miR-19b在骨肉瘤组织中的表达及其通过调控KLF13影响骨肉瘤Saos-2细胞侵袭和迁移的研究

Expression of miR-19b in osteosarcoma and its effect on invasion and migration of osteosarcoma Saos-2 cells by regulating KLF13

目的:研究miR-19b在骨肉瘤组织中的表达及其通过调控KLF13影响骨肉瘤细胞侵袭和迁移的作用。方法:Realtime PCR方法检测骨肉瘤组织和对应正常癌旁组织中的miR-19b表达差异。在骨肉瘤Saos-2细胞中转染miR-19b inhibitor,Transwell小室检测侵袭和迁移能力变化,Western blot检测MMP-9和MMP-2蛋白表达差异。靶基因预测软件预测KLF13可能为miR-19b的靶基因,荧光素酶报告系统鉴定靶向关系。Realtime PCR方法检测骨肉瘤组织和对应正常癌旁组织中的KLF13表达差异。将KLF13 siRNA和miR-19b inhibitor共转染到骨肉瘤细胞中,利用上述方法检测细胞侵袭、迁移和MMP-9、MMP-2蛋白表达差异。结果:骨肉瘤组织中miR-19b表达水平高于对应癌旁组织。转染miR-19b inhibitor的骨肉瘤细胞中miR-19b水平降低,细胞侵袭、迁移以及MMP-9、MMP-2蛋白水平下降。miR-19b靶向负调控KLF13表达。KLF13在骨肉瘤组织中的表达水平低于正常癌旁组织,并且其表达水平与miR-19b表达水平呈负相关。KLF13 siRNA可以逆转miR-19b inhibitor对骨肉瘤细胞侵袭、迁移和MMP-9、MMP-2蛋白表达的抑制作用。结论:miR-19b在骨肉瘤组织中高表达,下调其表达可以通过靶向促进KLF13表达抑制骨肉瘤Saos-2细胞侵袭和迁移。

AIM:To study the expression of miR-19b in osteosarcoma and its effect on invasion and migration of osteosarcoma by regulating KLF13.METHODS:Realtime PCR was used to detect the expression of miR-19b in osteosarcoma tissues and corresponding normal adjacent tissues.miR-19b inhibitor was transfected into osteosarcoma Saos-2 cells.Transwell chamber was used to detect changes in invasion and migration ability.Western blot was used to detect the difference of MMP-9 and MMP-2 protein expression.Target gene prediction software predicts that KLF13 may be a target gene of miR-19b.Luciferase reporting system was used to identify the targeting relationship.Realtime PCR was used to detect the difference of KLF13 expression between osteosarcoma tissues and corresponding normal adjacent tissues.KLF13 siRNA and miR-19b inhibitor were co-transfected into osteosarcoma cells,the differences of cell invasion,migration and expression of MMP-9 and MMP-2 were detected by the above methods.RESULTS:The expression of miR-19b in osteosarcoma tissues was higher than that in adjacent tissues.In osteosarcoma cells transfected with miR-19b inhibitor,the level of miR-19b decreased,cell invasion,migration and the levels of MMP-9 and MMP-2 protein decreased.miR-19b targeting negatively regulates the expression of KLF13.The expression of KLF13 in osteosarcoma was lower than that in normal adjacent tissues,the expression level of miR-19b was negatively correlated with that of miR-19b.KLF13 siRNA can reverse the inhibitory effect of miR-19b inhibitor on invasion,migration and expression of MMP-9 and MMP-2 in osteosarcoma cells.CONCLUSION:miR-19b is highly expressed in osteosarcoma,downregulation of KLF13 expression can inhibit the invasion and migration of osteosarcoma Saos-2 cells by targeting KLF13 expression.