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Hypertension. 2007;49:664-671
Published online before print January 2, 2007, doi: 10.1161/01.HYP.0000255233.56410.20
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(Hypertension. 2007;49:664.)
© 2007 American Heart Association, Inc.

Original Articles, Part 2

Mechanisms of Oxidative Stress-Induced Increase in Salt Sensitivity and Development of Hypertension in Sprague-Dawley Rats

Anees Ahmad Banday; Abdul Bari Muhammad; Fatima Rizwan Fazili; Mustafa Lokhandwala

From the Heart and Kidney Institute, College of Pharmacy, University of Houston, Tex.

Correspondence to Mustafa F. Lokhandwala, Heart and Kidney Institute, College of Pharmacy, University of Houston, 4800 Calhoun, S & R-2 Bldg, Houston, TX 77204. E-mail mlokhandwala{at}uh.edu


*    Abstract
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 *Abstract
down arrowIntroduction
 down arrowMethods
 down arrowResults
 down arrowDiscussion
 down arrowReferences
 
High salt intake produces vascular changes that contribute to the development of hypertension in salt-sensitive individuals. Because reactive oxygen species play a role in the pathogenesis of cardiovascular diseases, we investigated whether oxidative stress contributes to salt-sensitive hypertension. Sprague-Dawley rats were divided in different groups and received tap water (vehicle), 30 mmol/L of L-buthionine sulfoximine ([BSO] an oxidant), high salt ([HS] 1% NaCl), and BSO plus HS without and with antioxidant tempol (1 mmol/L) in drinking water for 12 days. Compared with vehicle, BSO treatment caused oxidative stress and mild increase in blood pressure. Thoracic aortic rings from BSO-treated rats exhibited decreased response to endothelium-independent vasorelaxants. In HS-treated rats, the response to vasoactive agents, as well as blood pressure, was unaffected. Concomitant treatment of rats with BSO and HS produced a marked increase in blood pressure and a decreased response to both endothelium-dependent and endothelium-independent vasorelaxants with an increase in EC50. Incubation of aortic tissue from BSO-treated rats with sodium nitroprusside showed decreased cGMP accumulation, whereas HS rats had decreased basal NO synthase activity. Tempol decreased oxidative stress, normalized blood pressure, and restored NO signaling and responses to vasoactive compounds in BSO and BSO plus HS rats. We conclude that BSO increases oxidative stress and reduces NO signaling, whereas HS reduces NO levels by decreasing the NO synthase activity. These phenomena collectively result in reduced responsiveness to both endothelium -dependent and endothelium- independent vasorelaxants and may contribute to salt-sensitive hypertension.


Key Words: acetylcholine • hypertension • oxidative stress • salt sensitivity • tempol


*    Introduction
up arrowTop
 up arrowAbstract
 *Introduction
down arrowMethods
 down arrowResults
 down arrowDiscussion
 down arrowReferences
 
Endothelial cells modulate the reactivity of the underlying vascular smooth muscle cells by releasing endothelium-derived relaxing factors.1,2 Previous studies have demonstrated that elevated dietary salt intake leads to an impaired relaxation of blood vessels to endothelium-dependent relaxations induced by a variety of vasodilator agents.3–9 A possible contributor to impaired vascular relaxation to dilator stimuli in animals on a high-salt diet is an impaired function of the endothelium.3–9 Impaired endothelium-dependent dilation in vessels of animals on a high-salt diet could occur either because the acetylcholine (Ach)–mediated production of NO by the endothelium is impaired or because of the failure of NO to cause vasodilation.3–9

NO is a major regulator of vascular tone in humans.10 Decreased production or bioavailability, or decreased vascular response to NO, has been implicated in the pathogenesis of human hypertension.11,12 An increase in blood pressure (BP) in response to dietary sodium (salt sensitivity) is a well-documented phenomenon in humans and is considered to be an important factor in the pathogenesis of hypertension.4 In animal models of salt-sensitive hypertension, the increase in BP after salt loading is characterized by reduced NO production.8,13,14 Similarly, in humans with salt-sensitive hypertension, salt loading was associated with decreased plasma and urinary levels of NO metabolites.3,13,14

