RNA sequencing of single allergen-specific memory B cells after grass pollen immunotherapy: Two unique cell fates and CD29 as a biomarker for treatment effect.

BACKGROUND: Sublingual immunotherapy (SLIT) for grass pollen allergy can modify the natural history of allergic rhinitis and is associated with increased allergen-specific IgG4 . IgG4 competitively inhibits functional IgE on the surface of effector cells, such as mast cells and basophils, from binding to allergens. To further understand the important role memory B-cell (Bmem) responses play in mediating the beneficial effects of SLIT, we assessed changes in allergen-specific Bmem subsets induced by SLIT for grass pollen allergy. METHODS: Blood samples were collected twice outside the pollen season from twenty-seven patients with sensitization to ryegrass pollen (RGP; Lolium perenne) and seasonal rhinoconjunctivitis. Thirteen received 4-month pre-seasonal SLIT for grass pollen allergy, and 14 received standard pharmacotherapy only. Single-cell RNA sequencing was performed on FACS-purified Lol p 1-specific Bmem before and after SLIT from four patients, and significant genes were validated by flow cytometry on the total cohort. RESULTS: Four months of SLIT increased RGP-specific IgE and IgG4 in serum and induced two Lol p 1-specific Bmem subsets with unique transcriptional profiles. Both subsets had upregulated expression of beta 1 integrin ITGB1 (CD29), whereas IGHE (IgE), IGHG4 (IgG4 ), FCER2 (CD23), and IL13RA1 were upregulated in one subset. There was an increase in the proportion of Lol p 1+ Bmem expressing surface IgG4 , CD23, and CD29 after SLIT. CONCLUSIONS: A clinically successful 4 months course of SLIT for grass pollen allergy induces two transcriptionally unique Bmem fates. Associated changes in surface-expressed proteins on these Bmem subsets can be used as early biomarkers for treatment effects.

Delivery of Acetate to the Peripheral Blood after Consumption of Foods High in Short-Chain Fatty Acids.

SCOPE: To promote local and systemic benefits of short-chain fatty acids (SCFA), methods of increasing their delivery to the gastrointestinal tract are needed. SCFA in foods and beverages represents a poorly characterized source. Main aims of this study are: 1) quantify SCFA in commonly consumed foods and beverages, and 2) explore the pharmacokinetics of consuming oral SCFA from dietary sources. METHODS AND RESULTS: Gas-chromatography coupled to flame ionization detection is used measure SCFA in 38 commonly consumed foods and beverages. Acetate is the most abundant SCFA detected, with kombucha and vinegar found to provide >1000 mg of acetate per serve. An acute pharmacokinetic study is conducted in 10 participants. Acetate is stable across the 2-h sampling period after consumption of a control drink, with consumption of a vinegar drink containing 25 mmol acetate significantly increasing plasma acetate concentration after 60 min and increasing acetate delivery to the blood upon assessment of the area under the pharmacokinetic curve. CONCLUSION: Fermented foods and beverages are a natural source of dietary SCFA that acutely deliver SCFA to the blood. If systemic delivery is needed for immunological and metabolic effects to occur, these may be achieved if delivered over a longer period of time.

Induction of IgG2 and IgG4 B-cell memory following sublingual immunotherapy for ryegrass pollen allergy.

BACKGROUND: While treatment for atopic rhinitis is aimed mostly to relieve symptoms, only allergen-specific immunotherapy (AIT) is targeted to modify the natural history of allergic diseases. This results in sustained clinical tolerance, even when treatment has stopped. The immunomodulatory effects of AIT are attributed mainly to increased regulatory T-cell function and increased allergen-specific IgG4 , yet little is known about the effect on the memory B-cell compartment. OBJECTIVE: We aimed to examine the effects of AIT on the IgE- and IgG subclass-expressing memory B cells. METHODS: We recruited 29 patients with atopic seasonal rhinoconjunctivitis and performed a longitudinal analysis of the peripheral immune compartment before, during, and after sublingual immunotherapy (SLIT) for allergy to temperate grass pollen, predominantly to ryegrass pollen (RGP; Lolium perenne). Using flow cytometry on peripheral blood mononuclear cells and serum immunoassays, we analyzed the effects of a 4 months preseasonal treatment regimen comprising two or three courses in consecutive years on circulating IgE+ and IgG+ memory B cells and allergen-specific Ig levels. RESULTS: SLIT increased RGP-specific serum IgG2 and IgG4 , as well as the frequencies of IgG2+ and IgG4+ memory B cells, whereas no effect was observed on the IgE+ memory B-cell compartment. Furthermore, SLIT enhanced proportions of regulatory T cells specific to RGP. These changes were associated with clinical improvement. CONCLUSION: Our data provide evidence for immunological effects of SLIT on B-cell memory. Skewing responses toward IgG2 and IgG4 subclasses might be a mechanism to suppress IgE-mediated allergic responses.

An Artemis polymorphic variant reduces Artemis activity and confers cellular radiosensitivity.

Artemis is required for V(D)J recombination and the repair of a subset of radiation-induced DNA double strand breaks (DSBs). Artemis-null patients display radiosensitivity (RS) and severe combined immunodeficiency (SCID), classified as RS-SCID. Strongly impacting hypomorphic Artemis mutations confer marked infant immunodeficiency and a predisposition for EBV-associated lymphomas. Here, we provide evidence that a polymorphic Artemis variant (c.512C > G: p.171P > R), which has a world-wide prevalence of 15%, is functionally impacting. The c.512C > G mutation causes an approximately 3-fold decrease in Artemis endonuclease activity in vitro. Cells derived from a patient who expressed a single Artemis allele with the polymorphic mutational change, showed radiosensitivity and a DSB repair defect in G2 phase, with Artemis cDNA expression rescuing both phenotypes. The c.512C > G change has an additive impact on Artemis function when combined with a novel C-terminal truncating mutation (p.436C > X), which also partially inactivates Artemis activity. Collectively, our findings provide strong evidence that monoallelic expression of the c.512C > G variant impairs Artemis function causing significant radiosensitivity and a G2 phase DSB repair defect. The patient exhibiting monoallelic c.512C > G-Artemis expression showed immunodeficiency only in adulthood, developed bilateral carcinoma of the nipple and myelodysplasia raising the possibility that modestly decreased Artemis function can impact clinically.