2c1k hipertensão e atividade física

Prévia do material em texto

INTRODUCTION
Epidemiological studies verify that hypertension is a serious
public health problem because it is a major and well-known risk
factor for the occurrence of several cardiovascular (peripheral
arterial disease, ischemic heart disease, aortic aneurysm) and
cerebrovascular diseases resulting in multi-organ damage (1).
Renovascular hypertension (RVH), which is a form of secondary
hypertension resulting from fibromuscular dysplasia or
atherosclerotic renal artery disease, involves the activation of
renin-angiotensin-aldosterone (2) and sympathetic nervous
systems (3) and the presence of chronic oxidative stress (4).
Although RVH constitutes a small percentage of hypertensive
patients, it is the most common cause of hypertension that can be
surgically corrected (5). During the initial stages of RVH,
increased angiotensin II activity, which maintains the high blood
pressure, contributes to the progression of atherogenesis and
glomerulosclerosis by stimulating vasoconstriction, increasing
endothelin release, enhancing extracellular matrix deposition
and vascular remodeling (6). Reactive oxygen species (ROS)
further trigger the release of vasoconstrictor substances and play
an important role in the pathogenesis of hypertension-induced
organ failure (7-10). A number of studies have also demonstrated
that increased oxidative stress results in endothelial dysfunction,
causing reduced bioavailability of nitric oxide (NO), one of the
most powerful endothelium-derived vasodilators (11-13).
Apart from various pharmacological treatments that target
high blood pressure, physical exercise as a non-pharmacological
approach was previously shown to delay the generation of
hypertension or to reduce the level of high blood pressure by
central and peripheral neurohumoral mechanisms (14-16) and by
correcting the imbalance between the production of ROS and the
antioxidant defense systems (17). A number of well-controlled
epidemiologic studies with large patient groups have reported
JOURNAL OF PHYSIOLOGY AND PHARMACOLOGY 2016, 67, 1, 45-55
www.jpp.krakow.pl
Z.N.O. KUMRAL1, G. SENER2, S. OZGUR3, M. KOC4, S. SULEYMANOGLU5, C. HURDAG6, B.C. YEGEN1
REGULAR EXERCISE ALLEVIATES RENOVASCULAR HYPERTENSION-INDUCED
CARDIAC/ENDOTHELIAL DYSFUNCTION AND OXIDATIVE INJURY IN RATS
1Department of Physiology, School of Medicine, Marmara University, Istanbul, Turkey; 2Faculty of Pharmacy, Department of
Pharmacology, Faculty of Pharmacy, Marmara University, Istanbul, Turkey; 3Department of Biochemistry, Faculty of Dentistry,
School of Medicine, Marmara University, Istanbul, Turkey; 4Department of Nephrology, School of Medicine, Marmara University,
Istanbul, Turkey; 5Department of Cardiology, Gulhane Military Academy, Ankara, Turkey; 6Department of Histology and
Embryology, Istanbul Bilim University, Istanbul, Turkey
The importance of physical activity in the management of renovascular diseases is well-known, but lacks evidence of
underlying mechanisms. The purpose of the study was to elucidate the protective/therapeutic effects of regular exercise on
experimental renovascular hypertension (RVH)-induced oxidative stress and cardiac dysfunction. Wistar albino rats
underwent a RVH surgery (2K1C, Goldblatt). Three weeks later half of the rats started swimming exercise for 9 weeks (n
= 15), while the sedentary RVH group (n = 15) had no exercise during that period. Sham-operated control rats (n = 10),
had the similar surgical procedures but the left renal artery was left unclipped. Body weights were monitored, and blood
pressures were measured weekly using tail-cuff. Echocardiographic evaluation was performed on the 3rd week and on the
12th week of the experiment before the rats were decapitated. Heart and thoracic aorta were removed and serum was
collected, while aortic samples were put in a 10% formaldehyde solution for immunochemistry. Cardiac tissue samples
obtained from each animal were used for the determination of tissue myeloperoxidase (MPO) and catalase (CAT) activities,
malondialdehyde (MDA), and glutathione (GSH) levels. In the sedentary RVH group, aortic contractile response
(contraction/relaxation in isolated organ bath), left ventricular diastolic and systolic dimensions, and immunohistochemical
staining of aortic inducible nitric oxide synthase (iNOS) were increased, while ejection fraction and aortic endothelial nitric
oxide synthase (eNOS) staining were decreased. RVH in the sedentary rats resulted in increased pro-inflammatory
cytokines (TNF-α, IL-2, IL-6), lipid peroxidation (malondialdehyde) and neutrophil infiltration (myeloperoxidase activity)
along with reductions in antioxidant glutathione and catalase levels in the cardiac tissue. Exercise after RVH increased the
immunhistochemical staining of aortic eNOS, decreased iNOS staining and reversed the alterations in echocardiographic
and oxidative parameters. Regular exercise commenced after RVH surgery alleviated renovascular hypertension-induced
oxidative injury, by modulating oxidant-antioxidant balance via the involvement of the endothelial NO system.
