SphK2促进人卵巢癌细胞侵袭转移及其机制探讨

目的:研究鞘氨醇激酶2(SphK2)对人卵巢癌细胞系SKOV3和HO8910细胞侵袭和迁移能力的影响,探讨其可能调控机制。方法:设计并体外合成靶向SphK2的shRNA(short hairpin RNA)序列(shRNA SphK2)及阴性对照序列(shRNA NC),并与LV3(H1/GFP&Puro)慢病毒载体重组形成shRNA表达载体,包装shRNA慢病毒颗粒后分别转染SKOV3和HO8910细胞系。分别用Real-time PCR和Western blot法检测shRNA SphK2病毒颗粒转染后在mRNA及蛋白水平的沉默效率。Transwell小室实验检测细胞迁移和侵袭能力变化;Western blot法检测SphK2敲降后细胞中MMP-9蛋白表达。结果:用构建的shRNA慢病毒颗粒感染SKOV3和HO8910细胞后,SphK2 mRNA和蛋白表达水平显著下降。SKOV3和HO8910细胞中,shRNA SphK2转染组的迁移和侵袭细胞数均显著少于shRNA NC组和未转染组,差异均有统计学意义(P<0.05)。Western blot结果显示,转染后48h,shRNA SphK2转染组中MMP-9蛋白表达明显下降(P<0.05)。结论:SphK2可促进卵巢癌细胞的迁移和侵袭,其分子机制可能与调节MMP-9表达有关,提示SphK2在卵巢癌的发展转移中起着重要的作用。

Effect and mechanism of SphK2 on migration and invasion in ovarian cancer cells

Objective:To investigate the effect of sphingosine kinase 2(SphK2)on invasion and migration of human ovarian cancer cells(SKOV3 and HO8910)and to explore its possible regulatory mechanism.Methods:The shRNA(short hairpin RNA)sequence specific targeting SphK2 and the negative control sequence were designed and synthesized,then recombined with LV3(H1/GFP&Puro)lentiviral shRNA expression vector.After transfected with shRNA lentiviral particles,the silencing efficiency at mRNA and protein levels were detected by real-time PCR and Western blot in SKOV3 and HO8910 cells.Transwell experiments were preformed to detect the ability of invasion and migration for ovarian cancer cells,and Western blot was used to analyze the expression of MMP-9 in ovarian cancer cells.Results:In SKOV3 and HO8910 cells after transfected with shRNA lentiviral particles targeting SphK2,the expression level of SphK2 mRNA was significantly decreased,the expression of protein was reduced.The ability of invasion and migration were significantly inhibited in SKOV3 and HO8910 cells transfected with shRNA lentiviral particles targeting SphK2.Expression of MMP-9 protein in cells transfected with shRNA lentiviral particles targeting SphK2 decreased significantly.Conclusion:SphK2 could promote invasion and migration in ovarian cancer cells via regulating MMP-9,suggesting that SphK2 plays an important role in the development of ovarian cancer.