miR-378通过靶向RAB11A促进乳腺癌细胞系增殖和迁移

miR-378 promotes breast cancer cell proliferation and migration by targeting Ras-related protein RAB-11A expression

目的探讨miR-378在乳腺癌细胞系表达及作用机制。方法应用TargetScan分析miR-378的作用靶基因RAB11A。应用RT-PCR检测乳腺癌细胞系miR-378和RAB11A mRNA表达水平。转染MCF-7和MDA-MB-231细胞miR-378,NC(normal control)对照,采用MTT法和Transwell法检测乳腺癌细胞增殖和迁移能力的变化;采用RT-PCR和Western blot检测RAB11A表达。荧光素酶报告分析miR-378与靶基因RAB11A之间的信号。结果miR-378在乳腺癌细胞MCF-7、MDA-MB-231表达上调[(4.23±0.15)和(4.67±0.12)比(0.98±0.03),P<0.01];RAB11A在乳腺癌细胞MCF-7、MDA-MB-231中表达下调[(0.49±0.09)和(0.45±0.08)比(1.01±0.02),P<0.01];在MCF-7、MDA-MB-231细胞中敲降miR-378后,乳腺癌细胞增殖活力降低[MCF-7:(1.28±0.01)比(1.75±0.10);MDA-MB-231:(1.23±0.07)比(1.89±0.07),P均<0.01];细胞侵袭数目减少[MCF-7:(19.64±3.56)比(65.01±5.12);MDA-MB-231:(32.65±4.36)比(83.09±7.45),P均<0.01];迁移数目减少[MCF-7:(23.78±2.43)比(73.24±6.16);MDA-MB-231:(37.42±3.24)比(91.68±6.42),P均<0.01]。RAB11A蛋白表达水平明显增加[MCF-7:(0.88±0.12)比(1.32±0.11),P<0.01;MDA-MB-231:(1.56±0.09)比(0.80±0.14),P均<0.01],荧光素酶活性增加[MCF-7:(1.39±0.12)比(0.78±0.14);MDA-MB-231:(1.47±0.09)比(0.80±0.11),P均<0.01]。结论miR-378可能通过靶向RAB11A促进乳腺癌细胞增殖和迁移。

Objective To assess expression and functions of miR-378 in breast cancer cells.Methods Level of miR-378 was assessed using qRT-PCR,while the targeting gene of miR-378 was searched using the TargetScan and verified using qRT-PCR.Breast cancer MCF-7 and MDA-MB-231 cells were gene transfected and changes in cell viability and migration were detected using cell viability CCK-8 and Transwell assays.The luciferase reporter assay was used to confirm miR-378 binding to the target gene RAB11A.Results Level of miR-378 was higher in MCF-7 and MDA-MB-231 cells than in normal MCF-10A cells(0.98±0.03 vs.4.23±0.15 and 4.67±0.12;P<0.001),whereas level of Ras-related protein RAB11A,a targeting gene of miR-378,was lower in MCF-7 and MDA-MB-231 cells than in normal cells(1.01±0.02 vs.0.49±0.09 and 0.45±0.08;P<0.001).Moreover,after knockdown of miR-378 expression,tumor cell proliferation and migration capacity was significantly reduced in MCF-7(1.75±0.10 vs.1.28±0.01,65.01±5.12 vs.19.64±3.56,and 73.24±6.16 vs.23.78±2.43,respectively;P<0.001)and MDA-MB-231 cells(1.89±0.07 vs.1.23±0.07,83.09±7.45 vs.32.65±4.36,and 91.68±6.42 vs.37.42±3.24,respectively;P<0.001).Levels of and RAB11A mRNA and protein were all significantly increased in MCF-7(0.88±0.12 vs.1.32±0.11;P<0.01)and MDA-MB-231 cells(0.80±0.14 vs.1.56±0.09;P<0.01),while the luciferase activity was increased in MCF-7(0.78±0.14 vs.1.39±0.12;P<0.01)and MDA-MB-231 cells(0.80±0.11 vs.1.47±0.09;P<0.01)vs.the control cells.Conclusion miR-378 was able to promote breast cancer cell proliferation and migration by targeting of RAB11A expression.