The overall goal of the proposed studies is to elucidate cellular and molecular regulatory mechanisms that underlie acquisition of the invasive phenotype in a human skin organ culture model of invasion. In this model, exposure of organ-cultured skin to certain exogenous growth factors (notably epidermal growth factor or hepatocyte growth factor) results in erosion of the dermal-epidermal basement membrane and invasion of the underlying stroma by the epithelium. Acquisition of the invasive phenotype is accompanied by elaboration of increased amounts of matrix metalloproteinase-9 (MMP-9) (92-kD gelatinase B/type IV collagenase) and induction of motility in the epithelial cell population, and by up-regulation of collagenolytic activity in the adjacent stroma. It is hypothesized that exposure of the target tissue to the exogenous growth factors initiates signal-transduction through two mitogen-activated protein kinase (MAPK) cascades in the epidermis, and that the activation of these pathways results in MMP-9 elaboration and in the induction of keratinocyte motility. It is hypothesized further that stimulated epithelial cells elaborate factors that induce production of MMP-1 (interstitial collagenase) by adjacent fibroblasts. Together, these events bring about invasion.
In Specific Aim I of the proposed studies, the goal will be to define the intracellular signaling events that are required for up-regulation of MMP-9 production and for stimulation of cell motility in the epidermis. A combination of analytical experiments and interventional approaches will be used to define the relevant signaling events and to interfere with their expression.
In Specific Aim II, studies will be carried out to demonstrate that invasion-associated MMP-1 elaboration by dermal fibroblasts is a result of interaction of dermal fibroblasts with products of growth factor-stimulated epithelial cells (rather than with the exogenous growth factors themselves); and to assess intracellular signaling events that are required for fibroblast up-regulation of MMP-1.
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