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Surface availability of beta-glucans is critical determinant of host immune response to Cladosporium cladosporioides  Rachael A. Mintz-Cole, PhD, Eric.

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Presentation on theme: "Surface availability of beta-glucans is critical determinant of host immune response to Cladosporium cladosporioides  Rachael A. Mintz-Cole, PhD, Eric."— Presentation transcript:

1 Surface availability of beta-glucans is critical determinant of host immune response to Cladosporium cladosporioides  Rachael A. Mintz-Cole, PhD, Eric B. Brandt, PhD, Stacey A. Bass, AS, Aaron M. Gibson, BS, Tiina Reponen, PhD, Gurjit K. Khurana Hershey, MD, PhD  Journal of Allergy and Clinical Immunology  Volume 132, Issue 1, Pages e2 (July 2013) DOI: /j.jaci Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

2 Fig 1 Heat killing of C cladosporioides prevents the development of murine asthma. A, BALB/c mice were exposed i.t. to 106 live or heat-killed C cladosporioides (C clad or C clad HK) spores 3 times a week for 3 weeks and 48 hours after the last challenge, AHR was measured by flexiVent. B, BALF was analyzed for total and differential counts. C, Total serum IgE. Data are representative of 3 independent experiments and are expressed as mean and SEM. n = 4 to 6 mice per group. Eos, Eosinophils; HK, heat killed; i.t., intratracheally; Lymp, lymphocytes; Mac, macrophages; Neut, neutrophils. *P < .05, **P < .01, and ***P < .001 as compared with saline. #P < .05 and ###P < .001 as compared with C clad HK. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

3 Fig 2 Induction of a TH17 response by heat-killed C cladosporioides (C clad HK). Contour plots (A) and quantification of intracellular cytokine staining for IL-13 or IL-17A in CD4+ lung cells (B). C, Levels of IL-17A and IL-4 in BALF. D, Relative levels of mRNA for IL-17A and IL-4 in whole lung. Data are representative of 3 independent experiments and are expressed as mean and SEM. n = 4 to 6 mice per group. HPRT, Hypoxanthine phosphoribosyltransferase. *P < .05, **P < .01, and ***P < .001 as compared with saline. #P < .05, ##P < .01, and ###P < .001 as compared with C clad HK. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

4 Fig 3 Heat-killed C cladosporioides have increased surface exposure of beta-glucans and binding to Dectin-1. A, Immunofluorescence images of beta-glucan staining taken at ×1000 for live and heat-killed C cladosporioides spores. Scale bar is 10 μm. B, Quantification of beta-glucan fluorescence intensity per spore. C, Number of spores bound to recombinant Dectin-1. Data are representative of 2 independent experiments and are expressed as mean and SEM. n = 3 samples per group. C clad HK, Heat-killed C cladosporioides. *P < .05. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

5 Fig 4 Heat-killed C cladosporioides prevents murine asthma in a Dectin-1–dependent manner. A, Wild-type C57Bl/6 or Dectin-1−/− mice were exposed to heat-killed C cladosporioides spores, and AHR was assessed 48 hours later. B, Total and differential counts of BALF. Data are representative from 2 independent experiments and are expressed as mean and SEM. n = 4 to 6 mice per group. C clad HK, Heat-killed C cladosporioides; Eos, eosinophils; KO, knockout; Lymp, lymphocytes; Mac, macrophages; Neut, neutrophils; WT, wild type. **P < .01 and ***P < .001 as compared with saline. ##P < .01 and ###P < .001 as compared with wild-type mice exposed to C cladosporioides. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

6 Fig 5 In the absence of Dectin-1, heat-killed C cladosporioides induces a TH2 response. Contour plots (A) and quantification of intracellular cytokine staining for IL-13 or IL-17A in CD4+ lung cells (B). C, Levels of IL-17A in BALF and mRNA in whole lung. D, Levels of IL-4 in BALF and mRNA in whole lung. Data are representative from 2 independent experiments and are expressed as mean and SEM. n = 4 to 6 mice per group. C clad HK, Heat-killed C cladosporioides; KO, knockout; WT, wild type. **P < .01 and ***P < .001 as compared with saline. #P < .05, ##P < .01, and ###P < .001 as compared with wild-type mice exposed to Ccladosporioides. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

7 Fig 6 Expression of TH17-skewing cytokines in the lungs is dependent on Dectin-1. A and C, Expression of IL-6 in the lungs of mice exposed to live (Fig 6, A) or heat-killed (Fig 6, C) C cladosporioides. B and D, Expression of TNF-α in the lungs of mice exposed to live (Fig 6, B) or heat-killed (Fig 6, D) C cladosporioides. Data are representative from 2 independent experiments and are expressed as mean and SEM. n = 4 to 6 mice per group. C clad HK, Heat-killed C cladosporioides; HPRT, hypoxanthine phosphoribosyltransferase; KO, knockout; WT, wild type. *P < .05, **P < .01, and ***P < .001 as compared with saline. #P < .05 and ###P < .001 as compared with Dectin-1−/− mice exposed to C cladosporioides. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

8 Fig 7 Proinflammatory cytokine production in BMDCs is dependent on Dectin-1. Expression of IL-6 (A), TNF-α (B), and IL-23 (C) in BMDCs. Level of IL-6 (D) and TNF-α (E) in culture supernatants of BMDCs. Data are representative of 3 independent experiments and are expressed as mean and SEM. n = 4 samples per group. C clad HK, Heat-killed C cladosporioides; HPRT, hypoxanthine phosphoribosyltransferase; KO, knockout; WT, wild type. *P < .05, **P < .01, and ***P < .001 as compared with saline. ##P < 0.01 and ###P < .001 as compared with Dectin-1−/− BMDCs exposed to C cladosporioides. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

9 Fig E1 A, Live cells gating with live dead stain (Invitrogen). B, Gating of lymphocytes in the live population based on forward scatter (FSC) and side scatter (SSC). C, CD4 gating from the lymphocyte population. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

10 Fig E2 C clad or C clad HK spores were stained with an AF488 dye (green) or not stained and then incubated with anti-beta glucan and a secondary antibody (red). C clad HK, Heat-killed C cladosporioides. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

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