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Versican in the wound healing matrix : cellular interactions and degradation by matrix metalloproteinases Pourmalek, Saloumeh

Abstract

In wound healing, versican is a component of the provisional matrix laid down at the site of injury by proliferating myofibroblasts. Versican interacts with a variety of matrix molecules and is believed to interact with the cell surface. The mechanism of interaction of versican with the cell surface, however, is not well documented. Return to normal tissue structure, at late stages of wound healing, involves degradation of versican and concomitant fibroblast apoptosis. Macrophage enzymes are candidates for versican degradation; however, the mechanisms of actions of these enzymes on versican and the rates and cleavage sites are not yet known. This thesis tests several hypotheses: 1) Versican interacts with cell surface receptors of myofibroblasts and macrophages; 2) Versican influences myofibroblast cell morphology during wound contraction; and 3) Macrophage matrix metalloproteinases degrade versican during wound resolution. We first attempted to identify macrophage and fibroblast versican-binding cell surface ligands. Using biotinylated constructs of the C-terminal domain of versican as baits, we identified versican and versican fragments as the main ligands for the C-terminal construct. However, we found that most versican could be released from the cell surface by hyaluronidase treatment, and concluded that versican is held at the fibroblast cell surface mainly through its interaction with hyaluronan. Next, we examined the influence of versican and hyaluronan on the physical properties of a collagenous matrix, and the cells embedded within the matrix, using a novel 3-dimensional collagen/versican/hyaluronan matrix model. We found that fibroblast cells in matrices containing versican express smooth muscle actin and take on a contractile morphology. Finally, we hypothesized that macrophage metalloproteinases degrade versican. The macrophage matrix metalloproteinases (MMPs), MMP 2, MMP-7, and MMP-12 were chosen as candidate enzymes, which we localized to the resolving phase of wound healing in the human lung. We found that MMP-7 and MMP-12 cleave versican at multiple sites in vitro, whereas MMP-2 cleaves versican at a limited number of sites. These macrophage enzymes may be important in clearing versican in vivo. A better understanding of the mechanism of versican degradation could enable therapeutic modification of the disease process in fibrosis, cancer, and nervous system regeneration.

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