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Group 2 Innate Lymphoid Cells Directly Induce B Cell Activation in Humans
1
Group 2 Innate Lymphoid Cells Directly Induce B Cell Activation in Humans
Richard Kasjanski, MS1, Atsushi Kato, PhD1, Julie A. Poposki, MS1, Bruce S. Bochner, MD FAAAAI1, Yun Cao, BSc1, James E. Norton, MS1, Lydia Suh, BSc1, Roderick G. Carter, BSc1, Robert C. Kern, MD2, Stephanie S. Smith, MD2, David B. Conley, MD2, Anju T. Peters, MD1, Leslie C. Grammer, MD1, Whitney W. Stevens, MD PhD1, Kathleen E. Harris, BSc1, Bruce Tan, MD2, Robert P. Schleimer, PhD1, Kathryn E. Hulse, PhD3; 1Department of Medicine, Division of AllergyImmunology, Northwestern University Feinberg School of Medicine, Chicago, IL, 2Department of Otolaryngology, Northwestern University Feinberg School of Medicine, Chicago, IL, 3Division of Allergy-Immunology, Department of Medicine, Northwestern University Feinberg School of Medicine, Chicago, IL. RATIONALE: Chronic rhinosinusitis with nasal polyps (CRSwNP) is characterized by type 2 inflammation and increased group 2 innate lymphoid cells (ILC2). We have also found elevated B cells and overexpression of the extrafollicular B cell marker Epstein-Barr virusinduced protein 2 (EBI2). We investigated whether B cells in polyps expressed EBI2, indicating an extrafollicular phenotype, and whether ILC2s could regulate B cell activation. METHODS: EBI2 expression on cells from polyps of CRSwNP patients or tonsils from non-CRS patients was assessed by flow cytometry. Antibody production by tissue B cells was assessed by Luminex assay. In other experiments, B cells and ILC2s were isolated from peripheral blood by magnetic bead- and flow cytometry-based methods, respectively. B cells were cultured for 5 days under IgG-promoting conditions (IL-2 and R848 (a TLR7/8 agonist)), or IgE-promoting conditions (IL-4 and antiCD40), or with autologous ILC2s at a 1:1 ratio, or with IL-2 alone. RESULTS: Polyps contained elevated levels of plasmablasts (PB) and EBI2+ PB compared to tonsil (>5 fold, p<0.001 and >2.5 fold, p<0.01, respectively). In vitro, polyps also produced significantly higher levels of antibodies (p<0.01). IL-4 and anti-CD40 increased B cell survival and EBI2 expression (>5 fold), indicating in vitro development of extrafollicular PB, while IL-2 and R848 had no effect on EBI2. Co-culture of B cells with ILC2s increased B cell survival and promoted a significant increase of EBI2+ B cells (>5 fold, p<0.05). CONCLUSIONS: These findings suggest that type 2 inflammation, particularly ILC2s, may play an important role in B cell activation during chronic airway inflammation.
2
Novel IL-9-Producing Mucosal Mast Cells Promote IgEMediated Food Allergy
Yui-Hsi Wang, PhD; Cincinnati Children’s Hospital Medical Center, Cincinnati, OH. RATIONALE: IgE-mediated food allergy is manifested by an overactive T-helper 2 immune response to dietary antigens in the gastrointestinal tract. Frequency of newly identified IL-9-producing mast cells (MMC9s) correlates positively with levels of intestinal mastocytosis. We hypothesize that induction of MMC9s is a pivotal step to acquire the susceptibility to IgE-mediated food allergy. METHODS: Involvements of MMC9s in regulating susceptibility to food allergy were examined in mouse models of food allergy. Expression levels of MMC9-associated transcripts were analyzed in duodenal biopsies of atopic patients developed food allergy. RESULTS: MMC9s secrete prodigious amounts of IL-9 and mast cell protease-1 in response to IL-33 and antigen/IgE complex crosslinking, respectively. By producing significant amounts of IL-9, MMC9s cause pronounced intestinal mastocytosis and the production of mast cells. Repeated intragastric antigen challenge induces significant accumulations of both MMC9s and CD4+TH2 cells, which correlate positively with symptoms and susceptibility of murine strains to develop experimental food allergy. The development of MMC9s requires the presence of CD4+TH2 cells and the proteins of IL-4 and STAT6. Mice ablated or deficient of
MMC9 induction fail to develop intestinal mastocytosis, which resulted in decreased food allergy symptoms that can be restored by adoptively transferred MMC9s. Finally, atopic patients that develop food allergy display increased intestinal expression of Il9 and MC-specific transcripts. CONCLUSIONS: MMC9s, the primary cellular source of IL-9, function to amplify intestinal allergic inflammation and perpetuates anaphylactic response to allergenic dietary proteins. The acquisition of MMC9s may be a key cellular checkpoint for the susceptibility to develop IgE-mediated food allergy.
