Acute systemic lupus erythematosus (SLE) courses with surges of antibody-secreting cells (ASCs) whose origin, diversity and contribution to serum autoantibodies remain unknown. Here, deep sequencing, ...proteomic profiling of autoantibodies and single-cell analysis demonstrated highly diversified ASCs punctuated by clones expressing the variable heavy-chain region VH4-34 that produced dominant serum autoantibodies. A fraction of ASC clones contained autoantibodies without mutation, a finding consistent with differentiation outside the germinal centers. A substantial ASC segment was derived from a distinct subset of newly activated naive cells of considerable clonality that persisted in the circulation for several months. Thus, selection of SLE autoreactivities occurred during polyclonal activation, with prolonged recruitment of recently activated naive B cells. Our findings shed light on the pathogenesis of SLE, help explain the benefit of agents that target B cells and should facilitate the design of future therapies.
B-1 B cells derive from a developmental program distinct from that of conventional B cells, through B cell receptor (BCR)-dependent positive selection of fetally derived precursors. Here, we used ...direct labeling of B cells reactive with the N-acetyl-D-glucosamine (GlcNAc)-containing Lancefield group A carbohydrate of Streptococcus pyogenes to study the effects of bacterial antigens on the emergent B-1 B cell clonal repertoire. The number, phenotype, and BCR clonotypes of GlcNAc-reactive B-1 B cells were modulated by neonatal exposure to heat-killed S. pyogenes bacteria. GlcNAc-reactive B-1 clonotypes and serum antibodies were reduced in germ-free mice compared with conventionally raised mice. Colonization of germ-free mice with a conventional microbiota promoted GlcNAc-reactive B-1 B cell development and concomitantly elicited clonally related IgA+ plasma cells in the small intestine. Thus, exposure to microbial antigens in early life determines the clonality of the mature B-1 B cell repertoire and ensuing antibody responses, with implications for vaccination approaches and schedules.
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•GAC-reactive B cells exhibit the phenotype and localization patterns of B-1 B cells•Neonatal S. pyogenes immunization expands minor IGHV7-3 GAC-reactive BCR clonotypes•Establishment of IGHV6-3 GAC-reactive B cell clonal dominance is microbiota dependent•Microbiota-dependent expansion of GAC-reactive B cells seeds the SI LP with ASCs
Innate-like B-1 B cells express canonical B cell receptors and originate through specialized developmental programs. Through immunoglobulin gene sequencing of single antigen-reactive B cells, New et al. demonstrate a significant role for environmental antigen-dependent selection events in shaping the GAC-reactive B-1 B cell clonal repertoire.
Understanding antibody repertoires and in particular, the properties and fates of B cells expressing potentially pathogenic antibodies is critical to define the mechanisms underlying multiple ...immunological diseases including autoimmune and allergic conditions as well as transplant rejection. Moreover, an integrated knowledge of the antibody repertoires expressed by B cells and plasma cells (PC) of different functional properties and longevity is essential to develop new therapeutic strategies, better biomarkers for disease segmentation, and new assays to measure restoration of B‐cell tolerance or, at least, of normal B‐cell homeostasis. Reaching these goals, however, will require a more precise phenotypic, functional and molecular definition of B‐cell and PC populations, and a comprehensive analysis of the antigenic reactivity of the antibodies they express. While traditionally hampered by technical and ethical limitations in human experimentation, new technological advances currently enable investigators to address these questions in a comprehensive fashion. In this review, we shall discuss these concepts as they apply to the study of Systemic Lupus Erythematosus.
Crohn’s disease and ulcerative colitis are characterized by dysregulated adaptive immune responses to the microbiota in genetically susceptible individuals, but the specificity of these responses ...remains largely undefined. Therefore, we developed a microbiota antigen microarray to characterize microbial antibody reactivity, particularly to human-derived microbiota flagellins, in inflammatory bowel disease.
Sera from healthy volunteers (n = 87) at the University of Alabama at Birmingham and from patients recruited from the Kirklin Clinic of University of Alabama at Birmingham Hospital, including patients with Crohn’s disease (n = 152) and ulcerative colitis (n = 170), were individually probed against microbiota bacterial flagellins of both mouse and human origin and analyzed for IgG and IgA antibody responses. Circulating flagellin-reactive T effector (CD4+CD154+) and T regulatory (CD4+CD137+) cells were isolated and evaluated in selected patients. Resulting adaptive immune responses were compared with corresponding clinical data to determine relevancy to disease behavior.
We show that patients with IBD express selective patterns of antibody reactivity to microbiota flagellins. Patients with Crohn’s disease, but not patients with ulcerative colitis, display augmented serum IgG to human ileal-localized Lachnospiraceae flagellins, with a subset of patients having high responses to more than 10 flagellins. Elevated responses to CBir1, a mouse Lachnospiraceae flagellin used clinically to diagnose CD, correlated with multi-Lachnospiraceae flagellin reactivity. In this subset of patients with CD, multi-flagellin reactivity was associated with elevated flagellin-specific CD154+CD45RA– T memory cells, a reduced ratio of flagellin-reactive CD4+ T regulatory to T effector cells, and a high frequency of disease complications.
