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Am J Physiol Heart Circ Physiol 294: H2473-H2479, 2008. First published April 4, 2008; doi:10.1152/ajpheart.91533.2007
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Angiotensin II type 1 receptor blocker preserves tolerance to ischemia-reperfusion injury in Dahl salt-sensitive rat heart

Seiji Matsuhisa,1 Hajime Otani,1 Toru Okazaki,1 Koji Yamashita,1 Yuzo Akita,1 Daisuke Sato,1 Akira Moriguchi,1 Hiroji Imamura,2 and Toshiji Iwasaka1

1Second Department of Internal Medicine and 2Thoracic and Cardiovascular Surgery, Kansai Medical University, Moriguchi City, Japan

Submitted 31 December 2007 ; accepted in final form 1 April 2008


   ABSTRACT
TOP
 ABSTRACT
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Oxidative stress is involved in the tolerance to ischemia-reperfusion (I/R) injury. Because angiotensin II type 1 receptor blockers (ARBs) inhibit oxidative stress, there is concern that ARBs abolish the tolerance to I/R injury. Dahl salt-sensitive (DS) hypertensive and salt-resistant (DR) normotensive rats received an antioxidant, 2-mercaptopropionylglycine (MPG), or an ARB, losartan, for 7 days. Losartan and MPG significantly inhibited oxidative stress as determined by tissue malondialdehyde + 4-hydroxynoneal and increased expression of inducible nitric oxide synthase (iNOS) in the DS rat heart. However, losartan but not MPG activated endothelial nitric oxide synthase (eNOS) as assessed by phosphorylation of eNOS on Ser1177. Infarct size after 30-min left coronary artery occlusion followed by 2-h reperfusion was comparable between DS and DR rat hearts. Although MPG and losartan had no effect on infarct size in the DR rat heart, MPG but not losartan significantly increased infarct size in the DS rat heart. A selective iNOS inhibitor, 1400W, increased infarct size in the DS rat heart, but it had no effect on infarct size in the losartan-treated DS rat heart. However, a nonselective NOS inhibitor, N{omega}-nitro-L-arginine methyl ester, increased infarct size in the losartan-treated DS rat heart. These results suggest that losartan preserves the tolerance to I/R injury by activating eNOS despite elimination of redox-sensitive upregulation of iNOS and iNOS-dependent cardioprotection in the DS rat heart.

antioxidant; nitric oxide synthase

OXIDATIVE STRESS has been implicated in the pathogenesis of cardiac hypertrophy, congestive heart failure, and ischemia-reperfusion (I/R) injury. Oxidative stress was initially thought to be a detrimental event precipitating in cellular dysfunction by attacking a variety of biomolecules (18). Recently, a new paradigm of redox signaling has emerged whereby reactive oxygen species (ROS) and oxidants can function as intracellular signaling molecules that convert a death signal into a survival signal (7). This concept is exemplified by ischemic preconditioning (IPC), in which mild and transient oxidative stress confers tolerance to subsequent lethal cellular stress (23). The redox-sensitive nature of IPC has been substantiated by the use of antioxidants, such as N-acetylcysteine and 2-mercaptopropionylglycine (MPG), which blocked IPC-induced cardioprotection by eliminating oxidative stress (2, 6). In addition, it has been revealed that ROS and nitric oxide generated during the preconditioning stimuli trigger late cardioprotection by upregulating inducible nitric oxide synthase (iNOS) (4, 5). Upregulation of iNOS is also observed in the hypertensive heart (13) and the cardiomyopathic heart, in which the redox-sensitive upregulation of iNOS plays an obligatory role in the tolerance to I/R injury (16). Therefore, it is possible that the use of antioxidants compromises tolerance to I/R injury in diseased hearts by abrogating the redox-sensitive upregulation of iNOS.

The renin-angiotensin system is locally activated in the heart under various disease conditions (10, 27). Angiotensin II acting through high-affinity cell surface angiotensin II type 1 (AT1) receptors modulates cardiovascular homeostasis by exerting vasoconstriction and promoting myocardial hypertrophy and fibrosis, thereby contributing to pathological remodeling during the development of heart failure (19). Accumulating evidence suggests that such detrimental effects of AT1 receptor activation are mediated at least in part by oxidative stress through the activation of the NADPH oxidase system (7). It has been demonstrated that AT1 receptor blockers (ARBs) inhibit oxidative stress in the hypertensive heart (3, 9). This raises a concern that the use of ARBs may eliminate the tolerance to I/R injury in the hypertensive heart that is chronically exposed to oxidative stress by activation of AT1 receptors. On the contrary, accumulating evidence has indicated that preischemic treatment with ARBs confers cardioprotection against I/R injury in normotensive hearts, as demonstrated by improved postischemic cardiac function and reduced infarct size (11, 14, 24). Nevertheless, the effect of ARBs on I/R injury in the hypertensive heart has not been investigated. Therefore, the present study was undertaken to address the question as to whether inhibition of oxidative stress by a genuine antioxidant and an ARB modulates the tolerance to I/R injury in the hypertensive heart.

The Dahl salt-sensitive (DS) hypertensive rat is a useful model for investigating the effect of local activation of the renin-angiotensin system in the heart. The circulatory renin-angiotensin system is suppressed during the development of hypertension in these rats on high salt intake (26, 33), and this model of "low-renin hypertension" is in general considered to be unresponsive to treatment with blockers of the renin-angiotensin system (12). Accordingly, treatment with ARBs prevents the effect of locally elevated angiotensin II without affecting blood pressure. Using this rat model, we were able to show that a genuine antioxidant and an ARB inhibit oxidative stress and upregulation of iNOS but only an ARB preserves the tolerance to I/R injury in the DS rat heart.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Animals. All experiments were conducted in accordance with the Guide for the Care and Use of Laboratory Animals (NIH Pub. No. 85-23, revised 1996) and approved by the Kansai Medical University Institutional Animal Care and Use Committee.

Male inbred DS rats and Dahl salt-resistant (DR) rats were purchased from Japan SLC (Shizuoka, Japan). After they were weaned, the rats were fed a diet containing 0.3% NaCl until the age of 6 wk. Thereafter, the rats were fed a diet containing 8% NaCl. The rats were weighed, and their systolic blood pressure (SBP) was measured by the tail-cuff system (BP-98A, Softron, Tokyo, Japan) just before echocardiography and surgical preparation. Transthoracic echocardiography was performed with a SONOS-7500 (Philips Medical Systems, Andover, MA) equipped with a 6- to 15-MHz transducer (model 21390A, Philips) as described previously (16). Briefly, from the cardiac short axis (papillary muscle level), an M-mode trace of the left ventricle (LV) was obtained and septal diastolic wall thickness as well as LV end-diastolic dimension (LVEDD) and LV end-systolic dimension (LDESD) were measured at the widest and the narrowest dimensions, respectively. LV fractional shortening (FS) was calculated as %FS = (LVEDD – LVESD)/LVEDD x 100.

Surgical preparation. Pentobarbital-anesthetized (60 mg/kg ip) rats were intubated for positive-pressure ventilation with oxygen-enriched room air. A catheter was positioned in the abdominal cavity to allow intraperitoneal administration of pentobarbital for maintenance of anesthesia. Rectal temperature was continuously measured and maintained at 36.5–37.5°C. A Millar-tip catheter (2 F; Millar Instrument, Houston, TX) was inserted into the LV via the left carotid artery to monitor LV pressure. A left thoracotomy was performed at the fifth intercostal space, and the pericardium was opened to expose the heart. The left coronary artery was ligated 1–2 mm from its origin with a 7-0 silk suture and an atraumatic needle, and ends of this ligature were passed through a small vinyl tube to form a snare. After the completion of the surgical procedure, the heart was returned to its normal position in the thorax. The thoracic cavity was covered with saline-soaked gauze to prevent the heart from drying. The animals were then allowed to stabilize for 15 min before left anterior descending coronary artery (LAD) ligation. Myocardial ischemia was induced by one-stage occlusion of the LAD by pressing the polyethylene tubing against the ventricular wall and then fixing it in place by clamping the vinyl tube with a hemostat. The animals then underwent 30 min of ischemia, confirmed visually in situ by the appearance of regional epicardial cyanosis and ST segment elevation. The myocardium was reperfused by releasing the snare gently for a period of 2 h. Successful reperfusion was confirmed by visualization of arterial blood flow through the artery and appearance of hyperemia over the surface of the previously ischemia-cyanotic segment.

Experimental protocol. DR and DS rats fed the high-salt diet for 5 wk were randomly divided into 14 groups (Fig. 1). The control groups of rats were fed the high-salt diet for an additional 7 days without any drug treatment and subjected to coronary artery occlusion and reperfusion. The second groups of rats were treated with the antioxidant MPG (100 mg·kg–1·day–1 ip) for 7 days before coronary artery occlusion and reperfusion. The third groups of rats were administered the ARB losartan (30 mg·kg–1·day–1) in the drinking water for 7 days before coronary artery occlusion and reperfusion. The fourth groups of rats were treated with the iNOS-selective inhibitor 1400W (10 mg/kg ip) or the nonselective NOS inhibitor N{omega}-nitro-L-arginine methyl ester (L-NAME; 100 mg/kg ip) 30 min before coronary artery occlusion and reperfusion. The fifth groups of rats were administered losartan for 7 days and treated with 1400W or L-NAME 30 min before coronary artery occlusion and reperfusion.


Figure 1
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  		Fig. 1. Experimental protocol. Dahl salt-resistant (DR) and Dahl salt-sensitive (DS) rats fed the high-salt diet (HSD) for 5 wk were randomly divided into 14 groups. The control groups of rats were fed with HSD for an additional 7 days without any drug treatment and subjected to coronary artery occlusion (CAO) and reperfusion. The second groups of rats were treated with the antioxidant 2-mercaptopropionylglycine (MPG; 100 mg·kg–1·day–1 ip) for 7 days before CAO and reperfusion. The third groups of rats were administered the angiotensin II type 1 receptor blocker (ARB) losartan (Los; 30 mg·kg–1·day–1) in the drinking water for 7 days before CAO and reperfusion. The fourth groups of rats were treated with the inducible nitric oxide synthase (iNOS) selective inhibitor 1400W (10 mg/kg ip) or the nonselective NOS inhibitor N{omega}-nitro-L-arginine methyl ester (L-NAME; 100 mg/kg ip) 30 min before CAO and reperfusion. The fifth groups of rats were administered Los for 7 days and treated with 1400W or L-NAME 30 min before CAO and reperfusion.

 
Measurements of infarct size. The rats were euthanized with pentobarbital sodium (100 mg/kg ip) after 2 h of reperfusion, and the LAD was religated in the same location as above. Evans blue dye (1.2 ml of a 2% solution) was injected into the carotid artery catheter inserted into the heart to delineate the ischemic zone from the nonischemic zone. The heart was rapidly excised and serially sectioned along the long axis in 1-mm-thick sections, which were then incubated in 1.0% 2,3,5-triphenyltetrazolium chloride for 5 min at 37°C to demarcate the viable and nonviable myocardium within the risk zone. Each of the five 1-mm-thick myocardial slices was weighed and digitally photographed for determination of infarct size as a percentage of risk region, and the areas of infarction, risk, and nonischemic LV were assessed by a blinded observer using computer-assisted planimetry (Win Roof, Mitani, Fukui, Japan). All of the procedures for area at risk and infarct size determination have been described previously (15).

Determination of malondialdehyde + 4-hydroxynoneal in tissue preparation. The rats were euthanized as described above 6 wk after the high-salt diet, and the heart was rapidly excised and stored in liquid nitrogen until the assay. On the day of analysis, tissue samples were washed in ice-cold 0.9% NaCl, blotted on absorbent paper, and weighed. Each sample was then ground in a small volume of ice-cold 20 mM Tris·HCl buffer (pH 7.4), in a ratio of 1:10 (wt/vol), and homogenized with a Teflon pestle. After centrifugation at 3,000 g for 10 min at 4°C, the supernatant was used for the determination of malondialdehyde (MDA) plus 4-hydroxynoneal (HNE) with an assay kit (BIOXYTECH LPO-586, OXIS Research, Foster City, CA). The protein concentration was determined with a Bio-Rad protein assay kit (Bio-Rad Laboratories, Hercules, CA).

Western blot analysis. Heart samples stored in liquid nitrogen were ground with lysis buffer containing 30 mM Tris pH 7.4, 150 mM NaCl, 1% NP-40, 0.25% sodium deoxycholate, 1 mM EDTA, 0.1 mM phenylmethylsulfonyl fluoride, and a protease inhibitor cocktail (Complete, Roche Diagnostics, Mannheim, Germany). The protein concentration was determined as described above. The lysate samples were separated by 7.5% SDS-PAGE, and the separated proteins were transferred to a polyvinylidene difluoride membrane with a transfer buffer containing 25 mM Tris, 192 mM glycine, and 10% methanol. The membranes were blocked with 5% skim milk, incubated in primary antibodies specific for iNOS (Santa Cruz Biotechnology, Santa Cruz, CA), endothelial NOS (eNOS) (Cell Signaling Technology, Beverly, MA), and phospho-eNOS (Ser1177) (Cell Signaling Technology), and subsequently incubated with peroxidase-conjugated secondary antibodies and developed with an enhanced chemiluminescence detection system (Amersham Biosciences, Tokyo, Japan) according to the manufacturer's instructions. The immunolabeling was quantified with a densitometric analysis with the image analyzing software Win Roof (Mitani). Consistency in the data analysis was ensured by normalization of each immunoblot signal to the corresponding Coomassie blue stain signal as described previously (22).

Statistical analysis. All numerical data are expressed as means ± SE. Statistical analysis of data within and between groups was performed with one-way ANOVA followed by the Bonferroni post hoc test or two-way repeated-measures ANOVA when comparisons were made at different time points.


   RESULTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
Baseline characteristics of DS and DR rat heart. Baseline hemodynamic characteristics of DS and DR rats fed the high-salt diet for 6 wk in the presence or absence of MPG or losartan for the last 7 days are shown in Table 1. SBP was significantly higher in the DS rats compared with the DR rats. Treatment with MPG and losartan had no significant effect on SBP and heart rate in the DS and the DR rats. Echocardiography showed that septal wall thickness was significantly greater in the DS rats compared with the DR rats. Although there was no significant difference in end-diastolic diameter (EDD) between the DR and the DS rats, end-systolic diameter (ESD) was significantly smaller in the DS rats compared with the DR rats. Consequently, FS was significantly greater in the DS rats. Treatment with MPG and losartan did not affect septal wall thickness, EDD, or ESD in DS and DR rats.


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  		Table 1. Baseline hemodynamic characteristics

 
Effect of MPG and losartan on generation of MDA and HNE. The amount of MDA + HNE was significantly increased in the DS rat heart compared with the DR rat heart (Fig. 2). MPG and losartan inhibited the increase in MDA + HNE in the DS rat heart.


Figure 2
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  		Fig. 2. Malondialdehyde (MDA) + 4-hydroxynoneal (HNE) content in the heart. DR and DS rats were treated with or without MPG or Los as described in Fig. 1. Values are means ± SE of 5 experiments. *P < 0.05 compared with DR control; #P < 0.05 compared with DS control.

 
Effect of MPG and losartan on expression of iNOS. The expression of iNOS was significantly increased in the DS rat heart compared with the DR rat heart (Fig. 3). MPG and losartan inhibited the increase in iNOS expression in DS rat hearts.


Figure 3
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  		Fig. 3. iNOS expression in the heart. DR and DS rats were treated with or without MPG or Los as described in Fig. 1. Values are means ± SE of 5 experiments. *P < 0.05 compared with DR control; #P < 0.05 compared with DS control.

 
Effect of MPG and losartan on expression of total eNOS and phospho-eNOS. Phosphorylation of eNOS on Ser1177 was thought to be an indicator of activation of eNOS (8). MPG and losartan had no significant effect on the expression of total eNOS and phospho-eNOS and the ratio of phospho-eNOS to total eNOS in the DR rat heart (Fig. 4). There was no difference in total eNOS expression between the DR and DS rat hearts. However, losartan but not MPG increased the expression of phospho-eNOS and the ratio of phospho-eNOS to total eNOS in the DS rat heart.


Figure 4
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  		Fig. 4. Ratio of endothelial NOS (eNOS) to phospho-eNOS (p-eNOS). DR and DS rats were treated with or without MPG or Los as described in Fig. 1. Western blot analysis for eNOS and p-eNOS was performed as described in MATERIALS AND METHODS. Values are means ± SE of 5 experiments. *P < 0.05 compared with DR control; #P < 0.05 compared with DS control.

 
Effect of MPG and losartan on LV function. LV systolic pressure (LVSP) was significantly higher in the DS rats before coronary artery occlusion compared with the DR rats (Table 2). Neither MPG nor losartan affected LVSP and LV end-diastolic pressure (LVEDP) in the DS and the DR rats. LVSP remained significantly higher in the DS rats during coronary artery occlusion and reperfusion. However, LVSP significantly decreased and LVEDP significantly increased during reperfusion in the DS rats treated with MPG, although the same treatment had no effect on LVSP and LVEDP during reperfusion in the DR rats. In contrast, losartan did not affect LVSP and LVEDP during reperfusion in the DS rats.


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  		Table 2. Effect of MPG and losartan on left ventricular function

 
Effect of MPG and losartan on infarct size. Infarct size 2 h after reperfusion was comparable between the DS and the DR rat hearts (Fig. 5). MPG had no effect on infarct size in the DR rat hearts but significantly increased infarct size in the DS rat hearts. In contrast, losartan did not affect infarct size in the DS rat hearts.


Figure 5
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  		Fig. 5. DR and DS rats were treated with or without MPG or Los as described in Fig. 1. Infarct size after 30 min of left coronary artery occlusion and 2 h of reperfusion is expressed as % area at risk. Values are means ± SE of 5 experiments. *P < 0.05 compared with DR control; #P < 0.05 compared with DS control.

 
Effect of 1400W and L-NAME on losartan-induced cardioprotection. We next investigated whether NOS plays any roles in I/R injury or protection in the DS rat heart. Brief treatment with 1400W or L-NAME before coronary artery occlusion had no effect on LV function during reperfusion and infarct size in the DR rats (Table 3 and Fig. 6). However, 1400W and L-NAME significantly decreased LVSP and increased LVEDP during reperfusion and significantly increased infarct size in the DS rats. In contrast, 1400W had no effect on LV function during reperfusion and infarct size in the DS rats treated with losartan (Table 4 and Fig. 7). However, brief treatment with L-NAME before coronary artery occlusion significantly decreased LVSP and increased LVEDP during reperfusion and significantly increased infarct size in the DS rats treated with losartan.


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  		Table 3. Effect of 1400W and L-NAME on left ventricular function

 

Figure 6
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  		Fig. 6. DR and DS rats were treated with or without 1400W or L-NAME as described in Fig. 1. Infarct size after 30 min of left coronary artery occlusion and 2 h of reperfusion is expressed as % area at risk. Values are means ± SE of 5 experiments. *P < 0.05 compared with DR control; #P < 0.05 compared with DS control.

 

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  		Table 4. Effect of losartan, 1400W, and L-NAME on left ventricular function

 

Figure 7
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  		Fig. 7. DR and DS rats were treated with or without Los and 1400W or L-NAME as described in Fig. 1. Infarct size after 30 min of left coronary artery occlusion and 2 h of reperfusion is expressed as % area at risk. Values are means ± SE of 5 experiments. *P < 0.05 compared with DR control; #P < 0.05 compared with DS control; {dagger}P < 0.05 compared with DS Los.

 

   DISCUSSION
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
Although oxidative stress has been implicated in the pathophysiology of I/R injury, it also promotes redox-sensitive signal transduction that protects cells from lethal I/R injury (7, 21). Because oxidative stress is increased in the hypertensive heart (25), we attempted to address the question as to whether any protective mechanisms against I/R injury are installed in the hypertensive heart through oxidative stress and whether inhibition of oxidative stress by a genuine antioxidant and an antihypertensive drug ARB modulates the tolerance to I/R injury in the hypertensive heart. The present study demonstrated that DS rats fed a high-salt diet for 6 wk were exposed to oxidative stress as shown by an increase in MDA + 4-HNE in the heart. Moreover, oxidative stress promoted iNOS expression in the DS rat heart, because MPG abrogated the increase in MDA + 4-HNE and the expression of iNOS. Such redox-sensitive upregulation of iNOS appears to be crucial in acquisition of tolerance to I/R injury in the DS rat heart because MPG and 1400W deteriorated LV function during reperfusion and increased infarct size in this heart.

In contrast to MPG, inhibition of oxidative stress and upregulation of iNOS by losartan did not affect LV function and infarct size after I/R in the DS rat heart. These results suggest that losartan preserved the tolerance to I/R injury through a mechanism independent of oxidative stress and iNOS in the DS rat heart. On the other hand, losartan increased phosphorylation of eNOS in the DS heart, suggesting that eNOS was activated by losartan in the DS rat heart. Although preischemic treatment with 1400W had no effect on LV function and infarct size after coronary artery occlusion and reperfusion in DS rats treated with losartan, L-NAME deteriorated LV function and increased infarct size in these rat hearts, indicating that eNOS activation is involved in the cardioprotection mediated by losartan. However, because L-NAME is a nonselective NOS inhibitor, the role of neuronal NOS cannot be eliminated at present.

The mechanism of cardioprotection against I/R injury afforded by ARBs has not been completely understood. Blocking AT1 receptors renders local angiotensin II more accessible to angiotensin II type 2 (AT2) receptors. Resultant activation of AT2 receptors increases generation of bradykinin, which has been implicated as a potential mechanism for ARB-mediated cardioprotection (14, 17). Activation of eNOS is a downstream effect of increased bradykinin generation (1, 28). eNOS activation has been shown to be indispensable for cardioprotection against I/R injury mediated by late IPC (30), a dipyridamole-atorvastatin combination (31), and the volatile anesthetic agent isoflurane (29). The results of the present study are consistent with a potential role of eNOS in preservation of the tolerance to I/R injury in the DS rat heart. The reason why losartan significantly increased eNOS phosphorylation and conferred cardioprotection only in the DS rat heart and not in the DR rat heart is probably less angiotensin II generation and less activation of AT2 receptors in the DR rat heart. Although we did not investigate cardioprotective signals other than eNOS, Akt and protein kinase C-{varepsilon} may constitute a cardioprotective signaling module with eNOS (32).

The DS rats were the choice for the experimental model because hypertension in these rats is known not to be affected by ARBs. This lack of antihypertensive effect is important to compare the effect of elimination of oxidative stress on cardioprotection of the ARB because lowering blood pressure by itself is protective against I/R injury. Although infarct size after I/R was comparable between DR and DS rat hearts, it is considered that the DS rat heart may be a more cardioprotective phenotype, and infarct size would be smaller in the DS rat heart if blood pressure were comparable between these rats. This assumption was confirmed by the observation that when the DS rats were fed a low-salt diet for 4 days before experiments blood pressure at the time of coronary artery occlusion was significantly decreased in these rats, associated with significant reduction of infarct size compared with the DS rats continuously fed a high-salt diet (Matsuhisa S and Otani H, unpublished observation). Moreover, iNOS expression remained increased after 4 days of a low-salt diet, and 1400W or L-NAME increased infarct size in the DS rat heart, indicating that iNOS-dependent tolerance to I/R injury was preserved for at least 4 days after a low-salt diet and a decrease of blood pressure.

DS rats fed a high-salt diet for 6 wk developed LV hypertrophy associated with hypertension. Although LVEDD was not different between DR and DS rats, LVESD was significantly smaller in the DS rats. Consequently LV FS was significantly higher in the DR rats, indicating that the DS rat heart remained hyperdynamic after 6 wk of high-salt diet. Although the DS rat hearts had acquired tolerance to I/R injury at this compensated stage of hypertension, it is unknown whether these hearts retained a cardioprotective phenotype at the decompensated stage of hypertension when DS rats are deteriorated to heart failure. There is no definite answer to this question; however, it is possible that these hearts may not be tolerant to I/R injury after deterioration to heart failure. In line with this idea, Miki and associates (20) investigated whether or not postinfarction ventricular remodeling interferes with the preconditioning mechanism, a representative feature of ischemic tolerance. The investigators created myocardial infarction to induce remodeling by permanently ligating the left coronary artery in rabbits 2 wk before isolation of the hearts. When the isolated buffer-perfused hearts were subjected to I/R injury, IPC with two episodes of 5-min ischemia protected sham-operated but not remodeled hearts, indicating that the failing heart was not adequately preconditioned. Further studies are necessary to investigate whether or not ischemic tolerance and the preconditioning phenomenon are similarly lost after heart failure in the DS rat heart.

In conclusion, the DS rat heart at a compensated stage of hypertension is promoted to a cardioprotective phenotype through oxidative stress and upregulation of iNOS. Losartan preserves the tolerance to I/R injury by activating eNOS despite elimination of redox-sensitive upregulation of iNOS and iNOS-dependent cardioprotection in the DS rat heart.


   FOOTNOTES
 

Address for reprint requests and other correspondence: H. Otani, Cardiovascular Center, Kansai Medical Univ., 10-15 Fumizono-cho, Moriguchi City, 570-8507, Japan (e-mail: otanih{at}takii.kmu.ac.jp)

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.


   REFERENCES
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 

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