3
Follicular Helper T (Tfh) Cells Are Indispensable for IgE Antibody Responses to Airborne Allergens
Takao Kobayashi, PhD1, Koji Iijima, PhD1, Chien-Chang Chen1, Alexander L. Dent, PhD2, Hirohita Kita, MD1; 1Mayo Clinic, Rochester, MN, 2Indiana University, Indianapolis, IN. RATIONALE: Th2 cells have long been believed to play a pivotal role in regulating IgE antibody production. A new T cell subset, Tfh cells, is specialized in supporting B cell maturation and antibody production. The goal of this project was to investigate the roles of Tfh cells in allergic immune responses by using mouse models. METHODS: Na€ıve mice were exposed intranasally to natural allergens. Development of allergic immune responses was analyzed by collecting draining lymph nodes (LNs) and sera and by challenging mice with an antigen. RESULTS: Airway exposure of IL-4 reporter 4get mice to natural allergens, such as Alternaria and house dust mite, induced IL-4-competent Tfh and Th2 cells in various frequencies. Exposure of wild-type mice to Alternaria spiked with ovalbumin (OVA) induced OVA-specific IgE in the sera, and cells from LNs produced type 2 cytokines (i.e. IL-5, IL-13) when cultured with OVA. ICOS knockout and CD4-specific Bcl6 conditional knockout mice (both of which are deficient in Tfh cells while Th2 cells remain intact) developed significantly reduced serum levels of IgE antibody; however, LN cells from these mice produced robust type 2 cytokines when cultured with OVA. In contrast, OX40L knockout mice were deficient in type 2 cytokine responses and eosinophilic airway inflammation, but they developed levels of IgE antibody comparable to those of wild-type mice. CONCLUSIONS: Tfh cells, but unlikely Th2 cells, are indispensable for IgE antibody production to airborne allergens. Type 2 cytokine responses and IgE antibody responses can be regulated independently by two different subsets of CD4+ T cells.
FRIDAY
Abstracts AB1
J ALLERGY CLIN IMMUNOL VOLUME 137, NUMBER 2
Group 2 Innate Lymphoid Cells Directly Induce B Cell Activation in Humans
Richard Kasjanski, MS1, Atsushi Kato, PhD1, Julie A. Poposki, MS1, Bruce S. Bochner, MD FAAAAI1, Yun Cao, BSc1, James E. Norton, MS1, Lydia Suh, BSc1, Roderick G. Carter, BSc1, Robert C. Kern, MD2, Stephanie S. Smith, MD2, David B. Conley, MD2, Anju T. Peters, MD1, Leslie C. Grammer, MD1, Whitney W. Stevens, MD PhD1, Kathleen E. Harris, BSc1, Bruce Tan, MD2, Robert P. Schleimer, PhD1, Kathryn E. Hulse, PhD3; 1Department of Medicine, Division of AllergyImmunology, Northwestern University Feinberg School of Medicine, Chicago, IL, 2Department of Otolaryngology, Northwestern University Feinberg School of Medicine, Chicago, IL, 3Division of Allergy-Immunology, Department of Medicine, Northwestern University Feinberg School of Medicine, Chicago, IL. RATIONALE: Chronic rhinosinusitis with nasal polyps (CRSwNP) is characterized by type 2 inflammation and increased group 2 innate lymphoid cells (ILC2). We have also found elevated B cells and overexpression of the extrafollicular B cell marker Epstein-Barr virusinduced protein 2 (EBI2). We investigated whether B cells in polyps expressed EBI2, indicating an extrafollicular phenotype, and whether ILC2s could regulate B cell activation. METHODS: EBI2 expression on cells from polyps of CRSwNP patients or tonsils from non-CRS patients was assessed by flow cytometry. Antibody production by tissue B cells was assessed by Luminex assay. In other experiments, B cells and ILC2s were isolated from peripheral blood by magnetic bead- and flow cytometry-based methods, respectively. B cells were cultured for 5 days under IgG-promoting conditions (IL-2 and R848 (a TLR7/8 agonist)), or IgE-promoting conditions (IL-4 and antiCD40), or with autologous ILC2s at a 1:1 ratio, or with IL-2 alone. RESULTS: Polyps contained elevated levels of plasmablasts (PB) and EBI2+ PB compared to tonsil (>5 fold, p<0.001 and >2.5 fold, p<0.01, respectively). In vitro, polyps also produced significantly higher levels of antibodies (p<0.01). IL-4 and anti-CD40 increased B cell survival and EBI2 expression (>5 fold), indicating in vitro development of extrafollicular PB, while IL-2 and R848 had no effect on EBI2. Co-culture of B cells with ILC2s increased B cell survival and promoted a significant increase of EBI2+ B cells (>5 fold, p<0.05). CONCLUSIONS: These findings suggest that type 2 inflammation, particularly ILC2s, may play an important role in B cell activation during chronic airway inflammation.
