Critical comparative analyses of anti–α‐actinin and glomerulus‐bound antibodies in human and murine lupus nephritis

Abstract

Objective. Although anti-double-stranded DNA (anti-dsDNA) antibodies are important in lupus nephritis, the question regarding which glomerular structures (␣-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 vivobound 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 ؋ NZW)F 1 mice were analyzed for activity against proteins identified by monoclonal nephritogenic antibodies, and against ␣-actinin, dsDNA, nucleosomes, histone H1, heparan sulfate, DNase I, and type IV collagen. Immunoelectron microscopy was used to determine the glomerular localization of ␣-actinin and in vivo-bound autoantibodies in nephritic (NZB ؋ NZW)F 1 mouse kidneys.Results. Anti-␣-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 ؋ NZW)F 1 mouse kidneys revealed that antibodies eluted from kidneys, but not anti-␣-actinin antibodies, bound to distinct nephritis-associated electrondense structures linked to glomerular basement membranes. Conclusion.Cross-reactive anti-dsDNA/antihistone H1 antibodies, but not anti-␣-actinin antibodies, are central among those deposited in nephritic glomeruli.

Type I interferon signaling is involved in the spontaneous development of lupus-like disease in B6.Nba2 and (B6.Nba2 × NZW)F1 mice

“…[20][21][22][23][24] These antibodies were found to bind mainly nuclear antigens such as chromatin, histones and dsDNA, and/or components of the glomerular basement membrane. In particular, anti-dsDNA autoantibodies were found to be pathogenic and it has been shown that some anti-dsDNA autoantibodies can by themselves alter the gene expression profile of mesangial cell lines, turning on the expression of inflammatory molecules.…”

Heparin exerts a dual effect on murine lupus nephritis by enhancing enzymatic chromatin degradation and preventing chromatin binding in glomerular membranes

“…Serum antibodies against dsDNA were detected and titrated by ELISA exactly as described previously (15,36). An anti-dsDNA mAb was included in each ELISA for intraassay validation (15,36).…”

“…Preparation of kidney tissue and immune electron microscopy were performed as described previously (15). Sections were examined at 10,000-40,000ϫ magnification using a JEM-1010 transmission electron microscope (Jeol).…”

“…These immune complexes were observed as electron-dense structures in GBMs and the mesangial matrix, and bound IgG was confined to electron-dense structures in both murine (12,14) and human (13) lupus nephritis. In vivo bound IgG was not observed in regular glomerular structures (15), although anti-DNA antibodies have been shown to cross-react in vitro with other glomerular antigens (16)(17)(18)(19). It is unclear whether antibodies form complexes with chromatin in circulation or in situ in glomeruli.…”

Glomerular Autoimmune Multicomponents of Human Lupus Nephritis In Vivo

“…[6][7][8][9][10][11][12] They consider a pre-eminent role of circulating anti-double stranded DNA (dsDNA) antibodies that they can interact, for mimicry, with glomerular antigens expressed at the cell surface of podocytes and mesangial cells 7,[13][14][15] or in the glomerular basement membrane. 12,[16][17][18][19][20] Chromatin and nucleosomes deriving from ineffective fragmentation may interact with negatively charged constituents of the basement membrane 7,16 and become the planted antigen for anti-dsDNA (and autoantibodies of other specificities).…”