A variety of evidence suggests that reactive oxygen species contribute to impaired endothelial function in several forms of hypertension and that there is increased oxidative stress in the microvessels of spontaneously hypertensive rats and Dahl salt-sensitive hypertensive rats.15–17 A recent report by Lenda et al7 has suggested that reactive oxygen species can also contribute to a reduced endothelium-dependent dilation in normotensive rats on a high-salt diet. Despite the potential importance of reactive oxygen species in contributing to impaired endothelium-dependent vasodilation and reduced NO production during elevated dietary salt intake, the nature and mechanisms of the impaired vascular relaxation with the high-salt diet and the role of enhanced oxidative stress in contributing to salt-induced changes in vascular function and hypertension are not completely understood.18–20

The aim of this study was to examine the hypothesis that oxidative stress modulates the blood pressure responses to salt loading in normotensive animals. We sought to determine whether oxidative stress-mediated salt sensitivity involves changes in NO production and/or NO-mediated vasodilation. The study was performed in normotensive animals so as to exclude the possibility that any observed changes were secondary to pre-existing cardiovascular disorders.


*    Methods
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 up arrowAbstract
 up arrowIntroduction
 *Methods
down arrowResults
 down arrowDiscussion
 down arrowReferences
 
The BCA protein assay kit (Pierce), cGMP assay kit (R&D Systems), NO synthase assay kit (Calbiochem), RNeasy mini kit (QIAGEN), Advantage cDNA PCR kit (BD Biosciences), Glutathione (GSH) colorimetric determination kit (OXIS International Inc), 8-Isoprostane enzyme immunoassay kit (Chemicon International), and Complete protease inhibitor mixture (Roche Diagnostic Gmbh) were used in this study. Rabbit anti- endothelial NO synthase ([NOS] eNOS)/neuronal NOS and anti-nitrotyrosine polyclonal antibodies were purchased from Transduction Laboratories and Upstate Biotechnology, respectively. 14C-Arginine monohydrochloride was purchased from Amersham. 8-Bromo-cGMP was obtained from Calbiochem and L-buthionine-sulfoximine, tempol (4-hydroxy-2,2,6,6-tetramethylpiperidine 1-oxyl), Inactin (thiobutabarbital sodium salt hydrate), phenylephrine, Ach, adenosine diphosphate, calcium ionophore A23187, sodium nitroprusside (SNP), papaverine, NG-nitro-L-arginine methyl ester, nitroglycerin (NG), and all other reagents were purchased from Sigma-Aldrich.

Animals
Male Sprague-Dawley (SD) rats (Harlan) were fed normal rat chow and dived into following weight matching 8 groups: V, animals maintained on tap water (vehicle); HS, animals receiving high salt (1% NaCl); BSO, animals provided with 30 mmol/L of L-buthionine sulfoximine; HS+BSO, animals receiving both high salt and L-buthionine sulfoximine; T, animals provided with 1 mmol/L tempol; HS+T, animals kept on high salt and receiving tempol; BSO+T, animals receiving L-buthionine sulfoximine with tempol; and HS+BSO+T, animals receiving high salt plus L-buthionine sulfoximine and tempol. High salt, L-buthionine sulfoximine (a {gamma}-glutamylcysteine synthetase inhibitor), and tempol (an antioxidant) were provided in drinking water for 12 days. All of the experiments were performed according to the National Institutes of Health and University of Houston guidelines and protocols for care and use of laboratory animals.

