Fibronectin, a 440,000 dalton glycoprotein produce by fibroblasts, monocytes, and epithelial and endothelial cells, circulates in human plasma at concentrations of 300 g/ml; associates with fibrin upon activation of the clotting system; and is deposited at sites of inflammation and wound healing. The fibronectin deposition in these sites is both plasma-derived and cell-derived and is associated with proliferating migrating and localizing cell populations including monocyte/macrophage, endothelial cells, epidermal cells and fibroblasts. In vitro plasma or cellular fibronectin can bind to a substratum of collagen or fibrinogen and simultaneously bind to fibroblasts. These data taken together suggeest that fibronectin may provide a critical link between cells and their substrata especially during tissue remodeling. We propose to test this hypothesis by investigating how plasma and cellular fibronectin or fibronectin fragments affect normal human cell adherence and chemotaxis in vitro and cell localization in vivo. We have developed quantitative, reproducible assays for study of human cell adherence to determine whether normal human cells can utilize fibronectin for their attachment as do cell-lines and to study whether the requirement for fibronectin is the same among the various human cell types. To address the latter issue multiple parameters in the adherence assays will be varied to delineate time course for adherence, tenacity of adherence, optimal amounts of fibronectin for adherence and other optimal conditions for adherence of each cell type (Aim 1). We will also study epidermal cell adherence on fibroblast and epidermal cell derived substrates (Aim 1). Since we have already demonstrated that fibronectin enhances monocyte, and epidermal cell adherence, we will investigate mechanisms by which this may occur by altering the protein (Aim 2) and modifying the cell (Aim 3). In addition, we will investigate whether fibronectin or fibronectin fragments recruit cells by means other than cell adherence. Specifically, we have shown that the 120K and 210K fibronectin fragments stimulate monocyte chemotaxis and will study possible mechanism(s) by which chemoattractive fibronectin fragments may be generated in vivo and will define the limiting peptide sequence necessary for chemotactic activity (Aim 4). In addition, we will determine the effect of fibronectin and other ECM proteins on epidermal cell motility (Aim 5). Finally, we will investigate whether fibronectin or fibronectin fragments recruit cells in vivo (Aim 6). Our ultimate goal is to understand the role of fibronectin in wound healing.
|Wikner, N E; Elder, J T; Persichitte, K A et al. (1990) Transforming growth factor-beta modulates plasminogen activator activity and plasminogen activator inhibitor type-1 expression in human keratinocytes in vitro. J Invest Dermatol 95:607-13|
|Wikner, N E; Persichitte, K A; Baskin, J B et al. (1988) Transforming growth factor-beta stimulates the expression of fibronectin by human keratinocytes. J Invest Dermatol 91:207-12|