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Alveolar Macrophages from House Dust Mite-Tolerant Mice Induce Foxp3+ Regulatory T Cells in an IL-10-Independent Manner
AB70 Abstracts
SATURDAY
220
Alveolar Macrophages from House Dust MiteTolerant Mice Induce Foxp3+ Regulatory T Cells in an IL-10-Independent Manner
Sonali J. Bracken, Alexander J. Adami, Linda A. Guernsey, Robert R. Clark, Evan R. Jellison, Craig R. Schramm, and Roger S. Thrall; University of Connecticut Health, Farmington, CT. RATIONALE: Alveolar macrophages (AMs) are critical for maintaining immune tolerance in the lung. Previous studies have shown that naive AMs can induce Foxp3+ regulatory T cells (Tregs) but lose this ability when initially exposed to inhaled allergens, thereby promoting the development of asthma. The purpose of this study was to determine whether long-term exposure to inhaled house dust mite (HDM) improves the ability of AMs to induce Tregs. METHODS: We have previously established that short-term (2 wk) HDM exposure induces allergic airway disease (AAD) whereas long-term (11 wk) exposure promotes tolerance to HDM (i.e. suppression of allergic inflammation with increases in pulmonary Tregs and IL-10+ AMs). AMs were sorted from the lungs of HDM-AAD (2 wk) and HDM-tolerant (11 wk) C57BL/6 mice and co-cultured with na€ıve CD4+Foxp3- T cells in the presence or absence of exogenous TGF-b and IL-10/IL-10R blockade for 72 hours. The percentage of induced Foxp3+ Tregs was then examined. RESULTS: AMs from HDM-tolerant mice induced significantly more Tregs than HDM-AAD mice (31% vs 14%, p <0.0001), suggesting that AMs adopt regulatory functions following long-term allergen exposure. This effect was dependent upon the addition of exogenous TGF-b and was not reversed following IL-10/IL-10R blockade (p 5 0.15). CONCLUSIONS: These findings suggest that the generation of regulatory AMs following long-term HDM exposure may play an important role in the suppression of HDM-induced asthma through induction of Tregs. RNA sequencing studies are currently underway to determine the players involved in regulatory AM-mediated Treg induction, as these cells are a promising immunotherapeutic target for asthma.
221
Airway Exposure to Staphylococcus Aureus Particles Promotes Development of AntigenSpecific Th2-Type Immunity to Inhaled Antigens through Production of Interleukin-1 Alpha (IL-1 alpha)
Kenichiro Hara; Gunma University Graduate School of Medicne, Department of Respiratory Medicine, Maebashi, Japan. RATIONALE: We have already reported that airway exposure to Staphylococcus aureus (S aureus) particles promotes development of ovalbumin (OVA)-specific Th2-type immunity to inhaled antigens through production of thymic stromal lymphopoietin (TSLP) in mice model of chronic bronchial allergic inflammation. In early phase, airway administration of S.aureus increased bronchoalveolar lavage (BAL) levels of IL-1a and IL-1b. Therefore, we tested the hypothesis that airway exposure to S.aureus facilitates Th2-type immune responses through IL-1a in the airway. METHODS: S aureus particles or recombinant IL-1a were administered intranasally (i.n.) to airways of na€ıve BALb/c mice or IL-1 receptor-1 knockout mice (IL-1R1KO). Alternatively, S aureus particles or IL-1a were adminstered together with endotoxin-free OVA (i.e. chronic exposure) on days 0 and 7, and mice were then exposed to OVA alone i.n. on days 21,22 and 23. RESULTS: Airway administration of S aureus particles to na€ıve mice increased BAL levels of IL-1a, and then did them of TSLP in early phase. In the chronic model, airway exposure to OVA alone did not sensitize na€ıve mice. In contrast, na€ıve mice treated with both IL-1a and OVA developed OVA-specific Th2-type immune responses in a TSLP-dependent manner, when exposed to OVA alone, they developed airway eosinophilia, peribronchial hypertrophy, and increased BAL levels of IL-5 and IL-13. Moreover, airway Th2 response to OVA was significantly inhibited in IL1R1KO mice. CONCLUSIONS: Airway exposure to S aureus promotes development of Th2-type immune responses to inhaled antigens through production of IL-a. Inhibition of IL-1a may be effective in the treatment of patients with allergic asthma.
