High pulsatility flow stimulates smooth muscle cell hypertrophy and contractile protein expression.

Proximal arterial stiffening is an important predictor of events in systemic and pulmonary hypertension, partly through its contribution to downstream vascular abnormalities. However, much remains undetermined regarding the mechanisms involved in the vascular changes induced by arterial stiffening. We therefore addressed the hypothesis that high pulsatility flow, caused by proximal arterial stiffening, induces downstream pulmonary artery endothelial cell (EC) dysfunction that in turn leads to phenotypic change of smooth muscle cells (SMCs). To test the hypothesis, we employed a model pulmonary circulation in which upstream compliance regulates the pulsatility of flow waves imposed onto a downstream vascular mimetic coculture composed of pulmonary ECs and SMCs. The effects of high pulsatility flow on SMCs were determined both in the presence and absence of ECs. In the presence of ECs, high pulsatility flow increased SMC size and expression of the contractile proteins, smooth muscle α-actin (SMA) and smooth muscle myosin heavy chain (SM-MHC), without affecting proliferation. In the absence of ECs, high pulsatility flow decreased SMC expression of SMA and SM-MHC, without affecting SMC size or proliferation. To identify the molecular signals involved in the EC-mediated SMC responses, mRNA and/or protein expression of vasoconstrictors [angiotensin-converting enzyme (ACE) and endothelin (ET)-1], vasodilator (eNOS), and growth factor (TGF-β1) in EC were examined. Results showed high pulsatility flow decreased eNOS and increased ACE, ET-1, and TGF-β1 expression. ACE inhibition with ramiprilat, ET-1 receptor inhibition with bosentan, and treatment with the vasodilator bradykinin prevented flow-induced, EC-dependent SMC changes. In conclusion, high pulsatility flow stimulated SMC hypertrophy and contractile protein expression by altering EC production of vasoactive mediators and cytokines, supporting the idea of a coupling between proximal vascular stiffening, flow pulsatility, and downstream vascular function.

[1]  C. N. Gillis,et al.  Lisinopril and ramiprilat protection of the vascular endothelium against free radical-induced functional injury. , 1992, The Journal of pharmacology and experimental therapeutics.

[2]  N. Westerhof,et al.  The arterial load in pulmonary hypertension , 2010, European Respiratory Review.

[3]  K. Stenmark,et al.  Vascular remodeling versus vasoconstriction in chronic hypoxic pulmonary hypertension: a time for reappraisal? , 2005, Circulation research.

[4]  W. Linz,et al.  ACE-inhibition induces NO-formation in cultured bovine endothelial cells and protects isolated ischemic rat hearts. , 1992, Journal of molecular and cellular cardiology.

[5]  G. Mitchell Effects of central arterial aging on the structure and function of the peripheral vasculature: implications for end-organ damage. , 2008, Journal of applied physiology.

[6]  Timothy M. Wick,et al.  Endothelial Cell–Smooth Muscle Cell Co-Culture in a Perfusion Bioreactor System , 2005, Annals of Biomedical Engineering.

[7]  J. Murphy,et al.  The structural basis of persistent pulmonary hypertension of the newborn infant. , 1981, The Journal of pediatrics.

[8]  K. Amann,et al.  Hypertrophy and hyperplasia of smooth muscle cells of small intramyocardial arteries in spontaneously hypertensive rats. , 1995, Hypertension.

[9]  P. Doevendans,et al.  Regulation and characteristics of vascular smooth muscle cell phenotypic diversity , 2007, Netherlands heart journal : monthly journal of the Netherlands Society of Cardiology and the Netherlands Heart Foundation.

[10]  L. Gold,et al.  Vascular remodeling in primary pulmonary hypertension. Potential role for transforming growth factor-beta. , 1994, The American journal of pathology.

[11]  T. Spector,et al.  Arterial Stiffening Relates to Arterial Calcification But Not to Noncalcified Atheroma in Women , 2011, Journal of the American College of Cardiology.

[12]  R. Klein,et al.  The association of cardiovascular disease with the long-term incidence of age-related maculopathy: the Beaver Dam eye study. , 2003, Ophthalmology.

[13]  D. Levy,et al.  Circulating Vascular Growth Factors and Central Hemodynamic Load in the Community , 2012, Hypertension.

[14]  C. Ahn,et al.  Measurement of Hemodynamic Energy at Different Vessels in an Adult Swine Model , 2010, ASAIO journal.

[15]  M. Swartz,et al.  Mechanisms of Interstitial Flow-Induced Remodeling of Fibroblast–Collagen Cultures , 2006, Annals of Biomedical Engineering.

