The decline in immunocompetence with age is accompanied by the increase in the incidence of autoimmune diseases. Aging of the immune system, or immunosenescence, is characterized by a decline of both T and B cell function, and paradoxically the presence of low grade chronic inflammation. There is growing evidence that epigenetics, the study of inherited changes in gene expression that are not encoded by the DNA sequence itself, changes with aging. Interestingly, emerging evidence suggests a key role for epigenetics in human pathologies, including inflammatory and neoplastic disorders. Here we will review the potential mechanisms that contribute to the increase in autoimmune responses in aging. In particular, we will discuss how epigenetic alterations, especially DNA methylation and histone acetylation, are accumulated during aging and how these events contribute to autoimmunity risk.
aging; immunity and epigenetics
Activation-induced cytidine deaminase (AID) is expressed in a B cell differentiation stage-specific fashion and is essential for immunoglobulin (Ig) gene class switch DNA recombination (CSR) and somatic hypermutation (SHM). CSR and SHM play a central role in the maturation of antibody and autoantibody responses. AID displays a mutagenic activity by catalyzing targeted deamination of deoxycytidine (dC) residues in DNA resulting in dU:dG mismatches, which are processed into point-mutations in SHM or double-strand breaks (DSBs) in CSR. Although AID specifically targets the Ig gene loci (IgH, Igκ and Igλ), it can also home into a wide array of non-Ig genes in B- and non-B-cell backgrounds. Aberrant expression of AID is associated with multiple diseases such as allergy, inflammation, autoimmunity and cancer. In autoimmune systemic lupus erythematosus, dysregulated AID expression underpins increased CSR, SHM and autoantibody production. As a potent mutator, AID is under stringent transcriptional, post-transcriptional and post-translational regulation. AID is also regulated in its targeting and enzymatic function. In resting naïve or memory B cells, AID transcripts and protein are undetectable. These, however, are readily and significantly upregulated in B cells induced to undergo CSR and/or SHM. Transcription factors, such as HoxC4 and NF-κB, which are upregulated in a B cell lineage- and/or differentiation stage-specific manner, regulate the induction of AID. HoxC4 induces AID expression by directly binding to the AID gene promoter through an evolutionarily conserved 5’-ATTT-3’ motif. HoxC4 is induced by the same stimuli that induce AID and CSR. It is further upregulated by estrogen through three estrogen responsive elements in its promoter region. The targeting of AID to switch (S) regions is mediated by 14-3-3 adaptor proteins, which specifically bind to 5′-AGCT-3′ repeats that are exist at high frequency in S region cores. Like HoxC4, 14-3-3 adaptors are induced by the same stimuli that induce AID. These include “primary” inducing stimuli, that is, those that play a major role in inducing AID, i.e., engagement of CD40 by CD154, engagement of Toll-like receptors (TLRs) by microbial-associated molecular patterns (MAMPs) and cross-linking of the BCR, as synergized by “secondary” inducing stimuli, that is, those that synergize for AID induction and specify CSR to different isotypes, i.e., switch-directing cytokines IL-4, TGF-β or IFN-γ. In this review, we focus on the multi-levels regulation of AID expression and activity. We also discuss the dysregulation or misexpression of AID in autoimmunity and tumorigenesis.
