Do follicular dendritic cells regulate lupus-specific B cells?

The factors that allow self-reactive B cells to escape negative selection and become activated remain poorly defined. In this review we describe recently published results in which a B cell receptor-knock-in mouse strain specific for nucleolar self-antigens was bred with mice deficient in complement C4 and discuss the implications for the lupus field. Absence of C4 leads to a breakdown in the elimination of autoreactive B cell clones at the transitional stage. This is characterized by a relative increase in their response to a range of stimuli, entrance into follicles and a greater propensity to form self-reactive germinal centers. In this review, a model is proposed in which, in the absence of complement C4, inappropriate clearance of apoptotic debris promotes chronic activation of myeloid cells and follicular dendritic cells, resulting in secretion of Type I interferon. This allows for the maturation and activation of self-reactive B cell clones leading to increased spontaneous formation of germinal centers and subsequent generation of autoantibodies.     

Do follicular dendritic cells regulate lupus-specific B cells?

The factors that allow self-reactive B cells to escape negative selection and become activated remain poorly defined. In this review we describe recently published results in which a B cell receptor-knock-in mouse strain specific for nucleolar self-antigens was bred with mice deficient in complement C4 and discuss the implications for the lupus field. Absence of C4 leads to a breakdown in the elimination of autoreactive B cell clones at the transitional stage. This is characterized by a relative increase in their response to a range of stimuli, entrance into follicles and a greater propensity to form self-reactive germinal centers. In this review, a model is proposed in which, in the absence of complement C4, inappropriate clearance of apoptotic debris promotes chronic activation of myeloid cells and follicular dendritic cells, resulting in secretion of Type I interferon. This allows for the maturation and activation of self-reactive B cell clones leading to increased spontaneous formation of germinal centers and subsequent generation of autoantibodies.     

Autoreactivity and the positive selection of B cells

The diversity among B‐cell antigen receptor (BCR) specificities is generated by random rearrangement of gene segments during early B‐cell development. While such stochastic recombination of gene segments is important for diversity, it introduces the risk of producing self‐reactive BCRs which might lead to the development of autoimmune diseases. Therefore, it has been proposed that negative selection of autoreactive BCR specificities during early B‐cell development are required to establish tolerance towards self. In fact, transgenic mouse models have identified a number of “tolerance mechanisms” such as receptor editing, clonal deletion and anergy, all of which prevent the development of autoreactive B cells. Recent data, however, reveal that self‐recognition is crucial for the generation of B cells, and that the precursor‐BCR (pre‐BCR), which is essential for early B‐cell development, basically plays the role of an autoreactive BCR. Moreover, although it has become clear that autoreactive B cells are present in the periphery of healthy individuals, the role of autoantigen in their development, persistence and regulation is unclear. This review outlines the important role of autoreactivity in early B‐cell development and presents potential models for the regulation of the activation of peripheral B cells by different forms of self or foreign antigens.     

Monocyte‐derived dendritic cells can induce autoreactive CD4+ T cells showing myeloid lineage directed reactivity in healthy individuals

T cells against self‐antigens can be detected in peripheral blood of healthy individuals, although intrathymic negative selection removes most high‐avidity T cells specific for self‐antigens from the peripheral repertoire. Moreover, spontaneous T‐cell proliferation following stimulation with autologous monocyte‐derived dendritic cells (autoDCs) has been observed in vitro. In this study, we characterized the nature and immunological basis of the autoDC reactivity in the T‐cell repertoire of healthy donors. We show that a minority of naive and memory CD4⁺ T cells within the healthy human T‐cell repertoire mediates HLA‐restricted reactivity against autoDCs, which behave like a normal antigen‐specific immune response. This reactivity appeared to be primarily directed against myeloid lineage cells. Although cytokine production by the reactive T cells was observed, this did not coincide with overt cytotoxic activity against autoDCs. AutoDC reactivity was also observed in the CD8⁺ T‐cell compartment, but this appeared to be mainly cytokine‐induced rather than antigen‐driven. In conclusion, we show that the presence of autoreactive T cells harboring the potential to react against autologous and HLA‐matched allogeneic myeloid cells is a common phenomenon in healthy individuals. These autoDC‐reactive T cells may help the induction of primary T‐cell responses at the DC priming site.     

