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(Hypertension. 2003;42:1150.)
© 2003 American Heart Association, Inc.

Scientific Contributions

Role of Superoxide in Modulating the Renal Effects of Angiotensin II

Bernardo López; Miguel García Salom; Begoña Arregui; Fernando Valero; Francisco J. Fenoy

From Departamento de Fisiología, Facultad de Medicina, Universidad de Murcia, 30100-Murcia, Spain.

Correspondence to Francisco J. Fenoy, Departamento de Fisiología, Facultad de Medicina, Campus Universitario de Espinardo, 30100-Murcia, Spain. E-mail fjfenoy{at}um.es

	Abstract

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Angiotensin II is known to stimulate NADPH oxidase-dependent superoxide (O₂^-) generation, which may contribute to the acute renal vasoconstrictor and antinatriuretic actions of this peptide. To evaluate this hypothesis, the effects of a superoxide dismutase mimetic (tempol) or a NADPH inhibitor (apocynin) on the angiotensin renal actions were studied. Renal cortical nitric oxide (NO) was measured electrochemically in vivo. Tempol increased sodium excretion and NO levels. Apocynin raised renal blood flow, glomerular filtration rate, sodium excretion, and NO levels. These results indicate the presence of an endogenous NADPH oxidase-dependent O₂^- generation that may modulate renal function by scavenging NO. Angiotensin II infusion reduced renal blood flow, glomerular filtration, sodium excretion, and NO levels in a dose-dependent manner. The angiotensin receptor antagonist valsartan, tempol, or apocynin blunted the angiotensin effects on renal excretion and NO, suggesting that angiotensin receptors stimulation induces the NADPH oxidase-dependent O₂^- generation that might reduce NO bioavailability. This idea is supported by the finding that angiotensin increased O₂^- generation in renal homogenates, and this effect was prevented by valsartan, apocynin, or tempol. These results indicate that some of the acute renal effects of angiotensin II may be enhanced by an increased NADPH oxidase-derived O₂^- production that reduces renal NO bioavailability.

Key Words: antioxidants • kidney • oxygen • nitric oxide • angiotensin II

	Introduction

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The renin-angiotensin system plays a critical role in the regulation of renal function and arterial pressure. In addition to the classic signal transduction mechanisms, the angiotensin II (Ang II) receptor type 1 (AT₁R) activates the NADPH oxidase-dependent generation of superoxide (O₂^-).¹ Although O₂^- modulates some of the Ang II actions, the relative contribution of increased superoxide generation to the acute renal effects of Ang II in vivo remains to be established.

In recent years, the physiological role of endogenous O₂^- generation has been examined mostly in relation to vascular function. In the normal vascular wall, O₂^- is produced primarily in vascular smooth muscle cells (VSMCs) and fibroblasts.¹ The major enzyme responsible for vascular O₂^- appears to be NADPH oxidase,² which catalyzes the production of O₂^- by the reduction of oxygen.³ O₂^- is known to avidly react with nitric oxide (NO) to form peroxynitrite,⁴ thus diminishing the half-life of NO. Ang II regulates⁵ and increases NADPH oxidase-dependent O₂^- production in cultured VSMCs,⁶ in aortic adventitial fibroblasts,⁷ and in mesangial cells.⁸

A role for O₂^- in the control of renal function seems likely because recent studies in rats indicate that O₂^- production in the renal medulla exerts vasoconstrictor as well as antidiuretic and antinatriuretic actions.⁹ The mRNA encoding the 5 subunits of the classic phagocyte NADPH oxidase and the proteins p22phox, p47phox, and p67phox have been shown to be present in the normal adult rat kidney.¹⁰ It has also been reported that treatment with an AT₁R antagonist, a superoxide dismutase (SOD) mimetic, or a NADPH oxidase inhibitor reduces the oxidative stress in spontaneously hypertensive rats,¹¹ inhibits the enhanced tubuloglomerular feedback,¹² blunts sodium reabsorption in the thick ascending limb of the loop of Henle,¹³ and decreases the NADPH oxidase-stimulated O₂^- production in rat arteries.¹⁴ In addition, the intrarenal infusion of a superoxide dismutase inhibitor produces marked vasoconstriction and antinatriuresis; because these effects are potentiated by NO synthesis inhibition, it has been postulated that the renal actions of superoxide are buffered by nitric oxide.¹⁵ Therefore, it seems that the balance between superoxide and nitric oxide is important in controlling renal function, but the in vivo effects of Ang II affecting this equilibrium and contributing to modulate renal excretion and hemodynamics are unknown.

