Antigen-specific B cells are required as APCs and autoantibody-producing cells for induction of severe autoimmune arthritis

B cells play an important role in rheumatoid arthritis, but whether they are required as autoantibody-producing cells as well as APCs has not been determined. We assessed B cell autoantibody and APC functions in a murine model of autoimmune arthritis, proteoglycan (PG)-induced arthritis, using both B cell-deficient mice and Ig-deficient mice (mIgM) mice that express an H chain transgene encoding for membrane-bound, but not secreted, IgM. The IgH transgene, when paired with endogenous lambda L chain, recognizes the hapten 4-hydroxy-3-nitro-phenyl acetyl and is expressed on 1-4% of B cells. B cell-deficient and mIgM mice do not develop arthritis after immunization with PG. In adoptive transfer of PG-induced arthritis into SCID mice, T cells from mIgM mice immunized with PG were unable to transfer disease even when B cells from PG-immunized wild-type mice were provided, suggesting that the T cells were not adequately primed and that Ag-specific B cells may be required. In fact, when PG was directly targeted to the B cell Ig receptor through a conjugate of 4-hydroxy-3-nitrophenyl acetyl-PG, T cells in mIgM mice were activated and competent to transfer arthritis. Such T cells caused mild arthritis in the absence of autoantibody, demonstrating a direct pathogenic role for T cells activated by Ag-specific B cells. Transfer of arthritic serum alone induced only mild and transient arthritis. However, both autoreactive T cells and autoantibody are required to cause severe arthritis, indicating that both B cell-mediated effector pathways contribute synergistically to autoimmune disease.     

Amplification of autoimmune disease by infection

Reports of infection with certain chronic persistent microbes (herpesviruses or Chlamydiae) in human autoimmune diseases are consistent with the hypothesis that these microbes are reactivated in the setting of immunodeficiency and often target the site of autoimmune inflammation. New experimental animal models demonstrate the principle. A herpesvirus or Chlamydia species can be used to infect mice with induced transient autoimmune diseases. This results in increased disease severity and even relapse. The evidence suggests that the organisms are specifically imported to the inflammatory sites and cause further tissue destruction, especially when the host is immunosuppressed. We review the evidence for the amplification of autoimmune inflammatory disease by microbial infection, which may be a general mechanism applicable to many human diseases. We suggest that patients with autoimmune disorders receiving immunosuppressing drugs should benefit from preventive antiviral therapy.     

Development of proteoglycan-induced arthritis is independent of IL-17

IL-17 is the hallmark cytokine for the newly identified subset of Th cells, Th17. Th17 cells are important instigators of inflammation in several models of autoimmune disease; in particular, collagen induced arthritis (CIA) and experimental autoimmune encephalomyelitis (EAE), which were previously characterized as Th1-mediated diseases. Although high levels of IFN-gamma are secreted in CIA and EAE, disease is exacerbated in IFN-gamma- or IFN-gamma receptor-deficient mice due to the ability of IFN-gamma to suppress IL-17 secretion. However, in proteoglycan-induced arthritis (PGIA), severe arthritis is dependent on the production of IFN-gamma. We were therefore interested in determining the role of IL-17 in PGIA. We assessed the progression of arthritis in IL-17-deficient (IL-17-/-) mice and found the onset and severity of arthritis were equivalent in wild-type (WT) and IL-17-/- mice. Despite evidence that IL-17 is involved in neutrophil recruitment, synovial fluid from arthritic joints showed a comparable proportion of Gr1+ neutrophils in WT and IL-17-/- mice. IL-17 is also implicated in bone destruction in autoimmune arthritis, however, histological analysis of the arthritic joints from WT and IL-17-/- mice revealed a similar extent of joint cellularity, cartilage destruction, and bone erosion despite significantly reduced RANKL (receptor activator of NK-kappaB ligand) expression. There were only subtle differences between WT and IL-17-/- mice in proinflammatory cytokine expression, T cell proliferation, and autoantibody production. These data demonstrate that IL-17 is not absolutely required for autoimmune arthritis and that the production of other proinflammatory mediators is sufficient to compensate for the loss of IL-17 in PGIA.     