Autoreactive T cells in normal mice: unrestricted recognition of self peptides on dendritic cell I-A molecules by CD4-CD8- T cell receptor alpha/beta+ T cell clones expressing V beta 8.1 gene segments

CD4-CD8- double-negative (DN) and CD4+CD8- T cell clones were derived from splenic precursors resistant to killing by anti-Thy-1, -CD5, -CD4 and -CD8 monoclonal antibodies and complement. Both DN and CD4+ clones express functional T cell receptor (TcR) alpha/beta and exhibit strong autoreactivity in vitro. DN cells can be induced to proliferate by dendritic cells (DC) of all haplotypes tested, although this activation is inhibited by antibodies specific for I-A determinants expressed on the stimulatory DC. In contrast, CD4+ clones only respond to syngeneic or I-Ad-compatible DC. Both DN and CD4+ autoreactive clones do not proliferate when cultured with class II+ H-2d normal or tumor macrophages and B cell lines or with class II-transfected L cells, suggesting that these cells recognize self peptides only present on the surface of DC. Despite their phenotype resembling that of immature thymocytes and their inability to interact directly with B lymphocytes, DN cloned T cells, like CD4+ T cells, exhibit nonspecific helper functions and can induce polyclonal B cell proliferation and differentiation. DN TcR alpha/beta+ peripheral T cells represent, like TcR gamma/delta+ lymphocytes, a new T cell subset physiological role whose remains to be defined.     

Aberrant antibody affinity selection in SHIP‐deficient B cells

The strength of the Ag receptor signal influences development and negative selection of B cells, and it might also affect B‐cell survival and selection in the GC. Here, we have used mice with B‐cell‐specific deletion of the 5′‐inositol phosphatase SHIP as a model to study affinity selection in cells that are hyperresponsive to Ag and cytokine receptor stimulation. In the absence of SHIP, B cells have lower thresholds for Ag‐ and interferon (IFN)‐induced activation, resulting in augmented negative selection in the BM and enhanced B‐cell maturation in the periphery. Despite a tendency to spontaneously downregulate surface IgM expression, SHIP deficiency does not alter anergy induction in response to soluble hen‐egg lysozyme Ag in the MDA4 transgenic model. SHIP‐deficient B cells spontaneously produce isotype‐switched antibodies; however, they are poor responders in immunization and infection models. While SHIP‐deficient B cells form GCs and undergo mutation, they are not properly selected for high‐affinity antibodies. These results illustrate the importance of negative regulation of B‐cell responses, as lower thresholds for B‐cell activation promote survival of low affinity and deleterious receptors to the detriment of optimal Ab affinity maturation.     

Autoreactive T cell clones isolated from normal and autoimmune-susceptible mice exhibit lymphokine secretory and functional properties of both Th1 and Th2 cells

