9G4+ autoantibodies are an important source of apoptotic cell reactivity associated with high levels of disease activity in systemic lupus erythematosus.

During homeostasis billions of cells die through apoptosis daily and as a potential source of autoantigens these cells must be efficiently cleared in an immunologically silent fashion to prevent pathological autoimmune reactions (1). Defective clearance of apoptotic cells has been demonstrated both in vitro and in vivo in systemic lupus erythematosus (SLE), an autoimmune disease characterized by the generation of antibodies against multiple nuclear antigens (2-4). This is demonstrated by the high incidence of SLE in patients with genetic deficiency of C1q, a complement component involved in the opsonization and clearance of apoptotic cells. Like SLE patients, transgenic mice deficient in complement C1q develop autoantibodies or a lupus-like disease (5) and mice deficient in tyrosine receptor kinases necessary for apoptotic cell phagocytosis also develop severe autoimmunity (6). Furthermore, immunization of mice with apoptotic cells results in autoantibody production and autoimmune disease (7). Anti-apoptotic cell antibodies (AACA) have been previously recognized in SLE serum and detected in lupus nephritis kidneys bound to glomerular apoptotic nucleosomes (8). SLE AACA can exercise pathogenic functions by promoting phagocytosis of apoptotic cells (9, 10), resulting in the engagement of intracellular TLR receptors, which leads to the release of type I IFN and other pro-inflammatory cytokines (11-13). While studies of IgG AACA in SLE have been focused on their influence on phagocytosis (3, 10, 14), systematic studies of their prevalence and significance are lacking. Similarly, the nature of IgG AACA and the processes leading to their generation and selection in SLE remain unclear. Of note, IgM AACA have been associated with protection against renal disease in SLE (15). In this study, we systematically investigated the presence of IgG and IgM antibody binding to apoptotic cells in SLE patients using a flow cytometry-based assay and determined the contribution of antibodies bearing the 9G4 idiotype (9G4+) to this autoreactivity. The study of intrinsically autoreactive 9G4+ antibodies encoded by the VH4-34 gene is informative in SLE as these antibodies represent 10-40% of all serum IgG (16) due to defective germinal center censoring of VH4-34 B cells (17). The relevance of understanding the antigenic forces underpinning the expansion of 9G4+ antibodies in SLE is further illustrated by their high SLE specificity and their correlation with disease activity and specific clinical manifestations including lupus nephritis (18-21). Our results indicate that the presence of 9G4+ AACA is common in SLE and show that patients with elevated 9G4+ AACA are more likely to have active disease. These findings demonstrate that reactivity with apoptotic cell antigens contributes significantly to the expansion of a major autoreactive B cell population that is specifically expanded in SLE and provide the experimental basis for a better understanding of the antigenic forces involved in the pathogenesis of this disease.