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Human mast cells arise from a common circulating progenitor

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1 Human mast cells arise from a common circulating progenitor
Katariina Maaninka, BSc, Jani Lappalainen, MSc, Petri T. Kovanen, MD, PhD  Journal of Allergy and Clinical Immunology  Volume 132, Issue 2, Pages e3 (August 2013) DOI: /j.jaci Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

2 Fig 1 Expression of neutral granule proteases in developing human peripheral blood progenitor–derived MCs. Expression of tryptase, chymase, CPA3, cathepsin G, and granzyme B in developing peripheral blood progenitor–derived MCs represented as percentages of protease-positive MCs (left), mRNA levels (middle), and cellular contents (right). Data are means ± SD from cells derived from at least 4 donors. Journal of Allergy and Clinical Immunology  , e3DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

3 Fig 2 Expression of δ- and γ-tryptase in developing human peripheral blood progenitor–derived MCs. Expression of tryptase isoforms is represented as fold increases relative to week 1. Data are means ± SD from cells derived from 4 donors. Journal of Allergy and Clinical Immunology  , e3DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

4 Fig 3 Quadrant plots of flow cytometric analysis presenting tryptase and chymase expression in human peripheral blood progenitor–derived MCs at week 3 of culture. Quadrant plots were created to evaluate the proportion of KIT-tryptase double-positive cells (A, D), KIT-chymase double-positive cells (B, E), and tryptase-chymase double-positive cells (C, F) cultured with KITLG alone (Fig 3, A-C) or with KITLG, IL-3, and IL-9 (Fig 3, D-F) for 3 weeks. Fluorescence thresholds for positive staining were set by using isotype-matched immunoglobulins. Data are representative of cells derived from 4 donors. Journal of Allergy and Clinical Immunology  , e3DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

5 Fig 4 Neutral granule protease content of developing human peripheral blood progenitor–derived MCs at week 3 of culture. Cellular levels of tryptase, chymase, CPA3, cathepsin G, and granzyme B in 3-week-old peripheral blood progenitor–derived MCs cultured with KITLG alone or with KITLG, IL-3, and IL-9. Data are means ± SD from cells derived from 3 donors. Journal of Allergy and Clinical Immunology  , e3DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

6 Fig 5 Quadrant plots of flow cytometric analysis presenting chymase expression in human peripheral blood progenitor–derived MCs at weeks 4 and 9 of culture in the absence and presence of serum. Quadrant plots were created to evaluate the proportion of KIT-chymase double-positive cells (A, C, E, G) and tryptase-chymase double-positive cells (B, D, F, H) cultured with KITLG alone (Fig 5, A, B, E, F) or with KITLG and 10% fetal bovine serum (Fig 5, C, D, G, H) for 4 and 9 weeks. Fluorescence thresholds for positive staining were set by using isotype-matched immunoglobulins. Data are representative of cells derived from 3 donors. Journal of Allergy and Clinical Immunology  , e3DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

7 Fig E1 Immunofluorescent staining of human peripheral blood progenitor–derived MCs at 9 weeks of culture. MCs stained positively with mouse mAbs against tryptase and chymase. A fluorochrome-conjugated goat antimouse Alexa-488 IgG was used as the secondary antibody, and nuclei were stained with DAPI. Staining with mouse immunoglobulin was used as a negative control. DAPI, 4′,6-Diamidino-2-phenylindole. Journal of Allergy and Clinical Immunology  , e3DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

8 Fig E2 Cellular heparin and heparin-to-protease ratio in developing human peripheral blood progenitor–derived MCs. Heparin content and heparin-to-protease ratios in developing peripheral blood progenitor–derived MCs at weeks 4 to 9 of culture. Data are means ± SD from cells derived from 4 donors. Journal of Allergy and Clinical Immunology  , e3DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

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