activation-induced cytidine deaminase (AID); 14-3-3; antibody; autoantibody; class switch DNA recombination (CSR); HoxC4; mutagenesis; NF-κB; somatic hypermutation (SHM)
Immunoglobulin (Ig) gene somatic hypermutation (SHM) and class switch DNA recombination (CSR) play important roles in the generation of autoantibodies in systemic lupus erythematosus. Systemic lupus is characterized by the production of an array of pathogenic high-affinity mutated and class-switched, mainly IgG, antibodies to a variety of self-antigens, including nuclear components, such as dsDNA, histones and chromatin. We previously found that MRL/Faslpr/lpr mice, which develop a systemic autoimmune syndrome sharing many features with human lupus, display greatly upregulated CSR, particularly to IgG2a, in B cells of the spleen, lymph nodes and Peyer’s patches. In MRL/Faslpr/lpr mice, the significant upregulation of CSR is associated with increased expression of activation-induced cytidine deaminase (AID), which is critical for CSR and SHM. We also found that HoxC4 directly activates the promoter of the AID gene to induce AID expression, CSR and SHM. Here, we show that in both lupus patients and lupus-prone MRL/Faslpr/lpr mice, the expression of HoxC4 and AID is significantly upregulated. To further analyze the role of HoxC4 in lupus, we generated HoxC4−/− MRL/Faslpr/lpr mice. In these mice, HoxC4-deficiency resulted in reduced AID expression, impaired CSR and decreased serum anti-dsDNA IgG, particularly IgG2a, autoantibodies, which were associated with a reduction in IgG deposition in kidney glomeruli. In addition, consistent with our previous findings that in MRL/Faslpr/lpr mice, upregulated AID expression is associated with extensive DNA lesions, comprising deletions and insertions in the IgH locus, we found c-Myc to IgH(c-Myc/IgH) translocations to occur frequently in B cells of MRL/Faslpr/lpr mice. The frequency of such translocations was significantly reduced in HoxC4−/− MRL/Faslpr/lpr mice. These findings suggest that in lupus B cells, upregulation of HoxC4 plays a major role in dysregulation of AID expression, thereby increasing CSR and autoantibody production, and promoting c-Myc/IgH translocations.
Activation-induced cytidine deaminase (AID); B cell lymphoma; cancer; class switch DNA recombination (CSR); c-Myc/IgH translocation; HoxC4; MRL/Faslpr/lpr mice; systemic lupus erythematosus (SLE)
B cells contribute to autoimmunity both as secretors of pathogenic antibodies and through the activation of autoreactive T cells. B cells and antibodies acquire higher affinity to self-antigen through a process known as immunoglobulin hypermutation or SHM. The contribution of SHM to pathogenic antibody development in lupus has been established in various autoimmune mouse models and by examining antibodies from patients. However, its role in the antibody-independent contribution of B cells to autoimmunity has not been examined. Herein, we generate lupus-prone MRL/lpr mice with a limited IgM-only B cell repertoire, no secreted antibodies and no SHM. This enabled us to isolate the role of somatic hypermutation in B cell-mediated autoimmunity and found that SHM-deficiency correlated with a reduction in autoreactive B cells, a decrease in T cell activation and in kidney lymphocytic infiltration. These data establish AID as an important contributor to the antibody-independent role of B cells in autoimmunity.
somatic hypermutation; AID; B cells; T cells; lupus
Our laboratory investigates systemic autoimmune disease in the context of mouse models of systemic lupus erythematosus (SLE). SLE is associated with high titers of serum autoantibodies of the IgG class that are predominantly directed against nuclear antigens, with pathological manifestations that are considered by many to be characteristic of an immune-complex mediated disease. In this review, we focus on the known and potential roles of somatic mutagenesis in SLE. We will argue that antinuclear antibodies (ANA) arise predominantly from nonautoreactive B cells that are transformed into autoreactive cells by the process of somatic hypermutation (SHM), which is normally associated with affinity maturation during the germinal center reaction. We will also discuss the role of SHM in creating antigenic peptides in the V region of the B cell receptor (BCR) and its potential to open an avenue of unregulated T cell help to autoreactive B cells. Finally, we will end this review with new experimental evidence suggesting that spontaneous somatic mutagenesis of genes that regulate B cell survival and activation is a rate-limiting causative factor in the development of ANA.
Somatic mutation; Autoimmunity; Anti-nuclear Antibodies; Lupus; Haplodeficiency
B lymphocytes contribute to physiological immunity through organogenesis of secondary lymphoid organs, presentation of antigen to T cells, production of antibodies, and secretion of cytokines. Their role in several autoimmune diseases, mainly as producers of pathogenic antibodies, is also well known. However, certain subsets of B cells are emerging as the important regulatory cell populations in both mouse and human. The regulatory functions of B cells have been demonstrated in a variety of mouse models of autoimmune diseases including collagen-induced arthritis (CIA), experiment autoimmune encephalomyelitis (EAE), anterior chamber-associated immune deviation (ACAID), diabetes, contact hypersensitivity (CHS), and intestinal mucosal inflammation. Accumulating evidence from both mouse and human studies confirms the existence of regulatory B cells, and is beginning to define their mechanisms of action. In this article, we first review the history of B cells with regulatory function in autoimmune diseases, and summarize the current understanding about the characterizations of such B-cell subsets. We then discuss the possible regulatory mechanisms of B cells, and specifically define the role of regulatory B cells in immune homeostasis in the intestine.
