Signaling pathways and cell mechanics involved in wound closure by epithelial cell sheets

Sheets of cells move together as a unit during wound healing and embryonic tissue movements, such as those occurring during gastrulation and neurulation. We have used epithelial wound closure as a model system for such movements and examined the mechanisms of closure and the importance of the Rho family of Ras-related small GTPases in this process. Wounds induced in Madin-Darby canine kidney (MDCK) epithelial cell monolayers close by Rac- and phosphoinositide-dependent cell crawling, with formation of lamellipodia at the wound margin, and not by contraction of a perimarginal actomyosin purse-string. Although Rho-dependent actin bundles usually form at the margin, neither Rho activity nor formation of these structures is required for wound closure to occur at a normal rate. Cdc42 activity is also not required for closure. Inhibition of Rho or Cdc42 results, however, in statistically significant decreases in the regularity of wound closure, as determined by the ratio of wound margin perimeter over the remaining denuded area at different times. The Rac-dependent force generation for closure is distributed over several rows of cells from the wound margin, as inhibition of motility in the first row of cells alone does not inhibit closure and can be compensated for by generation of motile force in cells behind the margin. Furthermore, we observed high levels of Rac-dependent actin assembly in the first few rows of cells from the wound margin. Wounds in MDCK cell sheets do not close by purse-string contraction but by a crawling behavior involving Rac, phosphoinositides and active movement of multiple rows of cells. This finding suggests a new distributed mode of signaling and movement that, nevertheless, resembles individual cell motility. Although Rho and Cdc42 activities are not required for closure, they have a role in determining the regularity of closure.