2
Novel IL-9-Producing Mucosal Mast Cells Promote IgEMediated Food Allergy
Yui-Hsi Wang, PhD; Cincinnati Children’s Hospital Medical Center, Cincinnati, OH. RATIONALE: IgE-mediated food allergy is manifested by an overactive T-helper 2 immune response to dietary antigens in the gastrointestinal tract. Frequency of newly identified IL-9-producing mast cells (MMC9s) correlates positively with levels of intestinal mastocytosis. We hypothesize that induction of MMC9s is a pivotal step to acquire the susceptibility to IgE-mediated food allergy. METHODS: Involvements of MMC9s in regulating susceptibility to food allergy were examined in mouse models of food allergy. Expression levels of MMC9-associated transcripts were analyzed in duodenal biopsies of atopic patients developed food allergy. RESULTS: MMC9s secrete prodigious amounts of IL-9 and mast cell protease-1 in response to IL-33 and antigen/IgE complex crosslinking, respectively. By producing significant amounts of IL-9, MMC9s cause pronounced intestinal mastocytosis and the production of mast cells. Repeated intragastric antigen challenge induces significant accumulations of both MMC9s and CD4+TH2 cells, which correlate positively with symptoms and susceptibility of murine strains to develop experimental food allergy. The development of MMC9s requires the presence of CD4+TH2 cells and the proteins of IL-4 and STAT6. Mice ablated or deficient of
MMC9 induction fail to develop intestinal mastocytosis, which resulted in decreased food allergy symptoms that can be restored by adoptively transferred MMC9s. Finally, atopic patients that develop food allergy display increased intestinal expression of Il9 and MC-specific transcripts. CONCLUSIONS: MMC9s, the primary cellular source of IL-9, function to amplify intestinal allergic inflammation and perpetuates anaphylactic response to allergenic dietary proteins. The acquisition of MMC9s may be a key cellular checkpoint for the susceptibility to develop IgE-mediated food allergy.
3
Follicular Helper T (Tfh) Cells Are Indispensable for IgE Antibody Responses to Airborne Allergens
Takao Kobayashi, PhD1, Koji Iijima, PhD1, Chien-Chang Chen1, Alexander L. Dent, PhD2, Hirohita Kita, MD1; 1Mayo Clinic, Rochester, MN, 2Indiana University, Indianapolis, IN. RATIONALE: Th2 cells have long been believed to play a pivotal role in regulating IgE antibody production. A new T cell subset, Tfh cells, is specialized in supporting B cell maturation and antibody production. The goal of this project was to investigate the roles of Tfh cells in allergic immune responses by using mouse models. METHODS: Na€ıve mice were exposed intranasally to natural allergens. Development of allergic immune responses was analyzed by collecting draining lymph nodes (LNs) and sera and by challenging mice with an antigen. RESULTS: Airway exposure of IL-4 reporter 4get mice to natural allergens, such as Alternaria and house dust mite, induced IL-4-competent Tfh and Th2 cells in various frequencies. Exposure of wild-type mice to Alternaria spiked with ovalbumin (OVA) induced OVA-specific IgE in the sera, and cells from LNs produced type 2 cytokines (i.e. IL-5, IL-13) when cultured with OVA. ICOS knockout and CD4-specific Bcl6 conditional knockout mice (both of which are deficient in Tfh cells while Th2 cells remain intact) developed significantly reduced serum levels of IgE antibody; however, LN cells from these mice produced robust type 2 cytokines when cultured with OVA. In contrast, OX40L knockout mice were deficient in type 2 cytokine responses and eosinophilic airway inflammation, but they developed levels of IgE antibody comparable to those of wild-type mice. CONCLUSIONS: Tfh cells, but unlikely Th2 cells, are indispensable for IgE antibody production to airborne allergens. Type 2 cytokine responses and IgE antibody responses can be regulated independently by two different subsets of CD4+ T cells.
FRIDAY
Abstracts AB1
J ALLERGY CLIN IMMUNOL VOLUME 137, NUMBER 2