression of matrix metalloproteinase-2（MMP-2） and matrix metalloproteinase-9（MMP-9） in GC cells transfected with ABCG2 siRNA was detected by gelatin zymography assay. Transwell invasion and migration assays were employed to detect the effect of miR-144-3p mimic, miR-144-3p inhibitor, and ABCG2 siRNA on the invasion and migration of GC cells.RESULTS Compared with GES-1 cells, the expression of miR-144-3p in HGC-27 cells was significantly decreased and the expression of ABCG2 was significantly increased（P〈0.05）. The target gene prediction software predicted the binding site of miR-144-3p in the ABCG2 3＇UTR, and the dual luciferase reporter gene experiment confirmed the targeted binding relationship of miR-144-3p and ABCG2. Compared with the control group, the expression of miR-144-3p was significantly increased, the expression of ABCG2 was significantly decreased, and the cell invasion and migration ability were significantly decreased in the miR-144-3p mimic transfected group（P〈0.05）, while transfection with miR-144-3p inhibitor showed the opposite effect. Gelatin zymography assay showed that ABCG2 siRNA transfection significantly inhibited the activity of MMP-2 and MMP-9 proteins in GC cells and suppressed the invasion and migration of GC cells（P�
作者
吕弢
俞兴旺
胡静
周东辉
Tao Lv;Xing-Wang Yu;Jing Hu;Dong-Hui Zhou(Outpatient Department,the First People＇s Hospital of Yongkang City,Yongkang 321300,Zhejiang Province,China;Department of Oncology,First Affiliated Hospital of Zhejiang University,Hangzhou 310003,Zhejiang Province,China)
出处
《世界华人消化杂志》
CAS
2018年第28期1635-1644,共10页
World Chinese Journal of Digestology
基金
国家自然科学基金面上项目
No.81272680~~
