Little is known about the function of the extracellular matrix in the progression of crescentic glomerulonephritis (GN) to sclerosis and irreversible loss of renal function. Thrombospondin (TSP) is an extracellular matrix component which is synthesized by several cell types and which appears to be important in the regulation of cellular proliferation. TSP has never been described to be a participant in the pathophysiology of crescentic GN or any inflammatory state. In a preliminary experiment, TSP was observed to be present in glomeruli in large amounts relative to control following induction of injury in a rabbit model of crescentic GN. This proposal represents a series of experiments aimed at understanding the role that TSP plays in mediating glomerular injury in crescentic GN. We will characterize the temporal and spacial pattern of distribution and synthesis of TSP in a rabbit model of crescentic GN, using the techniques of immunofluorescence, Northern analysis and dot blot hybridization, as well as immunocytochemistry and in situ hybridization. In order to investigate TSP's role in the GN environment on a cellular and molecular level, mesangial cell (MC) culture systems will be employed. Specific growth factors (GF) will be assessed for their ability to stimulate TSP production. Using TSP inhibitors we will investigate whether TSP-cell binding is important in regulation of mesangial cell proliferation and in doing so test whether TSP represent a mechanism by which heparin inhibits MC proliferation. Further, the molecular mechanism (increased transcription or stability) by which TSP mRNA accumulation occurs after GF stimulation will be investigated. Finally, to establish that TSP does play an essential role in the evolution of glomerular injury and sclerosis in crescentic GN, in vivo blocking experiments are proposed using non-anticoagulant heparin, an Fab of monoclonal anti-TSP antibodies, and a recombinant TSP-heparin binding site fragment. Understanding the role of TSP in crescentic GN and the affect of specific TSP inhibitors may lead to attempted clinical use of these agents to attenuate progression to sclerosis in inflammatory glomerular diseases in man.
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