doi: 10.1161/01.RES.0000066880.62205.B0
© 2003 American Heart Association, Inc.
Editorials |
A Radical Adventure
The Quest for Specific Functions and Inhibitors of Vascular NAPDH Oxidases
From the Institut für Kardiovaskuläre Physiologie, Klinikum der J.W. Goethe-Universität, Frankfurt am Main, Germany.
Correspondence to Ralf P. Brandes, Institut für Kardiovaskuläre Physiologie, Klinikum der J.W. Goethe-Universität, Theodor-Stern-Kai 7, D-60596 Frankfurt am Main, Germany. E-mail r.brandes{at}em.uni-frankfurt.de
Key Words: oxidative stress • NADPH oxidase • restenosis • neointima
Over the past two decades, the number of scientific publications addressing the role of oxidative stress in physiology and pathophysiology has increased exponentially. Almost all cardiovascular disease states including hypertension, hyperlipidemia, diabetes, arteriosclerosis, unstable angina, vasculitis and myocarditis, restenosis as well as ischemia/reperfusion have been linked to an enhanced generation of oxygen-derived free radicals.1 Consequently, oxidative stress is generally considered to be a major progression factor for cardiovascular disease, despite the fact that the majority of the large prospective, controlled trials have failed to uncover a beneficial effect of antioxidative treatment.2
Therefore, one is left to wonder, whether an unspecific antioxidative radical scavenging approach is suitable to substantially lower oxidative stress and to affect disease progression. Indeed, individual sources of oxidative stress and the contribution of these enzymes to disease progression have to be identified. This approach may ultimately lead to the development of "specific" inhibitors of oxidative stress to target the intracellular source of free radical generation.
An important source of oxygen-derived radical generation are the NADPH oxidases, ubiquitous to all vascular cells.3 These enzymes, which in their subunit composition are either similar or even identical to the leukocyte NADPH oxidase can be induced and activated by many factors involved in the initiation and progression of cardiovascular disease such as thrombin, tumor necrosis factor-
, platelet-derived growth factor, and proatherosclerotic lipids such as lysophosphatidyl choline and Lp(a). The best characterized stimulus for NADPH oxidase activation and induction is angiotensin II, and experiments utilizing mice lacking different NADPH oxidase subunits have demonstrated that the oxidases plays a central role in angiotensin II–induced hypertension and hypertrophy.3
In this issue of Circulation Research, Jacobson et al report that inhibition of vascular NADPH oxidases suppresses angioplasty-induced neointimal hyperplasia in the carotid artery of rats.4 By using the specific peptide inhibitor gp91ds-tat5 applied over an extended observation period, this study is one of the few clearly demonstrating involvement of NADPH oxidases in a specific disease state. Moreover, by demonstrating that this peptide inhibitor not only prevents neointima formation, inhibits stretch-induced superoxide anion release from distended vessels, as well as peroxynitrite formation after angioplasty, the authors have forged a clear link between neointima formation and NADPH oxidase–dependent radical formation.
Numerous studies have previously suggested that stretch,6 neointima formation,7 and restenosis8 are all associated with an increased vascular superoxide anion release. But although an increased expression and activity of NADPH oxidases has been documented previously in these studies,7,8 the lack of specific oxidase inhibitors has precluded mechanistic investigations. With the use of gp91ds-tat, a direct evidence for an involvement of NADPH oxidases has now been provided.
gp91ds-tat appears to be the only specific NADPH oxidase inhibitor currently available. This chimeric peptide consists of a tat site, derived from the tat peptide of the HIV virus, allowing uptake into the cell, and a fragment of gp91phox (Nox2), which has previously been shown to prevent the interaction of p47phox with the Nox subunits in cell-free preparations.9 As this p47phox-blocking peptide sequence is specific for the NADPH oxidases, it is likely that gp91ds-tat acts specifically on the oxidase, which makes it a unique tool for studying the involvement of the NADPH oxidase, particular in in vivo models. Several pharmacological, nonpeptide-based inhibitors of the oxidase are available, but the lack of specificity of these compounds has been a longstanding matter of concern (Figure 1).
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Nevertheless, gp91ds-tat has some limitations. For example, it is not possible to differentiate between the Nox-containing oxidases as the peptide sequence is conserved in all vascular homologues of the oxidase. Moreover, the sensitization against the peptide will most probably lead to the formation of antibodies, limiting the treatment duration to a couple of weeks. Finally, the tat sequence of the peptide has been shown to cause side effects affecting cellular activity and signaling.10 Consequently, for chronic studies, specifically addressing the contribution of individual NADPH oxidase isoforms, additional molecular tools and compounds have to be developed.