Recent studies have suggested the existence of two mutually exclusive subpopulations of T helper (Th) cells in the murine immune system, called Th1 which produces interleukin (IL)-2 and interferon (IFN)-gamma but not IL-4 and Th2 which secretes IL-4 and IL-5 but not IL-2. Also, functionally, Th1 cells generally activate the macrophages and mediate delayed-type hypersensitivity whereas Th2 cells provide help efficiently to B cells. In the present study, we investigated the lymphokine secretory properties of two well-characterized autoreactive (self-Ia reactive) T cell clones isolated from normal DBA/2 mice and autoimmune-susceptible MRL-lpr/lpr mice. It was observed that both the autoreactive T cell clones, following activation, produced IL-2, IL-4, and IFN-gamma. They induced hyper-Ia expression and cell proliferation in syngeneic B cells as well as activated the macrophages to exhibit tumoristatic properties. Both clones could also induce T-T network interaction in which syngeneic naive CD4+ T cells responded directly to stimulation with autoreactive T cell clones. The T-T interaction was demonstrable in 1-month-old MRL-lpr/lpr mice prior to the onset of the autoimmune disease but not in 6-month-old mice having lymphadenopathy and autoimmune disease. Unlike Th1 and Th2 cells which upon antigenic stimulation respond to exogenous IL-2 and IL-4, the autoreactive T cell clones responded only to IL-2 but not to IL-4. Our data suggest the existence of a unique subset of immunoregulatory CD4+ Th cells having the lymphokine secretory and functional properties of both the murine Th1 and Th2 subsets.     

The absence of immunoglobulin D B cell receptor-mediated signals promotes the production of autoantibodies and exacerbates glomerulonephritis in murine lupus

Immunoglobulin (Ig)D is the major antigen receptor isotype co-expressed with IgM on the surface of most peripheral B cells in mice and humans. However, the biological role of IgD as B cell receptor (BCR) has remained unclear. Previous studies have indicated that IgD may play a role in B cell tolerance. To understand the role of IgD in B cell tolerance and autoimmunity, we have examined the development of autoimmune syndrome in lpr mice deficient for IgD. The present study showed that IgD deficiency did not alter lymphoproliferation and lymphocyte activation in lpr mice. The survival and proliferation of B cells were not affected by the absence of IgD, indicating that IgD BCR-mediated signals do not have an important role in negative selection of autoreactive B cell clones. Interestingly, compared to IgD-competent littermates, lpr mice with IgD deficiency had elevated autoantibody production, increased deposition of immune complex in the kidney and more severe nephritis. Accumulation of abnormal CD4(-) CD8(-) αβ(+) T cells was accelerated in IgD(-/-) lpr mice compared to lpr mice. These results suggest that IgD BCR-mediated signals may be involved in the differentiation of autoreactive B cells into plasma cells and abnormal T cell expansion.     

The five dimensions of B cell tolerance

B cell tolerance has been generally understood to be an acquired property of the immune system that governs antibody specificity in ways that avoid auto‐toxicity. As useful as this understanding has proved, it fails to fully explain the existence of auto‐reactive specificities in healthy individuals and contribution these may have to health. Mechanisms underlying B cell tolerance are considered to select a clonal repertoire that generates a collection of antibodies that do not bind self, ie tolerance operates more or less in three dimensions that largely spare autologous cells and antigens. Yet, most B lymphocytes in humans and probably in other vertebrates are auto‐reactive and absence of these auto‐reactive B cells is associated with disease. We suggest that auto‐reactivity can be embodied by extending the concept of tolerance by two further dimensions, one of time and circumstance and one that allows healthy cells to actively resist injury. In this novel concept, macromolecular recognition by the B cell receptor leading to deletion, anergy, receptor editing or B cell activation is extended by taking account of the time of development of normal immune responses (4th dimension) and the accommodation (or tolerance) of normal cells to bound antibody, activation of complement, and interaction with inflammatory cells (fifth dimension). We discuss how these dimensions contribute to understanding B cell biology in health or disease.     

Chronic bacterial infection activates autoreactive B cells and induces isotype switching and autoantigen‐driven mutations

The links between infections and the development of B‐cell‐mediated autoimmune diseases are still unclear. In particular, it has been suggested that infection‐induced stimulation of innate immune sensors can engage low affinity autoreactive B lymphocytes to mature and produce mutated IgG pathogenic autoantibodies. To test this hypothesis, we established a new knock‐in mouse model in which autoreactive B cells could be committed to an affinity maturation process. We show that a chronic bacterial infection allows the activation of such B cells and the production of nonmutated IgM autoantibodies. Moreover, in the constitutive presence of their soluble antigen, some autoreactive clones are able to acquire a germinal center phenotype, to induce Aicda gene expression and to introduce somatic mutations in the IgG heavy chain variable region on amino acids forming direct contacts with the autoantigen. Paradoxically, only lower affinity variants are detected, which strongly suggests that higher affinity autoantibodies secreting B cells are counterselected. For the first time, we demonstrate in vivo that a noncross‐reactive infectious agent can activate and induce autoreactive B cells to isotype switching and autoantigen‐driven mutations, but on a nonautoimmune background, tolerance mechanisms prevent the formation of consequently dangerous autoimmunity.     