Regulatory B cells; immunoregulation; autoimmune diseases; molecular mechanism; mucosal homeostasis
SJL mice represent a mouse model in which young adult females are susceptible to autoimmune disease while age matched male are relatively resistant. T cells primed in female SJL mice secrete cytokines associated with a Th1 phenotype. By contrast, T cells primed in males secrete cytokines associated with a Th2 phenotype. Activation of Th2-type T cells in males versus Th1 cells in females correlates with increased CD4+CD25+ T regulatory cells (Treg) in males. T cells primed in males depleted of CD4+CD25+ T cells preferentially secrete IFN-γ and decreased IL-4 and IL-10 compared to CD4+CD25+ T cells sufficient males suggesting that Treg influence subsequent antigen specific cytokine secretion. Treg from males and females exhibit equivalent in vitro T cell suppression. Treg from males expressed increased CTLA-4 and CD62L and preferentially secrete IL-10. These data suggest that an increased frequency of IL-10 secreting Treg in male SJL mice may contribute to resistance to autoimmune disease by favoring development of Th2 immune responses.
autoimmunity; EAE; regulatory T cells; cytokines; mice
B cells have been implicated both with pathogenic as well as protective capabilities in induction and regulation of autoimmune diseases. Rheumatoid arthritis (RA) is an autoimmune disease that occurs more often in women than men. A significant role of B cells as antibody producing and antigen-presenting cells has been demonstrated in RA. Predisposition to RA is associated with the presence of certain HLA class II alleles that share sequences with DRB1*0401. To determine the role of HLA genes and B cells in vivo, we have generated transgenic mice carrying HLA genes, DRB1*0401 and DQ8, known to be associated with susceptibility to RA. Humanized mice can be induced to develop arthritis that mimics human disease in clinical, histopathological and sex bias. Effect of hormones on immune cells and their function has been described in humans and mice and has been suggested to be the major reason for female bias of autoimmune diseases. An immune response to an antigen requires presentation by HLA molecules thus suggesting a critical role of MHC in combination with sex hormones in susceptibility to develop rheumatoid arthritis. Based on our observations, we hypothesize that modulation of B cells by estrogen, presentation of modified antigens by DR4 and production of antigen-specific B cell modulating cytokines leads to autoreactivity in females. These data suggest that considering patient’s sex may be crucial in selecting the optimal treatment strategy. Humanized mice expressing RA susceptible and resistant haplotype provide a means to investigate mechanism sex-bias of arthritis and future strategies for therapy.
B regulatory cells; antigen presentation; MHC polymorphism; HLA transgenic mice; Rheumatoid arthritis
The present study uses the agent-based model IMMSIM to simulate immune responses to a viral infection, with a focus on the impact of preformed memory (homologous and heterologous) on the quality and the efficacy of the response. The in machina results confirm the observed thwarting of new, naïve responses exerted by cross-reacting memory, but they also reveal that the competitive inhibition is made possible by the different time frame used by the primary and the secondary response, a well-known fact, epitomized by the interval of about 75 time steps between their peaks. This novel finding justifies the depression of naïve responses and the long-term consequences it could bring about and the role of memory as a player in a survival of the fittest game.
Cross-reacting memory; dynamic competition; clonal dominance
The actin cytoskeleton plays a crucial role in in a variety of important cellular processes required for normal immune function, including locomotion, intercellular interactions, endocytosis, cytokinesis, signal transduction and maintenance of cell morphology. Recent studies have uncovered not only many of the components and mechanisms that regulate the cortical actin cytoskeleton, but have also revealed significant immunopathologic consequences associated with genetic alteration of actin cytoskeletal regulatory genes. These advances have provided new insights into the role of cortical actin cytoskeletal regulation in a number of immune cell functions, and have identified cytoskeletal regulatory proteins critical for normal immune system activity and susceptibility to autoimmunity.
