IGFB-2 is the only expressed IGFB in a murine kidney glomerular mesangial cell model, providing for a unique experimental system in which to study the biochemical and cell regulatory properties of this protein in the absence of interference by other IGFBPs. It has been found that IGFBP-2 associates with mesangial cell plasma membranes while simultaneously binding to IGF-1 (or IGF-II), suggesting that IGFBP-2 may regulate IGF peptide availability in close proximity to the cellular type-1 IGF receptors. Since the presence of IGFBP-2 on the cell membrane can be competitively inhibited in the presence of fibronectin or synthetic peptides containing the specific arg-gly-asp (RGD) integrin receptor recognition sequence, the membrane binding of IGFBP-2 appears to occur, at least in part, via integrin receptors. When the cells are cultured in the presence of a diabetic concentration of glucose (25 mmol/L), both basal and IGF-stimulated ECM production is stimulated concomitantly with substantial increases in extracellular levels of IGFBP- 2 distributed both on the cell membranes as well as in the medium. This increase in IGFBP-2 is not associated with altered mRNA levels, but rather with an inhibition of proteolytic breakdown of IGFBP-2 by a protease(s) produced by cells in high glucose, a mechanism which favors accumulation of IGFBP-2 protein. It is there hypothesized that IGFBP-2 has direct cell-regulatory effects mediated at the plasma membrane level that may include potentiation of IGF/IGF-receptor interaction and/or """"""""IGF-independent"""""""" pathways initiated by direct IGFBP-2 integrin receptor binding. A second, related hypothesis is that such direct IGFBP- 2 action may mediate, at least in part, activation of mesangial cell anabolic activity under diabetic glucose conditions and may therefore represent a novel mechanism of action pertinent to diabetic glomerulopathy. These hypotheses will be tested under three specific aims, which will: 1. define the underlying biochemical mechanisms- and resultant cell-regulatory impacts- of IGFBP-2 binding to mesangial cell membranes and its proteolysis by a cell-produced protease(s), 2. establish the degree to which IGFB-2 is directly involved in the activation of mesangial cell anabolic activity, particularly under diabetic glucose conditions, and 3. define the relative participation of IGF-R-versus integrin receptor-mediated events in mesangial cells activity by IGFBP-2 and diabetic glucose.
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