The discovery that lymphocyte subpopulations participate in distinct components of the immune response focused attention onto the origins and function of lymphocytes more than 40 years ago. Studies in the 1960s and 1970s demonstrated that B and T lymphocytes were responsible primarily for the basic functions of antibody production and cell-mediated immune responses, respectively. The decades that followed have witnessed a continuum of unfolding complexities in B-cell development, subsets, and function that could not have been predicted. Some of the landmark discoveries that led to our current understanding of B lymphocytes as the source of protective innate and adaptive antibodies are highlighted in this essay. The phenotypic and functional diversity of B lymphocytes, their regulatory roles independent of antibody production, and the molecular events that make this lineage unique are also considered. Finally, perturbations in B-cell development that give rise to certain types of congenital immunodeficiency, leukemia/lymphoma, and autoimmune disease are discussed in the context of normal B-cell development and selection. Despite the significant advances that have been made at the cellular and molecular levels, there is much more to learn, and cross-disciplinary studies in hematology and immunology will continue to pave the way for new discoveries.

Regulatory B cells control inflammation and autoimmunity in mice, including the recently identified IL-10–competent B10 cell subset that represents 1% to 3% of spleen B cells. In this study, a comparable IL-10–competent B10 cell subset was characterized in human blood. B10 cells were functionally identified by their ability to express cytoplasmic IL-10 after 5 hours of ex vivo stimulation, whereas progenitor B10 (B10pro) cells required 48 hours of in vitro stimulation before they acquired the ability to express IL-10. B10 and B10pro cells represented 0.6% and approximately 5% of blood B cells, respectively. Ex vivo B10 and B10pro cells were predominantly found within the CD24hiCD27+ B-cell subpopulation that was able to negatively regulate monocyte cytokine production through IL-10–dependent pathways during in vitro functional assays. Blood B10 cells were present in 91 patients with rheumatoid arthritis, systemic lupus erythematosus, primary Sjögren syndrome, autoimmune vesiculobullous skin disease, or multiple sclerosis, and were expanded in some cases as occurs in mice with autoimmune disease. Mean B10 + B10pro-cell frequencies were also significantly higher in patients with autoimmune disease compared with healthy controls. The characterization of human B10 cells will facilitate their identification and the study of their regulatory activities during human disease.

IL-21- and CD40-dependent cognate interactions with T cells are identified as key drivers for the generation of IL-10-producing regulatory B cells, which can protect against autoimmune disease. Subsets of regulatory B cells have been identified in both mice and humans, including the regulatory B10 cell subset that produces the inhibitory cytokine interleukin-10 (IL-10). B-cell derived IL-10 can protect against autoimmune disease in mice. Here, Thomas Tedder and colleagues identify IL-21- and CD-40-dependent cognate interactions with T cells as key drivers for the generation of IL-10-producing CD5+ regulatory B cells. Transfer of in vitro-expanded regulatory B cells is shown to suppress signs of disease in the experimental autoimmune encephalomyelitis mouse model of multiple sclerosis. This work suggests a novel strategy for the treatment of severe autoimmune diseases for which effective therapies are not available. B cells regulate immune responses by producing antigen-specific antibodies1. However, specific B-cell subsets can also negatively regulate T-cell immune responses, and have been termed regulatory B cells2,3,4. Human and mouse regulatory B cells (B10 cells) with the ability to express the inhibitory cytokine interleukin-10 (IL-10) have been identified2,3,4,5. Although rare, B10 cells are potent negative regulators of antigen-specific inflammation and T-cell-dependent autoimmune diseases in mice5,6,7. How B10-cell IL-10 production and regulation of antigen-specific immune responses are controlled in vivo without inducing systemic immunosuppression is unknown. Using a mouse model for multiple sclerosis, here we show that B10-cell maturation into functional IL-10-secreting effector cells that inhibit in vivo autoimmune disease requires IL-21 and CD40-dependent cognate interactions with T cells. Moreover, the ex vivo provision of CD40 and IL-21 receptor signals can drive B10-cell development and expansion by four-million-fold, and generate B10 effector cells producing IL-10 that markedly inhibit disease symptoms when transferred into mice with established autoimmune disease. The ex vivo expansion and reinfusion of autologous B10 cells may provide a novel and effective in vivo treatment for severe autoimmune diseases that are resistant to current therapies.