Surgical Procedures for Blood Pressure and Aortic Preparations
Rats were anesthetized with Inactin (100 mg/kg IP). Tracheotomy was performed to facilitate breathing. To measure the blood pressure and heart rate, the left carotid artery was catheterized with PE-50 tubing. This tubing was connected to a Statham P23AC pressure transducer. Blood pressure and heart rate were continuously recorded on a Grass polygraph (model 7D, Grass Instrument).21 For aortic preparations, a midline abdominal incision was made, and thoracic aortas were removed and immediately placed in Krebs-Henseleit buffer (mM: NaCl 118.4, KCl 4.7, CaCl2 2.5, KH2PO4 1.2, MgSO4 1.2, NaHCO3 25.0, and glucose 10.0 [pH 7.4]). The aortas were cleaned of adherent tissue and cut into rings with minimum damage to endothelium. Each ring was fixed under a resting tension of 1 g in a 10-mL organ bath filled with Krebs-Henseleit buffer (37°C) and continuously aerated with a 95% O2/5% CO2 gas mixture, and the rings were allowed to equilibrate for 90 minutes before the start of the experiments. In some rings, the endothelium was mechanically removed by gentle rubbing with moistened cotton. Isometric tension change was measured with a digital force isometric transducer (Harvard Apparatus) connected to a data acquisition system (AD Instruments).

Vascular Relaxation Studies
Aortic rings with intact endothelium were preconstricted with 10 µmol/L of phenylephrine. Ach (0.1 nM to 10 µmol/L), ADP (1 nM to 1 mol/L), or calcium ionophore A23187 (0.1 nM to 0.1 mol/L) in the presence or absence of NG-nitro-L-arginine methyl ester (100 µmol/L) was added to the bath solution after the phenylephrine-induced contractions reached a plateau. To determine the endothelium-independent vasorelaxation, endothelium-denuded aortic rings were preconstricted with phenylephrine, and SNP (0.1 nM to 1 µmol/L), NG (0.1 nM to 1 µmol/L), or 8-bromo-cGMP (1 to 100 µmol/L) was added to the bath medium. The relaxation responses obtained were expressed as a percentage of the maximal relaxation evoked by papaverine (100 µmol/L). Endothelium integrity was confirmed by achieving ≥40% relaxation with 10 µmol/L Ach, whereas denudation of the endothelium was confirmed by the disappearance of the 1 µmol/L ACh-induced relaxation response.

Assay of NOS Activity
Endothelium intact rings were snap frozen and maintained at –80°C. Thawed samples were lysed, centrifuged for 5 minutes, and the supernatant (5 to 10 µg/µL of protein) was used for NOS assay. NOS activity was quantified by assay of conversion of [14C]-arginine to citrulline, performed as described in the NOS Assay kit. Total NOS activity was measured in standard Ca2+-containing buffer. The activities of eNOS and neuronal NOS, but not inducible NO, are calcium dependent.

Western Blotting
Portions of isolated aortic samples were homogenized in lysis buffer containing 0.25 mol/L of sucrose, 50 mmol/L of dithiothreitol, 3 mmol/L of HEPES (pH 7.9), 0.5 mmol/L of EGTA, 0.4 mmol/L of PMSF, protease inhibitor mixture, and 1% Triton X-100. Samples were centrifuged, and supernatants were mixed with an equal volume of Laemmli buffer and subjected to SDS-PAGE. The proteins were transferred to polyvinylidene difluoride membranes, blocked, and incubated with corresponding antibodies. The blots were incubated with horseradish peroxidase-conjugated secondary antibodies and visualized with the enhanced chemiluminescence method. The density (arbitrary units) of the bands was quantified by Kodak Imaging software.

Determination of cGMP Level
Endothelium intact rings were preconstricted with phenylephrine for 5 minutes and then stimulated with SNP (0.01 mol/L) for 1 or 3 minutes, which allowed the generation of {approx}75% to 80% of the maximal relaxation. The tissues were snap frozen in liquid nitrogen, followed by homogenization in a glass/glass homogenizer in ice-cold 0.1 N HCl.22 The cGMP content was determined with an enzyme immunoassay kit according to the manufacturer’s protocol. Protein was determined by BCA protein assay kit using bicinchoninic acid for the colorimetric detection and quantitation of total protein with BSA as standard.