Scarcity of autoreactive human blood IgA+ memory B cells

Class‐switched memory B cells are key components of the “reactive” humoral immunity, which ensures a fast and massive secretion of high‐affinity antigen‐specific antibodies upon antigenic challenge. In humans, IgA class‐switched (IgA⁺) memory B cells and IgA antibodies are abundant in the blood. Although circulating IgA⁺ memory B cells and their corresponding secreted immunoglobulins likely possess major protective and/or regulatory immune roles, little is known about their specificity and function. Here, we show that IgA⁺ and IgG⁺ memory B‐cell antibodies cloned from the same healthy humans share common immunoglobulin gene features. IgA and IgG memory antibodies have comparable lack of reactivity to vaccines, common mucosa‐tropic viruses and commensal bacteria. However, the IgA⁺ memory B‐cell compartment contains fewer polyreactive clones and importantly, only rare self‐reactive clones compared to IgG⁺ memory B cells. Self‐reactivity of IgAs is acquired following B‐cell affinity maturation but not antibody class switching. Together, our data suggest the existence of different regulatory mechanisms for removing autoreactive clones from the IgG⁺ and IgA⁺ memory B‐cell repertoires, and/or different maturation pathways potentially reflecting the distinct nature and localization of the cognate antigens recognized by individual B‐cell populations.     

Impaired B‐cell tolerance checkpoints promote the development of autoimmune diseases and pathogenic autoantibodies

A role for B cells in autoimmune diseases is now clearly established both in mouse models and humans by successful treatment of multiple sclerosis and rheumatoid arthritis with anti‐CD20 monoclonal antibodies that eliminate B cells. However, the underlying mechanisms by which B cells promote the development of autoimmune diseases remain poorly understood. Here, we review evidence that patients with autoimmune disease suffer from defects in early B‐cell tolerance checkpoints and therefore fail to counterselect developing autoreactive B cells. These B‐cell tolerance defects are primary to autoimmune diseases and may result from altered B‐cell receptor signaling and dysregulated T‐cell/regulatory T‐cell compartment. As a consequence, large numbers of autoreactive naive B cells accumulate in the blood of patients with autoimmune diseases and may promote autoimmunity through the presentation of self‐antigen to T cells. In addition, new evidence suggests that this reservoir of autoreactive naive B cells contains clones that may develop into CD27^?CD21^?/lo B cells associated with increased disease severity and plasma cells secreting potentially pathogenic autoantibodies after the acquisition of somatic hypermutations that improve affinity for self‐antigens.     

High-frequency and adaptive-like dynamics of human CD1 self-reactive T cells

CD1 molecules present lipid antigens to T cells. An intriguing subset of human T cells recognize CD1‐expressing cells without deliberately added lipids. Frequency, subset distribution, clonal composition, naïve‐to‐memory dynamic transition of these CD1 self‐reactive T cells remain largely unknown. By screening libraries of T‐cell clones, generated from CD4⁺ or CD4^?CD8^? double negative (DN) T cells sorted from the same donors, and by limiting dilution analysis, we find that the frequency of CD1 self‐reactive T cells is unexpectedly high in both T‐cell subsets, in the range of 1/10–1/300 circulating T cells. These T cells predominantly recognize CD1a and CD1c and express diverse TCRs. Frequency comparisons of T‐cell clones from sorted naïve and memory compartments of umbilical cord and adult blood show that CD1 self‐reactive T cells are naïve at birth and undergo an age‐dependent increase in the memory compartment, suggesting a naïve/memory adaptive‐like population dynamics. CD1 self‐reactive clones exhibit mostly Th1 and Th0 functional activities, depending on the subset and on the CD1 isotype restriction. These findings unveil the unanticipated relevance of self‐lipid T‐cell response in humans and clarify the basic parameters of the lipid‐specific T‐cell physiology.     