Although the experimental evidence supports a role for O₂^- in the regulation of renal function, little is known about the renal effects of NADPH oxidase inhibitors or O₂^- scavengers in vivo. In addition, the contribution of NADPH oxidase-dependent generation of O₂^- to the acute renal actions of Ang II remains to be established. In the present study, the response to intrarenal infusions of a NADPH oxidase inhibitor (apocynin) and a SOD mimetic (tempol) were evaluated to exclude that sources of superoxide other than NADPH oxidase may regulate renal function. Furthermore, the effects of pretreating the kidney with tempol or apocynin on the renal actions of Ang II were studied.

	Methods

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Experiments were performed on 63 anesthetized male Sprague-Dawley rats (weight, 250 g) anesthetized with ketamine (30 mg/kg IM) and thiopental (50 mg/kg IP). The surgery and basic experimental techniques were previously described.¹⁶ The left kidney was exposed and stabilized in a renal cup to allow for measurement of NO, bathed in saline at 37°C. Renal cortical [NO] was measured with carbon fiber microelectrodes¹⁷ by differential normal pulse voltammetry. With this technique, NO produces an oxidation peak at 0.65 V.¹⁸

All drugs (tempol, apocynin, valsartan, and Ang II) were administered intrarenally¹⁶ in the same volume of fluid (isotonic NaCl solution, at a rate of 50 µL/min), and an intravenous infusion of saline containing [³H]inulin (1 µCi/mL, to allow for glomerular filtration rate measurements) at a rate of 1 mL/100 g per hour was given. The lucigenin-derived chemiluminescence assay⁵ was used to determine superoxide in renal cortex homogenates.

Protocol 1
Protocol 1 involved characterization of the response of the NO electrodes in vivo (n=5). L-Nitroarginine methylester (10 mg/kg) was given intravenously, and the NO peak current was measured. An intravenous infusion of sodium nitroprusside was started at a dose (20 µg/kg per minute) that normalizes arterial pressure, and the NO current was redetermined.

Protocol 2
Protocol 2 involved time course studies (n=7). Renal function and [NO] levels were evaluated during a 1-hour period.

Protocols 3 and 4
Protocols 3 and 4 involved characterization of the renal effects of tempol or apocynin (n=6). After a control period, increasing doses of tempol (4, 8, and 16 µmol/kg per minute to yield 250, 500, and 1000 µmol/L in renal arterial plasma at baseline renal blood flow [RBF]^19,12) or apocynin (5, 10, and 50 µmol/kg per minute to achieve 300, 600, and 3000 µmol/L renal arterial plasmatic concentrations at baseline RBF¹⁴) were consecutively infused into the renal artery for 10 minutes each, and renal function and [NO] were measured during a 10-minute clearance period for each dose. The highest dose of tempol or apocynin tended to decrease arterial pressure; this was avoided by occluding the mesenteric and celiac arteries and by using an aortic clamp.²⁰ Apocynin is a specific inhibitor of NADPH oxidase in blood vessels, with no other known actions.¹⁴

Protocol 5
Protocol 5 involved the renal effects of tempol plus apocynin (n=6). These experiments evaluated whether these compounds have additive effects, as an indication that they might produce actions other than suppressing O₂^-. The doses were the same used above.

Protocols 6 to 9
Protocols 6 to 9 involved the renal effects of Ang II alone (n=7) or in rats pretreated with valsartan (n=7), tempol (n=7), or apocynin (n=6). Either saline, valsartan (160 nmol/kg per minute to yield -10 µmol/L in renal arterial plasma), tempol (8 µmol/kg per minute to achieve 500 µmol/L in renal arterial plasma), or apocynin (10 µmol/kg per minute to obtain 600 µmol/L in renal arterial plasma) were infused. The doses of tempol or apocynin used were the maximum that alter renal function without affecting arterial pressure. Then, Ang II at doses of 1.6, 16, and 160 pmol/kg per minute was successively infused intrarenally (to obtain 0.1, 1, and 10 nmol/L renal arterial plasma concentrations at baseline RBF).

