title: Wound closure in sheared endothelial cells is enhanced by modulation of vascular endothelial-cadherin expression and localization.
abstract: We previously demonstrated that laminar shear stress enhances human coronary artery endothelial cell (HCAEC) wound closure via the mechanisms of cell spreading and migration. Because cell-cell junctional proteins such as vascular endothelial cell cadherin (VE-cadherin) are critical to cell-cell adhesion and motility, we tested the hypothesis that modulation of VE-cadherin expression under shear stress may be linked to this enhancement in wound closure. HCAEC monolayers were preconditioned to attain cellular alignment by shearing at 12 dynes/cm(2) for 18 hr in a parallel-plate flow chamber. Subsequently, they were divided into the following three groups: (i) control; (ii) treated with anti-cadherin-5 antibody; or (iii) treated with the calcium chelating agent EGTA. Next, the monolayers were wounded with a metal spatula and resheared at 20 dynes/cm(2) or left static. Time-lapse imaging was performed during the first 3 hr after imposition of these conditions. Immunocytochemistry or Western blot analyses for VE-cadherin expression were performed on all wounded monolayers. Deconvolution microscopy, three-dimensional cell-cell junctional reconstruction images, and histogram analyses of interendothelial junction signal intensities were performed on cells at the wound edge of a monolayer. Under shear, HCAEC demonstrated increased VE-cadherin immunofluorescence and protein expression despite an enhancement in wound closure compared with static conditions. In separate experiments, application with anti-cadherin-5 antibody or treatment with EGTA attenuated VE-cadherin expression and further enhanced wound closure compared with control shear and all static conditions. In addition, the pattern of VE-cadherin localization with these treatments became more intracellular and nuclear in appearance. These findings of changes in this junctional adhesion protein expression and localization may further our understanding of laminar shear stress-induced endothelial repair in the coronary circulation.
authors.affiliation: Critical Care Laboratory of Vascular Research, Division of Critical Care Medicine (Children's Memorial Hospital), Department of Pediatrics, Northwestern University, Feinberg School of Medicine, Chicago, Illinois 60611, USA. malbuquerque@northwestern.edu
authors.forename: Maria Luiza C
authors.initials: ML
authors.lastname: Albuquerque
authors.forename: Annette S
authors.initials: AS
authors.lastname: Flozak
mesh_terms: D015153:Blotting, Western; D015820:Cadherins / Q000378:metabolism*; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D008856:Microscopy, Fluorescence; D014945:Wound Healing
chemical_list: D015820:Cadherins