Brief Reports

Effects of changing the availability of the substrate for nitric oxide synthase by l-arginine administration on coronary vasomotor tone in angina patients with angiographically narrowed and in patients with normal coronary arteries

Due to ageing populations and improvements in survival, increasing numbers of patients suffering from ischemic cardiovascular disease are not amenable to revascularization. Hence, interests are currently focused on “therapeutic angiogenesis”, which is the clinical use of growth factors to enhance or promote the development of collateral blood vessels in ischemic tissue. Several growth factors (or genes encoding these growth factors) are now available for therapeutic vascular growth in normal and ischemic tissues. However, the successes of angiogenic therapy observed in pre-clinical studies have not been realized in clinical trials. Most animal studies demonstrating the physiologic effectiveness of angiogenic therapies have been performed in normal young animals, while clinical trials typically enroll older patients with various endothelial disruptive risk factors. The promising results of trials using endothelial function-improving strategies support the hypothesis that the decreased effectiveness of growth factor therapy due to endothelial dysfunction could be a principle reason for failure of trials using growth factors. We will have a retrospection of therapeutic angiogenesis trials and discuss the mechanisms that contribute to an impaired angiogenic response in the setting of endothelial dysfunction. We also briefly explore endothelial function-improving procedures that have the potentially therapeutic benefit of enhancing the angiogenic response.

Endothelial dysfunction and decreased nitric oxide bioavailability may explain why therapeutic angiogenesis and cell therapy have mostly failed in humans. Building from previous large animal work, the Phase I Endothelial Modulation in Angiogenic Therapy trial tested the hypothesis that L-arginine, a nitric oxide donor, may be safe and effective in potentiating surgical angiogenesis in humans.

Patients with surgical triple-vessel coronary disease and a severely diffusely diseased left anterior descending artery were randomized in 2 × 2 factorial fashion to receive ten 200-μg injections of vascular endothelial growth factor-165 plasmid DNA or placebo in the anterior myocardium along the proximal and mid-left anterior descending arteries, plus oral L-arginine supplementation at a dose of 6 g per day or placebo for 3 months. The distal left anterior descending artery and other coronary arteries were grafted. End points included 3-month changes in myocardial perfusion and contractility of the anterior myocardium, using ¹³N-ammonia positron emission tomography and echocardiography. Baseline scans were obtained 3 to 7 days postoperatively to delineate treatment effects from the effects of coronary artery bypass grafting.

Patient (N = 19) characteristics were equivalent between groups. There was no perioperative or late mortality. Patients who received the combination of vascular endothelial growth factor and L-arginine had improved anterior wall perfusion on positron emission tomography (P = .02), a trend toward smaller perfusion defects (P = .10), and better anterior wall contractility (P = .02, Kruskal–Wallis) at 3 months versus baseline. This was corroborated by a trend toward better disease perception at 3 months versus baseline on the Seattle Angina Questionnaire (score improvement of 47 ± 35, combination treatment group; P = .1, Kruskal–Wallis).

To our knowledge, this is the first study to examine concomitant substrate modification in patients undergoing new biosurgical therapies by using vascular endothelial growth factor angiogenesis. The results suggest safety and efficacy. Concomitant endothelial modulation with L-arginine not only has the potential to make angiogenesis effective but also may have implications for cell therapy trials.

Nitric oxide (NO) is a molecule that has gained recognition as a crucial modulator of vascular disease. NO has a number of intracellular effects that lead to vasorelaxation, endothelial regeneration, inhibition of leukocyte chemotaxis, and platelet adhesion. Endothelium damage induced by atherosclerosis leads to the reduction in bioactivity of endothelial NO synthase (eNOS) with subsequent impaired release of NO together with a local enhanced degradation of NO by increased generation of reactive oxygen species with subsequent cascade of oxidation-sensitive mechanisms in the arterial wall. Many commonly used vasculoprotective agents have their therapeutic actions through the production of NO. l-Arginine, the precursor of NO, has demonstrated beneficial effects in atherosclerosis and disturbed shear stress. Finally, eNOS gene polymorphism might be an additional risk factor that may contribute to predict cardiovascular events. However, further studies are needed to understand the possible clinical implications of these correlations.

Endothelial dysfunction has been shown in a wide range of vascular desorders including atherosclerosis and related diseases. Here, we examine and address the complex relationship among nitric oxide (NO)-mediated pathways and atherogenesis. In view of the numerous pathophysiological actions of NO, abnormalities could potentially occur at many sites: (a) impairment of membrane receptors in the arterial wall that interact with agonists or physiological stimuli capable of generating NO; (b) reduced concentrations or impaired utilization of $\text{l}$-arginine; (c) reduction in concentration or activity both of inducible and endothelial NO synthase; (d) impaired release of NO from the atherosclerotic damaged endothelium; (e) impaired NO diffusion from endothelium to vascular smooth muscle cells followed by decreased sensitivity to its vasodilator action; (f) local enhanced degradation of NO by increased generation of free radicals and/or oxidation-sensitive mechanisms; and (g) impaired interaction of NO with guanylate cyclase and consequent limitation of cyclic GMP production. Therefore, one target for new drugs should be the preservation or restoration of NO-mediated signaling pathways in arteries. Such novel therapeutic strategies may include administration of $\text{l}$-arginine/antioxidants and gene-transfer approaches.

Nitric oxide is formed from the N-guanido terminal of the amino acid l-arginine and from molecular oxygen by nitric oxide synthase enzymes. l-arginine administration improves the coronary blood flow response to acetylcholine in patients with normal coronary arteries and hypercholesterolemia, reverses the defective endothelium-dependent vasodilation associated with an elevated plasma low-density lipoprotein level or hypercholesterolemia, dilates coronary epicardial arteries and stenoses, enhances nitric oxide generation, and inhibits lesion formation after balloon angioplasty. Stimulation of endogenous nitric oxide production could inhibit atherogenesis, and therefore may be of benefit in patients with risk factors for atherosclerosis.