Protocol 10
Protocol 10 involved superoxide levels in renal cortex homogenates (n=6). Ang II (10 nmol/L), valsartan (10 µmol/L), apocynin (600 µmol/L), tempol (500 µmol/L), Ang II plus valsartan, Ang II plus tempol, or Ang II plus apocynin (same doses) were added to the samples and incubated in presence of lucigenin.

	Results

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Baseline values of renal function and [NO] levels in all groups are presented in the Table.

View this table: [in this window] [in a new window]   Control Values of Renal Function, Renal Cortex NO Concentration, and Urinary NOx Excretion

Characterization of the Response of the NO Electrode In Vivo
When the electrode is introduced in the renal cortex, an oxidation current peak can be measured at 650 mV (Figure 1). This voltage coincides with the oxidation peak found in vitro in anaerobic NO solutions. The peak height decreases after L-NAME is given (to 24% of control) and is restored during the infusion of the NO donor sodium nitroprusside (to 93% of control), indicating that it is probably due to NO.

View larger version (17K): [in this window] [in a new window]   Figure 1. Top, Voltammograms recorded in renal cortex during infusion of saline (control), L-NAME (10 mg/kg IV), or L-NAME plus sodium nitroprusside (SNP, 20 µg/kg per minute IV). Bottom, Changes in current peak height produced during infusion of saline (control), L-NAME (10 mg/kg IV), or L-NAME plus SNP (20 µg/kg per minute IV). #P<0.01 from control.

Renal Effects of Tempol, Apocynin, or Tempol Plus Apocynin
Tempol had no significant effects on RBF and slightly decreased glomerular filtration rate (GFR) (-13%) and the filtration fraction (FF, -15%) at the higher dose used (Figure 2). On the other hand, apocynin increased RBF (+31%) and GFR (+19%), having no effect on FF. The coadministration of tempol plus apocynin had no additive effects on renal hemodynamics.

View larger version (18K): [in this window] [in a new window]   Figure 2. Dose-dependent effects of intrarenal infusion of a SOD mimetic (tempol, 4, 8, and 16 µmol/kg per minute, experimental periods E1, E2, and E3, respectively), a NADPH oxidase inhibitor (apocynin, 5, 10, and 50 µmol/kg per minute, experimental periods E1, E2, and E3, respectively), or tempol plus apocynin (same doses coadministered in experimental periods E1, E2, and E3, respectively) on RBF, GFR, and FF. P<0.05, P<0.01, from corresponding value in the time-control group.

Tempol raised U_NaV (+33%) and FE_Na (31%, Figure 3). Apocynin also increased U_NaV (+122%) and FENa (+67%). The coadministration of tempol plus apocynin had no additive effects on renal sodium excretion.

View larger version (22K): [in this window] [in a new window]   Figure 3. Dose-dependent effects of intrarenal infusion of a SOD mimetic (tempol, 4, 8, and 16 µmol/kg per minute, experimental periods E1, E2, and E3, respectively), a NADPH oxidase inhibitor (apocynin, 5, 10, and 50 µmol/kg per minute, experimental periods E1, E2, and E3, respectively), or tempol plus apocynin (same doses coadministered in experimental periods E1, E2, and E3, respectively) on UNaV and FENa. P<0.01 from corresponding value in the time-control group.

Tempol increased [NO] (88%) and U_NOxV (+68%, Figure 4). In addition, apocynin raised [NO] (+114%) and U_NOxV (+98%). The infusion of tempol plus apocynin had no additive effects on renal [NO].

View larger version (22K): [in this window] [in a new window]   Figure 4. Dose-dependent effects of intrarenal infusion of a SOD mimetic (tempol, 4, 8, and 16 µmol/kg per minute, experimental periods E1, E2, and E3, respectively), a NADPH oxidase inhibitor (apocynin, 5, 10, and 50 µmol/kg per minute, experimental periods E1, E2, and E3, respectively), or tempol plus apocynin (same doses coadministered in experimental periods E1, E2, and E3, respectively) on renal cortical [NO] and UNOxV. P<0.05, P<0.01, from corresponding value in the time-control group.

Renal Effects of Ang II, Tempol Plus Ang II, and Apocynin Plus Ang II
Ang II similarly reduced RBF (-36%) and GFR (-35%), having no effect on FF (Figure 5). During the infusion of tempol or apocynin, Ang II still reduced RBF, but it had no effect on GFR, thus increasing FF (Figure 5).