K e y w o r d s : renovascular hypertension, nitric oxide synthase, echocardiography exercise, oxidative stress, endothelium,
proinflammatory cytokines, catalase
that the mortality risk of the hypertensive individuals is reduced
as their exercise capacity is increased (18-20). Exercise training
in hypertensive subjects was also shown to reduce left
ventricular hypertrophy or does not worsen the underlying
inflammatory burden associated with hypertension (21).
Similarly, exercise training in hypertensive animals was shown
to decrease blood pressure (22-24) while an increase in vessel
compliance (25) was reported.
Although the importance of physical activity in the
prevention of cardiovascular and renal diseases has been
demonstrated in many studies, the impact of exercise training on
RVH-induced vascular dysfunction and oxidative stress with its
underlying mechanisms has not been thoroughly elucidated yet.
Since hypertension is regarded as a chronic low-grade
inflammation with elevated plasma levels of the pro-
inflammatory cytokines in hypertensive patients (26, 27), the
impact of exercise on RVH-induced oxidative damage of cardiac
tissue needs to be investigated. Accordingly, through
hemodynamic measurements, biochemical and histological
analyses, we aimed to elucidate the therapeutic effect of exercise
training when performed after the onset of RVH.
MATERIALS AND METHODS
Animals
Male Wistar albino rats (300 – 350 g, 10 weeks old, n = 40),
supplied by the MU Animal Center (DEHAMER), were housed
in a humidity (65 – 70 %) and temperature-controlled room (22
± 2°C) with standardized light/dark (12/12 hour) cycles. Rats
were fed with standard rat pellets and tap water ad libitum.
All experimental protocols were approved by the Marmara
University (MU) Animal Care and Use Committee.
Surgery and experimental design
One day before the surgery, blood pressures of all rats were
measured and transthoracic echocardiography was made to
obtain the basal values. Following the anesthesia of the rats with
intraperitoneal injection of ketamine (100 mg/kg, i.p.) and
chlorpromazine (0.75 mg/kg, i.p.), two-kidney, one-clip (2K1C,
Goldblatt) procedure was used to induce RVH (28). Briefly, a
silver clip (internal diameter 0.25 mm) was placed around the
left renal artery of rats (n = 30), while the right artery has
remained untouched. Sham-operated control rats (n = 10) had
the similar surgical procedures but the left renal artery was left
unclipped.
Exercise was started three weeks after RVH surgery (n = 15)
and continued for 9 weeks without a prior training period and no
load was applied on the swimming rats. A moderate load
swimming exercise model was selected in the exercise groups
(29). Swimming sessions of 30-min were made in a cylindrical
glass pool (100 × 50 × 50 cm) filled with35-cm high lukewarm
water and were continued 5 days/week for nine consecutive
weeks. Sham-operated control rats and half of the RVH group
that had surgery on the 3rd week (n = 15) were put on their feet
in a separate glass tank filled with only 5-cm water and were left
sedentary.
Body weights were monitored, and blood pressures were
measured using tail-cuff on a weekly basis throughout the
experiment. RVH surgeries of sedentary and exercised groups
were performed on the 3rd week after the blood pressure
recordings were taken. On the 12th week of the experiment, the
second echocardiographic evaluation was performed and the rats
were decapitated. Heart and thoracic aorta were carefully
removed and blood was collected for the measurement of serum
tumor necrosis factor-alpha (TNF-α), interleukin-2 (IL-2) and
IL-6 levels. Thoracic aorta was placed in a dish containing
chilled Krebs-Henseleit buffer solution aerated with 95% O2 and
5% CO2, while aortic samples were put in a 10% formaldehyde
solution for immunochemistry. Heart weights were measured
and cardiac tissue samples obtained from each animal were
stored at –80°C until the determination of tissue
myeloperoxidase (MPO) and catalase (CAT) activities,
malondialdehyde (MDA) and glutathione (GSH) levels.
Blood pressure measurement
Following the swimming sessions that were repeated for 
5 days of the week, 2 days of rest period was given. After this
interval and before a swimming bout was started, indirect blood
pressure measurements were made by the tail-cuff method
(Biopac MP35 Systems, Inc. COMMAT Ltd., Ankara, Turkey)
on the first day of the week. Rats were placed for 10 min in a
chamber heated to 35°C, and then a cuff equipped with a
pneumatic pulse sensor was wrapped around the tail of the rat
that was placed in an individual plastic restrainer. Blood pressure
recorded from each rat during each measurement period was
averaged from at least three consecutive readings on that
occasion. Blood pressure recordings were made before the first
recording of the rats that had a two-day resting.