J ALLERGY CLIN IMMUNOL FEBRUARY 2017
222
Tr1 Cell Identification and Phenotype in Children with and without Food Allergy
Jenna R. Bergerson, MD, MPH1, Kristin Erickson2, and Anne Marie Singh, MD3; 1Northwestern University, Division of Allergy and Immunology, chicago, IL; Division of Allergy-Immunology, Department of Pediatrics, Ann & Robert H. Lurie Children’s Hospital of Chicago, Chicago, IL, 2Northwestern University Feinberg School of Medicine, Chicago, IL, 3Division of Allergy & Immunology, Department of Pediatrics, Ann & Robert H. Lurie Children’s Hospital of Chicago, Northwestern University Feinberg School of Medicine, Chicago, IL; Northwestern Feinberg School of Medicine, Chicago, IL. RATIONALE: Tr1 cells are a subtype of peripherally induced regulatory T cells and are thought to be pivotal in promoting and maintaining tolerance. As early life is a critical period for establishing oral tolerance, we sought to investigate the role of food allergy diagnosis and age on Tr1 cells in children with and without FA. METHODS: Peripheral blood mononuclear cells were isolated from 30 pediatric patients and analyzed using flow cytometry. Populations of Tr1 cells (CD4+CD45RA-Lag3+CD49b+) were determined in children with and without FA, and compared by Mann-Whitney test. RESULTS: Overall, the percentages of Tr1 were significantly higher (0.57 v. 0.39, p50.023) in healthy children compared to food allergic children. When examining the effect of age, percentages of Tr1 cells were significantly higher (0.55 v. 0.18, p5 0.038) in young healthy children <6 years old compared to food allergic children <6 years old. In addition, healthy children <6 years old had significantly more CD4+CD45RALag3+CD49b+ cells that express the putative gut homing marker CCR6+ than their peers with FA (0.28 v. 0.08, p50.0041). CONCLUSIONS: Overall, children without FA have a higher percentage of Tr1 cells than food allergic children, and younger control children have significantly more Tr1 cells. Furthermore, gut-homing Tr1 cells in younger children may be important in the development of oral tolerance. These data suggest that Tr1 cells are important in oral tolerance, particularly in younger children.
223
Follicular Helper T (Tfh) Cells Mediate Peanut Allergy
Takao Kobayashi, PhD1, Joseph J. Dolence, PhD2, Koji Iijima, PhD2, and Hirohito Kita, MD2; 1Mayo Clinic Rochester, Rochester, MN, 2 Mayo Clinic, Rochester, MN. RATIONALE: While peanut allergy is a major medical problem, the immunologic mechanisms of the disease have not been fully understood. Follicular helper T (Tfh) cells are specialized in supporting B cell maturation and antibody production. The goal of this study was to investigate the roles for Tfh cells in the development of peanut allergy by using mouse models. METHODS: To induce peanut allergy, na€ıve BALB/c mice were exposed intranasally (i.n.) to peanut flour for 4 weeks. FACS analyses, in vitro cell culture and gene-deficient mice were used to dissect the immunologic mechanisms. RESULTS: When na€ıve mice were exposed i.n. to peanut flour, they developed peanut-specific IgE antibody in 2 weeks, which continued to rise for 4 weeks. Intraperitoneal challenge of these mice with peanut extract induced clinical signs of systemic anaphylaxis, including itching, reduced motility, and hypothermia. In IL-4 reporter mice, a large number of IL-4competent Tfh cells and a smaller number of Th2 cells were identified in the draining lymph nodes. Tfh cells produced large quantities of IL-4 and IL-21 in vitro. Furthermore, TCRb-deficient mice transferred with Tfh cells produced >25x higher titers of peanut-specific IgE as compared to those transferred with Th2 cells. Finally, CD4-specific Bcl6 conditional knockout mice that are deficient in Tfh cells (but Th2-cell compartment is intact) were totally protected from developing peanut-specific IgE antibody and anaphylactic responses CONCLUSIONS: Tfh cells, but unlikely Th2 cells, mediate production of specific IgE antibody in peanut allergy. Tfh cells may serve as a potential therapeutic target for food allergy.