View larger version (21K): [in this window] [in a new window]   Figure 5. Dose-dependent effects of an intrarenal infusion of Ang II alone (ANG II, 1.6, 16, and 160 pmol/kg per minute) or Ang II (same doses) in kidneys pretreated with either a SOD mimetic (tempol, 8 µmol/kg per minute) or a NADPH oxidase inhibitor (apocynin, 10 µmol/kg per minute) on RBF, GFR, and FF. C indicates control; pret, pretreatment. *P<0.05, #P<0.01 from control period of the same group; P<0.05, P<0.01 from corresponding value in the Ang II group.

Ang II lowered U_NaV mainly because of the fall in filtered load, because it did not decrease FENa (Figures 5 and 6). Pretreatment with tempol blocked the antinatriuresis produced by Ang II by preventing the fall in GFR, with no effects on tubular sodium reabsorption. In rats given apocynin, however, Ang II increased U_NaV (+61%) by decreasing tubular sodium reabsorption (Figure 6).

View larger version (24K): [in this window] [in a new window]   Figure 6. Dose-dependent effects of an intrarenal infusion of Ang II alone (ANG II, 1.6, 16, and 160 pmol/kg per minute) or Ang II (same doses) in kidneys pretreated with either a SOD mimetic (tempol, 8 µmol/kg per minute) or a NADPH oxidase inhibitor (apocynin, 10 µmol/kg per minute) on UNaV and FENa. C indicates control; pret, pretreatment. *P<0.05, #P<0.01 from control period of the same group; P<0.05, P<0.01 from corresponding value in the Ang II group.

Ang II alone decreased [NO] (-23%) and U_NOxV (-24%). However, in rats pretreated with tempol or apocynin, both [NO] and U_NOxV rose during the Ang II administration (Figure 7).

View larger version (22K): [in this window] [in a new window]   Figure 7. Dose-dependent effects of an intrarenal infusion of Ang II alone (ANG II, 1.6, 16, and 160 pmol/kg per minute) or Ang II (same doses) in kidneys pretreated with either a SOD mimetic (tempol, 8 µmol/kg per minute) or a NADPH oxidase inhibitor (apocynin, 10 µmol/kg per minute) on renal cortex [NO] and UNOxV. C indicates control; pret, pretreatment. *P<0.05, #P<0.01 from control period of the same group; P<0.05, P<0.01 from corresponding value in the Ang II group.

Renal Effects of Ang II in Rats Pretreated With Valsartan
Valsartan lowered mean arterial pressure (MAP) (from 120±3 to 114±3 mm Hg) and raised RBF (from 7.67±0.31 to 8.73±0.40 mL/min per gram), but it had no effect on GFR, U_NaV, and [NO]. The subsequent intrarenal infusion of Ang II during the administration of valsartan had no effects on MAP, RBF, GFR, and [NO]. The actions of Ang II on U_NaV were blunted by valsartan, because only the highest dose of angiotensin used (160 pmol/kg per minute) decreased U_NaV (from 7.09±0.67 to 6.08±0.64 µEq/min per gram).

O₂^- Levels in Renal Cortex Homogenates
The levels of O₂^- in renal homogenates, as estimated by lucigenin-enhanced chemiluminescence, increased by +17% in Ang II-incubated (10 nmol/L) cortical homogenates compared with basal O₂^- generation (2283.94±188.21 photons/min per milligram of protein) (Figure 8). In contrast, valsartan (10 µmol/L), tempol (500 µmol/L), or apocynin (600 µmol/L) markedly reduced basal O₂^- levels (by -37%, -30%, and -60%, respectively). In renal homogenates preincubated with either valsartan, tempol, or apocynin, Ang II had no effect on O₂^- levels.

View larger version (22K): [in this window] [in a new window]   Figure 8. Superoxide (O2-) levels in renal cortex homogenates incubated with vehicle [C, control], Ang II (ANG II, 10 nmol/L), valsartan (V, 10 µmol/L), tempol (T, 500 µmol/L), apocynin (A, 600 µmol/L), valsartan plus Ang II (V+AII, same concentrations), tempol plus Ang II (T+AII, same concentrations), or apocynin plus Ang II (A+AII, same concentrations). P<0.05 from control value; P<0.05 from homogenates incubated with Ang II (n=6).

