Cell-extracellular matrix adhesion is necessary for rapid embryonic wound closure
Authors: Michelle Ly 1,2,3; Katheryn Rothenberg 1,2; Clara Schimmer 1,2; Raymond Hawkins 1,2; Rodrigo Fernandez-Gonzalez 1,2,3,4,5
Affiliations: 1) Institute of Biomedical Engineering, University of Toronto, Canada; 2) Translational Biology and Engineering Program, Ted Rogers Centre for Heart Research, University of Toronto, Canada; 3) Collaborative Specialization in Developmental Biology, University of Toronto, Canada; 4) Department of Cell and Systems Biology, University of Toronto, Canada; 5) Developmental and Stem Cell Biology Program, The Hospital for Sick Children, Canada
Keywords: s. extracellular matrix; a. cytoskeleton
Embryos repair wounds rapidly, with no inflammation or scarring. In embryos, wound repair is driven by the collective movement of the cells around the wound to seal the lesion. Embryonic wound healing is associated with reorganization of the actomyosin cytoskeleton and cell-cell adhesive structures in the cells adjacent to the wound. Cell migration often involves adhesion to and remodeling of the extracellular matrix (ECM) through integrin-based adhesions. However, the role of integrin-based adhesion in embryonic wound closure has not been investigated. To establish the role of cell-ECM adhesion in embryonic wound repair, we imaged and wounded the epidermis of embryos mid-way through development (12-14 hours) using a pulsed ultraviolet laser mounted on a spinning disk confocal microscope. Using embryos expressing fluorescently labeled talin, we found that upon wounding, the cells adjacent to the wound polarized talin to the wound edge, suggesting increased cell-ECM interactions at the wound margin. To determine the impact of cell-ECM adhesion on embryonic wound closure, we injected embryos with RGDS, a peptide that binds integrins and inhibits ECM binding. RGDS treatment delayed wound closure by 43% with respect to controls. Imaging of fluorescently labeled myosin II revealed that the delay in wound healing occurred with no significant effects on the myosin rearrangements associated with wound repair. We obtained similar results when we knocked down integrins using RNA interference against the β integrin subunit: wound closure displayed a 54% delay compared to controls with no changes in myosin dynamics around the wound. However, we found a 71% decrease in actin accumulation around the wound edge relative to controls. We also noticed a defect in the E-cadherin reorganization necessary for actomyosin remodelling during wound healing. In control embryos, E-cadherin accumulated at former tricellular junctions at the wound margin, displaying a 29% increase in tricellular junction E-cadherin fluorescence 15 minutes after wounding. In contrast, RGDS treatment resulted in a 13% decrease in tricellular junction E-cadherin at the same time point. We obtained similar results in integrin RNAi embryos, which displayed a 20% decrease in tricellular junction E-cadherin. Together, our results suggest that cell-ECM adhesion contributes to wound repair and reveal a previously unrecognized interplay between cell-cell and cell-ECM adhesion in the collective cell movements that drive embryonic wound closure.