FIBRINOGEN STRUCTURE, FUNCTION AND PHYSIOLOGY

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Fibroblast growth exert several important effects of endothelial cells including increasing proliferation, stimulating migration, up-regulating synthesis of several proteins and stimulation angiogenesis. Endothelial cell properties are also influenced by interaction with fibrinogen. and fibrin which can support adhesion, stimulate protein synthesis and secretion, alter inflammatory cell interactions and support angiogenesis. We have recently demonstrated that fibroblast growth factor-2 (FGF-2) binds specifically to fibrinogen and fibrin, and that the interaction with fibrinogen increases its capacity to stimulate endothelial cell proliferation. These findings form the basis for the studies proposed which have a goal of defining the structural basis of the interaction between fibrin(ogen) and FGF-2 and characterizing its functional implications. The first specific aim will be to characterize the interaction between FGF-2 and specific experiments will identify the binding sites involved and determine if other members of the FGF family also bind to fibrinogen. Such plasmon resistance techniques will be used to determine the association and dissociation rate constants and stoichiometry defining the binding. Preliminary experiments indicate that the binding of FGF-2 to fibrinogen provides protection from proteolytic degradation, and this may be important in prolonging action at inflammatory sites where thrombin and plasmin are active. Experiments in Aim 2 will explore the extent to which fibrinogen protects FGF-2 binds to plasmic derivatives of fibrinogen and retains activity when bound to such derivatives. The experiments in Aim 3 will extend the observation that the binding of FGF-2 to fibrinogen potentiates its capacity to stimulate endothelial cell proliferation. The involvement of known endothelial cell receptors in this potentiation of activity will be characterized using specific blocking monoclonal antibodies, peptides and disintegrins. The need for formation for an FGF-2-fibrinogen complex in potentiating proliferation will be examined using non-FGF-2-binding fibrinogen fragments of FGF-2 mutants. These will be used to determine if FGF-2 mutants lacking the ability to bind fibrin(ogen) can support angiogenesis in a fibrin matrix. Further experiments will determine if fibrinogen also alters effects of FGF-2 on endothelial cell migration and synthesis of u-PA. The results obtained will provide information regarding fundamental aspects of the association of FGF-2 and fibrin(ogen) which will be important for understanding normal vascular development and function, response to injury, and disease conditions in which endothelial cells are exposed to both fibrin(ogen) and fibroblast growth factors.