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. 2014 Mar 19;369(1638):20130105. doi: 10.1098/rstb.2013.0105

Figure 4.

Figure 4.

Upregulation of functional expression of Nav1.7 in human non-small-cell lung cancer H460 cells and consequent increase in invasiveness via ERK1/2 signalling. (a) Current–voltage (I–V) plots for control/untreated cells (open squares) and cells treated in the presence of serum for 24 h with 100 ng ml−1 EGF (closed squares), 1 μM gefitinib/Gef (open circles) or 10 μg ml−1 EGF receptor blocking antibody (filled circles). Currents were evoked using 30 ms depolarizing steps in 5 mV intervals (−90 to +70 mV) from a holding potential of −90 mV. (b) I–V plots for control/untreated cells (open squares) and cells treated with 10 μM U0126 (closed squares). (c) Relative Nav1.7 mRNA expression showing effect of serum starvation for 48 h and treatment for 24 h with EGF (100 ng ml−1), Gef (1 μM) and co-application of EGF + Gef. (d) Matrigel invasion measured after 48 h in control medium (CTL), 0.5 μM TTX, 100 ng ml−1 EGF, 1 μM Gef and EGF + TTX. (e) Effects of treatment with 10 μM U0126 or 100 nM wortmannin (WORT) for 24 h on relative Nav1.7 mRNA expression, compared with control/untreated (CTL) cells. (f) Matrigel invasion measured over 48 h in control/untreated cells (CTL), and following treatment with 10 μM U0126, 10 μM U0126/1 μM TTX, 100 nM WORT, and WORT + TTX. (g) Proposed model for EGF-mediated upregulation of Nav1.7 and consequent invasiveness of H460 cells. Stimulation of EGFR with EGF results in increased functional expression of Nav1.7 via ERK1/2. Following transcription and translation, the mature Nav1.7 protein is trafficked to the cell surface where it becomes functional. At the resting membrane potential, VGSCs are partially activated but not fully inactivated, resulting in a basal influx of Na+. This increase in [Na+]i then drives cell invasion through an, as yet, unknown mechanism. All data are presented as means ± s.e. (n = 6–13). Statistical analyses were with Student's t-test or one-way ANOVA and Student–Newman–Keuls correction, as appropriate; significance: *p < 0.05, **p < 0.01, ***p < 0.001. Adapted from [89]. (Online version in colour.)