Indexes (Biomarkers) of Oxidative Stress, GSH, and Nicotinamide-Adenine Dinucleotide Phosphate Oxidase Activity
Aortic tissue carboxymethyllysine was measured by ELIZA as described by Koo and Vaziri,23 and malondialdehyde was determined by the method of Mihara and Uchiyama.24 For nicotinamide-adenine dinucleotide phosphate oxidase activity, fluorescence spectrometric assay of O2– production was performed by Satoh et at.20 A biooxytech GSH kit from Oxis Health was used to measure glutathione (GSH). Plasma 8-isoprostane prostaglandin F2{alpha} was measured by 8-isoprostane prostaglandin F2{alpha} immunoassay kit from Cayman.

Statistical Analysis
Differences between means were evaluated using the unpaired t test or ANOVA with Newman-Keuls multiple test, as appropriate. P<0.05 was considered statistically significant.


*    Results
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 up arrowIntroduction
 up arrowMethods
 *Results
down arrowDiscussion
 down arrowReferences
 
As shown in Table 1, the BSO, HS, and T regimen had no effect on body weight or food intake of animals. The mean blood pressure of BSO-supplemented rats showed a mild increase, whereas animals provided with both BSO and HS showed marked increase in blood pressure compared with vehicle. Treatment with tempol normalized blood pressure in both BSO and BSO plus HS-treated rats. In the vehicle and HS groups, tempol did not have any effect on blood pressure (Table 1).


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  		TABLE 1. Effect of HS, BSO, and T on Body Weight, BP, and Oxidative Stress

Oxidative Stress
Animals treated with BSO or BSO plus HS showed a marked decrease in thoracic aorta tissue GSH levels and a significant increase in oxidative markers, such as malondialdehyde, carboxymethyl lysine, plasma 8-isoprostane, and nitrotyrosine levels (Table 1 and Figure 1A). Incubation of tissue homogenates with dihydroethidium and nicotinamide-adenine dinucleotide phosphate oxidase substrates caused increased O2– production in BSO and BSO plus HS rats (Figure 1B). HS and tempol showed a mild but nonsignificant increase and decrease, respectively, in superoxide formation compared with vehicle (Figure 1B). Treatment of these animals with tempol normalized tissue malondialdehyde, carboxymethyl lysine, plasma 8-isoprostane, and nitrotyrosine levels, as well as nicotinamide-adenine dinucleotide phosphate oxidase activity (Figure 1A and 1B and Table 1). Although tempol supplementation increased tissue GSH levels in BSO or BSO plus HS-treated rats, it remained significantly lower than vehicle or HS-treated animals (Table 1). HS or tempol did not cause any significant effect on oxidative stress in the absence of BSO (Table 1).


Figure 1
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  		Figure 1. Protein nitrotyrosination (A) and nicotinamide-adenine dinucleotide phosphate oxidase activity (B) in thoracic aortic tissues from vehicle, BSO-, HS-, and T-supplemented rats (n=6 to 8). A, top, representative nitrotyrosine band and bars represent density arbitrary units (mean±SE). B, Bars represent fluorescence normalized with protein (mean±SE). Data were analyzed by ANOVA followed by posthoc Newman-Keuls multiple comparison test. P<0.05 was considered statistically significant. *Significantly different from vehicle.

Endothelium-Dependent Relaxation
There was no difference in the maximum contraction of aortic rings to KCl (data not shown) or phenylephrine between the different groups (Table 2). Endothelium-dependent relaxations in response to Ach in endothelium-intact aortic rings from animals treated with HS or BSO alone showed a mild rightward shift. In animals treated with BSO plus HS, the response was significantly reduced, and concentration-dependent responsiveness to Ach showed a marked rightward shift with significant increase in EC50 (Figure 2). Similar to Ach, ADP and calcium ionophore A23187–induced relaxation curves showed significant increases in EC50 in BSO plus HS animals (Table 2). Treatment with tempol restored the ACh, ADP, and calcium ionophore A23187–dependent vasorelaxation and normalized the EC50 (Figure 2B and Table 2). These relaxations were completely inhibited by NG-nitro-L-arginine methyl ester, an NOS inhibitor (data not shown).


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  		TABLE 2. Effect of HS, BSO, and T on Phenylephrine, ADP, A23187, NG, and 8b-cGMP Response in Aortic Rings


Figure 2
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  		Figure 2. Endothelium-dependent relaxation in response to Ach in thoracic aortic rings from vehicle, BSO-, HS-, and T-supplemented rats (n=6 to 8). Results (EC50) are expressed as mean±SE and analyzed by ANOVA followed by posthoc Newman-Keuls multiple comparison test. P<0.05 was considered statistically significant. *Significantly different from vehicle.

Endothelium-Independent Relaxations
There was no significant difference between the HS group and the vehicle group in endothelium-independent relaxations in response to SNP and NG, which are NO donors, in endothelium-denuded aortic rings (Figure 3A and Table 2). However, the relaxation response was reduced in animals treated with BSO or BSO plus HS. The dose-response curve showed a significant rightward shift with marked increase in EC50 in these animals (Figure 3A and Table 2). On the other hand, the relaxation in response to 8-bromo-cGMP, a stable cyclic GMP molecule, was similar in all of the experimental groups (Table 2). Treatment with tempol restored the vasorelaxant response in BSO or BSO plus HS-treated rats (Figure 3B). Table 2 also shows the values of vasodilator potency of NG in various groups.


Figure 3
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  		Figure 3. Endothelium-independent relaxation in response to SNP in thoracic aortic rings from vehicle, BSO-, HS-, and T-supplemented rats (n=6 to 8). Results (EC50) are expressed as mean±SE and analyzed by ANOVA followed by posthoc Newman-Keuls multiple comparison test. P<0.05 was considered statistically significant. *Significantly different from vehicle.

NOS Activity and Protein Expression
The basal NOS activity was decreased in HS-treated animals compared with vehicle (Figure 4A). The concomitant treatment with BSO and HS caused further decrease in NOS activity, whereas BSO by itself showed no effect on NOS activity (Figure 4A). Tempol normalized the NOS activity in BSO plus HS-treated rats but failed to restore the NOS activity in animals provided with HS alone (Figure 4A). There was no difference in NOS activity between vehicle and tempol-treated rats (Figure 4A).


Figure 4
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  		Figure 4. NOS activity (A) and protein expression (B) in thoracic aortic tissues from vehicle, BSO-, HS-, and T-supplemented rats (n=6 to 8). A, Bars represent activity, and (B) top is a representative Western blot. Bars represent band densities (arbitrary units). Results are expressed as mean±SE and analyzed by ANOVA followed by posthoc Newman-Keuls multiple comparison test. P<0.05 was considered statistically significant. *Significantly increased compared with vehicle, #Significantly decreased compared with vehicle.

Western blot analysis showed higher eNOS protein in BSO or BSO plus HS animals (Figure 4B). Treatment with tempol normalized protein abundance in BSO and BSO plus HS rats (Figure 4B). On the other hand, HS caused a small but significant decrease in eNOS protein expression, which was not restored by tempol (Figure 4B).

SNP Induced cGMP Accumulation
In aortic rings from vehicle and HS-treated rats, SNP stimulation increased the cGMP levels. However, SNP-induced cGMP accumulation was reduced in aortas from BSO and BSO plus HS rats (Figure 5). The decrease in SNP-induced cGMP accumulation was much higher in BSO plus HS rats than in BSO animals (Figure 5). Tempol restored the SNP-induced cGMP accumulation in both BSO and BSO plus HS animals. Tempol showed no effect on SNP signaling in vehicle or HS rats (Figure 5).


Figure 5
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  		Figure 5. SNP-induced cGMP accumulation in thoracic aortic tissues from vehicle, BSO-, HS-, and T-supplemented rats (n=6 to 8). Results are expressed as mean±SE and analyzed by ANOVA followed by posthoc Newman-Keuls multiple comparison test. P<0.05 was considered statistically significant. *Significantly different from vehicle.


*    Discussion
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 up arrowAbstract
 up arrowIntroduction
 up arrowMethods
 up arrowResults
 *Discussion
down arrowReferences
 
An increase in blood pressure in response to increased dietary sodium intake is considered an important contributing factor in the pathogenesis of hypertension, particularly in humans or animals predisposed to salt sensitivity.25–28 In this study, we observed that in normotensive rats, high salt intake has no effect on response to vasodilatory factors or blood pressure despite reduced NOS activity and eNOS expression. On the other hand, treatment of animals with BSO, a {gamma}-glutamylcysteine synthetase inhibitor, reduced GSH, increased oxidative stress, nicotinamide-adenine dinucleotide phosphate oxidase activity and eNOS protein expression, reduced SNP-mediated guanylyl cyclase (GC) activation and vascular response, and caused a mild increase in blood pressure. Concomitant administration of HS and BSO led to increased oxidative stress, nicotinamide-adenine dinucleotide phosphate oxidase activity, decreased in NOS activity, and increased eNOS expression. These animals exhibited a decreased response to both endothelial-dependent and endothelial-independent vasodilators and marked elevation in blood pressure. Treatment with tempol mitigated oxidative stress and corrected eNOS expression, NOS activity, and NO signaling in BSO and BSO plus HS-treated rats. Furthermore, tempol normalized the response to vasodilatory agents and BP in these animals.

High Salt Intake, Vascular Function, and BP
Several studies have reported a moderate increase, whereas others have found no significant change in arterial BP with high salt intake in normotensive SD rats.28–32 In the present study, we found that high salt intake had no effect on either endothelium-dependent or endothelial-independent relaxations in normotensive SD rats, and there was no change in BP in the presence of decreased aortic NOS activity and expression. These relaxations were mediated mainly by NO, because they almost disappeared in the presence of NG-nitro-L-arginine methyl ester, an inhibitor of NOS. We also found that high salt intake does not affect the NO-GC coupling as evidenced by an increase in cGMP accumulation in response to SNP. Taken together, the observations that the decrease in NOS activity or expression during salt loading is not related to BP response and that vascular relaxation response is preserved imply that, in healthy subjects, other compensatory mechanisms are of primary importance in adjusting BP to salt loading.

Oxidative Stress, Vascular Function, and BP
We used SD rats to study the role of oxidative stress on vascular function and BP. Treatment of rats with BSO produced oxidative stress and decreased relaxant response to endothelial-independent agents. These animals exhibited a mild increase in BP despite normal NOS activity and intact response to endothelial-dependent vasoactive agents indicating that factors proximal to NO production also play an important role in hypertension. It is well known that NO activates the soluble isoform of GC to form cGMP.33,34 The increased cGMP level causes vascular smooth muscle relaxation and regulates vascular tone in various vascular beds.33,34 In hypertensive animal models, altered vascular activity of the NO-cGMP pathway, such as reduced GC activity and decreased cGMP accumulation, has been reported.35,36 We also observed that BSO reduced SNP-mediated cGMP accumulation in aortic tissue, indicating reduced NO-GC signaling. Therefore, in the presence of oxidative stress, a decreased NO-mediated GC stimulation followed by a reduced cGMP level may lead to impairment of smooth muscle relaxation and a subsequent increase in BP.

We observed a significant increase in eNOS expression in BSO-treated rats, although their NOS activity was similar to control animals. There are reports indicating that superoxide radicals can interact with NO and decrease its bioavailability.18,19 Although we did not measure the NO levels in the present study, it is possible that the increase in eNOS protein synthesis could be a compensatory mechanism to maintain the normal NO levels. This is supported by the finding that treatment of BSO rats with tempol decreased oxidative stress, restored the NO-mediated relaxation and signaling, corrected eNOS expression, and normalized BP. These studies are in agreement with Vaziri et al37 showing that BSO caused oxidative stress and increased BP, and these changes were reversed by antioxidant supplementation.