IL-23 Dependent and Independent Stages of Experimental Arthritis: No Clinical Effect of Therapeutic IL-23p19 Inhibition in Collagen-induced Arthritis

IL-23p19 deficient mice have revealed a critical role of IL-23 in the development of experimental autoimmune diseases, such as collagen-induced arthritis (CIA). Neutralizing IL-23 after onset of CIA in rats has been shown to reduce paw volume, but the effect on synovial inflammation and the immunological autoimmune response is not clear. In this study, we examined the role of IL-23 at different stages of CIA and during T cell memory mediated flare-up arthritis with focus on changes in B cell activity and Th1/Th17 modulation. Anti-IL-23p19 antibody (anti-IL23p19) treatment, starting 15 days after the type II collagen (CII)-immunization but before clinical signs of disease onset, significantly suppressed the severity of CIA. This was accompanied with significantly lower CII-specific IgG1 levels and lower IgG2a levels in the anti-IL-23p19 treated mice compared to the control group. Importantly, neutralizing IL-23 after the first signs of CIA did not ameliorate the disease. This was in contrast to arthritic mice that underwent an arthritis flare-up since a significantly lower disease score was observed in the IL-23p19 treated mice compared to the control group, accompanied by lower synovial IL-17A and IL-22 expression in the knee joints of these mice. These data show IL-23-dependent and IL-23-independent stages during autoimmune CIA. Furthermore, the memory T cell mediated flare-up arthritis is IL-23-mediated. These data suggest that specific neutralization of IL-23p19 after onset of autoimmune arthritis may not be beneficial as a therapeutic therapy for patients with rheumatoid arthritis (RA). However, T cell mediated arthritis relapses in patients with RA might be controlled by anti-IL-23p19 treatment.     

Immunization with glucose-6-phosphate isomerase induces T cell-dependent peripheral polyarthritis in genetically unaltered mice

Rheumatoid arthritis is a chronic inflammatory disease primarily affecting the joints. The search for arthritogenic autoantigens that trigger autoimmune responses in rheumatoid arthritis has largely focused on cartilage- or joint-specific Ags. In this study, we show that immunization with the ubiquitously expressed glycolytic enzyme glucose-6-phosphate isomerase (G6PI) induces severe peripheral symmetric polyarthritis in normal mice. In genetically unaltered mice, T cells are indispensable for both the induction and the effector phase of G6PI-induced arthritis. Arthritis is cured by depletion of CD4(+) cells. In contrast, Abs and FcgammaR(+) effector cells are necessary but not sufficient for G6PI-induced arthritis in genetically unaltered mice. Thus, the complex pathogenesis of G6PI-induced arthritis in normal mice differs strongly from the spontaneously occurring arthritis in the transgenic K/B x N model where Abs against G6PI alone suffice to induce the disease. G6PI-induced arthritis demonstrates for the first time the induction of organ-specific disease by systemic autoimmunity in genetically unaltered mice. Both the induction and effector phase of arthritis induced by a systemic autoimmune response can be dissected and preventive and therapeutic strategies evaluated in this model.     

Pituitary adenylate cyclase-activating polypeptide inhibits collagen-induced arthritis: an experimental immunomodulatory therapy.

Rheumatoid arthritis is a chronic, systemic, autoimmune, and inflammatory disorder that affects the synovial lining of the joints. We describe the beneficial effects of the pituitary adenylate cyclase-activating polypeptide (PACAP) in the collagen-induced arthritis experimental murine model being proposed as a novel therapeutic approach in the treatment of rheumatoid arthritis. PACAP greatly decreases arthritis frequency and severity in the studied mice by improving clinical symptoms, ameliorating joint damage, and blocking both the inflammatory and autoimmune mediators which are the main keys of the pathogenesis of this disease. With this study, PACAP emerges as a promising candidate for the treatment of a pathology with a high world incidence but currently no effective treatment.     

