Department of Pharmacology, Weifang Medical University, Weifang, Shandong 261053, P.R. China.
Department of Physics, Weifang Medical University, Weifang, Shandong 261053, P.R. China.
Mol Med Rep. 2019 Apr;19(4):2927-2934. doi: 10.3892/mmr.2019.9901. Epub 2019 Jan 28.
The aim of the current study was to investigate the effects and the molecular mechanisms of ARK5 in ovarian cancer cell invasion. The plasmid pGCsilencerU6/GFP/Neo‑RNAi‑ARK5 and the control vector with a scramble sequence were transfected into SKOV3 cells to establish ARK5‑deficient SKOV3 cells (siARK5/SKOV3) and a control cell line (Scr/SKOV3), respectively. Reverse transcription‑polymerase chain reaction (RT‑PCR) and Western blot analysis were used to determine the mRNA and protein expression levels of ARK5. Migration and invasion abilities of SKOV3 cells were determined in chemotaxis and invasion assays, respectively. The epidermal growth factor‑1 (EGF‑1)‑induced expression of matrix metallopeptidase (MMP)‑2 and MMP‑9, epithelial‑mesenchymal transition (EMT) and phosphorylation of mechanistic target of rapamycin kinase (mTOR) in siARK5/SKOV3 and Scr/SKOV3 cells were detected by western blot. RT‑PCR and western blot analyses demonstrated that the expression of ARK5 was significantly downregulated in siARK5/SKOV3 cells at the mRNA and protein levels (P<0.01). The migration and invasion abilities of siARK5/SKOV3 cells were markedly decreased compared with Scr/SKOV3 cells (P<0.01). In addition, the results demonstrated that EGF‑1‑induced expression of MMP‑2 and MMP‑9, EMT and phosphorylation of mTOR were suppressed in siARK5/SKOV3 cells as compared with Scr/SKOV3 cells (P<0.01). The current study demonstrated that ARK5 is a critical factor involved in SKOV3 cell invasion and ARK5 increases invasive potential by promoting EMT and activating the Akt‑mTOR‑MMPs pathway.
本研究旨在探讨 ARK5 在卵巢癌细胞侵袭中的作用及其分子机制。将 pGCsilencerU6/GFP/Neo-RNAi-ARK5 质粒和带有乱序序列的对照载体转染 SKOV3 细胞,分别建立 ARK5 缺陷型 SKOV3 细胞(siARK5/SKOV3)和对照细胞系(Scr/SKOV3)。采用逆转录-聚合酶链反应(RT-PCR)和 Western blot 分析检测 ARK5 的 mRNA 和蛋白表达水平。采用趋化和侵袭实验分别检测 SKOV3 细胞的迁移和侵袭能力。Western blot 检测 siARK5/SKOV3 和 Scr/SKOV3 细胞中表皮生长因子-1(EGF-1)诱导的基质金属蛋白酶(MMP)-2 和 MMP-9 表达、上皮-间充质转化(EMT)以及雷帕霉素靶蛋白激酶(mTOR)磷酸化。RT-PCR 和 Western blot 分析表明,siARK5/SKOV3 细胞中 ARK5 的表达在 mRNA 和蛋白水平均显著下调(P<0.01)。与 Scr/SKOV3 细胞相比,siARK5/SKOV3 细胞的迁移和侵袭能力明显降低(P<0.01)。此外,结果表明,与 Scr/SKOV3 细胞相比,siARK5/SKOV3 细胞中 EGF-1 诱导的 MMP-2 和 MMP-9 表达、EMT 以及 mTOR 磷酸化受到抑制(P<0.01)。本研究表明,ARK5 是 SKOV3 细胞侵袭的关键因素,通过促进 EMT 和激活 Akt-mTOR-MMPs 通路增加侵袭潜能。