Regulation of autoreactive B cells: checkpoints and activation

It has become clear that the autoreactive B cells are a part of the normal na?ve B cell repertoire in the periphery, despite the fact that they undergo a series of checkpoints, which include receptor editing (revision), clonal deletion, and anergy. However, most of those B cells reactive against self antigen remain functionally na?ve for autoantibody production by differential peripheral checkpoints. Therefore, the presence of autoreactive B cells does not always signify disease. Regulation of their activation and effector functions will determine the ultimate outcome. Although autoreactive B cell tolerance is well maintained in the healthy individual, the existence of pathogenic autoantibodies in autoimmune diseases indicates that these tolerogenic checkpoints are broken. Recent studies have demonstrated that autoreactive B cells are regulated by a composite of factors, such as genetic susceptibility and environmental triggers such as bacterial and viral infections as well as other immune cells. Interestingly, Toll-like receptors, previously considered as pattern-recognition receptors to detect and sense pathogens, may also have a potential to recognize self antigens and regulate autoreactive B cells for activation. Understanding the mechanisms of autoreactive B cell regulation and activation may help in identifying novel targets for the treatment of autoimmune diseases.     

Toll-like receptor driven B cell activation in the induction of systemic autoimmunity

Studies over the past decade have demonstrated a key role for pattern recognition receptors in the activation of autoreactive B cells. Self reactive B cells that manage to escape negative selection often express relatively low affinity receptors for self antigens (ignorant B cells), and can only be activated by integrating a relatively weak BCR signal with signals from additional receptors. Members of the toll-like receptor (TLR) gene family, and especially the nucleic acid binding receptors TLR 7, 8 and 9, appear to play a key role in this regard and promote the production of autoantibodies reactive with DNA- or RNA-associated autoantigens. These autoantibodies are able to form immune complexes with soluble or cell-bound ligands, and these immune complexes can in turn activate a second round of proinflammatory cells that further contribute to the autoimmune disease process. Recent data have emerged showing a pathogenic role for TLR7, with an opposing, protective role for TLR9. Targeting these disregulated pathways offers a therapeutic opportunity to treat autoimmune diseases without crippling the entire immune system. Further understanding of the role of specific receptors, cell subsets, and inhibitory signals that govern these TLR-associated pathways will enable future therapeutics to be tailored to specific categories of autoimmune disease.     

Multiple paths to loss of anergy and gain of autoimmunity

B cells and autoimmunity: cells of the immune system have the capacity to recognize/neutralize a myriad array of disease-causing pathogens, while simultaneously minimizing damage to self tissue. Obvious breakdowns in this ability to distinguish between self and non-self are evident in multiple forms of autoimmune disease, where B and T cells mount damaging attacks on cells and organs. B cells may directly damage tissue by producing pathogenic antibodies that bind self antigen, fix complement or form immune complexes. Recent evidence also suggests B cells indirectly induce autoimmunity by concentrating low avidity self antigen through the B cell receptor and presenting self-peptides to autoreactive T cells. B cells may also initiate autoimmunity when provided sufficient help from autoreactive T cells that have escaped deletion in the thymus. Here, we will review the role of anergy in maintenance of tolerance and how alterations in the normal balance of positive and negative signals may contribute to the development of autoimmune disease in mouse models and humans.     