Schistosoma mansoni infection reduces severity of collagen-induced arthritis via down-regulation of pro-inflammatory mediators

Various experimental and epidemiological studies have demonstrated that helminth infections affect outcomes of allergic or autoimmune disorders. Here, we examined the effects of Schistosoma mansoni infection on mouse collagen-induced arthritis, one of the most widely used animal models for rheumatoid arthritis. Male DBA/1 mice were infected with S. mansoni 2 weeks prior to being immunized with type II collagen (IIC). Cytokine mRNA expression in mouse paws, cytokine production by ConA-stimulated spleen cells, and anti-IIC antibodies were evaluated in addition to the severity of arthritis. S. mansoni infection significantly reduced the severity of arthritis. Anti-IIC IgG and IgG2a levels were lower in infected than uninfected mice. With regard to cytokine producing potentials in the infected mice, the down-regulation of Th1 (IFNγ) and pro-inflammatory cytokines (TNFα and IL-17A), and up-regulation of Th2 (IL-4) and an anti-inflammatory cytokine (IL-10) were observed. In addition, real-time PCR revealed that the augmentation of pro-inflammatory mediators such as IL-1β, IL-6 and receptor activator of NFκB in inflamed paws was abrogated by S. mansoni infection. In conclusion, schistosome infection reduced the severity of autoimmune arthritis via systemic and local suppression of pro-inflammatory mediators, suggesting the potential of parasite-derived materials as therapeutic agents against rheumatoid arthritis.     

Estradiol treatment redirects the isotype of the autoantibody response and prevents the development of autoimmune arthritis

A number of clinical and experimental observations have been made relating elevated estrogen levels with the amelioration of autoimmune diseases, yet questions remain about the levels required for efficacy as well as the mechanism of disease inhibition. Using the collagen-induced arthritis (CIA) model, we have studied the effects of physiological, sustained levels of 17beta-estradiol in preventing the development of autoimmune arthritis and analyzed the changes in the autoimmune response. Using time-release pellets of 17beta-estradiol, arthritis development was significantly inhibited in three different strains of CIA-susceptible mice compared with the effect of placebo treatment, and serum estradiol levels similar to those of mice in estrus were found to be equally effective as higher estradiol concentrations. Analysis of the autoimmune response in the estradiol-treated mice indicated that T cell production of IFN-gamma was markedly decreased, and significant decreases were also observed in levels of IL-10 and GM-CSF produced by lymph nodes cells from estradiol-treated mice. Although the total IgG anti-CII response was only minimally affected by estrogen treatment, a significant reduction in the levels of IgG2a anti-CII Abs and an increase in the levels of IgG1 anti-CII Abs were observed in estradiol-treated mice. These data indicate that estradiol treatment altered the Th profile of the autoimmune T cell response, which, in turn, altered the production of IgG Abs to an isotype that is poor at fixing complement, an important component in the immunopathogenesis of CIA.     

Estrogen regulates T helper 17 phenotype and localization in experimental autoimmune arthritis

IntroductionThe incidence and progression of many autoimmune diseases are sex-biased, which might be explained by the immunomodulating properties of endocrine hormones. Treatment with estradiol potently inhibits experimental autoimmune arthritis. Interleukin-17-producing T helper cells (Th17) are key players in several autoimmune diseases, particularly in rheumatoid arthritis. The aim of this study was to investigate the effects of estrogen on Th17 cells in experimental arthritis.MethodsOvariectomized DBA/1 mice treated with 17β-estradiol (E2) or placebo were subjected to collagen-induced arthritis (CIA), and arthritis development was assessed. Th17 cells in joints and lymph nodes were studied by flow cytometry. Lymph node Th17 cells were also examined in ovariectomized estrogen receptor α–knockout mice (ERα^−/−) and wild-type littermates, treated with E2 or placebo and subjected to antigen-induced arthritis.ResultsE2-treated mice with established CIA showed reduced severity of arthritis and fewer Th17 cells in joints compared with controls. Interestingly, E2-treated mice displayed increased Th17 cells in lymph nodes during the early phase of the disease, dependent on ERα. E2 increased the expression of C-C chemokine receptor 6 (CCR6) on lymph node Th17 cells as well as the expression of the corresponding C-C chemokine ligand 20 (CCL20) within lymph nodes.ConclusionsThis is the first study in which the effects of E2 on Th17 cells have been characterized in experimental autoimmune arthritis. We report that E2 treatment results in an increase of Th17 cells in lymph nodes during the early phase of arthritis development, but leads to a decrease of Th17 in joints during established arthritis. Our data suggest that this may be caused by interference with the CCR6-CCL20 pathway, which is important for Th17 cell migration. This study contributes to the understanding of the role of estrogen in the development of autoimmune arthritis and opens up new fields for research concerning the sex bias in autoimmune disease.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-015-0548-y) contains supplementary material, which is available to authorized users.     

