involves multiple cell signaling pathways, including those regulating cell-extracellular matrix adhesion. Previous work demonstrated that arachidonate oxidation to leukotriene B4 (LTB4) by 5-lipoxygenase (5-LOX) signals fibroblast spreading on fibronectin, whereas cyclooxygenase-2 (COX-2)-catalyzed prostaglandin E2 (PGE2) 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 LTB4 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 LTB4. Conversely, COX inhibition by indomethacin reduced directed migration into the wound but enhanced early cell spreading and bridge formation. Exogenous PGE2 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.

Transparent substrates having heterogeneities ranging from nanometer to micrometer lateral length scale were fabricated to study cell migration. The surfaces were generated using thin films of block copolymers and homopolymer blends on ultra smooth transparent polyethylene terephthalate films. Results show that the lateral size scale of the surface heterogeneities affects fibroblast (NIH-3T3) adhesion, spreading and motility. More specifically, fibroblasts migrate faster on micron-sized than on nanometer-sized heterogeneities. Cell movements and morphology on the micron patterned surfaces resemble cells cultured in a 3D environment. These surfaces, therefore, can potentially be utilized as models to study cell behavior in physiologically relevant conditions which can add to our fundamental understanding of cell-substrate interactions and facilitate development of surfaces for medical devices.