title: Cholesterol oxides mediated changes in cytoskeletal organisation involves Rho GTPases.
abstract: The small GTPases Rho, Rac, and Cdc42 regulate the actin cytoskeleton in all eukaryotic cells. In this study we have evaluated the effect of cholesterol oxides (7-ketocholesterol and 25-hydroxycholesterol) on cell migration, cell adhesion, and cytoskeletal organisation of lens epithelial cells (LEC). Effects of cholesterol oxides on cytoskeleton were evaluated by immunofluorescence confocal microscopy. The 7-ketocholesterol induced cell arborisation, with bundling of vimentin and tubulin in the cell processes and formation of filopodia and stress fibres. Cells treated with 25-hydroxycholesterol showed a collapse of vimentin filaments towards the nucleus and formation of lamellipodia. In addition, cells treated with 7-ketocholesterol or 25-hydroxycholesterol showed decreased migration. The effects of cholesterol oxides on cytoskeletal proteins involve the activation of the small GTPases Rho, Rac, and Cdc42. Indeed, formation of both filopodia and stress fibres induced by 7-ketocholesterol is inhibited by overexpressing dominant negatives forms of Cdc42 and RhoA, respectively. Similarly, the collapse of vimentin intermediate filament network and the formation of lamellipodia, induced by 25-hydroxycholesterol, is inhibited by overexpressing dominant negatives forms of Rac1. The effects of cholesterol oxides described in this study for LEC are also observed for at least two other cell lines (H36CE and U373), suggesting that this may represent a general mechanism whereby cholesterol oxides induces cytoskeletal disorganisation.
authors.affiliation: Centre of Ophthalmology, Biomedical Institute for Research in Light and Image (IBILI), Faculty of Medicine, University of Coimbra, 3000 Coimbra, Portugal.
authors.forename: Henrique
authors.initials: H
authors.lastname: Girão
authors.forename: Paulo
authors.initials: P
authors.lastname: Pereira
authors.forename: José
authors.initials: J
authors.lastname: Ramalho
authors.forename: Roy
authors.initials: R
authors.lastname: Quinlan
authors.forename: Alan
authors.initials: A
authors.lastname: Prescott
mesh_terms: D008841:Actin Cytoskeleton / Q000187:drug effects; D002448:Cell Adhesion / Q000187:drug effects; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects; D002784:Cholesterol / Q000494:pharmacology*; D003598:Cytoskeletal Proteins / Q000187:drug effects; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism*; D004847:Epithelial Cells / Q000166:cytology; D006801:Humans; D006888:Hydroxycholesterols / Q000494:pharmacology; D007653:Ketocholesterols / Q000494:pharmacology; D020741:rho GTP-Binding Proteins / Q000187:drug effects / Q000378:metabolism*
chemical_list: D003598:Cytoskeletal Proteins; D006888:Hydroxycholesterols; D007653:Ketocholesterols; C007997:25-hydroxycholesterol; D002784:Cholesterol; D020741:rho GTP-Binding Proteins; C003001:7-ketocholesterol