Identification of collagen-induced arthritis loci in aged multiparous female mice

Collagen-induced arthritis in mice is one of the most commonly used autoimmune experimental models, with many similarities to rheumatoid arthritis. Since collagen-induced arthritis is a complex polygenic disease there is a need for identification of several major disease-controlling genes. Because rheumatoid arthritis particularly affects aged women, we have in the present study identified new genetic regions critical for collagen-induced arthritis by studying aged female mice of a cross between NFR/N and B10.Q (H-2^q haplotype). The mice in the present study had different reproductive histories, which did not significantly affect the onset, incidence or severity of the disease. A total of 200 female mice were used in a total genome-wide screening with 125 microsatellite markers. We found one new significant quantitative trait locus affecting the arthritis incidence, severity and day of onset on chromosome 11 (denoted Cia40), which colocalizes with a locus controlling pregnancy failure. Furthermore, a quantitative trait locus of suggestive significance associated with the incidence, severity and day of onset was identified on chromosome 1. Finally, a suggestively significant quantitative trait locus associated with collagen type II antibody titers was identified on chromosome 13. This study indicates that several gene loci control arthritis in aged multiparous females, and that at least one of these loci coincides with pregnancy failure.     

Systemic autoimmune disease caused by autoreactive B cells that receive chronic help from Ig V region-specific T cells

B cells present BCR V region-derived Id-peptides on their MHC class II molecules to Id-specific CD4+ T cells. Prolonged Id-driven T-B collaboration could cause autoimmune disease, but this possibility is difficult to test in normal individuals. We have investigated whether mice doubly transgenic for an Id+ Ig L chain and an Id-specific TCR develop autoimmune disease. Surprisingly, T cell tolerance was not complete in these mice because a low frequency of weakly Id-reactive CD4+ T cells accumulated with age. These escapee Id-specific T cells provided chronic help for Id+ B cells, resulting in a lethal systemic autoimmune disease including germinal center reactions, hypergammaglobulinemia, IgG autoantibodies, glomerulonephritis, arthritis, skin affection, and inflammatory bowel disease. Inflamed tissues contained foci of Id-driven T-B collaboration, with deposition of IgG and complement. The disease could be transferred with B and T cells. The results demonstrate a novel mechanism for development of autoimmune disease in which self-reactive Id+ B cells receive prolonged help from Id-specific T cells, thus bypassing the need for help from T cells recognizing conventional Ag.     

A role for complement in antibody-mediated inflammation: C5-deficient DBA/1 mice are resistant to collagen-induced arthritis

Collagen-induced arthritis (CIA) represents an animal model of autoimmune polyarthritis with significant similarities to human rheumatoid arthritis that can be induced upon immunization with native type II collagen. As in rheumatoid arthritis, both cellular and humoral immune mechanisms contribute to disease pathogenesis. Genotypic studies have identified at least six genetic loci contributing to arthritis susceptibility, including the class II MHC. We have examined the mechanism of Ab-mediated inflammation in CIA joints, specifically the role of complement activation, by deriving a line of mice from the highly CIA-susceptible DBA/1LacJ strain that are congenic for deficiency of the C5 complement component. We show that such C5-deficient DBA/1LacJ animals mount normal cellular and humoral immune responses to native type II collagen, with the activation of collagen-specific TNF-alpha-producing T cells in the periphery and substantial intra-articular deposition of complement-fixing IgG Abs. Nevertheless, these C5-deficient mice are highly resistant to the induction of CIA. These data provide evidence for an important role of complement in Ab-triggered inflammation and in the pathogenesis of autoimmune arthritis.     

Immune and autoimmune disorders in HCV chronic liver disease: personal experience and commentary on literature

HCV chronic liver disease can be associated with a plethora of immune and autoimmune perturbations and many authors claim that HCV chronic infection can play an important role in the pathogenesis of these disorders. To compare our experience with literature reports, we performed a retrospective study on the case histories of 265 patients with HCV chronic liver disease, evaluating the type and prevalence of the associated immune and autoimmune manifestations. We found that the patients with HCV chronic liver disease can present arthromyalgias (7.1% of the patients), Sj?rgen's syndrome (5.2%), thyroiditis (4.1%), rheumatoid arthritis (2.2%), autoimmune thrombocytopenia (2.6%), mixed cryoglobulinemia (1.5%), autoimmune anemia (0.3%) and oral lichen planus (0.3%). We claim that HCV liver infection is able to induce immune and autoimmune perturbations, without playing a significant role in the pathogenesis of a well-defined disorder.     

