Nucleosomes and C1q bound to glomerular endothelial cells serve as targets for autoantibodies and determine complement activation.

Various studies indicate a role for both anti-nucleosome and anti-C1q autoantibodies in glomerulonephritis in patients with systemic lupus erythematosus. However, a causal relationship between these autoantibodies and the development of lupus nephritis has not been fully established. Since injury of the endothelium is a major target in lupus nephritis we assessed the interaction of C1q and nucleosomes with glomerular endothelial cells in vitro in the presence or absence of autoantibodies against these antigens. We demonstrate a direct and dose-dependent binding of both nucleosomes and C1q to immortalized human glomerular endothelial cells (GEnC) in vitro, which in part is mediated by cell surface heparan sulfate. We demonstrate that nucleosomes and C1q serve as targets for monoclonal and polyclonal antibodies as well as for anti-nuclear autoantibodies from patients with systemic lupus erythematosus. An additive effect of anti-C1q autoantibodies on anti-nucleosome mediated complement activation was observed. Furthermore, we showed that the activation of complement on glomerular endothelial cells is mediated by the classical pathway since the deposition of C3 on GEnC is abrogated by MgEGTA and does not occur in C1q-depleted serum. Taken together, our studies demonstrate a direct binding of both nucleosomes and C1q to glomerular endothelial cells in vitro. The subsequent binding of autoantibodies against nucleosomes in patients with systemic lupus erythematosus is potentially pathogenic and autoantibodies against C1q seem to have an additional effect.

The inhibiting Fc receptor for IgG, FcγRIIB, is a modifier of autoimmune susceptibility.

FcγRIIB-deficient mice generated in 129 background (FcγRIIB(129)(-/-)) if back-crossed into C57BL/6 background exhibit a hyperactive phenotype and develop lethal lupus. Both in mice and humans, the Fcγr2b gene is located within a genomic interval on chromosome 1 associated with lupus susceptibility. In mice, the 129-derived haplotype of this interval, named Sle16, causes loss of self-tolerance in the context of the B6 genome, hampering the analysis of the specific contribution of FcγRIIB deficiency to the development of lupus in FcγRIIB(129)(-/-) mice. Moreover, in humans genetic linkage studies revealed contradictory results regarding the association of "loss of function" mutations in the Fcγr2b gene and susceptibility to systemic lupus erythematosis. In this study, we demonstrate that FcγRIIB(-/-) mice generated by gene targeting in B6-derived ES cells (FcγRIIB(B6)(-/-)), lacking the 129-derived flanking Sle16 region, exhibit a hyperactive phenotype but fail to develop lupus indicating that in FcγRIIB(129)(-/-) mice, not FcγRIIB deficiency but epistatic interactions between the C57BL/6 genome and the 129-derived Fcγr2b flanking region cause loss of tolerance. The contribution to the development of autoimmune disease by the resulting autoreactive B cells is amplified by the absence of FcγRIIB, culminating in lethal lupus. In the presence of the Yaa lupus-susceptibility locus, FcγRIIB(B6)(-/-) mice do develop lethal lupus, confirming that FcγRIIB deficiency only amplifies spontaneous autoimmunity determined by other loci.

Constitutive activation of Bruton's tyrosine kinase induces the formation of autoreactive IgM plasma cells.

B-cell receptor (BCR)-mediated signals provide the basis for B-cell differentiation in the BM and subsequently into follicular, marginal zone, or B-1 B-cell subsets. We have previously shown that B-cell-specific expression of the constitutive active E41K mutant of the BCR-associated molecule Bruton's tyrosine kinase (Btk) leads to an almost complete deletion of immature B cells in the BM. Here, we report that low-level expression of the E41K or E41K-Y223F Btk mutants was associated with reduced follicular B-cell numbers and significantly increased proportions of B-1 cells in the spleen. Crosses with 3-83 mu delta and VH81X BCR Tg mice showed that constitutive active Btk expression did not change follicular, marginal zone, or B-1 B-cell fate choice, but resulted in selective expansion or survival of B-1 cells. Residual B cells were hyperresponsive and manifested sustained Ca(2+) mobilization. They were spontaneously driven into germinal center-independent plasma cell differentiation, as evidenced by increased numbers of IgM(+) plasma cells in spleen and BM and significantly elevated serum IgM. Because anti-nucleosome autoantibodies and glomerular IgM deposition were present, we conclude that constitutive Btk activation causes defective B-cell tolerance, emphasizing that Btk signals are essential for appropriate regulation of B-cell activation.

