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目的:探讨血管内皮生长因子C(vascular endothelial growth factor-C,VEGF-C)促进宫颈癌ME180细胞的侵袭和转移是否涉及电压门控钠通道(voltage-gated sodium channels,VGSCs)Nav1.6亚型的表达及其他可能的信号转导分子。方法:应用RT-PCR和免疫荧光定位的方法分别检测VEGF-C受体3(亦称Flt-4)和Nav1.6mRNA及其蛋白在ME180细胞中的表达和定位,Transwell法检测VEGF-C和Nav1.6对细胞侵袭的作用,实时荧光定量-PCR和蛋白质印迹法检测VEGF-C对Nav1.6mRNA和蛋白表达的影响及信号转导分子p38丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)在其中的作用。结果:ME180细胞表达Flt-4和Nav1.6mRNA,Flt-4和Nav1.6蛋白主要定位于细胞膜。50和100ng/L VEGF-C可分别增加细胞的侵袭指数[(27.2±5.2)%和(49.1±6.1)%],与对照组比较差异有统计学意义(P<0.05);VGSC抑制剂TTX(tetrodotoxin)可有效阻断VEGF-C促进的细胞侵袭作用(P<0.05)。VEGF-C可上调ME180细胞Nav1.6mRNA和蛋白的表达水平(P<0.05)。p38MAPK抑制剂PD169316可抑制VEGF-C促进的Nav1.6mRNA和蛋白的表达水平及细胞侵袭作用。结论:VEGF-C通过上调VGSC Nav1.6亚型的表达促进宫颈癌ME180细胞侵袭,而且p38MAPK参与了VEGF-C诱导的Nav1.6表达和细胞侵袭。
Objective: To investigate whether invasion and metastasis of cervical cancer cell line ME180 by vascular endothelial growth factor-C (VEGF-C) involve the voltage-gated sodium channels (VGSCs) Nav1.6 subtype Expression and other possible signal transduction molecules. Methods: The expression and localization of VEGF-C receptor 3 (also known as Flt-4) and Nav1.6 mRNA and their proteins in ME180 cells were detected by RT-PCR and immunofluorescence staining. The expressions of VEGF-C and Nav1.6 on cell invasion, the effects of VEGF-C on the expression of Nav1.6 mRNA and protein and the expression of the signal transduction molecule p38 mitogen-activated protein kinase (MAPK) were detected by real-time fluorescence quantitative PCR and Western blotting. MAPK) in which the role. Results: Flt-4 and Nav1.6 mRNA were expressed in ME180 cells. Flt-4 and Nav1.6 proteins mainly localized in the cell membrane. 50 and 100ng / L VEGF-C could increase the invasion index of cells (27.2 ± 5.2% vs 49.1 ± 6.1%, respectively) compared with the control group (P <0.05). The VGSC inhibitor TTX (tetrodotoxin) can effectively block the VEGF-C-promoted cell invasion (P <0.05). VEGF-C upregulated the expression of Nav1.6 mRNA and protein in ME180 cells (P <0.05). The p38 MAPK inhibitor PD169316 can inhibit the expression of VEGF-C-promoted Nav1.6 mRNA and protein and cell invasion. Conclusion: VEGF-C promotes the invasion of cervical cancer cell line ME180 by up-regulating the expression of VGSC Nav1.6 subtype, and p38MAPK is involved in the expression of VEGF-C-induced Nav1.6 and cell invasion.