Systemic lupus erythematosus is characterized by faulty B-cell tolerance in the germinal centers

Abstract

Studies of human B-cell tolerance have been hampered by the low frequency of antigen-specific autoreactive B cells in healthy subjects as well as by lack of access to secondary lymphoid tissue in autoimmune patients. We have resolved the first problem by studying the fate of a population of human B cells endowed with intrinsic autoreactivity owing to the expression of antibodies encoded by the VH4.34 gene segment (VH4.34 B-cells and VH4.34 antibodies, respectively) [1]. These pathogenic autoantibodies are very abundant in systemic lupus erythematosus (SLE), where they represent 10–40% of serum IgG, correlate with disease activity, renal and central nervous system involvement, and contribute substantially to the anti-dsDNA and anti-lymphocyte antibody repertoire [2-5]. Of note, increased production of VH4.34 antibodies is highly specific for SLE and has not been observed in other autoimmune diseases. Taking advantage of the abundance of VH4.34 B cells in healthy individuals and of the ability of the anti-idiotypic monoclonal antibody 9G4 to specifically recognize VH4.34 B cells, we previously proposed that healthy VH4.34 B cells are negatively selected during the germinal center (GC) reaction [1]. Herein, we have expanded our initial studies to further dissect this observation using tonsil biopsies obtained from patients with SLE (n = 8) and rheumatoid arthritis (n = 3) as a source of secondary lymphoid tissue. Using both flow cytometry, intracellular calcium analysis and immunocytochemistry, our studies demonstrate that in normal subjects 9G4+ autoreactive VH4.34 B cells display an anergic phenotype and undergo strong counter selection in the early phase of the GC reaction, as indicated by their decline in the transition from the GC founder stage to the centroblast stage and by their absence from proliferative GCs. In contrast, SLE 9G4+ B cells participate in approximately 20% of all productive GC reactions, as substantiated by flow cytometry and histological analyses and as reflected by a 10-fold to 20-fold expansion into the post-GC IgG memory and plasma cell compartments. Remarkably, in patients with RA, 9G4+ B cells behave like healthy 9G4+ B cells and are also strongly counter-selected in the GC. Overall, our results strongly indicate that a critical checkpoint in the maintenance of B-cell tolerance depends on effective negative selection of autoreactive B cells in the GCs and that this checkpoint is specifically defective in SLE. Detailed comparisons of the cellular, biochemical and genetic profiles of the cells targeted at this checkpoint (GC founders) between healthy control and SLE patients should shed considerable light into the mechanisms responsible for disease development, thereby suggesting new therapeutic approaches.