An alternative approach is to use transgenic animals. Indeed, mice lacking the NADPH oxidase subunits p47phox and gp91phox (Nox2) have been used to demonstrate the involvement of the oxidase in angiotensin II–induced hypertension11 and hypertrophy12,13 as well as in VEGF-induced angiogenesis.14 In contrast, studies investigating the role of the oxidase in the development of arteriosclerosis, using ApoE/NADPH oxidase subunit double knockout mice on a Western-type diet were negative15,16 or indicate that the oxidase is not the main progression factor for arteriosclerosis in ApoE-/- mice.17 Such reports contrast with those describing an increased expression of NADPH oxidase subunits as well as enhanced vascular superoxide anion formation in arteriosclerosis18–20 and highlight the need for novel approaches that differentially address the contribution of different sources of oxidative stress during cardiovascular disease. Indeed, for early stages of arteriosclerosis, particular in the setting of hyperlipidemia, xanthine oxidase,21 cytochrome P450 epoxygenases,22 and NO synthase isoforms23 might be of greater importance than NADPH oxidases (Figure 2).
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Certainly, the present study addresses only one particular aspect of the role of NADPH oxidases in pathophysiology; the convincing demonstration of the effectiveness of gp91ds-tat peptide for inhibiting the enzyme in vivo, however, will spur on this field toward its goal of elucidating the contribution of the oxidases in other disease conditions. Ultimately, these efforts will lead to a deeper understanding of the role of oxygen-derived free radicals in vascular homeostasis and may reveal vascular NADPH oxidases as a new target for drug development.
Regarding this aspect, it is noteworthy that two classes of compounds widely used in cardiovascular practice today can act as potent, although nonspecific inhibitors of NADPH oxidases. ACE inhibitors, by inhibiting angiotensin II formation, prevent the induction and activation of the NADPH oxidase. HMG-CoA inhibitors (statins), by depleting the cells of isoprenoids, prevent the integration of Rac in the plasma membrane, where it is involved in activating the oxidase.24 Consequently, it is tempting to speculate that some of the beneficial effects of statins and ACE inhibitors on vascular homeostasis and cardiovascular disease are a consequence of NADPH oxidase inhibition.
Footnotes
The opinions expressed in this editorial are not necessarily those of the editors or of the American Heart Association.
References
1. Kojda G, Harrison DG. Interactions between NO and reactive oxygen species: pathophysiological importance in atherosclerosis, hypertension, diabetes and heart failure. Cardiovasc Res. 1999; 43: 562–571.[CrossRef][Medline] [Order article via Infotrieve]
2. Kritharides L, Stocker R. The use of antioxidant supplements in coronary heart disease. Atherosclerosis. 2002; 164: 211–219.[CrossRef][Medline] [Order article via Infotrieve]
3. Griendling KK, Sorescu D, Ushio-Fukai M. NAD(P)H oxidase: role in cardiovascular biology and disease. Circ Res. 2000; 86: 494–501.
4. Jacobson GM, Dourron HM, Liu J, Carretero OA, Reddy DJ, Andrzejewski T, Pagano PJ. Novel NAD(P)H oxidase inhibitor suppresses angioplasty-induced superoxide and neointimal hyperplasia of the rat carotid artery. Circ Res. 2003; 92: 637–643.
5. Rey FE, Cifuentes ME, Kiarash A, Quinn MT, Pagano PJ. Novel competitive inhibitor of NAD(P)H oxidase assembly attenuates vascular O2- and systolic blood pressure in mice. Circ Res. 2001; 89: 408–414.
6. Oeckler RA, Kaminski PM, Wolin MS. Stretch enhances contraction of bovine coronary arteries via an NAD(P)H oxidase-mediated activation of the extracellular signal-regulated kinase mitogen-activated protein kinase cascade. Circ Res. 2003; 92: 23–31.
7. Paravicini TM, Gulluyan LM, Dusting GJ, Drummond GR. Increased NADPH oxidase activity, gp91phox expression, and endothelium-dependent vasorelaxation during neointima formation in rabbits. Circ Res. 2002; 91: 54–61.
8. Szocs K, Lassegue B, Sorescu D, Hilenski LL, Valppu L, Couse TL, Wilcox JN, Quinn MT, Lambeth JD, Griendling KK. Upregulation of Nox-based NAD(P)H oxidases in restenosis after carotid injury. Arterioscler Thromb Vasc Biol. 2002; 22: 21–27.
9. DeLeo FR, Yu L, Burritt JB, Loetterle LR, Bond CW, Jesaitis AJ, Quinn MT. Mapping sites of interaction of p47-phox and flavocytochrome b with random-sequence peptide phage display libraries. Proc Natl Acad Sci U S A. 1995; 92: 7110–7114.