The complement system facilitates clearance of Enterococcus faecium during murine peritonitis.

BACKGROUND: Infections with multidrug-resistant enterococci are a growing problem worldwide. Little is known about the host defense against enterococcal diseases. In vitro studies have demonstrated an important role played by complement proteins in neutrophil-mediated phagocytosis. In this study, we investigated the importance of complement in an in vivo model of Enterococcus faecium peritonitis. METHODS: Peripheral neutrophils and peritoneal macrophages were incubated with E. faecium that had been preincubated with decomplemented or normal plasma, and phagocytosis and killing were examined. E. faecium peritonitis was induced in C57BL/6 mice rendered complement deficient by intraperitoneal injection with cobra venom factor (CVF) and in complement 3 (C3) knockout mice. The course of the infection was compared with that in saline control and wild-type mice, respectively, at several time points up to 48 h after infection. RESULTS: Opsonization by complement enhanced phagocytosis by neutrophils and macrophages. CVF-treated and C3 knockout mice were severely hampered in clearing E. faecium from all organs and tissues under study (peritoneal fluid, blood, lungs, and liver). Higher peritoneal cytokine and chemokine levels were measured in decomplemented mice, whereas no differences in systemic or peritoneal cell kinetics were detected. CONCLUSION: Complement deficiency severely hampers the clearance of E. faecium peritonitis and subsequent systemic infection.

Both complement and IgG fc receptors are required for development of attenuated antiglomerular basement membrane nephritis in mice.

To elucidate the mechanisms of glomerulonephritis, including Goodpasture's syndrome, mouse models are used that use heterologous Abs against the glomerular basement membrane (GBM) with or without preimmunization with foreign IgG from the same species. These studies have revealed the requirement of either FcgammaR or complement, depending on the experimental model used. In this study, we provide evidence that both FcgammaR and complement are obligatory for a full-blown inflammation in a novel attenuated passive model of anti-GBM disease. We demonstrate that administration of subnephritogenic doses of rabbit anti-GBM Abs followed by a fixed dose of mouse mAbs to rabbit IgG, allowing timing and dosing for the induction of glomerulonephritis, resulted in reproducible complement activation via the classical pathway of complement and albuminuria in wild-type mice. Because albuminuria was absent in FcR-gamma-chain(-/-) mice and reduced in C3(-/-) mice, a role for both FcgammaR and complement is postulated. Because C1q(-/-) and C4(-/-) mice lacking a functional classical and lectin pathway did develop albuminuria, we suggest involvement of the alternative pathway of complement. Anti-GBM glomerulonephritis occurs acutely following the administration of mouse anti-rabbit IgG, and proceeds in a chronic fashion dependent on both FcgammaR and complement. This novel attenuated model allows elucidating the relative contribution of different mediator systems of the immune system to the development of renal injury, and also provides a platform for the assessment of different treatment protocols and evaluation of drugs that ultimately may be beneficial for the treatment of anti-GBM mediated glomerulonephritides.

Cutting edge: TNFR-shedding by CD4+CD25+ regulatory T cells inhibits the induction of inflammatory mediators.

