Novel Frontiers in Regulatory B cells
Claudia Mauri
Abstract
The number of regulatory B cell “followers” has increased steadily in the last few years confirming the rising interest in understanding the relevance of this subset of B cells in controlling immune responses in a variety of immune-related diseases. Regulatory B cells (Breg) are potent modulators of immune responses, which prevent excessive inflammation and maintain immune homeostasis after infection or tissue-injury.1 Abnormalities in Breg number and function have been identified in immune-related pathologies such as autoimmune disease, chronic infections, cancer, and in the rejection of transplants.1, 2 Thus, it is of utmost clinical importance to understand the ontogeny of these populations, to phenotypically characterize Breg populations, and to understand the cellular signals and the molecular “cues,” which drive Breg differentiation. By better characterizing Breg populations and the processes leading to their differentiation, we can identify target molecules/pathways for therapeutic interventions in immune-related pathologies. The first manuscript describing the existence of B cells with suppressive function dates back to the 1970’s, where the suppressive function of B cells was first described in studies examining delayed-type hypersensitivity (DTH) reactions using either 2,4 dinitrofluorobenzene (DNFB) or ovalbumin (OVA) in incomplete Freund's adjuvant (IFA) or paraeminobenzoic acid-Hen egg albumin conjugate (PABA-HEA) in guinea pigs. Increased intensity and prolonged DTH reactions were observed in B cell-depleted DNFB-sensitized guinea pigs.3 Later studies showed that adoptive transfer of lymphocytes or total splenocytes suppressed the intensity of the DTH response.4, 5 These cells were dubbed “suppressor B cells.” However, the mechanisms behind the suppressive response were never investigated and the study of B cell regulation was not renewed until the 1990’s, when the late Charles Janeway and his team observed that B cell-deficient mice developed an exacerbated form of experimental autoimmune encephalomyelitis (EAE).6 It was not until 10 years later that almost simultaneously three papers revisited the concept of B cells as suppressors of immune responses in different autoimmune diseases and the term regulatory B cells was coined.7-9 Nevertheless, the field remained stalled for several years until B cell-depletion therapies were trialed and proved efficacious in rheumatic diseases. B cell therapy (BCT) originates from the cancer field where B cell-depletion therapies are used in the clinic to remove B cells in hematological B cell malignancies.10 A pioneering study conducted at University College, London, back in the early 2000’s, confirmed the efficacy that rituximab had in ameliorating disease in a small number of patients with refractory rheumatoid arthritis (RA).11 This initial study was followed by several clinical trials showing the therapeutic efficacy of this treatment in rheumatoid arthritis, systemic lupus erythematosus, and more recently in multiple sclerosis (MS).12-14 The majority of studies that have been focusing on understanding how B cell depletion therapies work showed, somewhat surprisingly, that autoantibodies known to play an important part in the pathogenesis of these diseases remain largely unaltered after B cell depletion. These results reignited an interest in understanding how B cells contribute to the pathogenesis of immune responses through antibody-independent mechanisms including antigen processing cells and cytokine/chemokine production. Twenty years later, we thought that it was timely to dedicate an entire volume to this underappreciated B cell subset, which regulates many of the immune responses in the body. In this volume, experts in this field discuss and reconcile issues related to the identification of Bregs, including whether Bregs are lineage-specific or if they arise at any stage of maturation in response to microenvironmental “cues”; the importance of tissue location and tissue-associated pathology in their differentiation; the need of interaction with other immune cells and how other immune cells shape Breg function and differentiation (Figure 1). Regulatory B cells (Bregs) play an important role in the control of inflammation, and IL-10 production is considered to be the hallmark for their identification. The identification of a universal surface marker, which captures the cellular and functional heterogeneity of human and murine Breg subsets, has so far evaded discovery.1, 15 In particular, we all know in the field that when we study Bregs by flow cytometry, for example, we are confronted with a highly heterogeneous cell population and that different B cell subsets, regardless of its developmental stage, can in response to an appropriate signal, become suppressive. Ma et al16 provide an phenotypical and functional overview of the different subsets of Bregs that have been so far reported. In particular, the authors describe how Bregs with different phenotypes arise in different allergic conditions including in food allergy and atopic dermatitis. They show that whereas CD9-expressing Bregs play a preventive role in