SATURDAY
220
Alveolar Macrophages from House Dust MiteTolerant Mice Induce Foxp3+ Regulatory T Cells in an IL-10-Independent Manner
Sonali J. Bracken, Alexander J. Adami, Linda A. Guernsey, Robert R. Clark, Evan R. Jellison, Craig R. Schramm, and Roger S. Thrall; University of Connecticut Health, Farmington, CT. RATIONALE: Alveolar macrophages (AMs) are critical for maintaining immune tolerance in the lung. Previous studies have shown that naive AMs can induce Foxp3+ regulatory T cells (Tregs) but lose this ability when initially exposed to inhaled allergens, thereby promoting the development of asthma. The purpose of this study was to determine whether long-term exposure to inhaled house dust mite (HDM) improves the ability of AMs to induce Tregs. METHODS: We have previously established that short-term (2 wk) HDM exposure induces allergic airway disease (AAD) whereas long-term (11 wk) exposure promotes tolerance to HDM (i.e. suppression of allergic inflammation with increases in pulmonary Tregs and IL-10+ AMs). AMs were sorted from the lungs of HDM-AAD (2 wk) and HDM-tolerant (11 wk) C57BL/6 mice and co-cultured with na€ıve CD4+Foxp3- T cells in the presence or absence of exogenous TGF-b and IL-10/IL-10R blockade for 72 hours. The percentage of induced Foxp3+ Tregs was then examined. RESULTS: AMs from HDM-tolerant mice induced significantly more Tregs than HDM-AAD mice (31% vs 14%, p <0.0001), suggesting that AMs adopt regulatory functions following long-term allergen exposure. This effect was dependent upon the addition of exogenous TGF-b and was not reversed following IL-10/IL-10R blockade (p 5 0.15). CONCLUSIONS: These findings suggest that the generation of regulatory AMs following long-term HDM exposure may play an important role in the suppression of HDM-induced asthma through induction of Tregs. RNA sequencing studies are currently underway to determine the players involved in regulatory AM-mediated Treg induction, as these cells are a promising immunotherapeutic target for asthma.
221
Airway Exposure to Staphylococcus Aureus Particles Promotes Development of AntigenSpecific Th2-Type Immunity to Inhaled Antigens through Production of Interleukin-1 Alpha (IL-1 alpha)
Kenichiro Hara; Gunma University Graduate School of Medicne, Department of Respiratory Medicine, Maebashi, Japan. RATIONALE: We have already reported that airway exposure to Staphylococcus aureus (S aureus) particles promotes development of ovalbumin (OVA)-specific Th2-type immunity to inhaled antigens through production of thymic stromal lymphopoietin (TSLP) in mice model of chronic bronchial allergic inflammation. In early phase, airway administration of S.aureus increased bronchoalveolar lavage (BAL) levels of IL-1a and IL-1b. Therefore, we tested the hypothesis that airway exposure to S.aureus facilitates Th2-type immune responses through IL-1a in the airway. METHODS: S aureus particles or recombinant IL-1a were administered intranasally (i.n.) to airways of na€ıve BALb/c mice or IL-1 receptor-1 knockout mice (IL-1R1KO). Alternatively, S aureus particles or IL-1a were adminstered together with endotoxin-free OVA (i.e. chronic exposure) on days 0 and 7, and mice were then exposed to OVA alone i.n. on days 21,22 and 23. RESULTS: Airway administration of S aureus particles to na€ıve mice increased BAL levels of IL-1a, and then did them of TSLP in early phase. In the chronic model, airway exposure to OVA alone did not sensitize na€ıve mice. In contrast, na€ıve mice treated with both IL-1a and OVA developed OVA-specific Th2-type immune responses in a TSLP-dependent manner, when exposed to OVA alone, they developed airway eosinophilia, peribronchial hypertrophy, and increased BAL levels of IL-5 and IL-13. Moreover, airway Th2 response to OVA was significantly inhibited in IL1R1KO mice. CONCLUSIONS: Airway exposure to S aureus promotes development of Th2-type immune responses to inhaled antigens through production of IL-a. Inhibition of IL-1a may be effective in the treatment of patients with allergic asthma.