In myasthenia gravis (MG) and experimental autoimmune MG (EAMG) many pathologically significant autoantibodies are directed at the main immunogenic region (MIR), a conformation-dependent region at the extracellular tip of α1 subunits of muscle nicotinic acetylcholine receptors (AChRs). Human muscle AChR α1 MIR sequences were integrated into Aplysia ACh binding protein (AChBP). The chimera was potent at inducing both acute and chronic EAMG, though less potent than Torpedo electric organ AChR. Wild-type AChBP also induced EAMG but was less potent, and weakness developed slowly without an acute phase. AChBP is more closely related in sequence to neuronal α7 AChRs which are also homomeric, however autoimmune responses were induced to muscle AChR, but not to neuronal AChR subtypes. The greater accessibility of muscle AChRs to antibodies, compared to neuronal AChRs, may allow muscle AChRs to induce self-sustaining autoimmune responses. The human α1 subunit MIR is a potent immunogen for producing pathologically significant autoantibodies. Additional epitopes in this region or other parts of the AChR extracellular domain contribute significantly to myasthenogenicity. We show that an AChR-related protein can induce EAMG. Thus, in principle, an AChR-related protein could induce MG. AChBP is a water soluble protein resembling the extracellular domain of AChRs, yet rats which developed EAMG had autoantibodies to AChR cytoplasmic domains. We propose that an initial autoimmune response, directed at the MIR on the extracellular surface of muscle AChRs, leads to an autoimmune response sustained by muscle AChRs. Autoimmune stimulation sustained by endogenous muscle AChR may be a target for specific immunosuppression.
myasthenia gravis; autoantibodies; AChBP; AChR; MIR
heterologous immunity; T cell cross-reactivity; autoimmunity; viruses; immune modeling
The relationship between biological research and mathematical modeling is complex, critical, and vital. In this review, we summarize the results of the collaboration between two laboratories, exploring the interaction between mathematical modeling and wet-lab immunology. During this collaboration several aspects of the immune defence against viral infections were investigated, focusing primarily on the subject of heterologous immunity. In this manuscript, we emphasize the topics where computational simulations were applied in conjunction with experiments, such as immune attrition, the growing and shrinking of cross-reactive T cell repertoires following repeated infections, the short and long-term effects of cross-reactive immunological memory, and the factors influencing the appearance of new clonal specificities. For each topic, we describe how the mathematical model used was adapted to answer specific biological questions, and we discuss the hypotheses that were generated by simulations. Finally, we propose rules for testing hypotheses that emerge from model experimentation in the wet lab, and vice-versa.
IMMSIM simulator; heterologous memory; attrition; repertoire changes
EBV infection and the immune response may be involved in the pathogenesis of rheumatoid arthritis (RA). Past studies have suggested an association between EBV and RA.
We studied the association between EBV serologies and RA risk in a nested case-control study in the Nurses’ Health Study cohorts. We confirmed incident RA cases from 1990–2002 by questionnaire and medical record review. Each incident case with blood collected prior to RA symptoms was matched to a healthy participant by time of day and date of blood collection, birth year, menopausal status and postmenopausal hormone use. Immunofluorescence assays measured serologic EBV responses: viral capsid antigen (VCA), early-antigen-diffuse (EA-D) and early antigen-complex (EA-restricted and diffuse), Epstein Barr nuclear antigen (EBNA)-1, EBNA-2 and cytomegalovirus (CMV), as control. All were reported as titers, except BZLF-1 and CMV, which were reported as positive or negative. ANA positive samples were excluded. Elevated EBV antibody titers were defined as the upper 20% (or nearest titer) among controls. Conditional logistic regression analyses modeled RA risk associated with elevated EBV titers or the presence/absence CMV, further adjusted for pack-years smoking and alcohol intake.
