Orientation and directed migration of cultured corneal epithelial cells in small electric fields are serum dependent.

Reorientation and migration of cultured bovine corneal epithelial cells (CECs) in an electric field were studied. Electric field application was designed to model the laterally directed, steady direct current electric fields which arise in an injured corneal epithelium. Single cells cultured in media containing 10% foetal bovine serum showed significant galvanotropism, reorienting to lie perpendicular to electric field vector with a threshold field strength of less than 100 mV/mm. Cells cultured in serum-free medium showed no reorientation until 250 mV/mm. Addition of EGF, bFGF or TGF-beta 1 singly or in combination to serum free medium significantly restored the reorientation response at low field strengths. Both the mean translocation rate and directedness of cell migration were serum dependent. Cultured in medium with serum or serum plus added EGF, single cells showed obvious cathodal migration at 100 mV/mm. Increasing electric field strength enhanced the cathodal directedness of single cell migration. Supplementing serum free medium with growth factors restored the cathodal directed migration of single cells and highest directedness was found for the combination of EGF and TGF-beta 1. Corneal epithelial sheets also migrated towards the cathode in electric fields. Serum or individual growth factors stimulated CEC motility (randomly directed). Applied fields did not further augment migration rates but added a vector to stimulated migration. Electric fields which are present in wounded cornea interact with other environmental factors and may impinge on CECs migration during wound healing. Therapies which combine the application of growth factors and electric fields may be useful clinically.

[1]  Jeffrey L. Wrana,et al.  Mechanism of activation of the TGF-β receptor , 1994, Nature.

[2]  J. Forrester,et al.  P 139 Serum dramatically enhances orientation and migration of cultured bovine corneal epithelial cells in an applied electric field (EF) , 1995, Vision Research.

[3]  M. Poo,et al.  Orientation of neurite growth by extracellular electric fields , 1982, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[4]  N. Joyce,et al.  In situ injury-induced release of basic-fibroblast growth factor from corneal epithelial cells. , 1991, The American journal of pathology.

[5]  A. Ridley,et al.  The small GTP-binding protein rac regulates growth factor-induced membrane ruffling , 1992, Cell.

[6]  J. Curray,et al.  The Analysis of Two-Dimensional Orientation Data , 1956, The Journal of Geology.

[7]  H. Soong,et al.  Effects of electric fields on cytoskeleton of corneal stromal fibroblasts. , 1990, Current eye research.

[8]  R. Nuccitelli,et al.  Protein kinases are required for embryonic neural crest cell galvanotaxis. , 1993, Cell motility and the cytoskeleton.

[9]  K. Hotary,et al.  Endogenous electrical currents and voltage gradients in Xenopus embryos and the consequences of their disruption. , 1994, Developmental biology.

[10]  B. Abbott,et al.  EGF receptor expression in the developing tooth is altered by exogenous retinoic acid and EGF. , 1988, Developmental biology.

[11]  L M Loew,et al.  Electric field-directed fibroblast locomotion involves cell surface molecular reorganization and is calcium independent , 1994, The Journal of cell biology.

[12]  S. Wilson,et al.  Epidermal growth factor, transforming growth factor alpha, transforming growth factor beta, acidic fibroblast growth factor, basic fibroblast growth factor, and interleukin-1 proteins in the cornea. , 1994, Experimental eye research.

[13]  H. Haigler,et al.  Electric field-induced redistribution and postfield relaxation of epidermal growth factor receptors on A431 cells , 1987, The Journal of cell biology.

[14]  J. W. Vanable,et al.  Electrical fields in the vicinity of epithelial wounds in the isolated bovine eye. , 1992, Experimental eye research.

[15]  R. Nuccitelli,et al.  Embryonic fibroblast motility and orientation can be influenced by physiological electric fields , 1984, The Journal of cell biology.

[16]  Timo Tervo,et al.  EPIDERMAL GROWTH FACTOR (EGF) IN OCULAR FLUIDS: PRESENCE, ORIGIN AND THERAPEUTICAL CONSIDERATIONS , 1992, Acta ophthalmologica. Supplement.

[17]  L. Frati,et al.  Selective binding of the epidermal growth factor and its specific effects on the epithelial cells of the cornea. , 1972, Experimental eye research.

[18]  R. Weinberg,et al.  Biochemical evidence for the autophosphorylation and transphosphorylation of transforming growth factor beta receptor kinases. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[19]  A. Wells,et al.  Cell movement elicited by epidermal growth factor receptor requires kinase and autophosphorylation but is separable from mitogenesis , 1994, The Journal of cell biology.

[20]  S. Wilson,et al.  EGF, basic FGF, and TGF beta-1 messenger RNA production in rabbit corneal epithelial cells. , 1992, Investigative ophthalmology & visual science.

[21]  T. Nishida,et al.  Transforming growth factor-β modulates effects of epidermal growth factor on corneal epithelial cells , 1992 .

[22]  P. Gunning,et al.  Serum-induced signal transduction determines the peripheral location of beta-actin mRNA within the cell , 1994, The Journal of cell biology.

[23]  J. Boonstra,et al.  Epidermal growth factor induces rapid reorganization of the actin microfilament system in human A431 cells. , 1991, Journal of cell science.

[24]  C. McCaig,et al.  Factors influencing perpendicular elongation of embryonic frog muscle cells in a small applied electric field. , 1991, Journal of cell science.

[25]  S. Coleman,et al.  Inhibition of mouse mammary ductal morphogenesis and down-regulation of the EGF receptor by epidermal growth factor. , 1990, Developmental biology.

