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 mesangial proliferative glomerulonephritis (MPGN). Recent evidence has implicated 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, levels of mRNA, TGF-beta1 increased steady-state fibronectin RNA in cultured mesangial cells by 1.9 times 24 hr after treatment of cycling mesangial cells and by 11.8 times in growth-arrested cells. There was, however, no alteration in fibronectin pre-mRNA splicing in either the EIIIA or IIICS regions. Fibronectin protein concentrations in cell culture 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, however, in the deoxycholate-insoluble extracellular matrix (ECM) of cells stimulated with TGF-beta1. TGF-beta1 altered the physicochemical properties of fibronectin in ECM and supernatant such that the isoelectric point of fibronectin, determined from Western blots of 2D SDS-PAGE gels, was reduced so that both became more acidic. These studies demonstrate, therefore, that in addition to increasing its synthesis, TGF-beta1 increases incorporation of fibronectin into the ECM. Because fibronectin possesses binding sites for other ECM proteins, greater incorporation of fibronectin following TGF-beta1 treatment may be an important pathogenetic mechanism in mesangial sclerosis. Moreover, the altered charge of fibronectin may increase localization of serum immunoglobulins to the mesangium. (C) 1993 Academic Press, Inc.