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K. Kawamoto; Cell adhesion and migration of human corneal epithelial cells on fibronectin mediated by small GTPase Rac1 . Invest. Ophthalmol. Vis. Sci. 2004;45(13):4893.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Fibronectin–integrin system plays essential rolls in human cornea epithelial wound healing through cell adhesion and migration. Rho GTPases regulate cell migration, adhesion, polarity through the remodeling of actin cytoskeleton. In the present study, we examined the involvement of Rac1, one of Rho GTPases, in fibronectin–integrin system to clarify signal transduction responsible for cell adhesion and cell migration on fibronectin. Methods: Simian virus 40–transformed human corneal epithelial cells were cultured for 24 h in medium containing 1% fetal bovine serum, dissociated from the culture dish by exposure to trypsin–EDTA, and then replated on cover glasses coated with fibronectin or bovine serum albumin. Cell behavior was monitored by time–lapse videography. Alternatively, the cells were stained with rhodamine–conjugated Phalloidin and subjected to indirect immunostaining with antibodies to phosphotyrosine for visualization of the actin cytoskeleton and focal adhesions, respectively, by confocal microscopy. The effects of transfection of cells with an expression plasmid for a Myc epitope–tagged dominant negative mutant of Rac1 (Rac1–Asn17) were also determined. Results: Fibronectin promoted cell adhesion and migration when compared with cells plated on bovine serum albumin. The numbers of focal adhesions and membrane ruffles were thus both greater for cells plated on fibronectin. Expression of Rac1–Asn17 markedly inhibited the effects of fibronectin both on the formation of focal adhesions and on membrane ruffling. Results: Fibronectin facilitated cell adhesion and migaration compared with BSA as a control. The number of focal adhesion increased and membrane ruffles were promoted in cells on fibronectin. Rac1Asn17 overexpressed cells on fibronectin downregulated the formation of focal adhesion and membrane ruffling. Conclusions: Rac1 is essential for cornea wound healing through cell adhesion and migration.
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