Role of BCR affinity in T cell dependent antibody responses in vivo

Antibody affinity for antigen is believed to govern B lymphocyte selection during T-dependent immune responses. To examine antibody affinity in T cell dependent immune responses, we compared mice that carry targeted V(H)B1-8 antibody genes with high or low antigen-binding affinity. We found that high- and low-affinity B cells had the same intrinsic capacity to respond to antigen, but in experiments where limiting numbers of high- and low-affinity B cells were mixed in wild-type recipient mice, only the high-affinity B cells accumulated in germinal centers (GCs). In GCs, high-affinity B cells accumulated fewer V(H) somatic mutations than low affinity B cells. This effect was due to selections as the frequency of mutation in noncoding immunoglobulin gene DNA is the same in high- and low- affinity B cells. Thus, B cells recruited to the GC appeared to undergo a fixed mutation program, regardless of initial B cell receptor affinity. We conclude that in addition to the selection that occurs in GCs, stringent selection for high-affinity clones is also imposed in the early stages of the T cell dependent immune response in vivo.     

The immunoregulatory effects of Qa-1

Summary: The immune system is not only well equipped to control infections but also tightly controlled to prevent autoimmune disease. Despite the negative selection of T‐cell clones in the thymus, mature T cells capable of recognizing self‐antigens are present in every individual. Several types of specialized regulatory cells maintain homeostasis and prevent expansion of autoreactive T cells. In this issue of Immunological Reviews, the role of CD4⁺ regulatory T cells is extensively discussed. Suppression of T‐cell responses by CD8⁺ T cells has received less attention. Here, we review research on Qa‐1‐restricted CD8⁺ regulatory T cells. We focus on the role of this class Ib major histocompatibility complex (MHC) molecule in both CD8⁺ regulatory T‐cell activity and protection of activated T cells.     

Multiple paths to loss of anergy and gain of autoimmunity

B cells and autoimmunity: cells of the immune system have the capacity to recognize/neutralize a myriad array of disease-causing pathogens, while simultaneously minimizing damage to self tissue. Obvious breakdowns in this ability to distinguish between self and non-self are evident in multiple forms of autoimmune disease, where B and T cells mount damaging attacks on cells and organs. B cells may directly damage tissue by producing pathogenic antibodies that bind self antigen, fix complement or form immune complexes. Recent evidence also suggests B cells indirectly induce autoimmunity by concentrating low avidity self antigen through the B cell receptor and presenting self-peptides to autoreactive T cells. B cells may also initiate autoimmunity when provided sufficient help from autoreactive T cells that have escaped deletion in the thymus. Here, we will review the role of anergy in maintenance of tolerance and how alterations in the normal balance of positive and negative signals may contribute to the development of autoimmune disease in mouse models and humans.     

T cell interactions with B cells during germinal center formation,a three‐step model

Establishment of effective immunity against invading microbes depends on continuous generation of antibodies that facilitate pathogen clearance. Long‐lived plasma cells with the capacity to produce high affinity antibodies evolve in germinal centers (GCs), where B cells undergo somatic hypermutation and are subjected to affinity‐based selection. Here, we focus on the cellular interactions that take place early in the antibody immune response during GC colonization. Clones bearing B‐cell receptors with different affinities and specificities compete for entry to the GC, at the boundary between the B‐cell and T‐cell zones in lymphoid organs. During this process, B cells compete for interactions with T follicular helper cells, which provide selection signals required for differentiation into GC cells and antibody secreting cells. These cellular engagements are long‐lasting and depend on activation of adhesion molecules that support persistent interactions and promote transmission of signals between the cells. Here, we discuss how interactions between cognate T and B cells are primarily maintained by three types of molecular interactions: homophilic signaling lymphocytic activation molecule (SLAM) interactions, T‐cell receptor: peptide‐loaded major histocompatibility class II (pMHCII), and LFA‐1:ICAMs. These essential components support a three‐step process that controls clonal selection for entry into the antibody affinity maturation response in the GC, and establishment of long‐lasting antibody‐mediated immunity.