Epicutaneous immunization with type II collagen inhibits both onset and progression of chronic collagen-induced arthritis

Epicutaneous immunization is a potential non-invasive technique for antigen-specific immune-modulation. Topical application of protein antigens to barrier-disrupted skin induces potent antigen-specific immunity with a strong Th2-bias. In this study, we investigate whether the autoimmune inflammatory response of chronic collagen-induced arthritis (CCIA) in DBA/1-TCR-beta Tg mice can be modified by epicutaneous immunization. We show that epicutaneous immunization with type II collagen (CII) inhibited development and progression of CCIA and, importantly, also ameliorated ongoing disease as indicated by clinical scores of disease severity, paw swelling and joints histology. Treated mice show reduced CII-driven T cell proliferation and IFN-gamma production, as well as significantly lower levels of CII-specific IgG2a serum antibodies. In contrast, CII-driven IL-4 production and IgE antibody levels were increased consistent with skewing of the CII response from Th1 to Th2 in treated mice. IL-4 production in treated mice was inversely correlated with disease severity. Moreover, T cells from treated mice inhibited proliferation and IFN-gamma production by T cells from CCIA mice, suggesting induction of regulatory T cells that actively inhibit effector responses in arthritic mice. The levels of CD4(+)CD25(+) T cells were however not increased following epicutaneous CII treatment. Together, these results suggest that epicutaneous immunization may be used as an immune-modulating procedure to actively re-programme pathogenic Th1 responses, and could have potential as a novel specific and simple treatment for chronic autoimmune inflammatory diseases such as rheumatoid arthritis.     

Experimental African Trypanosome Infection by Needle Passage or Natural Tsetse Fly Challenge Thwarts the Development of Collagen-Induced Arthritis in DBA/1 Prone Mice via an Impairment of Antigen Specific B Cell Autoantibody Titers

Collagen-induced arthritis is a B cell-mediated autoimmune disease. Recently published studies have demonstrated that in some rare cases pathogens can confer protection from autoimmunity. Trypanosoma brucei parasites are tsetse fly transmitted extracellular protozoans causing sleeping sickness disease in humans and Nagana in livestock in sub-Saharan endemic areas. In the past, we demonstrated that trypanosome infections impair B cell homeostasis and abolish vaccine-induced protection against unrelated antigens. Hence, here we hypothesized that trypanosome infection can affect the onset of CIA by specifically dampening specific B-cell responses and type II collagen antibody titers in DBA/1 prone mice. We observed a substantial delay in the onset of collagen-induced arthritis in T. brucei-infected DBA/1 mice that correlates with a drastic decrease of type II collagen titers of the different IgG isotypes in the serum. Treatment of infected mice with Berenil, a trypanocidal drug, restored the development of CIA-associated clinical symptoms. Interestingly, these data were confirmed by the challenge of immunized DBA/1 prone mice with T. brucei-infected tsetse flies. Together, these results demonstrate that T. brucei infection is impairing the maintenance of the antigen specific plasma B cell pool driving the development of CIA in DBA/1 prone mice.     

CD44 Antibodies and Immune Thrombocytopenia in the Amelioration of Murine Inflammatory Arthritis

