Kinin-Mediated Coronary Nitric Oxide Production Contributes to the Therapeutic Action of Angiotensin-Converting Enzyme and Neutral Endopeptidase Inhibitors and Amlodipine in the Treatment in Heart Failure1

Increasing evidence suggests that angiotensin-converting enzyme (ACE) inhibitors can increase vascular nitric oxide (NO) production. Recent studies have found that combined inhibition of ACE and neutral endopeptidase (NEP) may have a greater beneficial effect in the treatment of heart failure than inhibition of ACE alone. Amlodipine, a calcium channel antagonist, has also been reported to have a favorable effect in the treatment of patients with cardiac dysfunction. The purpose of this study was to determine whether and the extent to which all of these agents used in the treatment of heart failure stimulate vascular NO production. Heart failure was induced by rapid ventricular pacing in conscious dogs. Coronary microvessels were isolated from normal and failing dog hearts. Nitrite, the stable metabolite of NO, was measured by the Griess reaction. ACE and NEP inhibitors and amlodipine significantly increased nitrite production from coronary microvessels in both normal and failing dog hearts. However, nitrite release was reduced after heart failure. For instance, the highest concentration of enalaprilat, thiorphan, and amlodipine increased nitrite release from 85 6 4 to 156 6 9, 82 6 7 to 139 6 8, and 74 6 4 to 134 610 pmol/mg (all *p , .01 versus control), respectively, in normal dog hearts. Nitrite release in response to the highest concentration of these two inhibitors and amlodipine was reduced by 41% and 31% and 32% (all #p , .01 versus normal), respectively, in microvessels after heart failure. The increase in nitrite induced by either ACE or NEP inhibitors or amlodipine was entirely abolished by N-nitro-L-arginine methyl ester, HOE 140 (a B2-kinin receptor antagonist), and dichloroisocoumarin (a serine protease inhibitor) in both groups. Our results indicate that: 1) there is an impaired endothelial NO production after pacing-induced heart failure; 2) both ACE and NEP are largely responsible for the metabolism of kinins and modulate canine coronary NO production in normal and failing heart; and 3) amlodipine releases NO even after heart failure and this may be partly responsible for the favorable effect of amlodipine in the treatment of heart failure. Thus, the restoration of reduced coronary vascular NO production may contribute to the beneficial effects of these agents in the treatment of heart failure. Nitric oxide (NO) derived from vascular endothelium plays an important role in the regulation of many biological functions including vasodilation and mitochondrial respiration (Moncada et al., 1991). Increasing evidence indicates that there is an impaired endothelial NO production during the development of heart failure in humans and animals (Treasure et al., 1990; Katz et al., 1993; Drexler et al., 1994; Wang et al., 1994). Experimental and clinical studies have demonstrated that angiotensin-converting enzyme (ACE) inhibition has an antihypertensive and cardioprotective action, partly by preventing kinin degradation and consequently increasing endothelial NO production (Schwelk et al., 1993; Linz et al., 1995; Scholkens, 1996). Numerous studies have shown the existence of the kallikrein-kinin system in cardiac and vascular tissue in animals (Nolly et al., 1993, 1994; Wiemer et al., 1994; Linz et al., 1995; Scholkens 1996; Zhang et al., 1997). A variety of endoand exopeptidases, which are widely distributed in various tissue and cell types contribute to the degradation of kinins (Erdos and Skidgel, 1989; Skidgel, 1992). ACE is the primary enzyme responsible for this catabolism. However, growing data from recent studies suggest that neutral endopeptidase (NEP) is also at least partially responsible for regulating the metabolism of kinins in the tissue from a variety of species (Graf et al., 1993; Trippodo et al., 1995a,b; Dragovic et al., 1996). Yang et al. (1997) Received for publication March 16, 1998. 1 These studies were supported by PO-1-HL 43023, HL 50142, HL 18579, and HL 53053 from the National Heart, Lung and Blood Institute and by Fellowship 96-103 from the New York Affiliate of the American Heart Association (to X.Z.). ABBREVIATIONS: ACE, angiotensin-converting enzyme; NO, nitric oxide; NEP, neutral endopeptidase; PBS, phosphate-buffered saline; L-NAME, N-nitro-L-arginine methyl ester; DCIC, dichloroisocoumarin; ANF, atrial natriuretic factor. 0022-3565/99/2882-0742$03.00/0 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 288, No. 2 Copyright © 1999 by The American Society for Pharmacology and Experimental Therapeutics Printed in U.S.A. JPET 288:742–751, 1999 742 at A PE T Jornals on O cber 5, 2017 jpet.asjournals.org D ow nladed from recently reported that inhibition of NEP protects the heart against ischemia/reperfusion injury by a kinin-dependent mechanism. Furthermore, a study from our laboratory (Zhang et al., 1998a) found that NEP inhibitors can release nitrite from canine coronary microvessels. All of these data suggest that NEP might be one of the main peptidases participating in the metabolism of kinins. Accordingly, we hypothesized that inhibition of NEP may also have a beneficial effect on the treatment of heart failure by potentiation of kinins. In addition, although calcium antagonists have not been shown to be beneficial in the treatment of patients with heart failure, a recent clinical trial (Packer et al., 1996) has demonstrated a favorable effect of amlodipine on the survival of patients with heart failure resulting from nonischemic dilated cardiomyopathy. A new concept for the treatment of heart failure has been suggested to combine ACE inhibitors with calcium-channel antagonists (Lliceto, 1997; Waeber and Brunner, 1997), because calcium antagonists and ACE inhibitors exhibit additive antihypertensive efficacy and counterbalance the negative effects caused by neurohormonal activation when combined, and their safety profile is, if anything, improved. However, the mechanism of the favorable action of amlodipine has not been determined. A recent study by Lyons et al. (1994) found that like enalaprilat, amlodipine significantly restored forearm arterial vasoconstriction to local intra-arterial infusions of N-monomethyl-L-arginine. We inferred from that study that amlodipine may mimic the effect of ACE inhibitors and promote NO production. With this in mind, the present study was designed to directly measure the hydration product of NO, nitrite, to determine: 1) whether inhibition of NEP can increase coronary microvascular NO production, and whether the mechanism is similar to that of ACE inhibition; 2) whether amlodipine increases NO production and whether this effect can be affected by blockade of B2-kinin receptor or inhibition of local kinin formation; and 3) whether ACE or NEP inhibition or amlodipine can also increase NO production from coronary microvessels after the development of severe congestive heart failure. Materials and Methods