Induction of IL-6 by anti-DNA antibodies in proximal tubular epithelial cells varies with the activity of lupus nephritis

Abstract

BACKGROUND: Anti-DNA antibodies and IL-6 induction are implicated in the pathogenesis of lupus nephritis. While the immunopathogenesis of glomerular lesions in lupus nephritis has been extensively studied, limited data is available on the pathogenesis of tubulo-interstitial lesions despite the fact that the rate of renal function deterioration correlates best with the degree of renal tubulo-interstitial inflammation and fibrosis. Proximal tubular epithelial cells (PTEC) constitute the predominant cell type within the tubulo-interstitium. Although previously considered to be mainly involved in the transport of fluid and electrolytes, there is increasing evidence that PTEC play an important role in the immunopathogenesis of various renal parenchymal diseases. Thus, in this study we investigated the effect of human anti-DNA antibodies on IL-6 synthesis and cellular functions in PTEC. METHODS: 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-1β, and TNF-α were determined in culture supernatants using commercial ELISAs according to the manufacturer’s instructions. RESULTS: Incubation of PTEC with purified anti-DNA antibodies (10 mg IgG/ml) for time periods up to 24h induced IL-6 secretion in a time dependent manner (0.5 ± 0.1, 8.0 ± 3.1, and 12.4 ± 4.7ng/µg cellular protein for control IgG, and anti-DNA antibodies derived from inactive and active patients respectively after 24h stimulation, P < 0.01 compared to control), which was dependent on de novo synthesis of mRNA and protein. Induction of IL-6 secretion was accompanied by altered cell morphology, proliferation (P < 0.05), viability (P < 0.05) and increased secretion of IL-1β (P < 0.05) and TNF-α (P < 0.05). The ability of anti-DNA to induce IL-6 secretion 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). The mechanism that mediated anti-DNA antibody induction of IL-6 secretion in PTEC was assessed using neutralizing antibodies to IL-6 and TNF-α, and IL-1 receptor antagonist. During active disease, the induction of IL-6 by anti-DNA antibodies was primarily mediated via TNF-α. In contrast, the induction of IL-6 by anti-DNA antibodies obtained during clinical remission was a direct effect of the anti-DNA antibodies, independent of TNF-a. CONCLUSIONS: Our data demonstrates 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.