	Discussion

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The present study examined the renal responses to an intrarenal infusion of either a SOD mimetic or a NADPH oxidase inhibitor. Scavenging renal superoxide with tempol increased sodium excretion as well as renal cortex NO activity and nitrite plus nitrate excretion. In addition, the inhibition of NADPH oxidase with apocynin raised RBF, GFR, U_NaV, [NO], and U_NOxV. The fact that these two compounds have no additive effects is compatible with the hypothesis that both affect renal function by reducing superoxide. These results indicate the existence of a basal O₂^- production in renal tissue, which appears to be NADPH oxidase-dependent,¹⁰ exerting a tonic regulatory action on renal hemodynamics⁹ and tubular sodium transport that may be partly mediated by affecting renal NO bioavailability.^13,14 This physiological mechanism is compatible with the recent finding that inhibition of renal superoxide dismutase induces renal vasoconstriction and reduces urinary sodium excretion.¹⁵ Interestingly, tempol produced less renal vasodilation and natriuresis than apocynin. The reasons explaining this interesting phenomenon are unknown, but one may speculate that it may be related to the fact that tempol produces hydrogen peroxide from superoxide, and it might be responsible for decreasing GFR and limiting the vasodilation and increased natriuresis produced by NO during the infusion of tempol, as it has been recently shown.²¹ Also, because apocynin affects renal function more than tempol, it can be concluded that NADPH oxidase appears to be the only source of superoxide influencing renal function. An alternative explanation would be that the doses of tempol used were too low to affect renal function. This seems unlikely, because the concentrations reached in renal arterial plasma are in the range previously tested in vitro.^12,19 In any case, higher doses of tempol cause arterial pressure to drop, and therefore their effect on renal function cannot be evaluated in vivo.

When the rats were infused intrarenally with increasing doses of Ang II, RBF, GFR, U_NaV, [NO], and U_NOxV fell in a dose-dependent manner, as described previously in dogs²² or rats.²³ Ang II increases filtration fraction in dogs, and this has been interpreted as a preferential efferent arteriolar action²²; however, this effect does not take place in rats because the peptide causes in this species similar afferent and efferent vasoconstriction.²³ The generation of O₂^- induced by Ang II may contribute to the renal vasoconstriction and antinatriuresis either by a direct effect on vascular smooth muscle cells^1,24 or indirectly, by acting on the juxtaglomerular apparatus and enhancing the tubuloglomerular feedback response.¹² The Ang II-induced decrease in [NO] observed in the present study is consistent with previous reports that a chronic infusion of Ang II markedly depressed vascular NO production (despite the fact that Ang II increased NOS protein in the kidney) by inducing NADPH oxidase-dependent O₂^- generation²⁵ and lowered urinary NOx excretion.²⁶ This interaction between O₂^- and NO has been demonstrated in vitro because it has been shown that O₂^- decreases NO bioavailability in single renal tubules.¹³ However, these findings contrast with other reports suggesting that Ang II increases NO concentrations in the renal cortex²⁷ and that activation of the renin-angiotensin system during sodium depletion, or Ang II infusion in rats maintained on a normal sodium intake, stimulate a vasodilator signaling cascade that includes bradykinin, NO, and cGMP.²⁸ The reasons explaining these differences are unknown, but they may be related to the fact that in these studies Ang II was infused intravenously, and the concentration of the peptide reaching the kidney may have been insufficient to stimulate O₂^- generation.

The acute intrarenal infusion of valsartan lowered MAP and raised RBF without affecting GFR, U_NaV, and [NO]. Similarly, it has been reported that losartan had no effect on renal cortical cGMP levels in sodium-depleted or sodium-replete rats.²⁸ In addition, the effects of Ang II on RBF, GFR, U_NaV, [NO], and U_NOxV were totally abolished by pretreatment with the AT₁R antagonist. These results indicate that the physiological effects of Ang II are mainly mediated through the stimulation of the AT₁R. The fact that the fall observed in cortical NO concentration and U_NOxV is prevented by valsartan suggests that the increase in renal O₂^- levels is triggered by AT₁R stimulation, as previously suggested.²⁹ An alternative possibility would be that Ang II may act on AT₂ receptors during AT₁R blockade and increase NO levels.²⁸