[16]  Robert T Tranquillo,et al.  Transmural flow bioreactor for vascular tissue engineering , 2009, Biotechnology and bioengineering.

[17]  N. Weissmann,et al.  Cellular and molecular basis of pulmonary arterial hypertension. , 2009, Journal of the American College of Cardiology.

[18]  D. Pinsky,et al.  Pulmonary artery smooth muscle hypertrophy: roles of glycogen synthase kinase-3beta and p70 ribosomal S6 kinase. , 2010, American journal of physiology. Lung cellular and molecular physiology.

[19]  M. Rabinovitch Pulmonary hypertension: updating a mysterious disease. , 1997, Cardiovascular research.

[20]  N. Van Rooijen,et al.  Blood flow-dependent arterial remodelling is facilitated by inflammation but directed by vascular tone. , 2008, Cardiovascular research.

[21]  H. Struijker‐Boudier,et al.  Current Perspectives on Arterial Stiffness and Pulse Pressure in Hypertension and Cardiovascular Diseases , 2003, Circulation.

[22]  J. Panza,et al.  Role of Endothelial Nitric Oxide in Shear Stress—Induced Vasodilation of Human Microvasculature: Diminished Activity in Hypertensive and Hypercholesterolemic Patients , 2001, Circulation.

[23]  R. Roth,et al.  Hypertrophy and Prolonged DNA Synthesis in Smooth Muscle Cells Characterize Pulmonary Arterial Wall Thickening After Monocrotaline Pyrrole Administration to Rats , 1997, Toxicologic pathology.

[24]  B. Berk,et al.  Glutaredoxin Mediates Akt and eNOS Activation by Flow in a Glutathione Reductase-Dependent Manner , 2007, Arteriosclerosis, thrombosis, and vascular biology.

[25]  M. Oka,et al.  Rho kinase-mediated vasoconstriction in pulmonary hypertension. , 2010, Advances in experimental medicine and biology.

[26]  Walter L Murfee,et al.  Enhanced Smooth Muscle Cell Coverage of Microvessels Exposed to Increased Hemodynamic Stresses In Vivo , 2003, Circulation research.

[27]  D. Ingber,et al.  Extracellular matrix and pulmonary hypertension : control of vascular smooth muscle cell contractility , 1997 .

[28]  M. Frid,et al.  Mature Vascular Endothelium Can Give Rise to Smooth Muscle Cells via Endothelial-Mesenchymal Transdifferentiation: In Vitro Analysis , 2002, Circulation research.

[29]  Nico Westerhof,et al.  Noninvasively assessed pulmonary artery stiffness predicts mortality in pulmonary arterial hypertension. , 2007, Chest.

[30]  S. Chien,et al.  PDGF-BB and TGF-β1 on cross-talk between endothelial and smooth muscle cells in vascular remodeling induced by low shear stress , 2011, Proceedings of the National Academy of Sciences.

[31]  G. Owens,et al.  Molecular regulation of vascular smooth muscle cell differentiation in development and disease. , 2004, Physiological reviews.

[32]  P. Connell,et al.  Pulsatile flow increases the expression of eNOS, ET-1, and prostacyclin in a novel in vitro coculture model of the retinal vasculature. , 2005, Investigative Ophthalmology and Visual Science.

[33]  P. Cahill,et al.  Flow-mediated regulation of G-protein expression in cocultured vascular smooth muscle and endothelial cells. , 1998, Arteriosclerosis, thrombosis, and vascular biology.

[34]  S. Chien Mechanotransduction and endothelial cell homeostasis: the wisdom of the cell. , 2007, American journal of physiology. Heart and circulatory physiology.

[35]  S. Gairhe,et al.  Myoendothelial gap junctional signaling induces differentiation of pulmonary arterial smooth muscle cells. , 2011, American journal of physiology. Lung cellular and molecular physiology.

[36]  J. Tarbell,et al.  Shear Stress Modulation of Smooth Muscle Cell Marker Genes in 2-D and 3-D Depends on Mechanotransduction by Heparan Sulfate Proteoglycans and ERK1/2 , 2010, PloS one.

[37]  J. Tarbell,et al.  Interstitial flow through the internal elastic lamina affects shear stress on arterial smooth muscle cells. , 2000, American journal of physiology. Heart and circulatory physiology.

[38]  P. Connell,et al.  The role of pulsatile flow in controlling microvascular retinal endothelial and pericyte cell apoptosis and proliferation. , 2011, Cardiovascular research.

