Hereditary angioedema: Key role for kallikrein and bradykinin in vascular endothelial- cadherin cleavage and edema formation Laurence Bouillet, MD, PhD,

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Hereditary angioedema: Key role for kallikrein and bradykinin in vascular endothelial- cadherin cleavage and edema formation Laurence Bouillet, MD, PhD, Tiphaine Mannic, PhD, Mélanie Arboleas, Mariela Subileau, Christian Massot, MD, Christian Drouet, PhD, Philippe Huber, PhD, Isabelle Vilgrain, PhD Journal of Allergy and Clinical Immunology Volume 128, Issue 1, Pages 232-234 (July 2011) DOI: 10.1016/j.jaci.2011.02.017 Copyright © 2011 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 1 VE-cadherin (VE-cad) extracellular domain in HAE patient sample. A, Phase contrast microscopy of cultured endothelial cells. Scale bar: 50 μm. B, VE-cadherin immunostaining. Scale bar: 50 μm. C, VE-cadherin immunoblot on HUVECs. D, VE-cadherin immunoblot of HAE patient serum samples out of attack (lane 1), during the attack (lane 2), and after the attack (lane 3). E, Densitometric analysis of VE-cadherin immunoblots expressed as absorbance values (mean ± 2 SD; n = 4). F, Plasma kininogenase activity levels (mean ± 2 SD; n = 4). HUVEC, Human umbilical vascular endothelial cell; MW, molecular weight; WB, Western blot. Journal of Allergy and Clinical Immunology 2011 128, 232-234DOI: (10.1016/j.jaci.2011.02.017) Copyright © 2011 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 2 Bradykinin (BK) induced increased endothelial permeability through a process involving VE-cadherin proteolytic cleavage. A, In vitro vascular permeability assay on endothelial cells treated for 4 hours with BK (10−7 mol/L), kallikrein (KK; 10−7 mol/L), or EGTA (1 mmol/L; positive control; mean ± SD; n = 3). B, VE-cadherin content of conditioned media from unstimulated cells and BK-stimulated HUVECs (10−6 mol/L) for 5 to 60 minutes (HUVEC extracts were loaded as positive control; 5 μg; n = 3). C, VE-cadherin content of conditioned media from KK (0.6 U)–treated cells. D, In vitro VE-cadherin (0.5 μg) cleavage by KK (0.6 U; n = 3). E, VE-cadherin immunostaining after KK treatment (0.60 U for 60 minutes; n = 4). Scale bar: 50 μm. CTL, Cytotoxic T lymphocyte; MW, molecular weight; WB, Western blot. Journal of Allergy and Clinical Immunology 2011 128, 232-234DOI: (10.1016/j.jaci.2011.02.017) Copyright © 2011 American Academy of Allergy, Asthma & Immunology Terms and Conditions