Electric Field–directed Cell Motility Involves Up-regulated Expression and Asymmetric Redistribution of the Epidermal Growth Factor Receptors and Is Enhanced by Fibronectin and Laminin

Abstract

Wounding corneal epithelium establishes a laterally oriented, DC electric field (EF). Corneal epithelial cells (CECs) cultured in similar physiological EFs migrate cathodally, but this requires serum growth factors. Migration depends also on the substrate. On fibronectin (FN) or laminin (LAM) substrates in EF, cells migrated faster and more directly cathodally. This also was serum dependent. Epidermal growth factor (EGF) restored cathodal-directed migration in serum-free medium. Therefore, the hypothesis that EGF is a serum constituent underlying both field-directed migration and enhanced migration on ECM molecules was tested. We used immunofluorescence, flow cytometry, and confocal microscopy and report that 1) EF exposure up-regulated the EGF receptor (EGFR); so also did growing cells on substrates of FN or LAM; and 2) EGFRs and actin accumulated in the cathodal-directed half of CECs, within 10 min in EF. The cathodal asymmetry of EGFR and actin staining was correlated, being most marked at the cell-substrate interface and showing similar patterns of asymmetry at various levels through a cell. At the cell-substrate interface, EGFRs and actin frequently colocalized as interdigitated, punctate spots resembling tank tracks. Cathodal accumulation of EGFR and actin did not occur in the absence of serum but were restored by adding ligand to serum-free medium. Inhibition of MAPK, one second messenger engaged by EGF, significantly reduced EF-directed cell migration. Transforming growth factor ␤ and fibroblast growth factor also restored cathodal-directed cell migration in serum-free medium. However, longer EF exposure was needed to show clear asymmetric distribution of the receptors for transforming growth factor ␤ and fibroblast growth factor. We propose that up-regulated expression and redistribution of EGFRs underlie cathodaldirected migration of CECs and directed migration induced by EF on FN and LAM.

The epithelial polarity program: machineries involved and their hijacking by cancer

“…The EGFR, a critical protooncogene, contains two basolateral sorting signals, namely PXXP and LL-motifs , and has been shown to localize to the basolateral side in polarized epithelia (Borg et al, 2000) and to the leading edge in EGF stimulated and migrating cells (Diakonova et al, 1995;Zhao et al, 1999;Sorkin et al, 2000;Lynch et al, 2003;Orth et al, 2006). A critical feature of polarized epithelia is the different compartmentalization of GFs to the basolateral side and receptors to the apical side, which prevents receptor activation.…”

Harnessing the Electric Spark of Life to Cure Skin Wounds

“…35,36 EF exposure also up-regulates the expression of EGF receptors in epithelial cells. 35 Pharmacological inhibition of multiple tyrosine kinases, including EGFR, reduces the cell migration rate; whereas specific pharmacological inhibition of EGFR kinase activity reduces directed motility in EFs. 36 EGF receptor signaling is, therefore, considered an important signaling component of the EF-mediated contribution to wound closure.…”

“…For keratinocytes and corneal epithelial cells, this occurs within 5-10 min of EF exposure. 35,36 As a consequence, EGF signaling becomes polarized, causing greater cathodal activation of extracellular signal-regulated kinases 1/2 (ERK1/2) and downstream cathodal polymerization of Filamentous actin. 35,36 EF exposure also up-regulates the expression of EGF receptors in epithelial cells.…”

“…35,36 As a consequence, EGF signaling becomes polarized, causing greater cathodal activation of extracellular signal-regulated kinases 1/2 (ERK1/2) and downstream cathodal polymerization of Filamentous actin. 35,36 EF exposure also up-regulates the expression of EGF receptors in epithelial cells. 35 Pharmacological inhibition of multiple tyrosine kinases, including EGFR, reduces the cell migration rate; whereas specific pharmacological inhibition of EGFR kinase activity reduces directed motility in EFs.…”

Effects of Micro-Electrical Stimulation on Regulation of Behavior of Electro-Active Stem Cells

“…1997; Zhao et al 1999;Haddad et al 2007;Aaron et al 1999;Aaron et al 2002;Au et al 2009;Tandon et al 2010). Firstly, electrical stimulation (ES) alters the complex interplay between living cells and their physical environments, which plays an integral role in maintaining normal biological functions in the human body (Haddad et al 2007).…”