J ALLERGY CLIN IMMUNOL FEBRUARY 2017
222
Tr1 Cell Identification and Phenotype in Children with and without Food Allergy
Jenna R. Bergerson, MD, MPH1, Kristin Erickson2, and Anne Marie Singh, MD3; 1Northwestern University, Division of Allergy and Immunology, chicago, IL; Division of Allergy-Immunology, Department of Pediatrics, Ann & Robert H. Lurie Children’s Hospital of Chicago, Chicago, IL, 2Northwestern University Feinberg School of Medicine, Chicago, IL, 3Division of Allergy & Immunology, Department of Pediatrics, Ann & Robert H. Lurie Children’s Hospital of Chicago, Northwestern University Feinberg School of Medicine, Chicago, IL; Northwestern Feinberg School of Medicine, Chicago, IL. RATIONALE: Tr1 cells are a subtype of peripherally induced regulatory T cells and are thought to be pivotal in promoting and maintaining tolerance. As early life is a critical period for establishing oral tolerance, we sought to investigate the role of food allergy diagnosis and age on Tr1 cells in children with and without FA. METHODS: Peripheral blood mononuclear cells were isolated from 30 pediatric patients and analyzed using flow cytometry. Populations of Tr1 cells (CD4+CD45RA-Lag3+CD49b+) were determined in children with and without FA, and compared by Mann-Whitney test. RESULTS: Overall, the percentages of Tr1 were significantly higher (0.57 v. 0.39, p50.023) in healthy children compared to food allergic children. When examining the effect of age, percentages of Tr1 cells were significantly higher (0.55 v. 0.18, p5 0.038) in young healthy children <6 years old compared to food allergic children <6 years old. In addition, healthy children <6 years old had significantly more CD4+CD45RALag3+CD49b+ cells that express the putative gut homing marker CCR6+ than their peers with FA (0.28 v. 0.08, p50.0041). CONCLUSIONS: Overall, children without FA have a higher percentage of Tr1 cells than food allergic children, and younger control children have significantly more Tr1 cells. Furthermore, gut-homing Tr1 cells in younger children may be important in the development of oral tolerance. These data suggest that Tr1 cells are important in oral tolerance, particularly in younger children.
223
Follicular Helper T (Tfh) Cells Mediate Peanut Allergy
Takao Kobayashi, PhD1, Joseph J. Dolence, PhD2, Koji Iijima, PhD2, and Hirohito Kita, MD2; 1Mayo Clinic Rochester, Rochester, MN, 2 Mayo Clinic, Rochester, MN. RATIONALE: While peanut allergy is a major medical problem, the immunologic mechanisms of the disease have not been fully understood. Follicular helper T (Tfh) cells are specialized in supporting B cell maturation and antibody production. The goal of this study was to investigate the roles for Tfh cells in the development of peanut allergy by using mouse models. METHODS: To induce peanut allergy, na€ıve BALB/c mice were exposed intranasally (i.n.) to peanut flour for 4 weeks. FACS analyses, in vitro cell culture and gene-deficient mice were used to dissect the immunologic mechanisms. RESULTS: When na€ıve mice were exposed i.n. to peanut flour, they developed peanut-specific IgE antibody in 2 weeks, which continued to rise for 4 weeks. Intraperitoneal challenge of these mice with peanut extract induced clinical signs of systemic anaphylaxis, including itching, reduced motility, and hypothermia. In IL-4 reporter mice, a large number of IL-4competent Tfh cells and a smaller number of Th2 cells were identified in the draining lymph nodes. Tfh cells produced large quantities of IL-4 and IL-21 in vitro. Furthermore, TCRb-deficient mice transferred with Tfh cells produced >25x higher titers of peanut-specific IgE as compared to those transferred with Th2 cells. Finally, CD4-specific Bcl6 conditional knockout mice that are deficient in Tfh cells (but Th2-cell compartment is intact) were totally protected from developing peanut-specific IgE antibody and anaphylactic responses CONCLUSIONS: Tfh cells, but unlikely Th2 cells, mediate production of specific IgE antibody in peanut allergy. Tfh cells may serve as a potential therapeutic target for food allergy.