Pretreatment with tempol had no effect on the renal vasoconstriction produced by Ang II. However, the SOD mimetic prevented the Ang II-induced fall in GFR, therefore increasing filtration fraction, suggesting a preferential afferent dilatory effect. Tempol also maintained renal cortical NO, U_NOxV, and U_NaV during the infusion of Ang II. These results are compatible with the idea that the increase in O₂^- generation produced by Ang II reduces NO within the kidney, thus contributing to some of the excretory and glomerular effects of the peptide. It is known that Ang II reduces glomerular surface area³⁰ by promoting mesangial cells contraction³¹ and O₂^- production.⁸ It is also known that those mesangial effects of Ang II can be blunted by NO.³¹ Therefore, it seems reasonable to postulate that the effect of tempol preventing the fall in GFR produced by Ang II may be due to increased NO bioavailability, as previously suggested.²⁶ A role for O₂^- in potentiating the antinatriuresis produced by Ang II seems likely, since it is known that O₂^- reduces sodium excretion^9,15 and because this Ang II effect is also prevented by tempol. In addition, tempol precludes the increase in plasma endothelin and isoprostanes caused by Ang II, and this action is thought to participate in impairing some of the renal hemodynamic Ang II effects.²⁶

In the present study, pretreatment with the NADPH oxidase inhibitor apocynin raised RBF, GFR, U_NaV, [NO], and U_NOxV. Apocynin also blunted the renal vasoconstriction observed during the subsequent infusion of Ang II and increased filtration fraction, indicating a preferential afferent arteriolar effect. This suggests that the stimulation of NADPH oxidase-dependent O₂^- generation contributes to the vasoconstrictor effects of Ang II and is compatible with the recent finding that O₂^- is a potent renal vasoconstrictor.^9,15 Apocynin abolished the actions of Ang II on GFR, U_NaV, [NO], and U_NOxV. It has been reported that Ang II specifically activates the NADPH oxidase enzyme system and promotes O₂^- generation.⁶ Also, it is known that apocynin blocks the activation of NADPH oxidase³² and is capable of increasing NO bioavailability.¹⁴ The five subunits of the classic phagocyte NADPH oxidase are expressed in the normal adult rat kidney at specific sites such as the renal artery, afferent and efferent arterioles, glomeruli, distal tubules, and macula densa.¹⁰ This suggests that the NADPH oxidase-dependent increased renal O₂^- generation stimulated by Ang II may limit NO bioavailability in the kidney. Therefore, O₂^- appears to participate as a modulator in the Ang II regulation of Na⁺ transport and renal hemodynamics.

To further evaluate the idea that Ang II increases renal NADPH oxidase-induced O₂^- generation and reduces [NO], we measured the O₂^- levels in renal cortex homogenates incubated with Ang II, valsartan, tempol, or apocynin. We found that Ang II enhanced O₂^-5 production and that this effect was blocked by coincubation with valsartan,^2,24,33 tempol,⁵ or apocynin.^14,34 In addition, the basal levels of O₂^- were diminished by valsartan, tempol, and apocynin. These in vitro experiments indicate that there is a basal O₂^- production in renal homogenates that is dependent on the stimulation of the AT₁R and NADPH oxidase. Moreover, the stimulation of AT₁R increases the NADPH oxidase-dependent superoxide generation, and this effect may contribute to the renal excretory and hemodynamic actions of Ang II.

In summary, the data in the present study indicate that there is within the kidney a basal NADPH oxidase-dependent generation of O₂^- that reduces NO bioavailability and modulates renal function. In addition, this NADPH oxidase-dependent generation of O₂^- is stimulated by Ang II and contributes to the acute renal actions of this peptide.

Perspectives
In the past few years, increasing evidence indicates that oxygen-derived free radicals modulate vascular function and arterial pressure in several models of arterial hypertension. Because the kidney and specifically the pressure-natriuresis phenomenon is thought to play a central role in the long-term control of arterial pressure, a more precise understanding of the renal mechanisms involved in the effects of free radicals influencing blood pressure regulation will be necessary to improve our knowledge of the pathophysiology of arterial hypertension.

	Acknowledgments

 
This work was supported by grants from the Ministerio de Ciencia y Tecnología (PM98-0057 and BFI2001-0497) and from the Fundación Séneca (00728/CV/99 and PI-31/00895/01).

Received August 6, 2003; first decision August 19, 2003; accepted October 8, 2003.

