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PGE2 Deficiency Causes a Phenotype Of Aspirin Sensitive Asthma That Depends On Platelets and Cysteinyl Leukotrienes
AB170 Abstracts
586
SUNDAY
IL33 and Type 2 Innate Lymphoid Cells (ILC2) But Not Th2 Cells Are Essential For Persistence Of Chronic Experimental Asthma Dr. Christina Christianson, PhD, Ms. Chaoyu Irvin, MS, Magdalena Gorska, MD, PhD, Rafeul Alam, MD, PhD, FAAAAI; National Jewish Health, Denver, CO. RATIONALE: We have developed a mouse model of chronic asthma where the disease persists longer than 6 months after cessation of allergen exposure. The objective was to delineate the contribution of IL33, Th2 and type 2 innate lymphoid cells (ILC2) to the persistence of asthma. METHODS: Chronic asthma was induced by semi-weekly intranasal exposures to dust mite, ragweed, and Aspergillus for six weeks. Mice underwent immune ablation by irradiation 3 weeks after the last allergen exposure and received bone marrow from na€ıve, Rag1-/- , or Rag2-/-gc-/mice. Airway hyperreactivity and immunohistological features were measured 6 weeks after irradiation with and without anti-IL33 treatment. Lung ILC2 and their contribution to total IL5 and IL13 production was measured by flow cytometry. The direct effect of IL33 on airways was studied 4 weeks after administration. RESULTS: Immune ablation eliminated allergen-specific T cell responses but failed to resolve airway hyperreactivity and remodeling. All features of chronic asthma completely resolved in mice that received bone marrow from Rag2-/-gc-/- (lacking T and B cells, and ILC) but not in mice that received marrow from Rag1-/- mice (lacking T and B cells only). Chronic asthma resulted in a 2.5 fold increase in ILC2 (linCD25+Sca1+c-kit+CRTH2+ST2+IL5+IL13+). Anti-IL33 inhibited ILC2 and induced resolution of asthma. Conversely, intranasal IL33 upregulated ILC2 and induced airway inflammation and hyperreactivity, persisting for 4 weeks. IL33 was autoinduced, partially facilitated by IL13. CONCLUSIONS: Airway epithelial cell-derived IL33 is critical for ILC2 generation. ILC2, not Th2 cells, are essential for persistence of chronic asthma in the absence of allergen exposure.
587
PGE2 Deficiency Causes a Phenotype Of Aspirin Sensitive Asthma That Depends On Platelets and Cysteinyl Leukotrienes Dr. Tao Liu, PhD1,2, Dr. Joshua A. Boyce, MD, FAAAAI1,2, Dr. Tanya M. Laidlaw, MD, FAAAAI1,2, Dr. Howard Katz, PhD1,2; 1Brigham and Women’s Hospital, Division of Rheumatology, Immunology and Allergy, Boston, MA, 2Harvard Medical School, Boston, MA. RATIONALE: Aspirin exacerbated respiratory disease (AERD) is characterized by asthma, tissue eosinophilia, overproduction of cysteinyl leukotrienes (cysLTs), and respiratory reactions to nonselective cyclooxygenase (COX) inhibitors. Ex vivo studies suggest that functional abnormalities of the COX-2/microsomal prostaglandin (PG)E2 synthase (mPGES)-1 system may underlie AERD. Previously we have demonstrated that mPGES-1-null (ptges-/-) mice develop a remarkably AERDlike phenotype in a model of eosinophilic pulmonary inflammation. We hypothesized that the AERD-like phenotype in mice depends on platelets and cysLTs. METHODS: ptges-/- mice were treated intranasally with low-dose of house dust mite extract on 6 occasions, and then were challenged with aerosol Lys-aspirin (Lys-ASA) and airway resistance was measured. Montelukast, zileuton, a T prostanoid (TP) receptor antagonist, or anti-CD41 (to deplete platelets) was given to some mice to determine their effects on Lys-ASA induced airway resistance (RL). Lung mast cell (MC) activation and cysLT production were measured by ELISA. RESULTS: Lys-ASA-challenged ptges-/- mice exhibit sustained increases in RL, along with lung MC activation and cysLT overproduction. The increases in RL and MC products were blocked by montelukast or zileuton, implying that bronchoconstriction and MC activation were both cysLT-dependent. Lys-ASA induced cysLT
J ALLERGY CLIN IMMUNOL FEBRUARY 2014
generation and MC activation depended on platelet adherent granulocytes and TP receptors. CONCLUSIONS: Lesions that impair the inducible generation of PGE2 remove control of platelet/granulocyte interactions and TP receptor-dependent cysLT production, permitting MC activation in response to COX-1 inhibition. The findings suggest applications of anti-platelet drugs or TP receptor antagonists for the treatment of AERD.
