miR-149-3p靶向FOXM1抑制人胃癌细胞SGC-7901生长和迁移

miR-149-3p targeting FOXM1 in inhibition of growth and migration of human gastric cancer cell SGC-7901

目的:探究miR-149-3p靶向FOXM1抑制人胃癌细胞SGC-7901生长和迁移的机制。方法:体外培养人胃癌细胞SGC-7901,并分为:Ctrl组、miR-NC组、miR-149-3p mimic组、pLV-FOXM1组和pLV-FOXM1+miR-149-3p mimic组;使用RT-PCR分别检测正常组织、原位肿瘤组织和转移后肿瘤组织中miR-149-3p和FOXM1 mRNA的表达水平并分析miR-149-3p与FOXM1的线性关系;使用miR-NC、miR-149-3p mimic转染细胞后检测miR-149-3p和FOXM1 mRNA表达水平;双荧光素酶检测miR-149-3p与FOXM1的靶向关系;CCK-8检测细胞增殖情况,体外侵袭实验检测细胞侵袭情况,划痕实验检测细胞迁移情况;蛋白质印迹检测N-cadherin、E-cadherin、MMP-2、MMP-9、FOXM1、PCNA、p21的表达水平。结果:相比原位肿瘤组织和转移后肿瘤组织,正常组织中FOXM1 mRNA表达较低、miR-149-3p表达较高。荧光素酶报告实验表明miR-149-3p序列上有FOXM1的结合位点。相比空白对照组(Ctrl),miR-149-3p mimic组细胞增长受到显著抑制,FOXM1、P21、E-cadherin蛋白表达水平显著升高(P<0.05);单位面积细胞侵袭数目、细胞迁移能力、PCNA、N-cadherin、MMP-9、MMP-2蛋白表达水平显著降低(P<0.05)。相比Ctrl组,pLV-FOXM1组细胞生长受到促进,FOXM1、P21、E-cadherin蛋白表达水平显著降低(P<0.05);单位面积细胞侵袭数目、细胞迁移能力、PCNA、N-cadherin、MMP-9、MMP-2蛋白表达水平显著升高(P<0.05)。相比pLV-FOXM1组,miR-149-3p+pLV-FOXM1组细胞增长受到显著抑制,FOXM1、P21、E-cadherin蛋白表达水平显著升高(P<0.05);单位面积细胞侵袭数目、细胞迁移能力、PCNA、N-cadherin、MMP-9、MMP-2蛋白表达水平显著降低(P<0.05)。结论:miR-149-3p可以靶向FOXM1抑制人胃癌细胞SGC-7901的生长侵袭和迁移,其机制与调控细胞生长侵袭和迁移的蛋白有关。

Objective:To investigate the mechanism of miR-149-3 p targeting FOXM1 in the inhibition of growth and migration of human gastric cancer cell SGC-7901.Methods: Human gastric cancer cells SGC-7901 were cultured in vitro, and they were divided into Ctrl group, miR-NC group,miR-149-3 p mimic group,pLV-FOXM1 group and pLV-FOXM1+miR-149-3 p mimic group.RT-PCR was performed to detect expression levels of miR-149-3 p and FOXM1 mRNA in normal tissues,tumor tissues in situ and metastatic tumor tissues.The linear relationship between miR-149-3 p and FOXM1 was analyzed.After cells were transfected with miR-NC and miR-149-3 p mimic,expression levels of miR-149-3 p and FOXM1 mRNA were detected.The dual luciferase was performed to detect targeting relationship between miR-149-3 p and FOXM1.CCK-8 was performed to detect cell proliferation.In vitro invasion experiments were performed to detect cell invasion.The cell migration was detected by scratch assay.The expression levels of N-cadherin,E-cadherin,MMP-2,MMP-9,FOXM1,PCNA and p21 were detected by Western blot.Results: Compared with in situ tumor tissues and metastatic tumor tissues,expression of FOXM1 mRNA was significantly lower in normal tissues,while expression of miR-149-3 p was significantly higher.Luciferase reporter experiments indicated that there were binding sites of FOXM1 on miR-149-3 p sequence.Compared with blank control group(Ctrl),cell growth was significantly inhibited in miR-149-3 p mimic group,and expression levels of FOXM1,P21 and E-cadherin protein were significantly increased(P<0.05).The number of cell invasion per unit area,cell migration ability,expression levels of PCNA,N-cadherin,MMP-9 and MMP-2 protein were significantly decreased(P<0.05).Compared with Ctrl group,cell growth was promoted in pLV-FOXM1 group,and expression levels of FOXM1,P21 and E-cadherin proteins were significantly decreased(P<0.05).The number of cell invasion per unit area,cell migration ability,expression level of PCNA,N-cadherin,MMP-9 and MMP-2 proteins were significantly increased(P<0.05).Compared with pLV-FOXM1 group,cell growth was significantly inhibited in miR-149-3 p+pLV-FOXM1 group,expression levels of FOXM1,P21 and E-cadherin proteins were significantly increased(P<0.05).The number of cell invasion per unit area,cell migration ability,expression levels of PCNA,N-cadherin,MMP-9 and MMP-2 protein were significantly decreased(P<0.05).Conclusion: miR-149-3 p can target FOXM1 to inhibit invasion and migration of human gastric cancer cell SGC-7901.The mechanism is related to regulation of cell growth,invasion and migration related proteins.