Antibodies to CD44 have been used to successfully ameliorate murine models of autoimmune disease. The most often studied disease model has been murine inflammatory arthritis, where a clear mechanism for the efficacy of CD44 antibodies has not been established. We have recently shown in a murine passive-model of the autoimmune disease immune thrombocytopenia (ITP) that some CD44 antibodies themselves can induce thrombocytopenia in mice, and the CD44 antibody causing the most severe thrombocytopenia (IM7), also is known to be highly effective in ameliorating murine models of arthritis. Recent work in the K/BxN serum-induced model of arthritis demonstrated that antibody-induced thrombocytopenia reduced arthritis, causing us to question whether CD44 antibodies might primarily ameliorate arthritis through their thrombocytopenic effect. We evaluated IM7, IRAWB14.4, 5035-41.1D, KM201, KM114, and KM81, and found that while all could induce thrombocytopenia, the degree of protection against serum-induced arthritis was not closely related to the length or severity of the thrombocytopenia. CD44 antibody treatment was also able to reverse established inflammation, while thrombocytopenia induced by an anti-platelet antibody targeting the GPIIbIIIa platelet antigen, could not mediate this effect. While CD44 antibody-induced thrombocytopenia may contribute to some of its therapeutic effect against the initiation of arthritis, for established disease there are likely other mechanisms contributing to its efficacy. Humans are not known to express CD44 on platelets, and are therefore unlikely to develop thrombocytopenia after CD44 antibody treatment. An understanding of the relationship between arthritis, thrombocytopenia, and CD44 antibody treatment remains critical for continued development of CD44 antibody therapeutics.     

Oral administration of highly oligomeric procyanidins of Jatoba reduces the severity of collagen-induced arthritis

We have previously reported that highly oligomeric procyanidins (HOPC) purified from Jatoba, a South American herb, ameliorated experimental autoimmune encephalomyelitis (EAE) in mice. In this present study, we report that symptoms of arthritis were also significantly reduced by administering the Jatoba extract, when compared with the vehicle-alone-treated control. Interferon-gamma (IFN-gamma) production by the splenocytes from mice injected with procyanidins was also dramatically decreased. The oral administration of purified HOPC was significantly more effective in disease prevention than the ethanol (EtOH) extract of Jatoba. Green tea polyphenol administration, however, surprisingly facilitated disease development. Observation of the joint histopathology on whole paws derived from the HOPC-treated mice showed complete abrogation of collagen induced arthritis (CIA), a characteristic of chronic inflammation in the synovial tissue. These results demonstrate that HOPC administration had an inhibitory effect on both chronic arthritis and EAE and that the oral administration of HOPC exerted its effect after the induction of secondary immunity.     

SLAP deficiency enhances number and function of regulatory T cells preventing chronic autoimmune arthritis in SKG mice

To test if manipulating TCR complex-mediated signaling (TCR signaling) could treat autoimmune disease, we generated the double SKG Src-like adapter protein (SLAP) knockout (DSSKO) mouse model. The SKG mutation in ZAP70 and SLAP have opposing functions on the regulation of TCR signaling. The combination of these two mutations alters TCR signaling in the context of a defined genetic background, uniform environmental conditions, and a well-characterized signaling disruption. In contrast to SKG mice, DSSKO mice do not develop zymosan-induced chronic autoimmune arthritis. This arthritis prevention is not due to significant alterations in thymocyte development or repertoire selection but instead enhanced numbers of regulatory T cells (Tregs) and decreased numbers of Th17 cells skewing the ratio of Tregs to autoreactive effector T cells. Treg depletion and/or functional blockade led to the development of arthritis in DSSKO mice. In vitro suppression of effector T cell proliferation was also enhanced, demonstrating that DSSKO mice have increased numbers of Tregs with increased function. Understanding how TCR signals influence development, expansion, and function of Tregs in DSSKO mice could advance our ability to manipulate Treg biology to treat ultimately autoimmune disease.     

CD4+ T cells recognizing a single self-peptide expressed by APCs induce spontaneous autoimmune arthritis

We have examined processes leading to the spontaneous development of autoimmune inflammatory arthritis in transgenic mice containing CD4+ T cells targeted to a nominal Ag (hemagglutinin (HA)) and coexpressing HA driven by a MHC class II promoter. Despite being subjected to multiple tolerance mechanisms, autoreactive CD4+ T cells accumulate in the periphery of these mice and promote systemic proinflammatory cytokine production. The majority of mice spontaneously develop inflammatory arthritis, which is accompanied by an enhanced regional immune response in lymph nodes draining major joints. Arthritis development is accompanied by systemic B cell activation; however, neither B cells nor Ab is required for arthritis development, since disease develops in a B cell-deficient background. Moreover, arthritis also develops in a recombinase activating gene-deficient background, indicating that the disease process is driven by CD4+ T cells recognizing the neo-self HA Ag. These findings show that autoreactive CD4+ T cells recognizing a single self-Ag, expressed by systemically distributed APCs, can induce arthritis via a mechanism that is independent of their ability to provide help for autoantibody production.