both mouse models and patients with allergic asthma,17, 18 CD5-expressing and TGFβ-producing Bregs have been implicated in controlling food allergy and atopic dermatitis.19 They also touch upon how Breg responses change according to different stimuli and according to tissue residency, thus justifying the importance of gaining increased insight in this field, in order to develop more targeted therapies to allergic disease. An example of the clinical utility of harnessing Bregs for the treatment of allergies is allergen-specific immunotherapy. Beekeepers tolerized to bee venom allergen phospholipase A2, have increased frequencies of IL-10+IgG4+CD25hiCD71hiCD73- Bregs (BR1) compared to healthy controls. Remarkably, allergic patients have increased numbers of BR1 cells after receiving specific immunotherapy, thus linking BR1 cells to the maintenance of tolerance against bee venom allergens.20 Cherukuri et al21 also proposed TIM-1 as a shared surface marker for Breg subsets from murine models of transplantation. They explore in detail its unique role in promoting the upregulation of IL-10 and other inhibitory cytokines and receptors previously ascribed to Breg function, as well as the mechanisms implemented by Bregs to improve allograft survival. This group also points to the persistent challenge remaining in the identification of a universal human Breg marker, as only around 5% of peripheral blood B cells express TIM-1.16 Despite a continuous lack of a unifying markers, mechanistic and translational progress have been made. Rothstein and colleagues have reported recently encouraging findings on the clinical utility of using the IL-10/TNF (Breg:B effector) ratio as a biomarker for the prediction of clinical outcomes in patients undergoing transplantation; patients with allograft rejection had decreased numbers of IL-10+Bregs.21 The authors also provide interesting insight into the effects of immunosuppressive agents on Breg function and the current conundrum faced in the immune-suppressive field; that is how can we promote Breg function, while depleting pathogenic B cells? Further elucidation of the molecular signals required for Breg differentiation and the identification of unique surface receptors would help us to address this question. Multiple sclerosis (MS) has been historically considered a T cell-driven disease; however, the recent undeniable success of B cell-depletion therapy in the amelioration of MS clearly shows the involvement of B cells in the pathology of this disease.22, 23 Wang et al24 provide a balanced resume of recent findings showing the complex interplay of both pathogenic and protective effects that B cells have in this disease. The first evidence of a protective role of IL-10 producing B cells in neuroinflammation was first described in the EAE model.8 Since this initial study, multiple regulatory roles and functions of regulatory plasmablasts (PB) and plasma cells (PC) have been documented in EAE.25, 26 Wang et al nicely tie old findings with their latest discovery showing that the gut microbiota, known to shape immune responses, can promote the differentiation of regulatory IgA+ plasma cells. They also provide a “theoretical platform” discussing the potential that these cells have in the regulation of neuroinflammation and if and how regulatory PCs, that remain post-rituximab therapy, contribute to MS amelioration. A better understanding of the contributions of different B cell subsets to the regulation of neuroinflammation, and factors that impact the development, maintenance, and migration of such subsets, will be important for rationalizing next-generation B cell-directed therapies for the treatment of MS. The role played by B cells in different tissues/organs is still under-investigation. B cells are key sentinels at the mucosal sites and contribute to local immune responses through the release of antibodies and cytokines and through the presentation of antigen. Menon et al27 provide a detailed overview of the current state-of-knowledge on the role of B cells and Bregs in the mucosa-associated lymphoid tissue (MALT) in the lung in the healthy state and in immune-related lung pathologies. B cells are sparsely present in healthy lung tissue but arise as a result of infection or chronic inflammation and are predominantly found in ectopic lymphoid tissues (ELT).28 Importantly, B cells can have proinflammatory and anti-inflammatory roles, dependent on the microenvironmental cues these cells receive and abnormalities in Breg numbers and/or function are well-documented in various lung pathologies. Topically, the authors highlight recent research into the role of Bregs in severe Sars-Cov2 infection, where decreased numbers of IL-10+Bregs have been suggested to contribute to increased immunopathology.27, 29 Immune evasion by tumor cells is a major problem in the design of therapeutics and in the successful treatment of solid cancers.30 It is well-established that regulatory T cells promote an immunotolerant environment, which contributes to immune evasion.30 However, compared to autoimmunity, where it is well-established that they are numerically deficient and functionally impaired,2 the contribution of Bregs in