Human anti-dsDNA antibodies induce amplifi cation loop of Transforming Growth Factor-β1, fibronectin and collagen type I in proximal renal tubular epithelial cells

Abstract

BACKGROUND: Nephritis affects up to 60% of patients with systemic lupus erythematosus and is characterized by the production of anti-dsDNA antibodies and immune-mediated renal injury. Severity of tubulointerstitial fibrosis is a strong predictor of reduced renal survival. Fibronectin (FN) is amongst the matrix components showing early deposition during the process of tubulointerstitial fibrosis. This study investigates the mechanisms through which FN is induced in renal proximal tubular epithelial cells (PTEC) following stimulation with human anti-dsDNA antibodies, and its role in tubulointerstitial fibrosis of lupus nephritis. METHODS: Confluent growth-arrested PTEC were incubated with serum free medium (SFM), control IgG or human polyclonal anti-dsDNA antibodies (10µg/ml) for 24h in the presence or absence of Gö6976 or TGF-neutralizing antibody, and the synthesis of FN assessed. In separate studies, PTEC were stimulated with soluble FN to determine its effect on fibrotic processes. RESULTS: Control IgG had no effect on FN synthesis compared to SFM. Anti-dsDNA antibodies significantly increased cell-associated and soluble FN compared to SFM and control IgG (6.4- and 5.2-fold respectively for cell-associated FN; 1.85- and 1.62-fold respectively for soluble FN, P<0.01 for all). This was accompanied by significantly increased TGF-β1 secretion (P<0.05), and increased PKC-α (P<0.05) and PKC-βII (P<0.01) but not PKC-βI phosphorylation. Pre-treatment of PTEC with Gö6976 (20µM) and TGF-neutralizing antibody (100g/ml) significantly suppressed anti-dsDNA antibody-induced FN (P<0.01 for both cell-associated and soluble FN). Incubation of PTEC with soluble FN significantly increased TGF-β1 secretion and collagen type I synthesis in a dose-dependent manner (P<0.05 and P<0.01 respectively for 10µg/ml soluble FN). CONCLUSIONS: These results suggest that anti-dsDNA antibodies induce an amplification loop comprising TGF-β1, FN and collagen type I in PTEC through PKC activation.