Modulation of hyaluronan synthesis in human mesangial cells after stimulation with Anti-DNA antibodies from patients with lupus nephritis

Abstract

Prominent features of lupus nephritis include elevated levels of circulating anti-DNA antibodies, induction of pro-inflammatory cytokines and modulation of the extracellular matrix (ECM). Hyaluronan (HA) is a major ECM component that has a profound effect on wound healing, and inflammatory responses. In this study, we investigated the effects of anti-DNA antibodies on HA synthesis in human mesangial cells (HMC). Anti-DNA antibodies were purified serially from 10 patients during active and inactive disease using protein A-Sepharose and DNA-cellulose affinity chromatography. HMC were stimulated for 24h with either control IgG or anti-DNA antibodies at a final IgG concentration of 10μg/ml. Using RT-PCR a significant increase in mRNA for HAS II was observed in HMC exposed to active anti-DNA antibodies compared to control human IgG (P<0.05). Inactive anti-DNA antibodies also induced mRNA for HAS II but to a lesser extent. No change in gene expression for HAS III was observed with either control IgG or active/inactive antibody stimulation. HAS I was not expressed by HMC. De novo synthesis of HA was investigated by radiolabelling with [3H]-glucosamine (25μCi/ml). Gel filtration chromatography revealed qualitative but not quantitative changes in [3H]-labelled HA isolated from HMC under experimental conditions. Whilst no change was observed in the MW of HA after stimulation with inactive antibodies compared to control IgG, stimulation of HMC with active antibodies resulted in the synthesis of HA with a significantly lower MW. Changes in HA synthesis during active disease was associated with an increase in IL-1β secretion. Expression of HA in renal biopsies obtained from lupus patients during active disease showed increased HA deposition within the glomerulus compared to controls. Our data suggest that anti-DNA antibodies modulate HA synthesis during active lupus nephritis. Given that low MW HA plays a pivotal role in inflammatory processes and leukocyte recruitment during tissue injury, it is possible that the generation of low MW HA by HMC may initiate the induction of pro-inflammatory cytokines and immune response during pathogenesis of disease.