Expression and Distribution of Voltage-gated Sodium Channels in Cervical Cancer Cells and Its Role in Cell Proliferation and Invasion
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摘要:
目的 探讨电压门控钠离子通道(voltage-gated sodium channels, VGSCs)在宫颈癌细胞中的表达、分布及其对该细胞增殖、侵袭能力的影响。 方法 用Western blot了解VGSCs在不同宫颈癌细胞株中表达水平, 采用免疫荧光检测VGSCs蛋白在细胞中的定位。分别用MTT法和Matrigel法检测VGSCs对宫颈癌细胞的增生和侵袭作用。以半定量RT-PCR和RNAi方法确认宫颈癌细胞中VGSC表达亚型及其功能。 结果 在检测的4种宫颈癌细胞株中, ME180细胞表达较高水平VGSCs蛋白, 其存在于细胞膜和细胞质中。VGSCs抑制剂-河豚毒素(TTX)对ME180细胞增生无影响(P > 0.05), 但呈浓度依赖性抑制细胞的Matrigel侵袭(P < 0.05)。在ME180细胞中检测到Nav 1.2、Nav 1.6和Nav 1.7三种VGSCs亚型, 其中Nav 1.6 mRNA占总mRNA的78%, RNAi抑制Nav 1.6mRNA表达后可降低细胞侵袭达52%(P < 0.05)。 结论 VCSCs在宫颈癌ME180细胞株高表达, Nav 1.6为其主要表达亚型, 增加了体外癌细胞的侵袭转移。 Abstract:Objective The present study investigates the expression and distribution of voltage-gated sodium channels(VGSCs), as well as its effects on the proliferation and invasion of human cervical cancer cells. Methods Western blot analysis and immunofluorescence were used to detect the level and intracellular distribution of the VGSC protein in cervical cancer cell lines, respectively. Methyl thiazolyl tetrazolium assay and Matrigel methods tested the effect of VGSCs on the proliferation and invasion of the cervical cancer ME180 cells, respectively.The expression of the VGSC subtype in ME 180 cells was determined using semi-quantitative polymerase chain reaction and ribonucleic acid interference(RNAi). Results Cervical cancer ME180 cells expressed the highest level of VGSC protein in the four cervical cancer cell lines tested.VGSC protein was primarily found in the plasma membrane and cytoplasm of the cells.Tetrodotoxin, an inhibitor of VGSCs, had no effect on the proliferation of the ME180 cells, but decreased Matrigel invasion in a dose-dependent manner(P < 0.05).Three VGSC isoforms(Nav1.2, Nav1.6, and Nav1.7) were identified to be expressed in the ME180 cells, among which Navl.6 mRNA was at the highest level(P < 0.05).RNAi targeting Navl.6 mRNA decreased the invasion of the cells by 52%(P < 0.05). Conclusion VGSC subtype Navl.6 was found to be expressed predominantly in human cervical cancer ME180 cells.The expression of Nav1.6 in the cells increases the invasion of the cells in vitro. -
Key words:
- Cervical cancer /
- Voltage-gated sodium channels /
- Invasion /
- Tetrodotoxin /
- Nav1.6
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图 1 VGSCα蛋白在宫颈上皮细胞和不同宫颈癌细胞株中的表达水平
Figure 1. Expression level of the VGSCαprotein in normal cervical cells and in four cervical cancer cell lines
图 2 VGSCα在宫颈癌ME180和Caski细胞中的分布
Figure 2. Cellular distribution of VCSCαin cervical cancer cell lines of ME 180 and Caski cells (scale bar=10μm).ConA (FITC/green) marked the membrane of cells.Alexa567 (red) marked the VGSCαin the cells.Scale bar is applicable to all panels
图 3 Matrigel检测VGSC抑制剂TTX对ME180细胞侵袭作用的抑制
Figure 3. Effect of ITX on the migration of ME 180 cells tested using the Matrigel method
图 4 在ME180细胞中表达的VGSC亚型
Figure 4. Expression of the VGSC isoform in cervical cancer ME180 and Caski cells
表 1 TTX对ME180细胞增殖能力的影响
Table 1. Effects of TTX on the proliferation of ME180 cells
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[1] 潘惠艳, 黄秉仁. 电压门控钠离子通道与恶悱肿瘤的转移[J]. 生理科学进展, 2011, 42(3): 217-220. https://www.cnki.com.cn/Article/CJFDTOTAL-SLKZ201103017.htm [2] Patino GA, Isom LL. Electrophysiology and beyond: multiple roles of Na+ channel 3 subunit in development and disease[J]. Neurosci Lett, 2010, 486(2): 53-59. doi: 10.1016/j.neulet.2010.06.050 [3] House CD, Vaske CJ, Schwartz AM, et al. Voltage-gated Na + channel SCN5A is a key regulator of a gene. transcriptional network that controls colon cancer invasion[J]. Cancer Res, 2010, 70(17): 6957-6967. doi: 10.1158/0008-5472.CAN-10-1169 [4] Diaz D, Delgadillo DM, Hernandez-Gallegos E, et al. Functional expression of voltage-gated sodium channels in primary cultures of human cervical cancer[J]. J Cell Physiol, 2007, 210(2): 469-478. doi: 10.1002/jcp.20871 [5] Fogh J, Fogh TM, Orfeo J. One hundred and twenty-seven cultured human tumor cell line producing tumors in nude mice[J]. J Nad Cancer Inst, 1977, 59(1): 221-226. doi: 10.1093/jnci/59.1.221 [6] Fraser SP, Diss JK, Chioni AM, et al. Voltage-gated sodium channel expression and potentiation of human breast cancer metastasis[J]. Clin Cancer Res, 2005, 11(15): 5381-5389. doi: 10.1158/1078-0432.CCR-05-0327 [7] Bennett E, Smith B, Harper J. Voltage-gated Na+ channels confer invasive properties on human prostate cancer cells[J]. Pflügers Arch, 2004, 447(6): 908-914. doi: 10.1007%2Fs00424-003-1205-x.pdf [8] Chioni AM, Sliao D, Grose R, et al. Protein kinase A and regulation of neonatal Nav1.5 expression in human breast cancer cells: activity -dependent positive feedback and cellular migration[J]. Int J Biochem Cell Biol, 2010, 42(2): 346-358. doi: 10.1016/j.biocel.2009.11.021 [9] Brackenbury WJ, Djamgoz, MB. Nerve growth factor enhances voltage-gated sodium channel activity expression in Mat-LyLu rat prostate cancer cell line[J]. J Cell Physiol, 2007, 210(3): 602-608. doi: 10.1002/jcp.20846 [10] Andrikopoulos P, Fraser SP, Patterson L, et al. Angiogenic functions of voltage-gated Na+ channels in human endothelial cells[J]. J Biol Chem, 2011, 286(9): 16846-16860. http://europepmc.org/abstract/med/21385874 [11] Onkal R, Djamgoz MB. Molecular pharmacology of voltage-gated sodium expression in metastatic disease: Clinical potential of neonatal Navl. 5 in breast cancer[J]. EuroJ Pharmo, 2009, 625(1): 206-219. http://www.sciencedirect.com/science/article/pii/S0014299909008802 [12] Roger S, Rollin J, Barascu A, et al. Voltage-gated sodium cliannels potentiate the invasive capacities of human non-small-cell lung cancer cell lines[J]. Int J Biochem Cell Biol, 2007, 39(4): 774-786. doi: 10.1016/j.biocel.2006.12.007 -
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