87 incident RA cases were identified. Mean time to RA after blood draw was 6.2 (±3.5) years in NHS and 1.9 (±0.6) years in NHSII. Antibody titers against EBV were not significantly different between pre-RA cases and controls.
In this prospective study of women, we observed no association between EBV serologies and RA risk.
rheumatoid arthritis; Epstein Barr virus; virus; risk factors; epidemiology
Modifications of both DNA and protein by methylation are key factors in normal T and B cell immune responses as well as in the development of autoimmune disease. For example, the failure to maintain the methylation status of CpG dinucleotides in DNA triggers T cell autoreactivity. Methylated proteins are known targets of autoimmunity, including the symmetrical dimethylarginine residues of SmD1 and SmD3 in SLE. Herein, we demonstrate that altering the metabolism of S-adenosylmethionine (SAM), the major methyl donor for transmethylation reactions, can suppress T cell immunity. A by-product of SAM metabolism, 5′-Deoxy-5′-methylthioadenosine (MTA), and an indirect inhibitor of methyltransferases, inhibits T cell responses including T cell activation markers, Th1/Th2 cytokines and TCR-related signaling events. Moreover, treatment of the lupus-prone MRL/lpr mouse with MTA markedly ameliorates splenomegaly, lymphadenopathy, autoantibody titers as well as IgG deposition and cellular infiltration in the kidney. Incubation of cells with SAM, which increases intracellular MTA levels, inhibits both TCR-mediated T cell proliferation and BCR (anti-IgM)-triggered B cell proliferation in a dose-dependent manner. These studies define the central role of MTA and SAM in immune responses and provide a simple approach to altering lymphocyte transmethylation and T cell mediated autoimmune syndromes.
Transmethylation; methylthioadenosine; S-adenosylmethionine; CD4+ T cells; SLE
Histone H2B is a common target of autoantibodies in both spontaneous and drug-induced systemic lupus erythematosus (SLE). Recent studies demonstrate that Asp25 of histone H2B (H2B) spontaneously converts to an isoaspartic acid (isoAsp) in vivo. Our laboratory has demonstrated that the posttranslational modification of an aspartic acid to an isoaspartic acid within self-peptides renders otherwise ignored peptides immunogenic. Analysis of serum from lupus-prone mice and histone antibody positive SLE patients revealed antibodies specific to the Asp and isoAsp H2B21 –35 peptide, and that the expression of these antibodies is dependent on TLR9. IsoAsp H2B21–35 is immunogenic in non-autoimmune prone mice and mice lacking the ability to repair isoAsp have significantly reduced levels of antibodies to H2B. Asp H2B21– 35 incubated at physiological temperatures and pH acquires the isoAsp modification, demonstrating that H2B21 – 35 is prone to spontaneous isoAsp formation in vivo. Autoimmunity to isoAsp H2B suggests that this form of the autoantigen may be critical in the induction of anti-histone autoantibodies in human SLE and in murine models of disease.
Isoaspartyl; B cells; drug-induced lupus; autoantigen; autoantibodies
MΦ of mice from the major inbred models of systemic lupus erythematosus (SLE) have an identical defect affecting the activity of the cytoskeletal regulator and G-protein Rho. This abnormality is triggered by apo cells. Strikingly, SLE-prone MΦ show normal Rho activity when cultured in the absence of apo cells. We used gene arrays to identify adhesion-related gene products that are abnormally expressed by MΦ from prediseased 4–6-week-old SLE-prone MRL mice in the presence of serum lipids mimicking apo cells (SL-Apo). MΦ of MRL mice differentially expressed 42 adhesion-related genes in the presence of SL-Apo. Of these, 32 were expressed normally in the absence of SL-Apo. As adhesive interactions play a major role in lymphocyte activation, the detected apo cell-dependent abnormality could predispose to the development of autoimmunity. Indeed, several recent genetic studies support a role for adhesion-related genes in the pathogenesis of chronic autoimmunity.