Patients with Crohn’s disease display strong adaptive immune response to human-derived Lachnospiraceae flagellins, which may be targeted for prognosis and future personalized therapies.
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Patients with Crohn’s disease display strong adaptive immune responses to human intestinal bacterial flagellins, with a subset responding to multiple flagellins. These responses may be targeted for prognosis and personalized therapies.
Antibody responses to viral infections are sustained for decades by long-lived plasma cells (LLPCs). However, LLPCs have yet to be characterized in humans. Here we used CD19, CD38, and CD138 to ...identify four PC subsets in human bone marrow (BM). We found that the CD19(-)CD38(hi)CD138(+) subset was morphologically distinct, differentially expressed PC-associated genes, and exclusively contained PCs specific for viral antigens to which the subjects had not been exposed for more than 40 years. Protein sequences of measles- and mumps-specific circulating antibodies were encoded for by CD19(-)CD38(hi)CD138(+) PCs in the BM. Finally, we found that CD19(-)CD38(hi)CD138(+) PCs had a distinct RNA transcriptome signature and human immunoglobulin heavy chain (VH) repertoire that was relatively uncoupled from other BM PC subsets and probably represents the B cell response's "historical record" of antigenic exposure. Thus, our studies define human LLPCs and provide a mechanism for the life-long maintenance of anti-viral antibodies in the serum.
To advance our understanding and treatment of disease, research immunologists have been called-upon to place more centralized emphasis on impactful human studies. Such endeavors will inevitably ...require large-scale study execution and data management regulation ("Big Biology"), necessitating standardized and reliable metrics of immune status and function. A well-known example setting this large-scale effort in-motion is identifying correlations between eventual disease outcome and T lymphocyte phenotype in large HIV-patient cohorts using multiparameter flow cytometry. However, infection, immunodeficiency, and autoimmunity are also characterized by correlative and functional contributions of B lymphocytes, which to-date have received much less attention in the human Big Biology enterprise. Here, we review progress in human B cell phenotyping, analysis, and bioinformatics tools that constitute valuable resources for the B cell research community to effectively join in this effort.
Interleukin 4 (IL-4) plays a central role in the orchestration of Type 2 immunity. During T cell activation in the lymph node, IL-4 promotes Th2 differentiation and inhibits Th1 generation. In the ...inflamed tissue, IL-4 signals promote innate and adaptive Type-2 immune recruitment and effector function, positively amplifying the local Th2 response. In this study, we identify an additional negative regulatory role for IL-4 in limiting the recruitment of Th1 cells to inflamed tissues. To test IL-4 effects on inflammation subsequent to Th2 differentiation, we transiently blocked IL-4 during ongoing dermal inflammation (using anti-IL-4 mAb) and analyzed changes in gene expression. Neutralization of IL-4 led to the upregulation of a number of genes linked to Th1 trafficking, including CXCR3 chemokines, CCL5 and CCR5 and an associated increase in IFNγ, Tbet and TNFα genes. These gene expression changes correlated with increased numbers of IFNγ-producing CD4+ T cells in the inflamed dermis. Moreover, using an adoptive transfer approach to directly test the role of IL-4 in T cell trafficking to the inflamed tissues, we found IL-4 neutralization led to an early increase in Th1 cell recruitment to the inflamed dermis. These data support a model whereby IL-4 dampens Th1-chemokines at the site of inflammation limiting Th1 recruitment. To determine biological significance, we infected mice with Leishmania major, as pathogen clearance is highly dependent on IFNγ-producing CD4+ T cells at the infection site. Short-term IL-4 blockade in established L. major infection led to a significant increase in the number of IFNγ-producing CD4+ T cells in the infected ear dermis, with no change in the draining LN. Increased lymphocyte influx into the infected tissue correlated with a significant decrease in parasite number. Thus, independent of IL-4's role in the generation of immune effectors, IL-4 attenuates lymphocyte recruitment to the inflamed/infected dermis and limits pathogen clearance.