	References

Top Abstract Introduction Methods Results Discussion References

 

1. Griendling KK, Sorescu D, Ushio-Fukai M. NAD(P)H oxidase: role in cardiovascular biology and disease. Circ Res. 2000; 86: 494–501.[Abstract/Free Full Text]
2. Berry C, Hamilton CA, Brosnan MJ, Magill FG, Berg GA, McMurray JJ, Dominiczak AF. Investigation into the sources of superoxide in human blood vessels: angiotensin II increases superoxide production in human internal mammary arteries. Circulation. 2000; 101: 2206–2212.[Abstract/Free Full Text]
3. Baud L, Ardaillou R. Reactive oxygen species: production and role in the kidney. Am J Physiol. 1986; 251: F765–F776.[Medline] [Order article via Infotrieve]
4. Beckman JS, Koppenol WH. Nitric oxide, superoxide, and peroxynitrite: the good, the bad, and ugly. Am J Physiol. 1996; 271: C1424–C1437.[Medline] [Order article via Infotrieve]
5. Touyz RM, Chen X, Tabet F, Yao G, He G, Quinn MT, Pagano PJ, Schiffrin EL. Expression of a functionally active gp91phox-containing neutrophil-type NAD(P)H oxidase in smooth muscle cells from human resistance arteries: regulation by angiotensin II. Circ Res. 2002; 90: 1205–1213.[Abstract/Free Full Text]
6. Griendling KK, Minieri CA, Ollerenshaw JD, Alexander RW. Angiotensin II stimulates NADH and NADPH oxidase activity in cultured vascular smooth muscle cells. Circ Res. 1994; 74: 1141–1148.[Abstract/Free Full Text]
7. Pagano PJ, Chanock SJ, Siwik DA, Colucci WS, Clark JK. Angiotensin II induces p67phox mRNA expression and NADPH oxidase superoxide generation in rabbit aortic adventitial fibroblasts. Hypertension. 1998; 32: 331–337.[Abstract/Free Full Text]
8. Jaimes EA, Galceran JM, Raij L. Angiotensin II induces superoxide anion production by mesangial cells. Kidney Int. 1998; 54: 775–784.[CrossRef][Medline] [Order article via Infotrieve]
9. Zou AP, Li N, Cowley AW Jr. Production and actions of superoxide in the renal medulla. Hypertension. 2001; 37: 547–553.[Abstract/Free Full Text]
10. Chabrashvili T, Tojo A, Onozato ML, Kitiyakara C, Quinn MT, Fujita T, Welch WJ, Wilcox CS. Expression and cellular localization of classic NADPH oxidase subunits in the spontaneously hypertensive rat kidney. Hypertension. 2002; 39: 269–274.[Abstract/Free Full Text]
11. Welch WJ, Wilcox CS. AT1 receptor antagonist combats oxidative stress and restores nitric oxide signaling in the SHR. Kidney Int. 2001; 59: 1257–1263.[CrossRef][Medline] [Order article via Infotrieve]
12. Ren Y, Carretero OA, Garvin JL. Mechanism by which superoxide potentiates tubuloglomerular feedback. Hypertension. 2002; 39: 624–628.[Abstract/Free Full Text]
13. Ortiz PA, Garvin JL. Interaction of O(2)(-) and NO in the thick ascending limb. Hypertension. 2002; 39: 591–596.[Abstract/Free Full Text]
14. Hamilton CA, Brosnan MJ, Al Benna S, Berg G, Dominiczak AF. NAD(P)H oxidase inhibition improves endothelial function in rat and human blood vessels. Hypertension. 2002; 40: 755–762.[Abstract/Free Full Text]
15. Majid DS, Nishiyama A. Nitric oxide blockade enhances renal responses to superoxide dismutase inhibition in dogs. Hypertension. 2002; 39: 293–297.[Abstract/Free Full Text]
16. Lopez B, Moreno C, Salom MG, Roman RJ, Fenoy FJ. Role of guanylyl cyclase and cytochrome P-450 on renal response to nitric oxide. Am J Physiol. 2001; 281: F420–F427.
17. Friedemann MN, Robinson SW, Gerhardt GA. o-Phenylenediamine-modified carbon fiber electrodes for the detection of nitric oxide. Anal Chem. 1996; 68: 2621–2628.[Medline] [Order article via Infotrieve]
18. Escrig A, Gonzalez-Mora JL, Mas M. Nitric oxide release in penile corpora cavernosa in a rat model of erection. J Physiol. 1999; 516: 261–269.[Abstract/Free Full Text]
