THE EFFECT OF TRANSFORMING GROWTH FACTOR-BETA(1) ON MESANGIAL CELL FIBRONECTIN SYNTHESIS - INCREASED INCORPORATION INTO THE EXTRACELLULAR-MATRIX AND REDUCED PI BUT NO EFFECT ON ALTERNATIVE SPLICING

Fibronectin is a multidomain glycoprotein which accumulates in mesangi al proliferative glomerulonephritis (MPGN). Recent evidence has implic ated transforming growth factor beta (TGF-beta) in the pathogenesis of experimental MPGN. We have, therefore, examined the influence of TGF- beta1 on mesangial cell fibronectin synthesis. Considering, first, lev els of mRNA, TGF-beta1 increased steady-state fibronectin RNA in cultu red mesangial cells by 1.9 times 24 hr after treatment of cycling mesa ngial cells and by 11.8 times in growth-arrested cells. There was, how ever, no alteration in fibronectin pre-mRNA splicing in either the EII IA or IIICS regions. Fibronectin protein concentrations in cell cultur e supernatants, determined by immunoprecipitation of supernatants from cells labeled with [S-35]methionine and by ELISA, were not increased by treatment with TGF-beta1. Western blots and immunoprecipitation of metabolically labeled cells showed that fibronectin was increased, how ever, in the deoxycholate-insoluble extracellular matrix (ECM) of cell s stimulated with TGF-beta1. TGF-beta1 altered the physicochemical pro perties of fibronectin in ECM and supernatant such that the isoelectri c point of fibronectin, determined from Western blots of 2D SDS-PAGE g els, was reduced so that both became more acidic. These studies demons trate, therefore, that in addition to increasing its synthesis, TGF-be ta1 increases incorporation of fibronectin into the ECM. Because fibro nectin possesses binding sites for other ECM proteins, greater incorpo ration of fibronectin following TGF-beta1 treatment may be an importan t pathogenetic mechanism in mesangial sclerosis. Moreover, the altered charge of fibronectin may increase localization of serum immunoglobul ins to the mesangium. (C) 1993 Academic Press, Inc.