Lung-resident memory B cells (lung-BRMs) differentiate into plasma cells after reinfection, providing enhanced pulmonary protection. Here, we investigated the determinants of lung-BRM differentiation ...upon influenza infection. Kinetic analyses revealed that influenza nucleoprotein (NP)-specific BRMs preferentially differentiated early after infection and required T follicular helper (Tfh) cell help. BRM differentiation temporally coincided with transient interferon (IFN)-γ production by Tfh cells. Depletion of IFN-γ in Tfh cells prevented lung-BRM differentiation and impaired protection against heterosubtypic infection. IFN-γ was required for expression of the transcription factor T-bet by germinal center (GC) B cells, which promoted differentiation of a CXCR3+ GC B cell subset that were precursors of lung-BRMs and CXCR3+ memory B cells in the mediastinal lymph node. Absence of IFN-γ signaling or T-bet in GC B cells prevented CXCR3+ pre-memory precursor development and hampered CXCR3+ memory B cell differentiation and subsequent lung-BRM responses. Thus, Tfh-cell-derived IFN-γ is critical for lung-BRM development and pulmonary immunity, with implications for vaccination strategies targeting BRMs.
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•B6.Bcl6fl/flCd4cre/+ mice lack lung-resident memory B cells (BRMs)•Lung-BRMs fail to differentiate in bone marrow chimeras in which Tfh cells are Ifng−/−•IfngR1−/−, Tbx21−/−, and Stat1−/− B cells fail to differentiate into lung-BRMs•T-bet expression in GC B cells is required for the development of lung-BRM precursors
Lung-resident memory B cells (lung-BRMs) provide superior protection against respiratory viruses. Arroyo-Díaz et al. examine the requirements for lung-BRM development and reveal that IFN-γ production by Tfh cells is essential for the differentiation of memory B cell precursors that give rise to lung-BRMs and subsequent protection against heterosubtypic influenza infection.
Although viral infections elicit robust interferon-γ (IFN-γ) and long-lived antibody-secreting cell (ASC) responses, the roles for IFN-γ and IFN-γ-induced transcription factors (TFs) in ASC ...development are unclear. We showed that B cell intrinsic expression of IFN-γR and the IFN-γ-induced TF T-bet were required for T-helper 1 cell-induced differentiation of B cells into ASCs. IFN-γR signaling induced Blimp1 expression in B cells but also initiated an inflammatory gene program that, if not restrained, prevented ASC formation. T-bet did not affect Blimp1 upregulation in IFN-γ-activated B cells but instead regulated chromatin accessibility within the Ifng and Ifngr2 loci and repressed the IFN-γ-induced inflammatory gene program. Consistent with this, B cell intrinsic T-bet was required for formation of long-lived ASCs and secondary ASCs following viral, but not nematode, infection. Therefore, T-bet facilitates differentiation of IFN-γ-activated inflammatory effector B cells into ASCs in the setting of IFN-γ-, but not IL-4-, induced inflammatory responses.
•Differentiation of Th1-activated B (Be1) cells requires B intrinsic T-bet and IFN-γR•Be1 differentiation requires T-bet repression of NF-κB, TLR, and STAT pathways•Plasma cell formation in Be2 cultures and nematode infection is T-bet independent•T-bet controls formation of long-lived and secondary plasma cells in flu infection
T-bet-expressing B cells are expanded in autoimmunity and viral infection; however, whether these cells represent early plasma cells was unknown. Stone et al. identify T-bet-repressed pathways that permit plasma cell differentiation in the presence of IFN-γ. They show that T-bet-expressing B cells are required for long-lived immunity to flu.
Lipopolysaccharide (LPS) can either promote or prevent T helper 2 (Th2) cell allergic responses. However, the underlying mechanism remains unknown. We show here that LPS activity switches from ...pro-pathogenic to protective depending on the production of granulocyte-macrophage colony-stimulating factor (GM-CSF) by non-classical monocytes. In the absence of GM-CSF, LPS can favor pathogenic Th2 cell responses by supporting the trafficking of lung-migratory dendritic cells (mDC2s) into the lung-draining lymph node. However, when non-classical monocytes produce GM-CSF, LPS and GM-CSF synergize to differentiate monocyte-derived DCs from classical Ly6Chi monocytes that instruct mDC2s for Th2 cell suppression. Importantly, only allergens with cysteine protease activity trigger GM-CSF production by non-classical monocytes. Hence, the therapeutic effect of LPS is restricted to allergens with this enzymatic activity. Treatment with GM-CSF, however, restores the protective effects of LPS. Thus, GM-CSF produced by non-classical monocytes acts as a rheostat that fine-tunes the pathogenic and therapeutic functions of LPS.
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•LPS can promote or suppress Th2 cell sensitization depending on GM-CSF production•GM-CSF is produced by Ly6Clo monocytes in response to cysteine protease allergens•GM-CSF induces differentiation of LPS-sensitive moDCs that instruct for Th2 suppression•Without GM-CSF, Th2-dependent sensitization is favored due to the lack of moDCs
LPS can promote or prevent allergic sensitization. Kaur et al. demonstrate that GM-CSF segregates those different LPS-driven roles. Only allergens with cysteine protease activity trigger GM-CSF production by non-classical Ly6Clo monocytes, preventing sensitization to LPS-containing allergens by modulating the function of classical Ly6Chi monocytes and migratory dendritic cells.
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