Laki J. Rousou, MS, MD, Kristin B

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Saphenous Vein Conduits Harvested by Endoscopic Technique Exhibit Structural and Functional Damage Laki J. Rousou, MS, MD, Kristin B. Taylor, RN, Xiu-Gui Lu, MD, Nancy Healey, BS, Michael D. Crittenden, MD, Shukri F. Khuri, MD, Hemant S. Thatte, PhD The Annals of Thoracic Surgery Volume 87, Issue 1, Pages 62-70 (January 2009) DOI: 10.1016/j.athoracsur.2008.08.049 Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 1 Schematic representation of the location and techniques used to harvest the saphenous vein. Endoscopic harvest was started below the knee (A) and proceeded all the way proximal (B). Endoscopic harvest was then begun down the leg to a distance approximately mid calf. At this point, an open incision (C) was made approximately 3 cm below the level where the endoscopic harvest was stopped. (D indicates end of open harvest.) Each sample was taken from a section of vein that would not be needed for bypass. (endo = endoscopic; ESVH = endoscopic saphenous vein harvest; OSVH = open saphenous vein harvest.) The Annals of Thoracic Surgery 2009 87, 62-70DOI: (10.1016/j.athoracsur.2008.08.049) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 2 Multiphoton transmission imaging showing no evidence of endothelial breaks or tears in any of the samples. A stretched endothelial layer was visible in the endoscopic saphenous vein harvest samples. (Magnification = ×320.) (ESVH = endoscopic saphenous vein harvest; OSVH = open saphenous vein harvest.) The Annals of Thoracic Surgery 2009 87, 62-70DOI: (10.1016/j.athoracsur.2008.08.049) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 3 Multiphoton fluorescence imaging shows esterase activity and endothelial injury in the open saphenous vein harvest (OSVH) and endoscopic saphenous vein harvest (ESVH) vein endothelium. Representative images of esterase activity in the OSVH and ESVH samples show greater esterase activity and viability (green fluorescence) in the OSVH samples (a) when compared with the ESVH (b). Additionally, in comparison with OSVH samples (c), ESVH vessels show robust red fluorescence and membrane damage (d), in endothelial and smooth vessel regions of the veins. (Magnification = ×320.) The Annals of Thoracic Surgery 2009 87, 62-70DOI: (10.1016/j.athoracsur.2008.08.049) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 4 Quantitative representation of mean esterase activity (green fluorescence) from all samples in the open saphenous vein harvest (OSVH) versus endoscopic saphenous vein harvest (ESVH) greater saphenous veins (Fig 3a, b). As shown in the above images, the OSVH group has a significantly greater esterase activity than the ESVH group (p < 0.0001, Wilcoxon two-sample test; n = 250 measurements for each group). The Annals of Thoracic Surgery 2009 87, 62-70DOI: (10.1016/j.athoracsur.2008.08.049) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 5 Calcium mobilization and nitric oxide production in response to bradykinin. (A) Representative images of calcium mobilization before (pre) and after (post) bradykinin stimulation in the open saphenous vein harvest (OSVH) and endoscopic saphenous vein harvest (ESVH) groups. (B) Representative images of nitric oxide production before (pre) and after (post) bradykinin stimulation. (Magnification = ×320.) The Annals of Thoracic Surgery 2009 87, 62-70DOI: (10.1016/j.athoracsur.2008.08.049) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 6 Graphical representation of mean calcium (dark gray bars) mobilization and nitric oxide (light gray bars) production in the open saphenous vein harvest (OSVH) and endoscopic saphenous vein harvest (ESVH) groups shows that calcium mobilization and nitric oxide production increases to a greater extent in the OSVH group with respect to baseline (n = 50 measurements for each group). The Annals of Thoracic Surgery 2009 87, 62-70DOI: (10.1016/j.athoracsur.2008.08.049) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 7 Immunofluorescence images of endothelial proteins in vein segments harvested by the open saphenous vein harvest (OSVH) and endoscopic saphenous vein harvest (ESVH) techniques show absence or abnormal distribution of caveolin and endothelial nitric oxide synthase (eNOS), but not cadherin or von Willebrand factor (vWF), in the ESVH group. (Magnification = ×320.) The Annals of Thoracic Surgery 2009 87, 62-70DOI: (10.1016/j.athoracsur.2008.08.049) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions

Fig 8 Western blot analyisis of endothelial proteins in the open saphenous vein harvest (OSVH) and endoscopic saphenous vein harvest (ESVH) groups shows a relative decrease in caveolin, endothelial nitric oxide synthase (eNOS), and 24 kd von Willebrand factor (vWF) concentrations in the ESVH group. The Annals of Thoracic Surgery 2009 87, 62-70DOI: (10.1016/j.athoracsur.2008.08.049) Copyright © 2009 The Society of Thoracic Surgeons Terms and Conditions