PMID: 19811276 CAMSID: cams2351
Cytoskeleton; SLE; rodent; autoimmunity; gene array
Growing evidence exists for a new role for apoptotic cell recognition and clearance in immune homeostasis. Apoptotic cells at all stages, irrespective of membrane integrity, elicit a signature set of signaling events in responding phagocytes, both professional and non-professional. These signaling events are initiated by receptor-mediated recognition of apoptotic determinants, independently of species, cell type, or apoptotic stimulus. We propose that the ability of phagocytes to respond to apoptotic targets with a characteristic set of signaling events comprises a second distinct dimension of innate immunity, as opposed to the traditional innate discrimination of self vs. non-self. We further propose that a loss or abnormality of the signaling events elicited by apoptotic cells, as distinct from the actual clearance of those cells, may predispose to autoimmunity.
PMID: 17516209 CAMSID: cams2362
Innate immunity; apoptosis; autoimmunity; inflammation; phagocytosis; signal transduction
Anti-phospholipid syndrome (APS) is an autoimmune disorder characterized by the presence of autoantibody (AAb) to phospholipid (PL)-binding proteins, such as β2-glycoprotein I (β2GPI), and clinical manifestations including thrombosis and/or recurrent pregnancy loss. β2GPI-reactive T cells are clearly implicated in the generation of these AAb, but the mechanism responsible for their activation remains unclear. We hypothesized that immunization of mice with human β2GPI, in the context of a potent innate immune activator lipopolysaccharide (LPS), would generate not only high titers of anti-PL AAb, but also a strong β2GPI-specific T cell response. Healthy, nonautoimmune C57BL/6 mice were immunized repeatedly with human β2GPI in the presence of LPS. High titers of anti-PL to β2GPI appeared after the second immunization, with T cell reactivity to β2GPI detectable only after the fourth immunization. Splenic T cells from these mice proliferated in response to native β2GPI, alone or bound to anionic PL. These T cells produced IL-2 and IFN-γ, but not IL-4 or IL-10, indicating a Th1 bias of the β2GPI-specific response. These findings suggest that T cells responsive to β2GPI may become activated in APS patients by exposure to their cognate Ag in the context of innate immune activation and a pro-inflammatory environment.
PMID: 19811280 CAMSID: cams2310
β2-Glycoprotein I; TC; anti-phospholipid syndrome; systemic lupus erythematosus (SLE); rodent; autoimmunity
Sjögren’s syndrome (SjS) patients often have a variety of extraglandular manifestations including neurological and gastrointestinal involvement. In this study we evaluated the diagnostic performance of luciferase immunoprecipitation system (LIPS) that employs mammalian cell-produced recombinant antigens for analyzing SjS autoantibody responses. LIPS testing of mammalian cell-produced La, Ro60 and Ro52 recombinant antigens with defined commercial antibodies demonstrated highly specific immunoprecitation of each antigen without cross-reactivity. Next, sera from 57 SjS and 25 volunteers were evaluated by LIPS against a panel of human autoantigens. LIPS detected robust anti-La antibody levels in 43/57 SjS patients (75% sensitivity) and markedly outperformed an ELISA (46% sensitivity). Profiling of other SjS-associated autoantigens revealed the presence of anti-Ro60, anti-Ro52 in 63% and 61%, of SjS patients, respectively. Interestingly, a C-terminal fragment of Ro52 (Ro52-Δ2), a protein fragment not previously found to be antigenic by ELISA, also showed positive immunoreactivity in 42/57 SjS patients (65% sensitivity). Additional profiling of other autoantigens revealed that certain SjS patients also showed positive immunoreactivity with thyroid peroxidase (14%), AQP-4 (12%) and the H+/K+ gastric ATPase (16%) suggesting potential autoantibody attack of thyroid, neuronal and gastric parietal cells, respectively. These heterogeneous autoantibody responses detected by LIPS in SjS will likely be useful for diagnosis and for evaluating extraglandular manifestations.
