ABSTRACT
The discovery in 1980 of endothelium-derived relaxing factor, EDRF (later found to be nitric oxide), by Furchgott and Zawadzki1 emanated from the observation that denuded endothelium from isolated rabbit aortic vascular rings produced a paradoxical vasoconstriction in response to the receptor-mediated substance acetylcholine. This initial observation stimulated intense research in vascular biology in the ensuing decades to further delineate the various components and mechanisms of vascular diseases. The nitric oxide radical (NO) is formed from endothelial cells from its precursor, Larginine via the enzymatic action of NO synthase (eNOS), in the presence of essential cofactors oxygen, calmodulin, tetrahydrobiopterin, and reduced nicotinamide adenine dinucleotide phosphate (NADPH). eNOS (a product of the NOS3 gene) is located in the invaginations of cell
membranes called caveoli. The protein caveolin maintains the inactive state of eNOS through its binding to calmodulin. eNOS is activated by binding calcium to calmodulin, which displaces the inhibitor caveolin. Once formed, NO effects vasodilation through diffusion from the endothelium to the underlying smooth muscle cell, stimulating an increase in cyclic guanosine monophosphate (cGMP) and other important signal transduction mechanisms.2 NO, a potent endogenous vasodilator, is secreted by the endothelium in response to various pharmacologic agents, physiologic agonists, and physical stimuli, including shear stress from a high-blood-flow state. The basal secretion of NO maintains the vessel in a vasodilatory state, and assists in maintaining vascular health through its other anti-atherogenic properties. NO mediates the inhibition of platelet aggregation and adherence, leukocyte adhesion/ infiltration, and proliferation of vascular smooth muscle cells, and prevents the oxidative modification of low-density lipoprotein (LDL) cholesterol. NO bioavailability is reduced in a pro-oxidant milieu, both in the presence of cardiovascular risk factors and in disease states. Oxidative stress impedes the biosynthesis and accelerates the breakdown of NO by various mechanisms.2-5
The initial experimental observations formed the basis for translational research to study the characteristics and significance of endotheliumdependent vasodilation in various vascular beds, and in various vascular disease states. In the 1980s, Ludmer and colleagues6-9 studied endothelial vasomotor function using intracoronary injection of acetylcholine in humans, with various degrees of atherosclerosis noted by angiography. Like isolated vascular rings, human coronaries with normal-appearing coronaries angiographically or with mild stenosis vasoconstricted paradoxically to acetylcholine.6 Angiography is invasive and not conducive to repeated studies over time. Ultrasound techniques have long been used to study vessel physiology. Ultrasound assessment of brachial artery vasoreactivity using high-resolution B-mode ultrasound emerged as a clinical research tool in the early 1990s to noninvasively study endothelium-dependent vasomotor function. In 1992, Celemejer et al10 used B-mode ultrasound to study the brachial artery’s vasoactive response to increased shear stress induced by hyperemia in adults with coronary artery disease (CAD) or smoking exposure. The
key observation, termed flow-mediated vasodilation and expressed as %FMD – was the presence of impaired vasoactivity detectable non-invasively with ultrasound in individuals with risk factors, CAD, or both. Concurrent with observations from basic studies and invasive coronary studies, flowmediated vasodilation in the brachial artery is abolished by the eNOS inhibitor NG-monomethylL-arginine (L-NMMA).11