Echocardiography
Echocardiographic imaging and calculations were done
using a 12-MHz linear transducer and 5 – 8 MHz sector
transducer (Vivid 3, General Electric Medical Systems
Ultrasound, Tirat Carmel, Israel) according to the guidelines
published by the American Society of Echocardiography (30)
Under ketamine (50 mg/kg, i.p.) anesthesia, transthoracic
echocardiography was made from M-mode, and after observing
at least six cardiac cycles, two-dimensional images were
obtained in the parasternal long and short axes at the level of the
papillary muscles. Interventricular septal thickness (IVS), left
ventricular diameter (LVD) and left ventricular posterior wall
thickness (LVPW) were measured during systole (s) and diastole
(d). Ejection fraction, fractional shortening and left ventricular
mass and relative wall thickness were calculated from the M-
mode images using the following formulas:
% ejection fraction = (LVDd)3 - (LVDs)3/(LVDd)3 × 100; %
fractional shortening = LVDd - LVDs/LVDd × 100; left
ventricular mass = 1.04 × ((LVDd + LVPWd + IVSd)3 -
(LVDd)3) × 0.8 + 0.14; relative wall thickness = 2 ×
(LVPWd/LVDd) (30).
Vascular reactivity studies
After removal of the surrounding connective tissue, fresh
thoracic aorta was cut transversely into rings approximately 4-
mm wide and mounted in an organ bath (Biopac MP35 Systems,
Inc. COMMAT Ltd., Ankara, Turkey) containing 20 ml of aerated
(95% O2 and 5% CO2). Krebs-Henseleit buffer maintained at
37°C. The rings were placed under a resting tension of 1.0 g.
After a 60-min period of equilibration, the rings were exposed to
80 mM KCl. For the measurement of contractile response to
phenylephrine (10–9 to 10–3 M), cumulative concentration-
response curves were obtained in a stepwise manner, where the
consecutive concentration was added after the response has
reached the plateau. After completion of phenylephrine
concentration curves, tissues were washed 3 times in 30 minutes.
Then, on the rings that were pre-contracted with the submaximal
dose of phenylephrine (3X10–6 M), the relaxation responses were
evaluated by adding increasing cumulative concentrations of
46
carbachol (CCh; 10–9 - 10–3 M). Contractile responses to
phenylephrine are expressed as percentages of the maximal
contraction induced by 80 mM KCl, and the relaxation responses
to CCh are expressed as percentages of the contraction caused by
3X10–6 M phenylephrine.
Measurement of cytokines in the serum
Serum levels of tumor necrosis factor-alpha (TNF-α),
interleukin-2 (IL-2) and IL-6 were quantified according to the
manufacturer’s instructions and guidelines (Biosource Europe
S.A., Nivelles, Belgium) using enzyme-linked immunosorbent
assay (ELISA) kits specific for rat cytokines.
Measurement of cardiac myeloperoxidase activity
Tissue MPO activity, which is frequently utilized to estimate
tissue neutrophil accumulation in inflamed tissues, was shown to
correlate significantly with the number of neutrophils
determined by histochemical analysis (31). The method used to
determine MPO activity in the cardiac tissues was similar to that
previously described by others (31). The cardiac tissue samples
(0.2 – 0.3 g) were homogenized in 10 volumes of ice-cold
potassium phosphate buffer (50 mm K2HPO4, pH 6.0) containing
hexadecyltrimethylammonium bromide (HETAB; 0.5%, w/v).
The homogenate was centrifuged at 41,400 g for 10 min at 4°C,
and the supernatant was discarded. The pellet was then re-
homogenized with an equivalent volume of 50 mm
K2HPO4containing 0.5% (w/v) HETAB and 10 mm EDTA
(Sigma). Myeloperoxidase activity was assessed by measuring
the H2O2-dependent oxidation of o-dianizidine 2HCl. One unit
of enzyme activity was defined as the amount of MPO present
per gram of tissue weight that caused a change in absorbance of
1.0 min–1 at 460 nm and 37°C.
Measurement of malondialdehyde (MDA) and glutathione
(GSH) levels in the cardiac tissues
Cardiac tissue samples were homogenized in ice-cold
trichloracetic acid (1 g tissue plus 10 ml 10% TCA) in an Ultra
Turrax tissue homogenizer. Homogenized cardiac samples were
centrifuged at 2,000 g for 15 min at 4°C. The supernatant was
removed and re-centrifuged at 41,400 g for 8 min. MDA levels
were assayed for products of lipid peroxidation by monitoring
thiobarbituric acid reactive substance formation as previously
described (32). Lipid peroxidation was expressed in terms of
MDA equivalents using an extinction coefficient of 1.56 × 10–5
M–1 cm–1 and the results are expressed as nmol MDA/g tissue.
