Roles of wound geometry, wound size, and extracellular matrix in the healing response of bovine corneal endothelial cells in culture

Abstract

It has classically been accepted that the healing of narrow wounds in epithelia occurs by the formation of a contractile actin cable, while wide wounds are resurfaced by lamellipodia-dependent migration of border cells into the denuded area. To further investigate the general validity of this idea, we performed systematic experiments of the roles of wound geometry, wound size, and extracellular matrix (ECM) in wound healing in monolayers of bovine corneal endothelial cells, a system shown here to predominantly display any of the two healing mechanisms according to the experimental conditions. We found that, in this system, it is the absence or presence of the ECM on the wound surface that determines the specific healing mode. Our observations demonstrate that, independent of their size and geometry, wounds created maintaining the ECM heal by migration of cells into the wound area, while ECM removal from the wound surface determines the predominant formation of an actin cable. While the latter mechanism is slower, the actin cable permits the maintainance of the epithelial phenotype to a larger extent during the healing process, as also confirmed by our finding of a more conserved localization of cadherin and vinculin. We also introduce a model that simulates experimental findings about the dynamics of healing mechanisms, both for the maintenance or removal of the ECM on the wound surface. The findings of this study may contribute to the understanding of physiological and pathological aspects of epithelial wound healing and to the design of therapeutic strategies.

Soybean agglutinin binding to corneal endothelial cell surfaces disrupts in situ monolayer integrity and actin organization and interferes with wound repair

“…Our present study also demonstrates that in situ it co-localizes around the cell periphery with N-cadherin. Previous studies of confluent cultures of bovine endothelial cells have shown that N-cadherin localization is also confined to the cell periphery (Chifflet et al 2004;Chen et al 2007;Grasso et al 2007). During normal wound repair, ZO-1 redistributes as speckled focal points in cells entering the injury area.…”

Loss of p27 upregulates MnSOD in a STAT3-dependent manner, disrupts intracellular redox activity and enhances cell migration

“…When cells reached ,80% confluence, a scratch was generated on the confluent monolayer using a silicon-coated pipette tip, which preserved the extracellular matrix across the scratched area (Grasso et al, 2007). Photographs showing the areas of the wounds in the control group and test group were taken at specific time-points up to 24 h. The wound areas were quantified by using cell migration analysis software (Muscale LLC., Scottsdale, AZ).…”

A mathematical analysis of physiological and morphological aspects of wound closure

“…Thus, Brock et al [9] demonstrated that when the formation of the actin cable is disrupted in embryo chick wounds, cells develop lamellipodia and the healing process may be compromised. Alternatively, Grasso et al [16] showed that the actin cable is not formed when the extracellular matrix is preserved after wounding in in vitro cultures of corneal endothelial cells, and lamellipodiainduced migration was obtained independently of wound shape or size. The migration mechanism may as well be influenced by the size of the wound-significantly smaller in embryos [9,26]-and growth factor levels-much lower in embryonic wounds [36], which allows for a stronger influence of mechanical constrains.…”