CD4+CD25+ regulatory T (Treg) cells play an essential role in maintaining tolerance to self and nonself. In several models of T cell-mediated (auto) immunity, Treg cells exert protective effects by the inhibition of pathogenic T cell responses. In addition, Treg cells can modulate T cell-independent inflammation. We now show that CD4+CD25+ Treg cells are able to shed large amounts of TNFRII. This is paralleled by their ability to inhibit the action of TNF-alpha both in vitro and in vivo. In vivo, Treg cells suppressed IL-6 production in response to LPS injection in mice. In contrast, Treg cells from TNFRII-deficient mice were unable to do so despite their unhampered capacity to suppress T cell proliferation in a conventional in vitro suppression assay. Thus, shedding of TNFRII represents a novel mechanism by which Treg cells can inhibit the action of TNF, a pivotal cytokine driving inflammation.

The clearance of apoptotic cells by complement.

The complement system is implicated among others in the clearance of apoptotic cells, and deficiency for different components of the complement system is associated with aberrant handling of apoptotic material. As a result of this, it is thought that the complement system is involved in the pathogenesis of autoimmune diseases such as systemic lupus erythematosus (SLE) which affects multiple organs. In this review, the authors highlight the available data on complement and apoptosis, as well as the role of complement in severe autoimmune diseases like SLE.

Pathogenic role of anti-C1q autoantibodies in the development of lupus nephritis--a hypothesis.

A substantial proportion of patients with systemic lupus erythematosus (SLE) develop renal inflammatory disease, so-called lupus nephritis (LN). LN is a severe complication of SLE which is strongly associated with the presence of autoantibodies against C1q, the first component of the complement system, and other self-antigens (i.e. against DNA and nucleosomes) as well. In this review, the authors focus on anti-C1q autoantibodies and interpret the available data in order to explain how LN may develop and how anti-C1q autoantibodies contribute to its pathogenesis.

Effective treatment of collagen-induced arthritis by adoptive transfer of CD25+ regulatory T cells.

OBJECTIVE: Regulatory T cells play an important role in the prevention of autoimmunity and have been shown to be effective in the treatment of experimental colitis, a T cell-mediated and organ-specific disease. We previously demonstrated that intrinsic CD25+ regulatory T cells modulate the severity of collagen-induced arthritis (CIA), which, in contrast to colitis, is a systemic antibody-mediated disease and an accepted model of rheumatoid arthritis. We undertook this study to determine whether regulatory T cells have the potential to be used therapeutically in arthritis. METHODS: We transferred CD4+,CD25+ T cells into mice exhibiting arthritis symptoms, both immunocompetent mice and mice subjected to lethal irradiation and rescued with syngeneic bone marrow transplantation. RESULTS: A single transfer of regulatory T cells markedly slowed disease progression, which could not be attributed to losses of systemic type II collagen-specific T and B cell responses, since these remained unchanged after adoptive transfer. However, regulatory T cells could be found in the inflamed synovium soon after transfer, indicating that regulation may occur locally in the joint. CONCLUSION: Our data indicate that CD25+ regulatory T cells can be used for the treatment of systemic, antibody-mediated autoimmune diseases, such as CIA.

Control of systemic B cell-mediated autoimmune disease by nonmyeloablative conditioning and major histocompatibility complex-mismatched allogeneic bone marrow transplantation.

Systemic autoimmune disease (AID) can be controlled with conventional therapies in most patients. However, relapses are common, leading to progressive disability and premature death. Nonmyeloablative conditioning and allogeneic bone marrow transplantation (BMT) could be an effective treatment for severe AID, because of mild toxicity of the conditioning and the potential benefits of donor chimerism. We examined the effects of this treatment in experimental autoimmune arthritis. Our results demonstrate the induction of complete donor chimerism and significant suppression of disease activity. No clinical graft-versus-host disease (GVHD) was observed. The beneficial effects were most likely caused by the elimination of plasma cells producing pathogenic autoantibodies, because these antibodies disappeared rapidly after BMT. Although this type of treatment was effective in organ-specific T-cell-mediated AID, the present study provides convincing evidence that nonmyeloablative conditioning and allogeneic BMT can effectively treat severe B-cell-mediated AID with a systemic inflammatory component.