Activin and transforming growth factor-β signaling pathways are activated after allergen challenge in mild asthma Harsha H. Kariyawasam, MBBS, PhD, Sophie.

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Activin and transforming growth factor-β signaling pathways are activated after allergen challenge in mild asthma Harsha H. Kariyawasam, MBBS, PhD, Sophie Pegorier, PhD, Julia Barkans, Georgina Xanthou, PhD, Maxine Aizen, Sun Ying, MD, PhD, A. Barry Kay, MD, PhD, Clare M. Lloyd, PhD, Douglas S. Robinson, MD Journal of Allergy and Clinical Immunology Volume 124, Issue 3, Pages 454-462 (September 2009) DOI: 10.1016/j.jaci.2009.06.022 Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 1 pSmad2 expression at baseline and postallergen. A, pSmad2 expression in airway epithelium in response to allergen challenge at the 24-hour time point is summarized. B and C, Representative photomicrographs from a paired volunteer at baseline and 24 hours postallergen (×20 magnification). D, Representative photomicrograph of activin-A expression in the airway (×40). E, Colocalization (brown via DAB) to elastase+ neutrophils (Fast Red; ×100). BM, Basement membrane. Journal of Allergy and Clinical Immunology 2009 124, 454-462DOI: (10.1016/j.jaci.2009.06.022) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 2 TGF-β and activin receptor modulation after allergen provocation in asthma. Epithelial ALK-5 (A) and ALK-1 (B) (submucosa, C) expression before and after allergen challenge is summarized. Submucosal cell expression of TβRII is also presented (D). ALK-4 (E and F) and ActRIIA (G and H) expression is summarized. Epithelial cells are expressed as cells/mm BM and submucosal cells as cells per mm2. BM, Basement membrane. Journal of Allergy and Clinical Immunology 2009 124, 454-462DOI: (10.1016/j.jaci.2009.06.022) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 3 Receptor modulation in response to allergen challenge. Representative photomicrographs of ALK-5 expression (A and B), ALK-1 (C and D), ALK-4 (E and F), and ActRIIA (G and H) at baseline and postallergen (×40 magnification). Submucosal cells expressing ALK-1, ALK-4, and ActRIIA are arrowed. Journal of Allergy and Clinical Immunology 2009 124, 454-462DOI: (10.1016/j.jaci.2009.06.022) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 4 ALK-1 and ALK-4 modulation of expression CD3+ T cells postallergen. Numbers of T cells expressing receptors for TGF-β (ALK-1, A) and activin-A (ALK-4, B) were enumerated before and after allergen challenge by double-staining for CD3 and ALK-1 or ALK-4. Counts are expressed as cells per mm2 biopsy. C, A representative photomicrograph for double-staining for CD3 (red) and ALK-4 (DAB). Double-stained cells are darker red-brown. The epithelium is indicated as Ep to orientate the section. N = 6. Journal of Allergy and Clinical Immunology 2009 124, 454-462DOI: (10.1016/j.jaci.2009.06.022) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 5 Effect of activin-A on human bronchial epithelial cells. A, Activin-A caused dose-dependent proliferation of NHBE cells in culture. Data are for 24 hours of culture and are means and SEs for 5 separate experiments. B, TNF-α induced-dose dependent release of activin-A from NHBE cells (24-hour time point; means and SEs; n = 5). C, IL-13 at 10 ng/mL induced dose-dependent release of IL-8 from NHBE cells, and this was significantly increased in the presence of 100 ng follistatin (24-hour time point; n = 5). D and E, TNF-α alone did not induce release of CXCL10/IP-10 or CCL2/MCP-1 from NHBE cells, but this was significantly induced in the presence of the activin-inhibitor follistatin at 100 ng/mL (n = 5). ∗P <.05 for comparison with medium-alone values. CT, Control. Journal of Allergy and Clinical Immunology 2009 124, 454-462DOI: (10.1016/j.jaci.2009.06.022) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions