ps1 17 lupus nephritis severely reduced urinary dnase i levels reflect loss of renal dnase i disease progression and may reduce the need for renal biopsies

Loss of renal DNase I leads to progression of lupus nephritis. Therefore, we determined if loss of renal DNase I reflects a concurrent loss of urinary DNase I, and whether absence of urinary DNase I predicts disease progression, which thus may reduce the need for renal biopsies. Here, mouse renal DNase I mRNA was determined by qPCR, whereas mouse and human DNase I protein and DNase I endonuclease activity levels were determined by Western blots, and gel and radial zymography assays, respectively, during different stages of the murine and human forms of the disease. Cellular localization of DNase I was analysed by immunohistochemistry, immunofluorescence, confocal microscopy and immune electron microscopy. We further compared DNase I levels in human native and transplanted kidneys to determine if the disease depended on autologous renal genes, or whether the nephritic process proceeded also in transplanted kidneys. We also analysed if DNase I levels in urine samples reflected expression levels in the kidneys, and if the mouse data were translatable to humans. The data indicates that silencing of the renal DNase I gene expression level relates to serious progression of lupus nephritis in murine, human native, and transplanted kidneys. Notably, silencing of renal DNase I correlates with loss of DNase I protein and endonuclease activity in the urine samples. Thus, urinary DNase I levels reflects the renal DNase I expression and activity levels, and may therefore be used as a marker of lupus nephritis disease progression and reduce the need for renal biopsies

Anti-dsDNA Antibodies Promote Initiation, and Acquired Loss of Renal Dnase1 Promotes Progression of Lupus Nephritis in Autoimmune (NZBxNZW)F1 Mice

BACKGROUND:Lupus nephritis is characterized by deposition of chromatin fragment-IgG complexes in the mesangial matrix and glomerular basement membranes (GBM). The latter defines end-stage disease. METHODOLOGY/PRINCIPALS: In the present study we determined the impact of antibodies to dsDNA, renal Dnase1 and matrix metalloprotease (MMP) mRNA levels and enzyme activities on early and late events in murine lupus nephritis. The major focus was to analyse if these factors were interrelated, and if changes in their expression explain basic processes accounting for lupus nephritis. FINDINGS:Early phases of nephritis were associated with chromatin-IgG complex deposition in the mesangial matrix. A striking observation was that this event correlated with appearance of anti-dsDNA antibodies and mild or clinically silent nephritis. These events preceded down-regulation of renal Dnase1. Later, renal Dnase1 mRNA level and enzyme activity were reduced, while MMP2 mRNA level and enzyme activity increased. Reduced levels of renal Dnase1 were associated in time with deficient fragmentation of chromatin from dead cells. Large fragments were retained and accumulated in GBM. Also, since chromatin fragments are prone to stimulate Toll-like receptors in e.g. dendritic cells, this may in fact explain increased expression of MMPs. SIGNIFICANCE:These scenarios may explain the basis for deposition of chromatin-IgG complexes in glomeruli in early and late stages of nephritis, loss of glomerular integrity and finally renal failure

Critical comparative analyses of anti-alpha-actinin and glomerulus-bound antibodies in human and murine lupus nephritis

Objective. Although anti-double-stranded DNA (anti-dsDNA) antibodies are important in lupus nephritis, the question regarding which glomerular structures (a-actinin, nucleosomes, or others) are recognized by nephritogenic anti-dsDNA antibodies is still controversial. In this study, we determined which glomerular structures are recognized by monoclonal and in vivo-bound nephritogenic antibodies. Methods. Western blotting was used to analyze the ability of nephritogenic anti-dsDNA antibodies to recognize glomerular and nucleosomal structures. Sera from patients with lupus nephritis, sera from random antinuclear antibody-positive patients, and paired antibodies from sera and kidney eluates from nephritic (NZB X NZW)F-1 mice were analyzed for activity against proteins identified by monoclonal nephritogenic antibodies, and against a-actinin, dsDNA, nucleosomes, histone H1, heparan sulfate, DNase I, and type IV collagen. Immunoelectron microscopy was used to determine the glomerular localization of a-actinin and in vivo-bound autoantibodies in nephritic (NZB X NZW)F-1 mouse kidneys. Results. Anti-alpha-actinin antibodies were observed in human and murine lupus nephritis sera and in sera from patients without systemic lupus erythematosus and were not detected in kidney eluates from nephritic mice. Antibodies to dsDNA and histone H1 were detected in all eluates. Western blot analyses revealed that nephritogenic anti-dsDNA antibodies recognized a 32-kd band, identified as histone H1. Competitive enzyme-linked immunosorbent assay demonstrated that nephritogenic monoclonal antibodies, and dominant antibodies eluted from nephritic kidneys, cross-reacted with dsDNA and H1. This cross-reactive anti-H1 specificity was largely absent in sera from those mice. Immunoelectron microscopic analysis of nephritic (NZB X NZW)F-1 mouse kidneys revealed that antibodies eluted from kidneys, but not anti-alpha-actinin antibodies, bound to distinct nephritis-associated electron-dense structures linked to glomerular basement membranes. Conclusion. Cross-reactive anti-dsDNA/anti-histone H1 antibodies, but not anti-alpha-actinin antibodies, are central among those deposited in nephritic glomeruli

Autoantibodies in lupus: culprits or passive bystanders?

Several autoantibodies are culprits in the pathogenesis of organ damage in systemic lupus erythematosus, by means of established or postulated mechanisms, whereby inducing a perturbation of cell structure and function, with consequent tissue-organ impairment. Common autoantibody-mediated mechanisms of damage include cell surface binding with or without cytolysis, immune complex-mediated damage, penetration into living cells, binding to cross-reactive extracellular molecules. Experimental data from both murine models and humans have recently clarified the key role of autoantibodies in severe organ involvements, including nephritis, neuropsychiatric (NP) dysfunction, and cerebrovascular disease (CVD). In lupus nephritis early and late phases are distinguishable and mediated by different processes in which anti-chromatin antibodies are both inducing and perpetuating agents, by immune-complex formation and massive deposition in mesangial matrix at first, and in glomerular basement membrane at end-stage. Also NP abnormalities occur very early, much earlier than other systemic manifestations, and exacerbate with the increase in autoantibody titers. Among the autoantibodies mainly implicated in neurolupus, anti-β2 glycoprotein I (β2GPI) antibodies are preferentially involved in focal NP events which are a consequence of non-inflammatory microangiopathy; otherwise, anti-ribosomal P protein antibodies and N-methyl-d-aspartate receptor (NMDAR) antibodies cause diffuse NP events through a direct cytotoxic effect on neuronal cells at specific brain zones