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Type 2 innate lymphoid cells in induced sputum from children with severe asthma  Prasad Nagakumar, MBBS, Laura Denney, PhD, Louise Fleming, MD, Andrew.

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Presentation on theme: "Type 2 innate lymphoid cells in induced sputum from children with severe asthma  Prasad Nagakumar, MBBS, Laura Denney, PhD, Louise Fleming, MD, Andrew."— Presentation transcript:

1 Type 2 innate lymphoid cells in induced sputum from children with severe asthma 
Prasad Nagakumar, MBBS, Laura Denney, PhD, Louise Fleming, MD, Andrew Bush, MD, Clare M. Lloyd, PhD, Sejal Saglani, MD  Journal of Allergy and Clinical Immunology  Volume 137, Issue 2, Pages e6 (February 2016) DOI: /j.jaci Copyright © 2015 American Academy of Allergy, Asthma & Immunology Terms and Conditions

2 Fig 1 Patients with asthma (n = 11) have significantly more ILC2s in BAL and PBMCs than do children with LRTI (n = 16). PBMC data were available in 7 of 16 patients with LRTIs. A, Proportion of Lin− CD45+ cells in the lymphoid gate. B, Higher proportion of Lin− CD45+CD127+. C, Higher proportion of ILC2s (Lin− CD45+ CD127+CRTH2+) in patients with asthma than in patients with LRTIs. D, Higher proportion of CD3+CD4+ cells in BAL of patients with asthma than in patients with LRTIs. E and F, Representative histogram showing higher proportion of ILC2s in BAL of a 14-year-old patient with STRA than in age- and sex-matched patient with LRTIs. Horizontal bar represents median. Group differences were analyzed using Man-Whitney U test or Wilcoxon matched pair test. ns, Not significant. *P < .05, **P < .01, and ***P < .001. Journal of Allergy and Clinical Immunology  , e6DOI: ( /j.jaci ) Copyright © 2015 American Academy of Allergy, Asthma & Immunology Terms and Conditions

3 Fig 2 ILC2s and TH2 cells in induced sputum from patients with STRA (n = 13). A, Proportion of Lin−CD45+, Lin−CD45+CD127−, and Lin−CD127+ cells in induced sputum. Lin−CD127+ cells account for a small proportion of total Lin− cells. B, Proportion of sputum ILC2s in the lymphoid gate. C, Higher proportion of CD4+ cells compared with CD8+ cells. D, Significantly more TH2 cells (CD4+CRTH2+) than ILC2s in induced sputum. E, Significantly more ILC2s (Lin−CD45+CD127+CRTH2+) in sputum than in PBMCs in patients with severe asthma (n = 7). Differences between 2 groups detected by Mann-Whitney U test or Wilcoxon matched pair test. ns, Not significant. *P < .05, **P < .01, and ***P < .001. Journal of Allergy and Clinical Immunology  , e6DOI: ( /j.jaci ) Copyright © 2015 American Academy of Allergy, Asthma & Immunology Terms and Conditions

4 Fig E1 Gating strategy to identify ILCs in BAL. Representative flow cytometry plots from a patient with severe asthma. A, Forward and side scatter showing lymphoid gate. B, Viable CD45+cells are identified in the lymphoid gate using fixable viability stain and anti-CD45 mAb. C, CD45+CD3− cells gated from viable CD45+ cells to identify Lin− cells. D, Further characterization of Lin− cells (CD45+CD19−CD3−CD2CD14−CD19−CD56−) using lineage cocktail antibody. E, Showing viable Lin−CD45+ cells expressing CD127 and CRTH2 (ILC2). F and G, Isotope controls for CD127 and CRTH2. FSC-A, Forward-scattered light; SSC, Side-scattered light. Journal of Allergy and Clinical Immunology  , e6DOI: ( /j.jaci ) Copyright © 2015 American Academy of Allergy, Asthma & Immunology Terms and Conditions

5 Fig E2 Gating strategy to identify ILCs in sputum. Representative flow cytometry plots from a patient with severe asthma. A, Forward and side scatter showing lymphoid gate. B, Viable sputum cells identified using fixable viability stain. C, CD45+ cells gated from the viable population. D, Further characterization of Lin− (CD19−CD3−CD2−CD14−CD19−CD56−) CD45+ cells using lineage cocktail antibody. E, Lin− CD45+ sputum cells expressing CD127 and CRTH2. Journal of Allergy and Clinical Immunology  , e6DOI: ( /j.jaci ) Copyright © 2015 American Academy of Allergy, Asthma & Immunology Terms and Conditions

6 Fig E3 Gating strategy to identify T-lymphocyte subsets in sputum. Representative graphs from a patient with STRA. A, Forward and side scatter showing lymphocyte population. B, Viable cells identified using fixable viability stain. C, CD45+ cells gated from the viable population. D, CD3+ cells gated from the CD45+ cells. E, Population of CD3+CD4+ and CD3+CD8+ cells gated from the CD3+ cells. F, CD4+ cells expressing CRTH2 (TH2 cells). Journal of Allergy and Clinical Immunology  , e6DOI: ( /j.jaci ) Copyright © 2015 American Academy of Allergy, Asthma & Immunology Terms and Conditions

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