Anti-DNA antibodies from patients with lupus nephritis induce IL-6 expression in renal tubular epithelial cells

Abstract

There is limited data on the pathogenesis of tubulo-interstitial lesions in lupus nephritis. In this study we investigated the effect of human anti-DNA antibodies on IL-6 synthesis and cellular functions in proximal renal tubular epithelial cells (PTEC). PTEC were isolated from renal cortical tissue by differential sieving and digestion with collagenase. Paired polyclonal anti-DNA antibodies were isolated from 15 lupus nephritis patients during active disease and remission using DNA-cellulose affinity chromatography. Cell morphology was assessed by phase contrast microscopy, and cell proliferation and viability by MTT assay and lactate dehydrogenase release respectively. IL-6, IL-1beta, and TNF-alpha were determined in culture supernatants using commercial ELISAs. Incubation of PTEC with anti-DNA antibodies (10 micrograms IgG/ml) for time periods up to 24 h induced IL-6 secretion in a time-dependent manner (0.5 +/- 0.1, 8.0 +/- 3.1, and 12.4 +/- 4.7 ng/microgram cellular protein for control IgG, anti-DNA antibodies during inactive disease, and anti-DNA antibodies during active disease respectively, p 0.01 compared to control), which was dependent on de novo mRNA and protein synthesis. Induction of IL-6 secretion was accompanied by alterations in cell morphology, proliferation (p 0.05), and viability (p 0.05), as well as increased secretion of IL-1beta (p 0.05) and TNF-alpha (p 0.05). IL-6 induction by anti-DNA correlated with circulating anti-DNA antibody levels and their binding to cultured PTEC (Spearman r = 0.561 and 0.576 respectively, p 0.001 for both). Data from inhibition experiments using neutralizing antibodies to IL-6 and TNF-alpha, and IL-1 receptor antagonist showed that during active disease IL-6, IL-1beta, and TNF-alpha interacted synergistically with anti-DNA antibodies to induce PTEC IL-6 secretion, but TNF-alpha was not involved in such induction during remission. Our data demonstrate that anti-DNA antibodies from patients with lupus nephritis can induce IL-6 production in PTEC, through distinct mechanisms that vary according to disease activity.