The neonatal splice variant of Nav1.5 potentiates in vitro invasive behaviour of MDA-MB-231 human breast cancer cells.
The neonatal splice variant of Nav1.5 potentiates in vitro invasive behaviour of MDA-MB-231 human breast cancer cells.
复制标题
DOI：
10.1007/s10549-006-9281-1
复制
发表时间：
2007-01
影响因子：
3.8
通讯作者：
Djamgoz, Mustafa B. A.
中科院分区：
文献类型：
--
作者：
Brackenbury, William J.;Chioni, Athina-Myrto;Diss, James K. J.;Djamgoz, Mustafa B. A.
功能性电压门控Na+通道（VGSC）的上调发生在体外和体内转移性人乳腺癌（BCa）中。本研究旨在确定Nav1.5（nNav1.5）的“新生儿”剪接变体（被认为是主要的）在MDA-MB-231细胞的VGSC依赖性侵袭行为中的具体参与。通过靶向nNav1.5的两种不同方法抑制nNav1.5的功能活性：（i）小干扰RNA（siRNA）和（ii）多克隆抗体（NESO-pAb）;确定对迁移和侵袭的影响。nNav1.5 mRNA、蛋白质和信号分别用实时PCR、Western印迹和膜片钳记录进行测量。用siRNA处理迅速降低（约90%）nNav1.5（但不是成人Nav1.5）mRNA的水平，但蛋白质的降低要小得多（约30%），即使在13天后。然而，siRNA使VGSC峰值电流密度降低了33%，并显著增加了细胞对纳摩尔河豚毒素（TTX）的敏感性。重要的是，siRNA抑制了43%的体外迁移，并消除了TTX的正常抑制作用。迁移的MDA-MB-231细胞在质膜上表达的nNav1.5蛋白比未迁移的细胞多。此外，NESO-pAb以剂量依赖性方式减少迁移高达42%。NESO-pAb还降低了Matrigel侵袭而不影响增殖。TTX对已经用NESO-pAb处理的细胞没有影响。得出的结论是，nNav1.5主要负责MDA-MB-231细胞中侵袭行为的VGSC依赖性增强。因此，靶向nNav1.5表达/活性可用于转移性BCa的临床管理。
Upregulation of functional voltage-gated Na+ channels (VGSCs) occurs in metastatic human breast cancer (BCa) in vitro and in vivo. The present study aimed to ascertain the specific involvement of the ‘neonatal’ splice variant of Nav1.5 (nNav1.5), thought to be predominant, in the VGSC-dependent invasive behaviour of MDA-MB-231 cells. Functional activity of nNav1.5 was suppressed by two different methods targeting nNav1.5: (i) small interfering RNA (siRNA), and (ii) a polyclonal antibody (NESO-pAb); effects upon migration and invasion were determined. nNav1.5 mRNA, protein and signalling were measured using real-time PCR, Western blotting, and patch clamp recording, respectively. Treatment with the siRNA rapidly reduced (by ~90 %) the level of nNav1.5 (but not adult Nav1.5) mRNA, but the protein reduction was much smaller (~30 %), even after 13 days. Nevertheless, the siRNA reduced peak VGSC current density by 33 %, and significantly increased the cells’ sensitivity to nanomolar tetrodotoxin (TTX). Importantly, the siRNA suppressed in vitro migration by 43 %, and eliminated the normally inhibitory effect of TTX. Migrated MDA-MB-231 cells expressed more nNav1.5 protein at the plasma membrane than non-migrated cells. Furthermore, NESO-pAb reduced migration by up to 42 %, in a dose-dependent manner. NESO-pAb also reduced Matrigel invasion without affecting proliferation. TTX had no effect on cells already treated with NESO-pAb. It was concluded that nNav1.5 is primarily responsible for the VGSC-dependent enhancement of invasive behaviour in MDA-MB-231 cells. Accordingly, targeting nNav1.5 expression/activity may be useful in clinical management of metastatic BCa.
登录
查看更多内容
影响因子：
11.5
作者：
Alao, JP;Lam, EWF;Vigushin, DM
通讯作者：
Vigushin, DM
影响因子：
4.5
作者：
Bennett, ES;Smith, BA;Harper, JM
通讯作者：
Harper, JM
影响因子：
5.6
作者：
Fraser, SP;Salvador, V;Djamgoz, MBA
通讯作者：
Djamgoz, MBA
影响因子：
4.8
作者：
Diss, JKJ;Stewart, D;Djamgoz, MBA
通讯作者：
Djamgoz, MBA
影响因子：
5.5
作者：
Brackenbury, William J.;Djamgoz, Mustafa B. A.
通讯作者：
Djamgoz, Mustafa B. A.