EAE is a mouse T cell–mediated autoimmune disease of the CNS used to model the human condition MS. The contributions of B cells to EAE initiation and progression are unclear. In this study, we have shown that EAE disease initiation and progression are differentially influenced by the depletion of B cells from mice with otherwise intact immune systems. CD20 antibody–mediated B cell depletion before EAE induction substantially exacerbated disease symptoms and increased encephalitogenic T cell influx into the CNS. Increased symptom severity resulted from the depletion of a rare IL-10–producing CD1d^hiCD5⁺ regulatory B cell subset (B10 cells), since the adoptive transfer of splenic B10 cells before EAE induction normalized EAE in B cell–depleted mice. While transfer of regulatory B10 cells was maximally effective during early EAE initiation, they had no obvious role during disease progression. Rather, B cell depletion during EAE disease progression dramatically suppressed symptoms. Specifically, B cells were required for the generation of CD4⁺ T cells specific for CNS autoantigen and the entry of encephalitogenic T cells into the CNS during disease progression. These results demonstrate reciprocal regulatory roles for B cells during EAE immunopathogenesis. The therapeutic effect of B cell depletion for the treatment of autoimmunity may therefore depend on the relative contributions and the timing of these opposing B cell activities during the course of disease initiation and pathogenesis.

Anti-CD20 antibody immunotherapy effectively treats non-Hodgkin's lymphoma and autoimmune disease. However, the cellular and molecular pathways for B cell depletion remain undefined because human mechanistic studies are limited. Proposed mechanisms include antibody-, effector cell–, and complement-dependent cytotoxicity, the disruption of CD20 signaling pathways, and the induction of apoptosis. To identify the mechanisms for B cell depletion in vivo, a new mouse model for anti-CD20 immunotherapy was developed using a panel of twelve mouse anti–mouse CD20 monoclonal antibodies representing all four immunoglobulin G isotypes. Anti-CD20 antibodies rapidly depleted the vast majority of circulating and tissue B cells in an isotype-restricted manner that was completely dependent on effector cell Fc receptor expression. B cell depletion used both FcγRI- and FcγRIII-dependent pathways, whereas B cells were not eliminated in FcR common γ chain–deficient mice. Monocytes were the dominant effector cells for B cell depletion, with no demonstrable role for T or natural killer cells. Although most anti-CD20 antibodies activated complement in vitro, B cell depletion was completely effective in mice with genetic deficiencies in C3, C4, or C1q complement components. That the innate monocyte network depletes B cells through FcγR-dependent pathways during anti-CD20 immunotherapy has important clinical implications for anti-CD20 and other antibody-based therapies.

Autoimmunity and inflammation are controlled in part by regulatory B cells, including a recently identified IL-10-competent CD1d^highCD5⁺ B cell subset termed B10 cells that represents 1–3% of adult mouse spleen B cells. In this study, pathways that influence B10 cell generation and IL-10 production were identified and compared with previously described regulatory B cells. IL-10-competent B cells were predominantly CD1d^highCD5⁺ in adult spleen and were the prevalent source of IL-10, but not other cytokines. B10 cell development and/or maturation in vivo required Ag receptor diversity and intact signaling pathways, but not T cells, gut-associated flora, or environmental pathogens. Spleen B10 cell frequencies were significantly expanded in aged mice and mice predisposed to autoimmunity, but were significantly decreased in mouse strains that are susceptible to exogenous autoantigen-induced autoimmunity. LPS, PMA, plus ionomycin stimulation in vitro for 5 h induced B10 cells to express cytoplasmic IL-10. However, prolonged LPS or CD40 stimulation (48 h) induced additional adult spleen CD1d^highCD5⁺ B cells to express IL-10 following PMA plus ionomycin stimulation. Prolonged LPS or CD40 stimulation of newborn spleen and adult blood or lymph node CD1d^low and/or CD5⁻ B cells also induced cytoplasmic IL-10 competence in rare B cells, with CD40 ligation uniformly inducing CD5 expression. IL-10 secretion was induced by LPS signaling through MyD88-dependent pathways, but not following CD40 ligation. LPS stimulation also induced rapid B10 cell clonal expansion when compared with other spleen B cells. Thereby, both adaptive and innate signals regulate B10 cell development, maturation, CD5 expression, and competence for IL-10 production.