Glutathione measurements were performed using a modification
of the Ellman procedure (33). Briefly, after centrifugation at
2000 g for 10 min, 0.5 ml of supernatant was added to 2 ml of
0.3 mol/l Na2HPO4·2H2O solution. A 0.2 ml solution of
dithiobisnitrobenzoate (0.4 mg/ml 1% sodium citrate) was added
and the absorbance at 412 nm was measured immediately after
mixing. Glutathione levels were calculated using an extinction
coefficient of 1.36 × 105 M–1 cm–1. The results are expressed in
µmol GSH/g tissue.
Measurement of catalase (CAT) activity in the cardiac tissues
The method for the measurement of CAT activity is based on
the catalytic activity of the enzyme that catalyses the
decomposition reaction of H2O2 to give H2O and O2 (34).
Briefly, the absorbance of the tissue samples containing 0.4 ml
homogenate and 0.2 ml H2O2 was read at 240 nm and 20°C
against a blank containing 0.2 ml phosphate buffer and 0.4 ml
homogenate for about 1 min.
Immunohistochemical analysis of endothelial nitric oxide
synthase (eNOS) and inducible nitric oxide synthase (iNOS)
protein distributions in the aorta
Aortae from all groups were fixed in a 10 % formaldehyde
solution and washed in tap water for 2 hours. Then the tissues were
dehydrated with increasing concentrations (70,90, 96 and 100%)
of ethanol and cleared with xylene. Paraffin-embedded sections
that were cut at 3-µm thickness were de-paraffinized with xylene,
and rehydrated with ethanol and water. Antigen retrieval was
accomplished by Decloacking chamber (Bicare Medical DC2008)
in a citrate diva buffer (DV Sitogen 2004 LX, MX pH 6.2) for 40
min at 110°C. The endogenous peroxidase activity was blocked
with 3% H2O2 (ScyTek ACA 125) for 15 min at room temperature
and later rinsed with phosphate buffered saline (PBS). Blocking
reagent (UV Blocking, SycTek AA125) was applied to each slide
followed by 5 min incubation at room temperature in a humid
chamber. Sections were incubated for 1 hour at room temperature
with rabbit polyclonal eNOS antibody (1:100, eNOS Rabbit PAb,
RB-9279-P) or iNOS antibody (1:50, iNOS Ab-1, Rabbit PAb, RB-
9242-P1). Antibodies were diluted in a large volume of UltrAb
Diluent (TA-125-UD). The sections were biotinylated goat anti-
rabbit antibodies (TS-060-HR, SycTek ABF125). After slides were
washed in PBS, the streptavidin peroxidase label reagent (TS-060-
HR, HRP SycTek ABG125) was applied for 20 min at room
temperature in a humid chamber. The colored product was
developed by incubation with 3,3’-diaminobenzidine
tetrahydrochloride dihydrate (DAB) (Thermo Scientific RTU TA-
060-HD). The slides were counterstained with Mayer’s
hematoxylin (LabVision, TA-125-MH) and mounted in entellan
(Shandon Ref: 999040). Immunohistochemical staining was
photographed with light microscope (Leica 390-CU, Germany).
Statistical analysis
Statistical analysis was carried out using GraphPad Prism
6.0 (GraphPad Software, Inc. La Jolla, CA, USA). All data are
expressed as means ± S.E.M. Groups of data were analyzed
using one-way ANOVA followed by Tukey’s multiple
comparison test or Student’s t test where appropriate. Values of
P < 0.05 were regarded as significant.
RESULTS
Effect of exercise performed after the onset of renovascular
hypertension on blood pressure
The basal systolic blood pressures that were recorded before
starting the experiments were not different among three groups
(Fig. 1). As expected in the current RVH model, mean systolic
blood pressures measured on the 3rd week of clip placement were
elevated in the sedentary RVH and the exercised RVH groups
significantly as compared with the blood pressures of the control
group (P < 0.001), indicating the onset of hypertension. Even on
the 12th week, the high blood pressure of the RVH group that has
exercised for 9 weeks was not significantly different than that of
the sedentary RVH group.
Effect of exercise performed after the onset of renovascular
hypertension on cardiac hypertrophy and renal atrophy
In order to assess cardiac hypertrophy, the ratio of the heart
weight (mg) to the body weight (g) (HW/BW) measured on the
day of decapitation was used. The body weights of the rats were
increased gradually throughout the 12-week experimental period
without any significant differences among the experimental
47
groups. However, the HW/BW ratios calculated in the sedentary
and exercised RVH groups were significantly increased when
compared with the ratio of the control group (P < 0.001, Table
1), suggesting eccentric cardiac hypertrophy. Renal atrophy
index, which is the ratio of the clipped left kidney weight (mg)
to the body weight (g), was measured on the day of decapitation.
Renal atrophy indices of both the sedentary and exercised RVH
groups were decreased with respect to sham-operated control
group (P < 0.001, Table 1), confirming the experimental model
of hypertension.
Effect of exercise performed after the onset of renovascular
hypertension on echocardiographic measurements
Transthoracic echocardiography measurements (Fig. 2)
monitored on the 12th week are summarized in Table 1. When
compared to sham-operated control group, RVH caused
significant increases in IVS, LV end-diastolic and LV end-
systolic dimensions of the sedentary group (P < 0.01 – 0.001),
while these parameters were reduced in the exercised RVH
group (P < 0.05 – 0.01; Table 1). Furthermore, reductions in
48
Fig. 1. Systolic blood pressure measurements in the sedentary sham-operated control (n = 10), sedentary RVH (n = 15) and exercised
RVH (n = 15) groups. Systolic blood pressure recorded by tail-cuff from each rat was averaged from at least three consecutive readings
on that occasion. “Week 0” represents the recordings obtained before clip placement, “week 3” and “week 12” represent the respective
weeks following clip placement. “Week 3” corresponds to the time-point for the initiation of exercise sessions and “week 12”
corresponds to the 9th week of the exercise sessions.
***P < 0.001: compared to control group.
Sham-operated 
control 
Sedentary RVH Exercised RVH
Cardiac oxidative stress parameters 
Malondialdehyde (nmol/g tissue) 17.2 ± 1.9 51.8 ± 4.2
***
 17.4 ± 1.2
+++
Myeloperoxidase (U/g tissue) 4.8 ± 0.8 13.2 ± 1.6
***
 4.8 ± 0.6
+++
Glutathione (µmol/g tissue) 3.5 ± 0.2 1.5 ± 0.04
***
 3.09 ± 0.1
+++
Catalase (U/mg protein) 231.2 ± 9.2 159.3 ± 11.9
***
 194.3 ± 11.8
+
Serum cytokine levels 
TNF-� (pg/ml) 25.3 ± 7.7 80.3 ± 6.4
***
 48.1 ± 8.6
+
IL-2 (pg/ml) 54.1 ± 13.2 180.3 ± 18.1
***
 97.0 ± 11.5
++
IL-6 (pg/ml) 8.0 ± 1.3 18.5 ± 2.3
***
 15.9 ± 0.5
**
Echocardiographic measurements 
IVS (mm) 2.2 ± 0.1 3.9 ± 0.2
***
 2.9 ± 0.2
+
LVDs (mm) 2.7 ± 0.2 5.0 ± 0.2
***
 3.6 ± 0.2
+
LVDd (mm) 4.3 ± 0.2 6.0 ± 0.2
**
 3.7 ± 0.2
++
Ejection fraction (%) 80.3 ± 1.2 67.3 ± 2.3
**
 75.6 ± 3.2
++
Fractional shortening (%) 49.3 ± 2.1 25.8 ± 2.3
***
 39.6 ± 2.2
++ 
Morphological changes 
Heart weights (mg/g body weight) 2.8 ± 0.1 4.1 ± 0.2 
***
 4.2 ± 0.1
***
Renal weights (mg/g body weight) 2.56 ± 0.13 0.92 ± 0.3
***
 0.6 ± 0.05
***
Table 1. Cardiac oxidative stress parameters, serum proinflammatory cytokine levels, echocardiographic measurements and
morphological changes in the sham-operated control, sedentary renovascular hypertension (RVH) (n = 15) and exercised RVH (n = 15)
groups. IVS, interventricular septal thickness; LVDd, left ventricular diameter in diastole; LVDs, left ventricular diameter in systole. **P
< 0.01, ***P < 0.001 compared with the control group. +P < 0.05, ++P < 0.01, +++P < 0.001, compared with the sedentary RVH group.
ejection fraction and fractional shortening observed in the
sedentary RVH group (P < 0.01 – 0.001) were found to be
elevated in the exercised RVH group (P < 0.01).
Effect of exercise performed after the onset of renovascular
hypertension on the contraction and relaxation of the aortic
rings
In the aortic rings of the sham-operated control rats, addition
of 10–9 to10–3 M phenylephrine cumulatively into the organ bath
caused a concentration-dependent contraction, reaching the 50%
maximal response (EC50) at the 7.83 × 10–7 M concentration (Fig.
3A). Both in the sedentary RVH or exercised RVH groups that had
a 9-week duration of hypertension after its onset, the contractile
responses of the aortic rings to phenylephrine were significantly
amplified, demonstrating reduced EC50 values (6.87 × 10–7 M and
5.9 × 10–7 M, respectively).
CCh added cumulatively at doses of 10–9 to 10–3 M to aortic
rings of sham-operated control group, which were pre-
contracted with the submaximal (60 – 70% of maximal
contraction) dose of phenylephrine (3 × 10–5 M), caused a dose-
dependent relaxation response with an EC50 value of 1.84 ×
10–6 M (Fig. 3B). EC50 value in the sedentary RVH (1.42 ×
10–6 M) was not different than that of the control group, while in
the exercised RVH groups EC50 value of the relaxation
responses was significantly higher (3.62 × 10–6 M; P < 0.05) as
compared to the control group, indicating a decrease in the CCh-
induced aortic relaxation.