10. Jia H, Lohr M, Jezequel S, Davis D, Shaikh S, Selwood D, Zachary I. Cysteine-rich and basic domain HIV-1 Tat peptides inhibit angiogenesis and induce endothelial cell apoptosis. Biochem Biophys Res Commun. 2001; 283: 469–479.[CrossRef][Medline] [Order article via Infotrieve]
11. Landmesser U, Cai H, Dikalov S, McCann L, Hwang J, Jo H, Holland SM, Harrison DG. Role of p47phox in vascular oxidative stress and hypertension caused by angiotensin II. Hypertension. 2002; 40: 511–515.
12. Bendall JK, Cave AC, Heymes C, Gall N, Shah AM. Pivotal role of a gp91phox-containing NADPH oxidase in angiotensin II- induced cardiac hypertrophy in mice. Circulation. 2002; 105: 293–296.
13. Wang HD, Xu S, Johns DG, Du Y, Quinn MT, Cayatte AJ, Cohen RA. Role of NADPH oxidase in the vascular hypertrophic and oxidative stress response to angiotensin II in mice. Circ Res. 2001; 88: 947–953.
14. Ushio-Fukai M, Tang Y, Fukai T, Dikalov SI, Ma Y, Fujimoto M, Quinn MT, Pagano PJ, Johnson C, Alexander RW. Novel role of gp91phox-containing NAD(P)H oxidase in vascular endothelial growth factor-induced signaling and angiogenesis. Circ Res. 2002; 91: 1160–1167.
15. Kirk EA, Dinauer MC, Rosen H, Chait A, Heinecke JW, LeBoeuf RC. Impaired superoxide production due to a deficiency in phagocyte NADPH oxidase fails to inhibit atherosclerosis in mice. Arterioscler Thromb Vasc Biol. 2000; 20: 1529–1535.
16. Hsich E, Segal BH, Pagano PJ, Rey FE, Paigen B, Deleonardis J, Hoyt RF, Holland SM, Finkel T. Vascular effects following homozygous disruption of p47phox: an essential component of NADPH oxidase. Circulation. 2000; 101: 1234–1236.
17. Barry-Lane PA, Patterson C, van der MM, Hu Z, Holland SM, Yeh ET, Runge MS. p47phox is required for atherosclerotic lesion progression in ApoE-/- mice. J Clin Invest. 2001; 108: 1513–1522.[CrossRef][Medline] [Order article via Infotrieve]
18. Sorescu D, Weiss D, Lassegue B, Clempus RE, Szocs K, Sorescu GP, Valppu L, Quinn MT, Lambeth JD, Vega JD, Taylor WR, Griendling KK. Superoxide production and expression of nox family proteins in human atherosclerosis. Circulation. 2002; 105: 1429–1435.
19. Kalinina N, Agrotis A, Tararak E, Antropova Y, Kanellakis P, Ilyinskaya O, Quinn MT, Smirnov V, Bobik A. Cytochrome b558-dependent NAD(P)H oxidase-phox units in smooth muscle and macrophages of atherosclerotic lesions. Arterioscler Thromb Vasc Biol. 2002; 22: 2037–2043.
20. Azumi H, Inoue N, Ohashi Y, Terashima M, Mori T, Fujita H, Awano K, Kobayashi K, Maeda K, Hata K, Shinke T, Kobayashi S, Hirata K, Kawashima S, Itabe H, Hayashi Y, Imajoh-Ohmi S, Itoh H, Yokoyama M. Superoxide generation in directional coronary atherectomy specimens of patients with angina pectoris: important role of NAD(P)H oxidase. Arterioscler Thromb Vasc Biol. 2002; 22: 1838–1844.
21. Ohara Y, Peterson TE, Harrison DG. Hypercholesterolemia increases endothelial superoxide anion production. J Clin Invest. 1993; 91: 2546–2551.[Medline] [Order article via Infotrieve]
22. Fleming I, Michaelis UR, Bredenkotter D, Fisslthaler B, Dehghani F, Brandes RP, Busse R. Endothelium-derived hyperpolarizing factor synthase (cytochrome P450 2C9) is a functionally significant source of reactive oxygen species in coronary arteries. Circ Res. 2001; 88: 44–51.
23. Laursen JB, Somers M, Kurz S, McCann L, Warnholtz A, Freeman BA, Tarpey M, Fukai T, Harrison DG. Endothelial regulation of vasomotion in apoE-deficient mice: implications for interactions between peroxynitrite and tetrahydrobiopterin. Circulation. 2001; 103: 1282–1288.
24. Wagner AH, Kohler T, Ruckschloss U, Just I, Hecker M. Improvement of nitric oxide-dependent vasodilatation by HMG-CoA reductase inhibitors through attenuation of endothelial superoxide anion formation. Arterioscler Thromb Vasc Biol. 2000; 20: 61–69.
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