Mechanical Strain and the Aortic Valve: Influence on Fibroblasts, Extracellular Matrix, and Potential Stenosis Sven Lehmann, MD, Thomas Walther, MD,

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Mechanical Strain and the Aortic Valve: Influence on Fibroblasts, Extracellular Matrix, and Potential Stenosis Sven Lehmann, MD, Thomas Walther, MD, PhD, Jörg Kempfert, MD, Ardawan Rastan, MD, Jens Garbade, MD, Stefan Dhein, MD, PhD, Friedrich W. Mohr, MD, PhD The Annals of Thoracic Surgery Volume 88, Issue 5, Pages 1476-1483 (November 2009) DOI: 10.1016/j.athoracsur.2009.07.025 Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 1 Prolyl-4-hydroxylase beta and 3-amino-9-ethylcarbazol method with haemalaun from fibroblast subcultures (black arrow indicates cell nucleus of fibroblast; gray arrow indicates cytoplasm of fibroblast). The Annals of Thoracic Surgery 2009 88, 1476-1483DOI: (10.1016/j.athoracsur.2009.07.025) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 2 (A) Osteocalcin expression in relation to the preoperative maximum transvalvular pressure gradient. (B) Osteopontin expression in relation to the preoperative maximum transvalvular pressure gradient. (C) Alkaline phosphatase expression in relation to the preoperative maximum transvalvular pressure gradient. (D) Tenascin-C expression in relation to the preoperative maximum transvalvular pressure gradient. (E) Decorin expression in relation to the preoperative maximum transvalvular pressure gradient. (F) Biglycan expression in relation to the preoperative maximum transvalvular pressure gradient. The Annals of Thoracic Surgery 2009 88, 1476-1483DOI: (10.1016/j.athoracsur.2009.07.025) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 3 (A) Immunohistochemistry of decorin expression in human aortic valve with a preoperative maximum transvalvular pressure gradient from 60 mm Hg. (1 = inflow layer; 2 = central layer; 3 = outflow layer; black arrow = decorin.) (B) Immunohistochemistry of decorin expression in human aortic valve with a preoperative maximum transvalvular pressure gradient from 120 mm Hg. (1 = inflow layer; 2 = central layer; 3 = outflow layer; black arrow = decorin.) The Annals of Thoracic Surgery 2009 88, 1476-1483DOI: (10.1016/j.athoracsur.2009.07.025) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 4 (A) Immunohistochemistry of osteocalcinexpression after 72 hours without stress (double black arrow indicates fibroblast; black arrow indicates osteocalcin). (B) Immunohistochemistry of osteocalcinexpression after 72 hours with stress (double black arrow indicates fibroblast; black arrow indicates osteocalcin). The Annals of Thoracic Surgery 2009 88, 1476-1483DOI: (10.1016/j.athoracsur.2009.07.025) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 5 Protein expression without stress (black bars) and after 72 hours of stress (gray bars). The Annals of Thoracic Surgery 2009 88, 1476-1483DOI: (10.1016/j.athoracsur.2009.07.025) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 6 Quantitative Western blot analysis against biglycan and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). (Line 1 = after 72 hours of stress; line 2 = without stress.) The Annals of Thoracic Surgery 2009 88, 1476-1483DOI: (10.1016/j.athoracsur.2009.07.025) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 7 Protein expression without stress (black bars) and after 72 hours of stress (gray bars) in relation to glyceraldehyde-3-phosphate dehydrogenase (GAPDH). The Annals of Thoracic Surgery 2009 88, 1476-1483DOI: (10.1016/j.athoracsur.2009.07.025) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions