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Evidence for altered levels of IgD in the nasal airway mucosa of patients with chronic rhinosinusitis  Jin-Young Min, MD, PhD, Jayakar V. Nayak, MD, PhD,

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Presentation on theme: "Evidence for altered levels of IgD in the nasal airway mucosa of patients with chronic rhinosinusitis  Jin-Young Min, MD, PhD, Jayakar V. Nayak, MD, PhD,"— Presentation transcript:

1 Evidence for altered levels of IgD in the nasal airway mucosa of patients with chronic rhinosinusitis  Jin-Young Min, MD, PhD, Jayakar V. Nayak, MD, PhD, Kathryn E. Hulse, PhD, Whitney W. Stevens, MD, PhD, Paul A. Raju, PhD, Julia H. Huang, MS, Lydia A. Suh, BS, Griet A. Van Roey, PhD, James E. Norton, MS, Roderick G. Carter, BS, Caroline P.E. Price, BA, Ava R. Weibman, BA, Ali R. Rashan, MD, Eliver E. Ghosn, PhD, Zara M. Patel, MD, Tetsuya Homma, MD, PhD, David B. Conley, MD, Kevin C. Welch, MD, Stephanie Shintani-Smith, MD, MS, Anju T. Peters, MD, Leslie C. Grammer, MD, Kathleen E. Harris, BS, Atsushi Kato, PhD, Peter H. Hwang, MD, Robert C. Kern, MD, Leonore A. Herzenberg, PhD, Robert P. Schleimer, PhD, Bruce K. Tan, MD, MS  Journal of Allergy and Clinical Immunology  Volume 140, Issue 6, Pages e5 (December 2017) DOI: /j.jaci Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

2 Fig 1 sIgD levels in nasal tissue of patients with CRSsNP. Protein levels of sIgD in nasal tissue (A), nasal lavage fluid (B), and serum (C) were measured by using ELISA. Dotted line indicates the threshold based on the 95th percentile of IgD levels in UT from control subjects. Individual patient data points are shown, and solid lines represent means ± SEMs. *P < .05, Kruskal-Wallis test. Journal of Allergy and Clinical Immunology  , e5DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

3 Fig 2 Immunofluorescence analysis of IgD+ cells in nasal tissue using phycoerythrin-conjugated anti-IgD (red). A-D, Representative images of IgD+ cells in UT from control subjects (Fig 2, A), patients with CRSsNP (Fig 2, B), and patients with CRSwNP (Fig 2, C) and NPs from patients with CRSwNP (Fig 2, D; magnification ×200). E-H, IgD+ cells at the periglandular (Fig 2, E) and subepithelial areas in UT from patients with CRSsNP (Fig 2, F) and periglandular (Fig 2, G) and subepithelial (Fig 2, H) areas in NPs of patients with CRSwNP (magnification ×400). Nuclei were counterstained with 4,6-diamidino-2-phenylindole (blue). I, IgD+ cells in nasal tissue were counted semiquantitatively. J, Correlation between numbers of IgD+ cells from immunofluorescence assay and IgD protein concentrations in matched nasal tissue. *P < .05. HPF, High-power field. Journal of Allergy and Clinical Immunology  , e5DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

4 Fig 3 Representative phenotype of IgD+ plasmablasts in nasal tissue, as assessed by using flow cytometry. Of the CD19+CD20− B-cell population, IgD+CD38bright cells (IgD+ plasmablasts) and IgD+CD38− cells (naive B cells) were subdivided, and each population was further analyzed for CD27 (A and G), IgM (B and H), CD138 (C and I), intracellular IgD (D and J), BLIMP-1 (E and K), and forward scatter (FSC) and side scatter (SSC) profiles (F and L). Journal of Allergy and Clinical Immunology  , e5DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

5 Fig 4 Representative flow cytometric plots showing the IgD+CD38bright plasmablast population and calculated numbers of IgD+CD38bright cells in nasal tissue from control subjects, patients with CRSsNP, and those with CRSwNP (A) and in peripheral blood from the same patients (B). **P < .01, Kruskal-Wallis test. Numbers in flow cytometric plots indicate relative percentages. Journal of Allergy and Clinical Immunology  , e5DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

6 Fig 5 Correlation between levels of IgD and IL-2 in nasal tissue in vivo. Protein levels of IL-2 in tissue were measured by using multiplex immunoassay, and IgD levels were measured by using ELISA. Correlation was assessed by using the Spearman rank test. Journal of Allergy and Clinical Immunology  , e5DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

7 Fig 6 sIgD levels in ex vivo tissue explant culture. Whole tissue explants were cultured with IL-2 (60 U/mL) for 4 days, and sIgD levels in supernatants were measured by using ELISA. *P < .05, Kruskal-Wallis test. Journal of Allergy and Clinical Immunology  , e5DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

8 Fig 7 Preoperative antibiotic use and presence of pathogenic bacterial infection can influence IgD levels in nasal tissue. A, History of preoperative antibiotic use was obtained from electronic medical records. B, Nasal swab cultures were performed before sinus surgery. IgD levels in UT were measured by using ELISA. Dot plots illustrate individual data points, and solid lines represent means ± SEMs. *P < .05. Journal of Allergy and Clinical Immunology  , e5DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

9 Fig E1 Representative gating strategy used for flow cytometric analysis. A and B, Collected cells were gated for live leukocytes (Aqua−CD45+; Fig E1, A) and then gated based on forward scatter A (FSC-A) versus FSC-H for monodispersed cells or singlets (Fig E1, B). C, Next, cells were gated out based on a dump channel indicating positive staining for T cells (CD3+), monocytes (CD14+), granulocytes (CD16+), and natural killer cells (CD56+). D, The remaining cells (CD3−CD14−CD16−CD56−) were segmented into CD19+/CD20−, CD19+/CD20+, and CD19−/CD20− populations. Further characterization for IgD+ B cells was mined in the CD19+CD20− population. Journal of Allergy and Clinical Immunology  , e5DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

10 Fig E2 Levels of inflammatory mediators in nasal tissue. mRNA expression of IL-2 was quantitated by real-time PCR (A) and protein levels of IL-2 (B) and IL-10 (C) were measured by using multiplex immunoassay. D, BAFF mRNA expression was quantitated by real-time PCR. Dot plots illustrate individual data points, and solid lines represent means ± SEMs. *P < .05 and **P < .01. Journal of Allergy and Clinical Immunology  , e5DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

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