solid tumors is in its infancy. Mounting evidence increasingly show that Bregs play a role in immune evasion in cancer and increased numbers of Bregs can be found intra-tumor in multiple mouse models of cancer.31 Menon et al report findings showing that increased number of tumor-infiltrating IL-10+Breg, suppressing tumoricidal responses, are found in the lungs of cancer patients.32, 33 Michaud et al31 describe the pro and anti-tumorigenic role that regulatory B cell and B cell play respectively in the anti-tumor microenvironment. They minutely review the existing literature showing how B cells recruited in tumors take advantage of their functional armory to increase cytotoxity and the killing of the tumor-target. B cells contribute to the clearance of tumors via production of antibody-dependent cellular cytotoxicity (ADCC) facilitating phagocytosis by dendritic cells; by enhancing presentation of tumors antigens to T cells; and via the production of proinflammatory cytokines.34-36 On the other hand, Bregs via the production of IL-35 and IL-10 contribute to the enhancement of the already immune-tolerogenic environment present in tumors. Michaud and colleagues propose how Bregs could be exploited in cancer treatment, and how, for example, selectively targeting IL-35-expressing Bregs could increase efficacy of cancer immunotherapy. Bregs require a direct interaction with other immune cells to exert their suppressive function. Bregs interact with CD4+ T cells and inhibit their differentiation into Th1 and Th17 cells, while promoting the development of FoxP3+Tregs. Bregs also suppress IL-12 producing dendritic cells, TNFα producing monocytes, and cytotoxic CD8+ T cells.1 The most recently discovered cell type found to cross-talk with Bregs are invariant natural killer T cells (iNKT)37, 38; a subset of cells bridging innate and adaptive responses to pathogens while also providing key regulation of immune homeostasis.39 Leadbetter and Karlsson provide a thorough review covering several aspects of iNKT and B cell interaction and how the outcome of this encounter leads to the generation of iNKT regulatory cell (iNKTreg)15 or iNKT follicular helper (iNKTFH) cells, that in turn support pathogen-specific effector B cell differentiation.40 In addition, an extensive review of the role that iNKT-B cell interactions have in modulating several autoimmune diseases and in cancer is presented. iNKT cells provide cognate help to B cells, which helps to promote the generation of tumor-specific antibodies.41 Conversely, the downregulation of CD1d in B cell leukemia contributes to immune evasion of the lytic function of iNKT cells.42-44 Whether iNKT cells can induce Bregs in the context of cancer remains to be determined. Targeted therapies increasing CD1d expression on B cells could be used to increase the tumoricidal activity of iNKT cells. Examining the nature of the antigen presented by CD1d as well as the tissue location of B cell:iNKT cell interactions could also be exploited for therapy. The formation of antigen-specific B cell clones and the incremental increase in antigen receptor affinity for any given antigen are key process for the formation of humoral immunity against viral pathogens. In this review, Burton and Maini, using the example of hepatitis B core antigen (HBcAg) and surface antigen (HBsAg), outline how the antigen load and properties of the antigen lead to the formation of memory and describe what determines whether a B cell clone proceeds through the germinal center (GC) reaction or through the extrafollicular differentiation pathway.45 Chronic hepatitis B infection is characterized by a persistent antigen load and an exhaustion of virus-specific CD8+ T cells. The authors delve into the dysregulation of the GC reaction in chronic hepatitis B patients and the preference of B cell clones to enter the extrafollicular B cell pathway, as is evidenced by the lack of long-lived plasma cell and memory B cell formation. Key to this cell fate decision are prefollicular helper T cells. The authors elucidate the mechanism behind chronic viral infection and the impact this has on the skewing of CD4 T cells subsets away from an archetypical T follicular helper cell phenotype, thereby facilitating the generation of extrafollicular B cell responses. Lastly, the authors touch on ectopic germinal center formation and their role in forming intrahepatic atypical memory B cell responses in hepatitis B patients. Taken together, these articles will provide an extensive overview of how B cells beyond antibodies can control immune responses to autoantigens but also to viral infections and cancer. This issue also provides several insights into the mechanisms of action of B cell depletion and how novel therapies targeting the interaction between B cells and cells of the innate immune system are currently used to inform drug development. By better understanding the phenotype, ontogeny, and molecular pathways leading to Breg differentiation, researchers can overcome the caveats of broad-spectrum B cell depletion and take more targeted approaches for the development of therapeutics specifically depleting pathogenic B cells and leaving regulatory B cells intact.