[39]  Jeffrey A. Feinstein,et al.  Three-Dimensional Hemodynamics in the Human Pulmonary Arteries Under Resting and Exercise Conditions , 2010, Annals of Biomedical Engineering.

[40]  K. Stenmark,et al.  Mechanics and function of the pulmonary vasculature: implications for pulmonary vascular disease and right ventricular function. , 2012, Comprehensive Physiology.

[41]  G. Kassab,et al.  Biomechanical considerations in the design of graft: the homeostasis hypothesis. , 2006, Annual review of biomedical engineering.

[42]  N. Voelkel,et al.  Cellular and molecular biology of vascular smooth muscle cells in pulmonary hypertension. , 1997, Pulmonary pharmacology & therapeutics.

[43]  Jing Zheng,et al.  Effects of Pulsatile Shear Stress on Nitric Oxide Production and Endothelial Cell Nitric Oxide Synthase Expression by Ovine Fetoplacental Artery Endothelial Cells1 , 2003, Biology of reproduction.

[44]  Jay D Humphrey,et al.  Importance of pulsatility in hypertensive carotid artery growth and remodeling , 2009, Journal of hypertension.

[45]  J. Cohn,et al.  Age-related abnormalities in arterial compliance identified by pressure pulse contour analysis: aging and arterial compliance. , 1999, Hypertension.

[46]  M. Peach,et al.  Angiotensin II Induces Hypertrophy, not Hyperplasia, of Cultured Rat Aortic Smooth Muscle Cells , 1988, Circulation research.

[47]  Elaine Warner,et al.  Cross-Sectional Relations of Peripheral Microvascular Function, Cardiovascular Disease Risk Factors, and Aortic Stiffness: The Framingham Heart Study , 2005, Circulation.

[48]  K. Hunter,et al.  Pulmonary vascular input impedance is a combined measure of pulmonary vascular resistance and stiffness and predicts clinical outcomes better than pulmonary vascular resistance alone in pediatric patients with pulmonary hypertension. , 2008, American heart journal.

[49]  P. Cahill,et al.  Sustained pulsatile flow regulates endothelial nitric oxide synthase and cyclooxygenase expression in co-cultured vascular endothelial and smooth muscle cells. , 1999, Journal of molecular and cellular cardiology.

[50]  Min Li,et al.  High Pulsatility Flow Induces Adhesion Molecule and Cytokine mRNA Expression in Distal Pulmonary Artery Endothelial Cells. , 2009, ATS 2009.

[51]  D B Longmore,et al.  Magnetic resonance assessment of the pulmonary arterial trunk anatomy, flow, pulsatility and distensibility. , 1993, European heart journal.

[52]  S. Chien,et al.  Shear Stress Induces Synthetic-to-Contractile Phenotypic Modulation in Smooth Muscle Cells via Peroxisome Proliferator-Activated Receptor &agr;/&dgr; Activations by Prostacyclin Released by Sheared Endothelial Cells , 2009, Circulation research.

[53]  S. Reuben,et al.  Compliance of the Human Pulmonary Arterial System in Disease , 1971, Circulation research.

[54]  G. Truskey Endothelial Cell Vascular Smooth Muscle Cell Co-Culture Assay For High Throughput Screening Assays For Discovery of Anti-Angiogenesis Agents and Other Therapeutic Molecules. , 2010, International journal of high throughput screening.

[55]  J. Haga,et al.  Molecular basis of the effects of shear stress on vascular endothelial cells. , 2005, Journal of biomechanics.

[56]  V. Gudnason,et al.  Arterial stiffness, pressure and flow pulsatility and brain structure and function: the Age, Gene/Environment Susceptibility--Reykjavik study. , 2011, Brain : a journal of neurology.

[57]  F. Dekker,et al.  Arterial aging and arterial disease: interplay between central hemodynamics, cardiac work, and organ flow—implications for CKD and cardiovascular disease , 2011, Kidney international supplements.

[58]  S. Ito,et al.  Central Pulse Pressure and Aortic Stiffness Determine Renal Hemodynamics: Pathophysiological Implication for Microalbuminuria in Hypertension , 2011, Hypertension.

[59]  Michael F O'Rourke,et al.  Arterial stiffness, its assessment, prognostic value, and implications for treatment. , 2011, American journal of hypertension.

[60]  O. Hill A Twin Study , 1968, British Journal of Psychiatry.

[61]  Wei Tan,et al.  Development and evaluation of microdevices for studying anisotropic biaxial cyclic stretch on cells , 2008, Biomedical microdevices.