A Functional Fibroblast Growth Factor-1 Immunoglobulin Fusion Protein

Abstract

FGF-1 and FGF-2 potently induce proliferation and migration of these cells as well as angiogenesis, events that are required during normal embryogenesis and tissue repair. Although expression of FGF-1 and FGF-2 at sites of injury is therefore beneficial, some evidence suggests these growth factors may also contribute to vascular pathology by promoting excessive intimal hyperplasia (2-5). FGF-1 expression is increased at several sites of chronic immune injury, including the synovium in rheumatoid arthritis (6 -8), the myocardium in human hearts after transplantation (9 -11), and in transplanted kidneys undergoing chronic rejection (12, 13). The pathologic lesions in these sites demonstrate cellular responses typical of FGF effects on mesenchymal cells that result in vascular intimal hyperplasia, increased extracellular matrix deposition, and neoangiogenesis. In addition, these sites are characterized by chronic infiltration of T lymphocytes, suggesting there may be interactions between the immune system and fibroblast growth factors as demonstrated by the finding that T cells can produce FGF-2 (14, 15). Evidence that FGFs may have immunoregulatory effects on T cells was first provided in 1985 by Johnson and Torres (16), who showed that FGF, at physiologically relevant concentrations, could replace the requirement for IL-2 or helper cells in production of interferon-␥. Although FGF could activate intracellular signals necessary for interferon-␥ production, FGF alone could not stimulate proliferation of T cells. More recent studies show directly that some human T cells express receptors for FGF-1 and that the normally small subpopulation of FGF-responsive T cells is expanded in the peripheral blood of patients with rheumatoid arthritis and in patients after heart transplantation (8, 17). These data suggest that T cells bearing FGF receptors can be stimulated and expanded in the FGF-enriched environment at sites of immune injury and subsequently migrate to the peripheral blood. As found in the earlier studies (16), FGF alone does not stimulate proliferation of T cells but together with engagement of the T cell antigen receptor induces production of IL-2 and proliferation (17). In T cells, FGF thus functions in a manner analogous to other well described "costimulators" (18,19) to activate a second signal transduction pathway necessary for T cell proliferation and effector function.These findings suggest that FGF and FGF receptors in T cells may function quite differently than in cells in which FGF alone can stimulate proliferation, migration, and secretion of effector molecules such as plasminogen activator (20). Our efforts to investigate T cells that express FGF receptors and the mechanisms by which FGF signals in T cells have been hampered by the lack of reagents that can conveniently identify these cells and allow us to examine the fate of FGF and its receptors in T cells. To address this difficulty, the experiments reported here describe the production and characterization of a fusion protein that includes a por...

Immunolocalization of fibroblast growth factor-1 (FGF-1), its receptor (FGFR-1), and fibroblast-specific protein-1 (FSP-1) in inflammatory renal disease

“…The tissues were blocked with 1% (wt/vol) bovine serum albumin (BSA) and incubated with polyclonal antibodies directed against FGF-1 (1:100) (SigmaAldrich, St. Louis, MO, USA) or FGFR-1 (1:200) (Santa Cruz Biotechnology, Santa Cruz, CA, USA), directed at the high-affinity signaling isoform with active kinase domain, thus detecting functional receptors, for 90 minutes at room temperature, or against FSP-1 (polyclonal rabbit anti-FSP-1) (1:200), at 37 • C for 1 hour in a humidified chamber [1]. This specificity was further verified by using an FGF-Ig fusion protein to first immunoprecipitate FGFR-1 and then blot with the anti-FGFR-1 reagent [21]. In addition, monoclonal antibodies directed at the first Ig loop of FGFR-1 and inhibition with a FGFR-1 specific peptide also confirmed the specificity of the anti-FGFR-1 reagent (unpublished observations).…”

Pretreatment Procedure for the Microdetermination of Chondroitin Sulfate in Plasma and Urine

“…GAGs have been found in biological fluids such as blood plasma and urine (5,6). In blood plasma, GAGs interact with biologically important proteins, including protease inhibitors, coagulation factors, lipoproteins, and complement proteins (7); cells such as lymphocytes (8), monocytes (9), and platelets (10); and the vascular endothelium (11). The presence of GAGs in human plasma has been demonstrated in several reports (12,13); however, the structures and physicochemical properties of these GAGs have not been fully identified and characterized due to their low blood concentration and a general lack of adequate analytical methodology.…”

c-Src Regulation of Fibroblast Growth Factor-induced Proliferation in Murine Embryonic Fibroblasts

“…To confirm expression of FGFR1 protein capable of ligand binding, FACS analysis was performed using an Fc-FGF fusion protein (26). After incubation with Fc-FGF, the fibroblasts were washed with heparin to remove ligand from low affinity heparan sulfate proteoglycan binding sites on the cell surface.…”