Basic Study
Copyright ©The Author(s) 2015. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Cardiol. Jun 26, 2015; 7(6): 331-343
Published online Jun 26, 2015. doi: 10.4330/wjc.v7.i6.331
Bone morphogenetic protein-4 and transforming growth factor-beta1 mechanisms in acute valvular response to supra-physiologic hemodynamic stresses
Ling Sun, Philippe Sucosky
Ling Sun, Philippe Sucosky, Department of Aerospace and Mechanical Engineering, University of Notre Dame, Notre Dame, IN 46556, United States
Author contributions: Sun L performed the experiments and analyzed the data; Sun L and Sucosky P designed the research and wrote the paper.
Supported by American Heart Association Scientist Development Grant, No. 11SDG7600103.
Conflict-of-interest: The authors declare no conflict of interest.
Data sharing: Complete dataset and statistical analyses available from the corresponding author at philippe.sucosky@nd.edu.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Correspondence to: Philippe Sucosky, PhD, FAHA, Department of Aerospace and Mechanical Engineering, University of Notre Dame, 143 Multidisciplinary Research Building, Notre Dame, IN 46556-5637, United States. philippe.sucosky@nd.edu
Telephone: +1-574-6311898 Fax: +1-574-6312144
Received: January 20, 2015
Peer-review started: January 21, 2015
First decision: February 7, 2015
Revised: February 20, 2015
Accepted: April 16, 2015
Article in press: April 20, 2015
Published online: June 26, 2015
Abstract

AIM: To explore ex vivo the role of bone morphogenetic protein-4 (BMP-4) and transforming growth factor-beta1 (TGF-β1) in acute valvular response to fluid shear stress (FSS) abnormalities.

METHODS: Porcine valve leaflets were subjected ex vivo to physiologic FSS, supra-physiologic FSS magnitude at normal frequency and supra-physiologic FSS frequency at normal magnitude for 48 h in a double-sided cone-and-plate bioreactor filled with standard culture medium. The role of BMP-4 and TGF-β1 in the valvular response was investigated by promoting or inhibiting the downstream action of those cytokines via culture medium supplementation with BMP-4 or the BMP antagonist noggin, and TGF-β1 or the TGF-β1 inhibitor SB-431542, respectively. Fresh porcine leaflets were used as controls. Each experimental group consisted of six leaflet samples. Immunostaining and immunoblotting were performed to assess endothelial activation in terms of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 expressions, paracrine signaling in terms of BMP-4 and TGF-β1 expressions and extracellular matrix (ECM) remodeling in terms of cathepsin L, cathepsin S, metalloproteinases (MMP)-2 and MMP-9 expressions. Immunostained images were quantified by normalizing the intensities of positively stained regions by the number of cells in each image while immunoblots were quantified by densitometry.

RESULTS: Regardless of the culture medium, physiologic FSS maintained valvular homeostasis. Tissue exposure to supra-physiologic FSS magnitude in standard medium stimulated paracrine signaling (TGF-β1: 467% ± 22% vs 100% ± 6% in fresh controls, BMP-4: 258% ± 22% vs 100% ± 4% in fresh controls; P < 0.05) and ECM degradation (MMP-2: 941% ± 90% vs 100% ± 19% in fresh controls, MMP-9: 1219% ± 190% vs 100% ± 16% in fresh controls, cathepsin L: 1187% ± 175% vs 100% ± 12% in fresh controls, cathepsin S: 603% ± 88% vs 100% ± 13% in fresh controls; P < 0.05), while BMP-4 supplementation also promoted fibrosa activation and TGF-β1 inhibition reduced MMP-9 expression to the native tissue level (MMP-9: 308% ± 153% with TGF-β1 inhibition vs 100% ± 16% in fresh control; P > 0.05). Supra-physiologic FSS frequency had no effect on endothelial activation and paracrine signaling regardless of the culture medium but TGF-β1 silencing attenuated FSS-induced ECM degradation via MMP-9 downregulation (MMP-9: 302% ± 182% vs 100% ± 42% in fresh controls; P > 0.05).

CONCLUSION: Valvular tissue is sensitive to FSS abnormalities. The TGF-β1 inhibitor SB-431542 is a potential candidate molecule for attenuating the effects of FSS abnormalities on valvular remodeling.

Keywords: Aortic valve, Fluid shear stress, Calcification, Bone morphogenetic protein, Transforming growth factor beta

Core tip: Although flow abnormalities have been shown to promote valvular pathogenesis in a bone morphogenetic protein-4 (BMP-4)- and transforming growth factor-beta1 (TGF-β1)-dependent manner, the mode of action of those molecules in response to fluid shear stress (FSS) abnormalities remains unknown. This ex vivo study aimed at isolating the role played by those cytokines in the acute response of porcine leaflets to supra-physiologic FSS magnitude/frequency. The study reveals that: (1) valvular endothelial activation is weakly regulated by BMP-4 in response to FSS abnormalities; (2) TGF-β1 silencing attenuates FSS-induced extracellular matrix degradation via MMP-9 downregulation; and (3) BMP-4 and TGF-β1 do not synergistically interact in response to FSS abnormalities.