title: 5-lipoxygenase and cyclooxygenase regulate wound closure in NIH/3T3 fibroblast monolayers.
abstract: Wound healing involves multiple cell signaling pathways, including those regulating cell-extracellular matrix adhesion. Previous work demonstrated that arachidonate oxidation to leukotriene B(4) (LTB(4)) by 5-lipoxygenase (5-LOX) signals fibroblast spreading on fibronectin, whereas cyclooxygenase-2 (COX-2)-catalyzed prostaglandin E(2) (PGE(2)) formation facilitates subsequent cell migration. We investigated arachidonate metabolite signaling in wound closure of perturbed NIH/3T3 fibroblast monolayers. We found that during initial stages of wound closure (0-120 min), all wound margin cells spread into the wound gap perpendicularly to the wound long axis. At regular intervals, between 120 and 300 min, some cells elongated to project across the wound and meet cells from the opposite margin, forming distinct cell bridges spanning the wound that act as foci for later wound-directed cell migration and resulting closure. 5-LOX inhibition by AA861 demonstrated a required LTB(4) signal for initial marginal cell spreading and bridge formation, both of which must precede wound-directed cell migration. 5-LOX inhibition effects were reversible by exogenous LTB(4). Conversely, COX inhibition by indomethacin reduced directed migration into the wound but enhanced early cell spreading and bridge formation. Exogenous PGE(2) reversed this effect and increased cell migration into the wound. The differential effects of arachidonic acid metabolites produced by LOX and COX were further confirmed with NIH/3T3 fibroblast cell lines constitutively over- and underexpressing the 5-LOX and COX-2 enzymes. These data suggest that two competing oxidative enzymes in arachidonate metabolism, LOX and COX, differentially regulate sequential aspects of fibroblast wound closure in vitro.
authors.affiliation: Program in Molecular and Cellular Biology, University of Massachusetts-Amherst, Amherst, MA 01003, USA.
authors.forename: J Angelo
authors.initials: JA
authors.lastname: Green
authors.forename: Rebecca A
authors.initials: RA
authors.lastname: Stockton
authors.forename: Christopher
authors.initials: C
authors.lastname: Johnson
authors.forename: Bruce S
authors.initials: BS
authors.lastname: Jacobson
mesh_terms: D000818:Animals; D001094:Arachidonate 5-Lipoxygenase / Q000235:genetics* / Q000378:metabolism*; D051546:Cyclooxygenase 2; D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D015232:Dinoprostone / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D007213:Indomethacin / Q000494:pharmacology; D007527:Isoenzymes / Q000235:genetics* / Q000378:metabolism*; D007975:Leukotriene B4 / Q000378:metabolism; D051379:Mice; D041681:NIH 3T3 Cells; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics* / Q000378:metabolism*; D014945:Wound Healing / Q000502:physiology*
chemical_list: D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors; D007527:Isoenzymes; D007975:Leukotriene B4; D001094:Arachidonate 5-Lipoxygenase; D051546:Cyclooxygenase 2; D011451:Prostaglandin-Endoperoxide Synthases; D015232:Dinoprostone; D007213:Indomethacin