1. Date of download: 5/27/2016 Copyright © The American College of Cardiology. All rights reserved. From: Endothelium-Dependent Regulation of the Mechanical Properties of Aortic Valve Cusps J Am Coll Cardiol. 2009;53(16):1448-1455. doi:10.1016/j.jacc.2008.11.056 Biaxial Micromechanical System and Normal Load-Strain Curves (A) Porcine aortic cusp mounted to biaxial micromechanical testing device. The 4 dots are tracked using a 60 frames/s charge- coupled device camera to determine the strains on the specimen. (B) Normal load-strain curve of an aortic cusp in the radial direction (blue diamonds) and circumferential direction (red squares) showing the initial toe region, transition region, and linear region. The linear region was used as a measure of the elastic modulus of the valve. Figure Legend:

2. Date of download: 5/27/2016 Copyright © The American College of Cardiology. All rights reserved. From: Endothelium-Dependent Regulation of the Mechanical Properties of Aortic Valve Cusps J Am Coll Cardiol. 2009;53(16):1448-1455. doi:10.1016/j.jacc.2008.11.056 Percentage Change in Aortic Valve Areal Strains Versus Control in Response to Various Agents The percentage change in areal strain (± SE) of aortic cusps (normal and denuded) in response to the various vasoactive agents and their inhibitors. Areal strains were measured before and after addition of the compounds (each valve served as its own control). *Dunnett-adjusted p < 0.05 versus control. †p < 0.05 versus endothelin (ET)-1 alone. #Dunnett-adjusted p < 0.1 versus control. **Dunnett-adjusted p < 0.01 versus control. ††p < 0.01 versus 5-hydroxytryptamine (5-HT) alone. CyB = cytochalasin B; L-NAME = N- nitro-l-arginine-methyl-ester. Figure Legend:

3. Date of download: 5/27/2016 Copyright © The American College of Cardiology. All rights reserved. From: Endothelium-Dependent Regulation of the Mechanical Properties of Aortic Valve Cusps J Am Coll Cardiol. 2009;53(16):1448-1455. doi:10.1016/j.jacc.2008.11.056 Concentration-Response Curves to 5-HT Concentration-dependent changes in contraction or relaxation (± SE) of porcine aortic valve specimens in response to (A) 5- hydroxytryptamine (5-HT) alone, (B) 5-HT + endothelial denudation, and (C) 5-HT + N-nitro-l-arginine-methyl-ester (L-NAME) in the radial direction (squares) and circumferential direction (diamonds). *Dunnett-adjusted p < 0.05 versus control. **Dunnett-adjusted p < 0.01 versus control. Figure Legend:

4. Date of download: 5/27/2016 Copyright © The American College of Cardiology. All rights reserved. From: Endothelium-Dependent Regulation of the Mechanical Properties of Aortic Valve Cusps J Am Coll Cardiol. 2009;53(16):1448-1455. doi:10.1016/j.jacc.2008.11.056 Concentration-Response Curves to ET-1 Concentration-dependent changes in contraction or relaxation (± SE) of aortic cusps in response to (A) endothelin (ET)-1 and (B) ET-1 + cytochalasin B (CyB) in the radial (squares) and circumferential (diamonds) axes. *Dunnet-adjusted p < 0.05 versus control. **Dunnett-adjusted p < 0.01 versus control. Figure Legend:

5. Date of download: 5/27/2016 Copyright © The American College of Cardiology. All rights reserved. From: Endothelium-Dependent Regulation of the Mechanical Properties of Aortic Valve Cusps J Am Coll Cardiol. 2009;53(16):1448-1455. doi:10.1016/j.jacc.2008.11.056 Cellular Integrity (A) Scanning electron micrograph of aortic cusp before biaxial testing, showing preserved endothelial monolayer. (Note: areas of apparent endothelial denudation are due to air drying of specimens with hexamethyldisilazane. Both ridges of the endothelium are juxtaposable.) (B) Histological section (original magnification ×40) showing preservation of the endothelial cells after biaxial testing by positive von Willebrand factor staining (brown). (C) Scanning electron micrograph of a denuded porcine aortic valve showing absence of endothelium and normal subendothelial structure. (D) Representative immunohistochemical analysis of smooth muscle α-actin (SMA) expression before stimulation with endothelin (ET)-1 (original magnification ×20). (E) SMA expression after stimulation with ET-1 (original magnification ×20) showing no changes in overall SMA expression. Figure Legend: