Bone Morphogenic Protein-4 Contributes to Venous Endothelial Dysfunction in Patients With Diabetes Undergoing Coronary Revascularization Jia Hu, MD,

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Bone Morphogenic Protein-4 Contributes to Venous Endothelial Dysfunction in Patients With Diabetes Undergoing Coronary Revascularization Jia Hu, MD, PhD, Jian Liu, MS, Micky W.T. Kwok, MRCS, Randolph H.L. Wong, FRCS, Yu Huang, PhD, Song Wan, MD, FRCS The Annals of Thoracic Surgery Volume 95, Issue 4, Pages 1331-1339 (April 2013) DOI: 10.1016/j.athoracsur.2012.12.028 Copyright © 2013 The Society of Thoracic Surgeons Terms and Conditions

Fig 1 (A) Endothelium-dependent relaxations (EDRs) in response to acetylcholine (ACh) in saphenous veins from normal controls (n = 3) and patients with diabetes (n = 15). (B) Concentration-response curves for sodium nitroprusside (SNP)-induced venous endothelium-independent relaxations in normal controls (n = 3) and patients with diabetes (n = 15). Data are mean ± standard error of the mean (*p < 0.05 compared with the normal controls). (DM = diabetes mellitus; Phe = phenylephrine.) The Annals of Thoracic Surgery 2013 95, 1331-1339DOI: (10.1016/j.athoracsur.2012.12.028) Copyright © 2013 The Society of Thoracic Surgeons Terms and Conditions

Fig 2 (A) Increased expression of bone morphogenic protein-4 (BMP4) in saphenous veins from patients with diabetes compared with those from patients without diabetes (n = 15 in both groups). (B) BMP4 levels in human umbilical venous endothelial cells (HUVECs) increased in a glucose concentration–dependent manner, and HUVECs subjected to 30 mmol/L glucose expressed a significantly higher level of BMP4. Data are mean ± standard error of the mean (*p < 0.05 compared with the controls). (DM = diabetes mellitus; GAPDH = glyceraldehyde-3-phosphate dehydrogenase; HG = high glucose; mM = mmol/L.) The Annals of Thoracic Surgery 2013 95, 1331-1339DOI: (10.1016/j.athoracsur.2012.12.028) Copyright © 2013 The Society of Thoracic Surgeons Terms and Conditions

Fig 3 Noggin-induced improvement in endothelium-dependent relaxations (EDRs) in veins from patients with diabetes (n = 15). (A) Representative traces. (B) Concentration-response curve showing that bone morphogenic protein-4 (BMP4) antagonist noggin (100 ng/mL) enhanced EDRs in saphenous veins from patients with diabetes. (C) The lack of effect of noggin on sodium nitroprusside (SNP)-induced relaxations in these veins. (D, E) Noggin reduced the increased production of ROS as measured by dihydroethidium (DHE) fluorescence in saphenous veins from patients with diabetes. Data are mean ± standard error of the mean (*p < 0.05 compared with the controls). (Ach = acetylcholine; Phe = phenylephrine.) The Annals of Thoracic Surgery 2013 95, 1331-1339DOI: (10.1016/j.athoracsur.2012.12.028) Copyright © 2013 The Society of Thoracic Surgeons Terms and Conditions

Fig 4 The effect of noggin on venous endothelial function in patients without diabetes. (A) Acetylcholine (Ach)-induced endothelium-dependent relaxations (EDRs) in noggin-treated saphenous veins from normal subjects and (B) from patients without diabetes (n = 15) were not different compared with their own controls. (C, D) Reactive oxygen species (ROS) levels in veins from these patients were not affected by noggin treatment. Data are mean ± standard error of the mean (DHE = dihydroethidium; DM = diabetes mellitus; Phe = phenylephrine.) The Annals of Thoracic Surgery 2013 95, 1331-1339DOI: (10.1016/j.athoracsur.2012.12.028) Copyright © 2013 The Society of Thoracic Surgeons Terms and Conditions

Fig 5 Bone morphogenic protein-4 (BMP4) impairs endothelium-dependent relaxations (EDRs) in cultured porcine saphenous veins. (A) Representative trace and (B) concentration-response curve showing the effect of BMP4 (25 ng/mL over 24 hours) and noggin (100 ng/mL over 24 hours) on acetylcholine (Ach)-induced EDRs in healthy porcine saphenous veins. (C, D) Noggin reversed the BMP4-stimulated reactive oxygen species (ROS) production. (E) The level of ROS in noggin-treated porcine veins remains unaffected. Data are mean ± standard error of the mean (*p < 0.05 compared with the controls, #p < 0.05 compared with the BMP4 + noggin group). The Annals of Thoracic Surgery 2013 95, 1331-1339DOI: (10.1016/j.athoracsur.2012.12.028) Copyright © 2013 The Society of Thoracic Surgeons Terms and Conditions