49
Fig. 2. Representativeechocardiographic scans
of (A) the sedentary sham-operated control
group with normal M-mode view; (B) sedentary
renovascular hypertension (RVH) group with
increased interventricular septum and left
ventricular posterior wall thickness; and (C)
exercised RVH group, demonstrating normal
M-mode view as the sham-operated group.
Effect of exercise performed after the onset of renovascular
hypertension on the serum levels of pro-inflammatory cytokines
Serum levels of pro-inflammatory cytokines TNF-α, IL-2
and IL-6 were increased in the sedentary RVH group as
compared to the control group (P < 0.001; Table 1). In the
exercised RVH group, TNF-α (P < 0.05) and IL-2 (P < 0.01)
levels, but not IL-6 levels, were significantly depressed.
Effect of exercise performed after the onset of renovascular
hypertension on the cardiac malondialdehyde and glutathione
levels, myeloperoxidase and catalase activities
In the sedentary RVH group, cardiac MDA level was
significantly higher than that of the sham-operated control
group, indicating increased lipid peroxidation (P < 0.001; Table
1). Exercise performed after the onset of RVH abolished this
increase and returned the cardiac MDA levels back to control
levels (P < 0.001). Similarly, MPO activity, which is accepted as
an indicator of neutrophil infiltration, was significantly higher in
the cardiac tissues of the sedentary RVH group (P < 0.001) as
compared to the control group (Table 1). However, exercise
significantly decreased the cardiac MPO activity when it was
performed after the onset of RVH (P < 0.001).
In accordance with increased MDA level and MPO
activity, induction of RVH in the sedentary rats caused
significant reductions in cardiac GSH levels and CAT
activities, when compared to those of the control rats (P <
0.001, Table 1). However, in the exercised RVH group, GSH
levels and CAT activities were replenished significantly (P <
0.001 and P < 0.05).
Effect of exercise performed after the onset of renovascular
hypertension on the immunohistochemical staining of inducible
nitric oxide synthase and endothelial nitric oxide synthase in
the aorta
Qualitative immunohistochemical analysis on the aortic
tissues of the sham-operated control rats revealed that the iNOS
immunoreactivity was evident in tunica media layer (Fig. 4A).
50
Fig. 3. (a) Concentration-response curves
obtained by cumulative addition of
phenylephrine (PE) to rat thoracic aorta. RVH
increased contractile activity of sedentary and
exercised RVH groups in comparison to
sedentary sham-operated control group (*P <
0.05). Points indicate percentage of contraction
induced by 124 mM KCl. (b) Concentration-
response curves obtained by cumulative
addition of carbachol (CCh) to rat thoracic aorta
strips pre-contracted with 30 mM
phenylephrine. RVH impaired relaxation
response in the sedentary group with respect to
sedentary sham-operated control group (*P <
0.05), and the relaxation response in the
exercised RVH group was higher than that of
the sedentary RVH group (+P < 0.05). Values are
shown as mean ± S.E.M. of eight experiments.
However, the iNOS immunostaining (dark brown) of aortic wall
layers appeared to be more intense in the sedentary RVH (Fig.
4B) and exercised RVH (Fig. 4C) groups as compared to the
control group. Normal eNOS immunoreactivity was observed in
the tunica media layer of the sham-operated control group (Fig.
4D). The dark brown immunostaining indicating eNOS activity
was more abundant in the exercised RVH group (Fig. 4F)
compared to the sham-operated control group. However, in the
sedentary group, the immunostaining of eNOS was not observed
in none of the layers of the vessel wall (Fig. 4E).
DISCUSSION
The present results demonstrate that blood pressure, aortic
contractile response, left ventricular dimensions, cardiac
oxidative damage and aortic iNOS staining were increased in the
sedentary rats, while eNOS staining in the aorta and ejection
fraction were decreased. Exercise after RVH reversed the
echocardiographic alterations and oxidative parameters and
increased the staining of aortic eNOS, indicating the positive
impact of exercise on RVH-induced oxidative damage and
51
Fig. 4. Representative photomicrographs of thoracic aortae cross sections showing immunohistochemical staining for iNOS and eNOS
at tunica media (TM) and tunica adventitia (TA) layers of the sedentary sham-operated control (A, D), sedentary RVH (B, E) and
exercised-RVH (C, F) groups. Immunohistochemical staining was observed as dark brown on the background staining with
hematoxylin. Original magnification: × 100, inset: × 400.
cardiac dysfunction. The reversal of RVH-induced increases in
the pro-inflammatory cytokines and the oxidant/antioxidant
imbalance in the cardiac tissue implicate the therapeutic effects
of exercise on hypertension-induced oxidative injury.
It has been shown that physical exercise delays the
generation of hypertension and reduces the level of high blood
pressure (16) when exercise is started before the development of
hypertension. However, our data suggests that implementing
exercise sessions after the development of hypertension -
starting by the third week of RVH surgery - had no effect on high
blood pressure, but improved oxidative damage and cardiac
dysfunction. This anti-oxidant effect of exercise was reached
even no load adjustments were made throughout the swimming
sessions. These data implicate that starting exercise as a life style
change following the development of hypertension cannot
reduce blood pressure, but can be of benefit in controlling the
oxidant/antioxidant balance in the cardiac tissue.
In the long term, hypertension often yields left ventricular
hypertrophy, which is a major risk factor for the development of
coronary heart disease, ventricular arrhythmia, and sudden
cardiac death (35). Despite any changes in the heart weight, our
data revealed that induction of RVH in sedentary rats resulted in
increased left ventricular end-diastolic/end-systolic dimensions
and thickened left interventricular septum with a concomitant
reduction in ejection fraction. The occurrence of hypertensive
vascular disease involves several hemodynamic mechanisms
including myocyte hypertrophy, collagen deposition, increased
ratio of arteriolar wall thickness/lumen and coronary arteriolar
compression by the left ventricle (36). In addition, increased
oxidative stress was shown to play an important part in the
pathogenesis of experimental renovascular hypertension (37). In
vivo and in vitro studies demonstrated that vascular endothelial
cells release cyclooxygenase-derived endothelium-dependent
contracting factors and ROS, resulting in facilitated
vasoconstriction along with impaired endothelium-dependent
relaxation mediated by the NO production (12, 38-42). Since
eNOS has a major role in determining blood flow and resistance
(43, 44), impairment in its expression or its increased
phosphorylation, as well as the presence of oxidative stress, may
result in endothelial dysfunction (45). Our results revealed that
decreased eNOS and increased iNOS staining were observed in
the aorta of sedentary rats with RVH. In accordance with these
results, we also demonstrated that aortic contractile response
was exaggerated along with a diminished relaxation response to
CCh, showing that altered iNOS/eNOS activities result in aortic
dysfunction. Moreover, exercise reversed the
contraction/relaxation responses in parallel with a possible
improvement in endothelial function.
Evidence obtained from animal (46, 47) and human studies
(48, 49) indicate that oxidative stress induced by hypertension
cause damage in several target organs (50). On the other hand,
several antioxidants, including GSH, vitamin C, and superoxide
dismutase were reported to improve blood pressure control and
vascular function in animals with hypertension andatherosclerosis (51, 52). As with the antioxidants, acute/chronic,
low/moderate-intensity exercise is well known to decrease the
elevated blood pressure and delay the onset of hypertension (53-
59) with a significant improvement in arterial and cardiac
remodeling (57, 60). Accordingly, when added to many
pharmaceutical antihypertensive therapies, exercise is widely
recommended to reduce high blood pressure (58, 61). However,
it has been also reported that the best strategies to enhance
endogenous antioxidant levels may themselves result in
oxidative stress (62), and high intensity exercise may induce
oxidative stress due to the generation of ROS exceeding the
defense capacity of the tissues (63, 64). On the other hand, high
levels of antioxidant enzymes with a greater resistance to
exercise-induced oxidative stress have been observed in
individuals undergoing exercise (65, 66). In spontaneously
hypertensive rats, exercise has improved endothelium function
with increased elastin, fibrillin and eNOS content in the aortic
wall (67). In keeping with these data, the current findings
revealed that oxidative stress due to RVH was ameliorated by
physical exercise, while cardiac dysfunction was significantly
improved, suggesting that exercise-induced improvements of
cardiac function in RVH may be associated with reduced ROS
production. Despite the beneficial effects of exercise on RVH-
induced oxidative stress, RVH-induced hypertension was not
affected. In the 2K1C renin-dependent RVH rats, oxidative
stress is not the only involved pathophysiological mechanism. It
was shown that depression of the central atrial natriuretic factor
in 2K1C hypertension may further promote the elevation of
blood pressure (68). Thus, the central adaptations following the
induction of RVH that result in the hyperactivity of the arteriolar
smooth muscle could not be reversed by exercise.
Recent data have suggested that one of the 3 polymorphisms
of endothelial NOS genotype (NOS3), which control the
synthesis of NO, could be associated with an enhanced risk for
essential hypertension in adults (69). Experimental studies have
also shown that endothelial injury aggravates the inflammatory
response through the loss of normal production of eNOS, while
its induction improves the injurious effects of oxidative stress
(70). In association with these data, the present study reveals that
the protection against oxidative stress afforded by regular
exercise in RVH involves eNOS upregulation and iNOS
suppression. Previously, different exercise protocols were shown
to reduce increased lipid peroxidation, xanthine oxidase activity,
nitrotyrosine and O2– levels, iNOS expression in various models
of hypertension (71-75). Although our present qualitative data
on the eNOS and iNOS staining do not provide concrete
evidence that could be confirmed by Western blotting, the
alterations in the overall staining of the tunica media along with
the contractility data suggest that both eNOS and iNOS activities
are modified by exercise in rats with RVH. Accordingly,
decreased iNOS staining and increased eNOS staining in the
aorta of exercised rats were accompanied with reductions in
neutrophil accumulation and lipid peroxidation, while GSH and
catalase were replenished in accomplishing the anti-oxidative
effect of exercise. In parallel to our results, a recent study has
shown that agents restoring vascular NO via the augmentation of
cytoprotective NO release were proved to reduce both
inflammation and hypertension in hypertensive rats with
diabetes (76).
