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T regulatory cells and IL-10 have been implicated in the mechanism of immunotherapy in patients with systemic anaphylaxis following bee stings. We studied the role of IL-10 in the induction of clinical, cellular, and humoral tolerance during immunotherapy for local mucosal allergy in subjects with seasonal pollinosis. Local and systemic IL-10 responses and serum Ab concentrations were measured before/after a double-blind trial of grass pollen (Phleum pratense, Phl P) immunotherapy. We observed local increases in IL-10 mRNA-positive cells in the nasal mucosa after 2 years of immunotherapy, but only during the pollen season. IL-10 protein-positive cells were also increased and correlated with IL-10 mRNA+ cells. These changes were not observed in placebo-treated subjects or in healthy controls. Fifteen and 35% of IL-10 mRNA signals were colocalized to CD3+ T cells and CD68+ macrophages, respectively, whereas only 1–2% of total CD3+ cells and 4% of macrophages expressed IL-10. Following immunotherapy, peripheral T cells cultured in the presence of grass pollen extract also produced IL-10. Immunotherapy resulted in blunting of seasonal increases in serum allergen Phl p 5-specific IgE, 60- to 80-fold increases in Phl p 5-specific IgG, and 100-fold increases in Phl p 5-specific IgG4. Post-immunotherapy serum exhibited inhibitory activity, which coeluted with IgG4, and blocked IgE-facilitated binding of allergen-IgE complexes to B cells. Both the increases in IgG and the IgG “blocking” activity correlated with the patients’ overall assessment of improvement. Thus, grass pollen immunotherapy may induce allergen-specific, IL-10-dependent “protective” IgG4 responses.

Despite their crucial role in health and disease, our knowledge of immune cells within human tissues remains limited. We surveyed the immune compartment of 16 tissues from 12 adult donors by single-cell RNA sequencing and VDJ sequencing generating a dataset of ~360,000 cells. To systematically resolve immune cell heterogeneity across tissues, we developed CellTypist, a machine learning tool for rapid and precise cell type annotation. Using this approach, combined with detailed curation, we determined the tissue distribution of finely phenotyped immune cell types, revealing hitherto unappreciated tissue-specific features and clonal architecture of T and B cells. Our multitissue approach lays the foundation for identifying highly resolved immune cell types by leveraging a common reference dataset, tissue-integrated expression analysis, and antigen receptor sequencing.

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BackgroundMost children with detectable peanut-specific IgE (P-sIgE) are not allergic to peanut. We addressed 2 non–mutually exclusive hypotheses for the discrepancy between allergy and sensitization: (1) differences in P-sIgE levels between children with peanut allergy (PA) and peanut-sensitized but tolerant (PS) children and (2) the presence of an IgE inhibitor, such as peanut-specific IgG4 (P-sIgG4), in PS patients.MethodsTwo hundred twenty-eight children (108 patients with PA, 77 PS patients, and 43 nonsensitized nonallergic subjects) were studied. Levels of specific IgE and IgG4 to peanut and its components were determined. IgE-stripped basophils or a mast cell line were used in passive sensitization activation and inhibition assays. Plasma of PS subjects and patients submitted to peanut oral immunotherapy (POIT) were depleted of IgG4 and retested in inhibition assays.ResultsBasophils and mast cells sensitized with plasma from patients with PA but not PS patients showed dose-dependent activation in response to peanut. Levels of sIgE to peanut and its components could only partially explain differences in clinical reactivity between patients with PA and PS patients. P-sIgG4 levels (P = .023) and P-sIgG4/P-sIgE (P < .001), Ara h 1–sIgG4/Ara h 1–sIgE (P = .050), Ara h 2–sIgG4/Ara h 2–sIgE (P = .004), and Ara h 3–sIgG4/Ara h 3–sIgE (P = .016) ratios were greater in PS children compared with those in children with PA. Peanut-induced activation was inhibited in the presence of plasma from PS children with detectable P-sIgG4 levels and POIT but not from nonsensitized nonallergic children. Depletion of IgG4 from plasma of children with PS (and POIT) sensitized to Ara h 1 to Ara h 3 partially restored peanut-induced mast cell activation (P = .007).ConclusionsDifferences in sIgE levels and allergen specificity could not justify the clinical phenotype in all children with PA and PS children. Blocking IgG4 antibodies provide an additional explanation for the absence of clinical reactivity in PS patients sensitized to major peanut allergens.

Protective humoral memory forms in secondary lymphoid organs where B cells undergo affinity maturation and differentiation into memory or plasma cells. Here, we provide a comprehensive roadmap of human B cell maturation with single-cell transcriptomics matched with bulk and single-cell antibody repertoires to define gene expression, antibody repertoires, and clonal sharing of B cell states at single-cell resolution, including memory B cell heterogeneity that reflects diverse functional and signaling states. We reconstruct gene expression dynamics during B cell activation to reveal a pre–germinal center state primed to undergo class switch recombination and dissect how antibody class–dependent gene expression in germinal center and memory B cells is linked with a distinct transcriptional wiring with potential to influence their fate and function. Our analyses reveal the dynamic cellular states that shape human B cell–mediated immunity and highlight how antibody isotype may play a role during their antibody-based selection.

Allergen immunotherapy (IT) has long-term efficacy in IgE-mediated allergic rhinitis and asthma. IT has been shown to modify lymphocyte responses to allergen, inducing IL-10 production and IgG Abs. In contrast, a putative role for IgA and local TGF-β-producing cells remains to be determined. In 44 patients with seasonal rhinitis/asthma, serum IgA1, IgA2, and polymeric (J chain-containing) Abs to the major allergen Phl p 5 were determined by ELISA before and after a 2-year double-blind trial of grass pollen (Phleum pratense) injection IT. Nasal TGF-β expression was assessed by in situ hybridization. Sera from five IT patients were fractionated for functional analysis of the effects of IgA and IgG Abs on IL-10 production by blood monocytes and allergen-IgE binding to B cells. Serum Phl p 5-specific IgA2 Abs increased after a 2-year treatment (∼8-fold increase, p = 0.002) in contrast to IgA1. Increases in polymeric Abs to Phl p 5 (∼2-fold increase, p = 0.02) and in nasal TGF-β mRNA (p = 0.05) were also observed, and TGF-β mRNA correlated with serum Phl p 5 IgA2 (r = 0.61, p = 0.009). Post-IT IgA fractions triggered IL-10 secretion by monocytes while not inhibiting allergen-IgE binding to B cells as observed with IgG fractions. This study shows for the first time that the IgA response to IT is selective for IgA2, correlates with increased local TGF-β expression, and induces monocyte IL-10 expression, suggesting that IgA Abs could thereby contribute to the tolerance developed in IT-treated allergic patients.