High Salt Intake Plus Oxidative Stress, Vascular Function, and BP
Based on our findings that animals treated with high salt had decreased NOS activity and were normotensive, whereas animals provided with BSO alone exhibited a mild increase in BP despite normal endothelium-dependent vasodilation, we hypothesized that oxidative stress may be a triggering factor for salt sensitivity. In support of our view, we found impaired vasorelaxation in response to both endothelium-dependent and endothelium-independent agents accompanied by a marked increase in BP in animals provided with HS and BSO. Treatment with tempol restored the vascular function and normalized BP. There are reports indicating that oxidative stress plays an important role in salt-sensitive hypertension.16,17,27,38,39 Swei et al17,39 found that hypertensive Dahl S rats have significantly higher microvascular superoxide and plasma H2O2 than Dahl R rats.

Although the exact mechanisms for decreased NO production in salt sensitivity are not known, it is suggested that reduced NO activity could be a contributing factor.40,41 Our data also revealed a significant decrease in NOS activity in aortic homogenates, confirming an association between reduced NOS activity and impaired endothelium-mediated relaxation in these animals. Similar to BSO rats, we also observed increased protein expression of eNOS in BSO plus HS rats, thus ruling out that a decrease in eNOS abundance could be a factor for reduced NOS activity in these hypertensive rats. Based on the report of Satoh et al,20 it is possible that increased oxidative stress caused NOS uncoupling by decreasing the cofactors that led to NOS inactivation. This is supported by our data showing that treatment of BSO plus HS rats with tempol normalized the NOS activity, as well as eNOS protein expression, thus reinforcing the role of oxidative stress in endothelium dysfunction.

In summary, the present study reveals that increased salt intake has no effect on vascular relaxation, BP, and NO signaling despite impaired NOS activity and expression in normotensive SD rats. On the other hand, oxidative stress reduces the vasorelaxation in response to NO donors, decreases the GC/cGMP signaling, and contributes to mild hypertension. However, the high salt intake in animals with oxidative stress leads to reduced vasorelaxation in response to both endothelium-dependent and -independent vasoactive compounds. These impairments are mediated by decreased NOS activity and NO-GC uncoupling, which leads to reduced levels of cGMP in smooth muscle. Finally, tempol, while reducing oxidative stress, restores the endothelium function and NO signaling and normalizes BP, providing unequivocal support for the role of oxidative stress in vascular diseases and salt sensitivity (Figure 6).


Figure 6
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  		Figure 6. A schematic presentation of oxidative stress, impaired endothelium dysfunction, decreased NO signaling, and its possible contribution to salt sensitivity. OS indicates oxidative stress. Effect (???), stimulation (arrow without arrowhead), decrease in protein activity or levels ({Downarrow}), increase ({Uparrow}), reduces or blocks stimulation ({perp}).

Perspectives
Our findings have important clinical relevance as they relate to the role of high salt intake in the pathogenesis of hypertension. Based on the results of the present study, it can be suggested that, in the absence of preexisting conditions, which may result in oxidative stress, an organism is able to adjust to high salt intake by triggering compensatory mechanisms, including renal compensation, and thereby maintaining BP. However, in situations associated with oxidative stress, where some of these compensatory mechanisms are impaired, the organism acquires salt sensitivity such that excess salt intake leads to the development of hypertension. Therefore, either lowering oxidative stress or preventing an increase in oxidative stress may be beneficial in affording protection against salt-induced hypertension.


*    Acknowledgments
 
Disclosures

None.

Received October 11, 2006; first decision November 5, 2006; accepted December 4, 2006.


*    References
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 up arrowAbstract
 up arrowIntroduction
 up arrowMethods
 up arrowResults
 up arrowDiscussion
 *References
 

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