588
CCR8 Is a Receptor For CCL18 On Human Th2 Cells Dr. Morris Ling, MD1, Dr. Sabina Islam, MD1,2, Dr. John Leung, MD3, Wayne G. Shreffler, MD, PhD, FAAAAI1,2, Dr. Andrew D. Luster, MD, PhD4,5; 1Massachusetts General Hospital, Boston, MA, 2 Harvard Medical School, Boston, MA, 3Tufts Medical Center, Boston, MA, 4Massachusetts General Hospital, Charlestown, MA, 5Harvard Medical School, Charlestown, MA. RATIONALE: The CC chemokine ligand 18 (CCL18) is one of the most highly expressed chemokines in chronic allergic diseases. The study of CCL18 has been hindered by the lack of an identified chemokine receptor. METHODS: We performed chemotaxis assays and calcium flux assays using a CCR8-transfected mouse pre B-cell line. We also isolated and cultured na€ıve CD4+ T cells in Th2 polarizing conditions and performed chemotaxis assays, calcium flux assays, RT-qPCR, and flow cytometry with intracellular cytokine staining. Expression of chemokine and chemokine receptor mRNA in esophageal biopsy specimens from patients with eosinophilic esophagitis (EoE), EoE in remission, or GERD was quantified by RT-qPCR. RESULTS: CCL18 induced chemotaxis and calcium flux of human CCR8-transfected cells. Highly polarized human Th2 cells expressed CCR8, secreted IL-4 and IL-5, and exhibited chemotaxis and calcium flux to CCL18. Furthermore, CCL1, a known CCR8 ligand, and CCL18 induced heterologous cross-desensitization of human Th2 cells, suggesting that they share a common receptor. Expression of PITPNM3, a previously identified CCL18 receptor relevant in breast cancer metastasis, was not detected on human na€ıve T cells or Th2 cells by PCR or flow cytometry. CCL18 and CCR8 were coexpressed in esophageal biopsy tissue from individuals with active EoE and were present at markedly higher levels compared with tissue from patients with EoE in remission or controls with GERD. CONCLUSIONS: Identifying CCR8 as a chemokine receptor for CCL18 will help clarify the biological role of this highly expressed chemokine in human allergic diseases.
586
SUNDAY
IL33 and Type 2 Innate Lymphoid Cells (ILC2) But Not Th2 Cells Are Essential For Persistence Of Chronic Experimental Asthma Dr. Christina Christianson, PhD, Ms. Chaoyu Irvin, MS, Magdalena Gorska, MD, PhD, Rafeul Alam, MD, PhD, FAAAAI; National Jewish Health, Denver, CO. RATIONALE: We have developed a mouse model of chronic asthma where the disease persists longer than 6 months after cessation of allergen exposure. The objective was to delineate the contribution of IL33, Th2 and type 2 innate lymphoid cells (ILC2) to the persistence of asthma. METHODS: Chronic asthma was induced by semi-weekly intranasal exposures to dust mite, ragweed, and Aspergillus for six weeks. Mice underwent immune ablation by irradiation 3 weeks after the last allergen exposure and received bone marrow from na€ıve, Rag1-/- , or Rag2-/-gc-/mice. Airway hyperreactivity and immunohistological features were measured 6 weeks after irradiation with and without anti-IL33 treatment. Lung ILC2 and their contribution to total IL5 and IL13 production was measured by flow cytometry. The direct effect of IL33 on airways was studied 4 weeks after administration. RESULTS: Immune ablation eliminated allergen-specific T cell responses but failed to resolve airway hyperreactivity and remodeling. All features of chronic asthma completely resolved in mice that received bone marrow from Rag2-/-gc-/- (lacking T and B cells, and ILC) but not in mice that received marrow from Rag1-/- mice (lacking T and B cells only). Chronic asthma resulted in a 2.5 fold increase in ILC2 (linCD25+Sca1+c-kit+CRTH2+ST2+IL5+IL13+). Anti-IL33 inhibited ILC2 and induced resolution of asthma. Conversely, intranasal IL33 upregulated ILC2 and induced airway inflammation and hyperreactivity, persisting for 4 weeks. IL33 was autoinduced, partially facilitated by IL13. CONCLUSIONS: Airway epithelial cell-derived IL33 is critical for ILC2 generation. ILC2, not Th2 cells, are essential for persistence of chronic asthma in the absence of allergen exposure.