Autoantibodies; Autoantigen; Diagnosis Sjögren’s syndrome; SSA; SSB
Epidermolysis bullosa acquisita (EBA) is a rare and acquired autoimmune subepidermal bullous disease of the skin and mucosa. EBA includes various distinct clinical manifestations resembling Bullous Pemphigus, Brunsting-Perry pemphigoid, or cicatricial pemphigoid. These patients have autoantibodies against type VII collagen, an integral component of anchoring fibrils, which are responsible for attaching the dermis to the epidermis. Destruction or perturbation of the normally functioning anchoring fibrils clinically results in skin fragility, blisters, erosions, scars, milia and nail loss, all features reminiscent of genetic dystrophic epidermolysis bullosa. These anti-type VII collagen antibodies are “pathogenic” because when injected into a mouse, the mouse develops an EBA-like blistering disease. Currently treatment is often unsatisfactory, however some success has been achieved with colchichine, dapsone, photopheresis, plasmaphresis, infliximab, rituximab and IVIG.
Antibodies recognizing the complement of the middle of PR3 (cPR3m) occur in ~30% of PR3-ANCA-vasculitis patients and immunization of animals with a peptide complementary to the middle of PR3 (cPR3m) induces not only anti-complementary PR3 antibodies, but also anti-PR3 antibodies derived through an anti-idiotypic response. PR3 epitopes recognized by patient ANCA however, are not restricted to the middle of PR3. This prompted us to test for antibodies that react with proteins complementary to the terminal regions of PR3 (cPR3C and cPR3N) in PR3-ANCA patients. Anti-cPR3C reactivity was detected in 28% of patients but anti-cPR3N reactivity in only 15%. Ranked anti-cPR3C and anti-cPR3m reactivity correlated in the cohort, whereas there was no significant relationship between cPR3C and cPR3N reactivity. Serial samples from three patients’ revealed that anti-cPR3C and anti-cPR3m reactivity followed a similar pattern over time. Serial samples from a fourth patient demonstrated an anti-cPR3N response without concurrent cPR3m or cPR3C reactivity. Epitope determination by mass spectrometry identified a thirteen amino acid sequence on cPR3C that contained a common binding site recognized by antibodies from three patients. This peptide sequence contains a “PHQ” motif which was reported to be the basis for cross-reactivity of anti-cPR3m antibodies with plasminogen. Why these antibodies are detected in only ~30% of the patients remains unclear. The data reveal it is not due to lack of inclusion of flanking regions of complementary PR3 during screening. Instead, quite unexpectedly, the data demonstrate that patients’ antibodies react with a restricted epitope that exists in both cPR3m and cPR3C.
Autoantigen Complementarity; ANCA vasculitis; anti-idiotypic antibodies; PR3; complementary protein pairs
FTY720 modulates lymphocyte trafficking through blood (PBL) and peripheral lymph nodes (PLN). Treatment with FTY720 causes retention of most blood lymphocytes in PLN. Long-term treatment can slow and/or prevent type 1 diabetes in the non-obese diabetic (NOD) mouse model. B and T cells are both affected by FTY720 binding to S1P1 (sphingosine-1-phosphate receptor 1). However, little has been done to elucidate which T cell subsets are differentially affected by FTY720 under healthy conditions, and how this affects disease pathogenesis in Type 1 Diabetes (T1D). In healthy C57BL/6J (B6) mice, total CD4+ and CD8+ T cell subsets were diminished by FTY720, but recently activated and memory subsets were spared and a constituted significantly higher percentage of remaining T cells in blood. FTY720 also lowered PBL counts in NOD mice, but less severely than in B6 mice. This is consistent with a different ratio of naïve, activated, and memory cells in NOD mice compared to B6 mice, as well as alterations in S1P1 and S1P (sphingosine-1-phosphate) levels in PBLs and blood of NOD mice, respectively. To address the functional consequences of PBL T cell depletion, we studied the effects of FTY720 on disease progression in a timed adoptive transfer model of T1D. Continuous treatment with FTY720 eliminated T1D if treatment was started before splenocyte transfer. FTY20 treatment started after disease onset slowed disease progression. The inability to fully suppress memory and effector T cell circulation may explain why FTY720 is only partially effective in the NOD adoptive transfer model of T1D.
Type 1 diabetes; T cells; immunomodulation; FTY720; adoptive transfer