[26]  P. W. Luther,et al.  Changes in cell shape and actin distribution induced by constant electric fields , 1983, Nature.

[27]  F. Maxfield,et al.  Ca2+- and calcineurin-dependent recycling of an integrin to the front of migrating neutrophils , 1995, Nature.

[28]  E. Gherardi,et al.  Regulation of cell movement: the motogenic cytokines. , 1991, Biochimica et biophysica acta.

[29]  R. Isseroff,et al.  Human keratinocytes migrate to the negative pole in direct current electric fields comparable to those measured in mammalian wounds. , 1996, Journal of cell science.

[30]  R. Singer,et al.  Beta-actin mRNA localization is regulated by signal transduction mechanisms , 1994, The Journal of cell biology.

[31]  K R Robinson,et al.  The responses of cells to electrical fields: a review , 1985, The Journal of cell biology.

[32]  J. D. Zieske,et al.  Regional variation in distribution of EGF receptor in developing and adult corneal epithelium. , 1993, Journal of cell science.

[33]  P S Binder,et al.  In vitro model for corneal wound healing; organ-cultured human corneas. , 1995, Current eye research.

[34]  S. Lloyd,et al.  Fibroblast Growth Factor-1 Receptor Messenger RNA Expression in Corneal Cells , 1993, Cornea.

[35]  P. Khaw,et al.  Effects of epidermal growth factor, fibroblast growth factor, and transforming growth factor-beta on corneal cell chemotaxis. , 1992, Investigative ophthalmology & visual science.

[36]  R. Nuccitelli,et al.  Neural crest cell galvanotaxis: new data and a novel approach to the analysis of both galvanotaxis and chemotaxis. , 1991, Cell motility and the cytoskeleton.

[37]  J. W. Vanable,et al.  Intrinsic electric fields promote epithelization of wounds in the newt, Notophthalmus viridescens. , 1991, Developmental biology.

[38]  M. Nakamura,et al.  Transforming growth factor-beta modulates effects of epidermal growth factor on corneal epithelial cells. , 1992, Current eye research.

[39]  Anne J. Ridley,et al.  The small GTP-binding protein rho regulates the assembly of focal adhesions and actin stress fibers in response to growth factors , 1992 .

[40]  J. Thiery,et al.  † Author for correspondence , 1693 .

[41]  S. Kinoshita,et al.  Immunohistochemical localization of transforming growth factor-beta 1, -beta 2, and -beta 3 latency-associated peptide in human cornea. , 1994, Investigative ophthalmology & visual science.

[42]  J. Forrester,et al.  2416 Cultured bovine corneal epithelial cells in a small applied electric field: Effects of EGF, bFGF and TGF , 1995, Vision Research.

[43]  A. Ullrich,et al.  Separate endocytic pathways of kinase-defective and -active EGF receptor mutants expressed in same cells , 1990, The Journal of cell biology.

[44]  S. Wilson,et al.  Hepatocyte growth factor, keratinocyte growth factor, their receptors, fibroblast growth factor receptor-2, and the cells of the cornea. , 1993, Investigative ophthalmology & visual science.

[45]  H. Peng,et al.  A role of tyrosine phosphorylation in the formation of acetylcholine receptor clusters induced by electric fields in cultured Xenopus muscle cells , 1993, The Journal of cell biology.

[46]  I. Gipson,et al.  A technique for obtaining sheets of intact rabbit corneal epithelium. , 1982, Investigative ophthalmology & visual science.

[47]  S. Klyce.  Electrical profiles in the corneal epithelium , 1972, The Journal of physiology.

[48]  Anne J. Ridley,et al.  The small GTP-binding protein rac regulates growth factor-induced membrane ruffling , 1992, Cell.

[49]  A. Ullrich,et al.  Growth factor signaling by receptor tyrosine kinases , 1992, Neuron.

[50]  J. Imanishi,et al.  Distribution of epidermal growth factor (EGF) receptors in rabbit corneal epithelial cells, keratocytes and endothelial cells, and the changes induced by transforming growth factor-β1 , 1992 .

[51]  D. Maurice.  Epithelial potential of the cornea. , 1967, Experimental eye research.

[52]  K. Hotary,et al.  Evidence of a role for endogenous electrical fields in chick embryo development. , 1992, Development.

[53]  H. Soong,et al.  Movements of cultured corneal epithelial cells and stromal fibroblasts in electric fields. , 1990, Investigative ophthalmology & visual science.

[54]  A. Neufeld,et al.  A tissue culture assay of corneal epithelial wound closure. , 1986, Investigative ophthalmology & visual science.

[55]  R. Nuccitelli.  Physiological Electric Fields can Influence Cell Motility, Growth, and Polarity , 1988 .

[56]  S. Wilson,et al.  EGF, EGF receptor, basic FGF, TGF beta-1, and IL-1 alpha mRNA in human corneal epithelial cells and stromal fibroblasts. , 1992, Investigative ophthalmology & visual science.

[57]  K. Hotary,et al.  Endogenous electrical currents and the resultant voltage gradients in the chick embryo. , 1990, Developmental biology.

[58]  S. Kinoshita,et al.  The mechanism of accelerated corneal epithelial healing by human epidermal growth factor. , 1990, Investigative ophthalmology & visual science.

[59]  W. C. Parkinson,et al.  Effects of steady electric fields on human retinal pigment epithelial cell orientation and migration in culture , 1992, Acta ophthalmologica.