Background: The activation of protein kinase C (PKC) and transforming growth factor-beta (TGF-beta) in glomerular mesangial cells has been linked to mesangial matrix expansion in diabetic nephropathy. The role of these mediators in affecting the changes associated with diabetes in the biology of glomerular endothelial cells (GEnCs), which synthesize components of the glomerular basement membrane, is not known. We postulated that the PKC and TGF-beta systems promote the increased endothelial cell synthesis of glomerular basement membrane that is evoked by Amadori-modified glycated albumin, which is present in elevated concentrations in diabetes.
Methods: We examined the effects of PKC inhibition on collagen IV and TGF-beta1 production by mouse GEnCs incubated with glycated albumin and the influence of glycated albumin on PKC activity, TGF-beta 1 production, and proliferation by these cells.
Results: In physiologic (5.5 mmol/L) glucose concentrations, glycated albumin caused an increase in type IV collagen production that was totally prevented by a general PKC inhibitor GF 109203X (GFX), but only partly prevented by a neutralizing anti-TGF-beta antibody. Glycated albumin increased the steady-state level of TGF-beta 1 mRNA and stimulated the production of TGF-beta 1 protein, which was also prevented by the PKC inhibitor GFX. Of note, glycated albumin significantly stimulated PKC activity, as measured by the phosphorylation of a PKC-specific substrate. Cell proliferation, measured by [(3)H]-thymidine incorporation and cell counting, was decreased in the presence of glycated albumin. This effect was completely prevented by GFX and partially reversed by anti-TGF-beta antibody. Exogenous TGF-beta 1 inhibited cell proliferation to a degree similar to that of glycated albumin.
Conclusions: PKC signaling and consequent TGF-beta 1 activation participate in the glycated albumin-induced stimulation of basement membrane collagen production by GEnC. By reducing the proliferative capacity, which is likely mediated by PKC and partly by TGF-beta, glycated albumin impedes the ability of the glomerular capillary endothelium to act as a first line of defense against deleterious circulating factors in the diabetic state.