587
PGE2 Deficiency Causes a Phenotype Of Aspirin Sensitive Asthma That Depends On Platelets and Cysteinyl Leukotrienes Dr. Tao Liu, PhD1,2, Dr. Joshua A. Boyce, MD, FAAAAI1,2, Dr. Tanya M. Laidlaw, MD, FAAAAI1,2, Dr. Howard Katz, PhD1,2; 1Brigham and Women’s Hospital, Division of Rheumatology, Immunology and Allergy, Boston, MA, 2Harvard Medical School, Boston, MA. RATIONALE: Aspirin exacerbated respiratory disease (AERD) is characterized by asthma, tissue eosinophilia, overproduction of cysteinyl leukotrienes (cysLTs), and respiratory reactions to nonselective cyclooxygenase (COX) inhibitors. Ex vivo studies suggest that functional abnormalities of the COX-2/microsomal prostaglandin (PG)E2 synthase (mPGES)-1 system may underlie AERD. Previously we have demonstrated that mPGES-1-null (ptges-/-) mice develop a remarkably AERDlike phenotype in a model of eosinophilic pulmonary inflammation. We hypothesized that the AERD-like phenotype in mice depends on platelets and cysLTs. METHODS: ptges-/- mice were treated intranasally with low-dose of house dust mite extract on 6 occasions, and then were challenged with aerosol Lys-aspirin (Lys-ASA) and airway resistance was measured. Montelukast, zileuton, a T prostanoid (TP) receptor antagonist, or anti-CD41 (to deplete platelets) was given to some mice to determine their effects on Lys-ASA induced airway resistance (RL). Lung mast cell (MC) activation and cysLT production were measured by ELISA. RESULTS: Lys-ASA-challenged ptges-/- mice exhibit sustained increases in RL, along with lung MC activation and cysLT overproduction. The increases in RL and MC products were blocked by montelukast or zileuton, implying that bronchoconstriction and MC activation were both cysLT-dependent. Lys-ASA induced cysLT
J ALLERGY CLIN IMMUNOL FEBRUARY 2014
generation and MC activation depended on platelet adherent granulocytes and TP receptors. CONCLUSIONS: Lesions that impair the inducible generation of PGE2 remove control of platelet/granulocyte interactions and TP receptor-dependent cysLT production, permitting MC activation in response to COX-1 inhibition. The findings suggest applications of anti-platelet drugs or TP receptor antagonists for the treatment of AERD.
588
CCR8 Is a Receptor For CCL18 On Human Th2 Cells Dr. Morris Ling, MD1, Dr. Sabina Islam, MD1,2, Dr. John Leung, MD3, Wayne G. Shreffler, MD, PhD, FAAAAI1,2, Dr. Andrew D. Luster, MD, PhD4,5; 1Massachusetts General Hospital, Boston, MA, 2 Harvard Medical School, Boston, MA, 3Tufts Medical Center, Boston, MA, 4Massachusetts General Hospital, Charlestown, MA, 5Harvard Medical School, Charlestown, MA. RATIONALE: The CC chemokine ligand 18 (CCL18) is one of the most highly expressed chemokines in chronic allergic diseases. The study of CCL18 has been hindered by the lack of an identified chemokine receptor. METHODS: We performed chemotaxis assays and calcium flux assays using a CCR8-transfected mouse pre B-cell line. We also isolated and cultured na€ıve CD4+ T cells in Th2 polarizing conditions and performed chemotaxis assays, calcium flux assays, RT-qPCR, and flow cytometry with intracellular cytokine staining. Expression of chemokine and chemokine receptor mRNA in esophageal biopsy specimens from patients with eosinophilic esophagitis (EoE), EoE in remission, or GERD was quantified by RT-qPCR. RESULTS: CCL18 induced chemotaxis and calcium flux of human CCR8-transfected cells. Highly polarized human Th2 cells expressed CCR8, secreted IL-4 and IL-5, and exhibited chemotaxis and calcium flux to CCL18. Furthermore, CCL1, a known CCR8 ligand, and CCL18 induced heterologous cross-desensitization of human Th2 cells, suggesting that they share a common receptor. Expression of PITPNM3, a previously identified CCL18 receptor relevant in breast cancer metastasis, was not detected on human na€ıve T cells or Th2 cells by PCR or flow cytometry. CCL18 and CCR8 were coexpressed in esophageal biopsy tissue from individuals with active EoE and were present at markedly higher levels compared with tissue from patients with EoE in remission or controls with GERD. CONCLUSIONS: Identifying CCR8 as a chemokine receptor for CCL18 will help clarify the biological role of this highly expressed chemokine in human allergic diseases.