19. Rhinehart KL, Pallone TL. Nitric oxide generation by isolated descending vasa recta. Am J Physiol. 2001; 281: H316–H324.
20. Fenoy FJ, Ferrer P, Carbonell L, Salom MG. Role of nitric oxide on papillary blood flow and pressure natriuresis. Hypertension. 1995; 25: 408–414.[Abstract/Free Full Text]
21. Chen YF, Cowley AW, Jr., Zou AP. Increased H2O2 counteracts the vasodilator and natriuretic effects of superoxide dismutation by tempol in renal medulla. Am J Physiol. 2003; 285: R827–R833.
22. Olsen ME, Hall JE, Montani JP, Guyton AC, Langford HG, Cornell JE. Mechanisms of angiotensin II natriuresis and antinatriuresis. Am J Physiol. 1985; 249: F299–F307.[Medline] [Order article via Infotrieve]
23. Mattson DL, Raff H, Roman RJ. Influence of angiotensin II on pressure natriuresis and renal hemodynamics in volume-expanded rats. Am J Physiol. 1991; 260: R1200–R1209.[Medline] [Order article via Infotrieve]
24. Rajagopalan S, Kurz S, Munzel T, Tarpey M, Freeman BA, Griendling KK, Harrison DG. Angiotensin II-mediated hypertension in the rat increases vascular superoxide production via membrane NADH/NADPH oxidase activation. Contribution to alterations of vasomotor tone. J Clin Invest. 1996; 97: 1916–1923.[Medline] [Order article via Infotrieve]
25. Mollnau H, Wendt M, Szocs K, Lassegue B, Schulz E, Oelze M, Li H, Bodenschatz M, August M, Kleschyov AL, Tsilimingas N, Walter U, Forstermann U, Meinertz T, Griendling KK, Munzel T. Effects of angiotensin II infusion on the expression and function of NAD(P)H oxidase and components of nitric oxide/cGMP signaling. Circ Res. 2002; 90: E58–E65.[CrossRef][Medline] [Order article via Infotrieve]
26. Ortiz MC, Manriquez MC, Romero JC, Juncos LA. Antioxidants block angiotensin II-induced increases in blood pressure and endothelin. Hypertension. 2001; 38: 655–659.[Abstract/Free Full Text]
27. Zou AP, Wu F, Cowley AW. Protective effect of angiotensin II–induced increase in nitric oxide in the renal medullary circulation. Hypertension. 1998; 31: 271–276.[Abstract/Free Full Text]
28. Carey RM, Wang ZQ, Siragy HM. Role of the angiotensin type 2 receptor in the regulation of blood pressure and renal function. Hypertension. 2000; 35: 155–163.[Abstract/Free Full Text]
29. Hannken T, Schroeder R, Stahl RA, Wolf G. Angiotensin II-mediated expression of p27Kip1 and induction of cellular hypertrophy in renal tubular cells depend on the generation of oxygen radicals. Kidney Int. 1998; 54: 1923–1933.[CrossRef][Medline] [Order article via Infotrieve]
30. Fujiwara Y, Kikkawa R, Kitamura E, Takama T, Shigeta Y. Angiotensin II effects upon glomerular intracapillary volume in the rat. Ren Physiol. 1984; 7: 344–348.[Medline] [Order article via Infotrieve]
31. Shultz PJ, Schorer AE, Raij L. Effects of endothelium-derived relaxing factor and nitric oxide on rat mesangial cells. Am J Physiol. 1990; 258: F162–F167.[Medline] [Order article via Infotrieve]
32. Meyer JW, Holland JA, Ziegler LM, Chang MM, Beebe G, Schmitt ME. Identification of a functional leukocyte-type NADPH oxidase in human endothelial cells: a potential atherogenic source of reactive oxygen species. Endothelium. 1999; 7: 11–22.[Medline] [Order article via Infotrieve]
33. Cifuentes ME, Rey FE, Carretero OA, Pagano PJ. Upregulation of p67(phox) and gp91(phox) in aortas from angiotensin II-infused mice. Am J Physiol. 2000; 279: H2234–H2240.
34. Stolk J, Hiltermann TJ, Dijkman JH, Verhoeven AJ. Characteristics of the inhibition of NADPH oxidase activation in neutrophils by apocynin, a methoxy-substituted catechol. Am J Respir Cell Mol Biol. 1994; 11: 95–102.[Abstract]

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