In hypertensive patients, the plasma levels of the pro-
inflammatory cytokines were reported to increase, verifying that
hypertension is a chronic low-grade inflammation (26, 27).
Inflammatory active state in hypertension induces the activation
of numerous pro-inflammatory genes (TNF-α, IL-1β, IL-6, and
monocyte chemoattractant protein-1 (MCP-1) (77). Pro-
inflammatory cytokines have been found to activate ROS, which
in turn can activate various intracellular signaling pathways (78).
In the current study, exercise depressed RVH-induced elevations
in TNF-α and IL-2, suggesting the inhibitory effect of exercise
on pro-inflammatory cytokine production. However, RVH-
induced elevations in IL-6 were not significantly reduced by
exercise, which may be explained by the direct stimulatory effect
of muscle contraction on the generation of IL-6 as a myokine
(79). Blunted endothelium-dependent relaxation of pre-
contracted aortic rings and decreased eNOS expression observed
as indicators of endothelial dysfunction in the current study may
be associated with the overproduction of ROS, which have a
major role in the development and progression of hypertension
(78, 79). Thus, the decreased bioavailability of NO, as well as
increased ROS production, accompanied with the generation of
52
inflammatory cytokines (82-84) could be contributing to blood
pressure elevation and the progression of renal disease by
facilitating the remodeling of the vascular wall. Although the
effects of exercise as a protective and therapeutic tool in
improving antioxidant capacity and reducing oxidative tissue
damage were extensively studied (85-91), impact of regular
moderate exercise on renovascular hypertension-induced
oxidative cardiac injury was not elucidated thoroughly yet. A
recent study has shown that exercise reversed the RVH-induced
blockade of endogenous NO generation within the
paraventricular nucleus (92). The present results indicate an
increment in cytokine levels in sedentary RVH (93), while
exercise exerted a protective effect on cardiac tissue concomitant
with a reduction in the levels of the pro-inflammatory cytokines.
In accordance with the pronounced activation of the
inflammatory markers, RVH in the sedentary rats elevated the
cardiac MDA levels and MPO activity and depleted the
antioxidant GSH and catalase contents of the cardiac tissue,
while regular swimming exercise performed after the onset of
RVH attenuated oxidative damage by up-regulating the cardiac
antioxidant defense system. Similarly, treadmill exercise has
resulted in the up-regulation of aortic and cardiac antioxidant
defense systems of hypertensive rats (72, 73). Meta-analysis of
randomized controlled intervention trials have shown that
exercise has more pronounced effects on the blood pressure of
hypertensive patients as compared to normotensive subjects that
have followed the similar exercise programs (55), and it is
widely recommended to all hypertensive patients (1, 94, 95).
Based on our results, it can be concluded that moderate
exercise ameliorates endothelium dysfunction and cardiac
oxidative stress even after emergence of hypertension, and
merits consideration as a preventive and a therapeutic tool for
renovascular hypertension. Physical activity, as a part of lifestyle
modification, should be considered as a common therapeutic
intervention for the control of renovascular hypertension and
associated cardiovascular outcomes.
Acknowledgement: The study was partially presented at the
The Physiological Society Meeting in Oxford and was supported
by Marmara University Research Fund (SAG-C-YLP-211009-
0305). The authors are grateful to Prof. Feriha Ercan for her critical
evaluation and re-organization of the histopathological data.
Conflict of interests: None declared.
REFERENCES
1. Chobanian AV, Bakris GL, Black HR, et al. The seventh
report of the Joint National Committee on Prevention,
Detection, Evaluation, and Treatment of High Blood
Pressure: the JNC 7 report. JAMA 2003; 289: 2560-2572.
2. Garovic V, Textor SC. Renovascular hypertension: current
concepts. Semin Nephrol 2005; 25: 261-271.
3. Campese VM, Ku E, Park J. Sympathetic renal innervation
and resistant hypertension. Int J Hypertens 2011; 2011:
814354. doi: 10.4061/2011/814354
4. Campos RR, Oliveira-Sales EB, Nishi EE, Boim MA,
DolnikoffMS, Bergamaschi CT. The role of oxidative stress
in renovascular hypertension. Clin Exp Pharmacol Physiol
2011; 38: 144-152.
5. Vertes V, Ghose MK. The pathophysiology of renovascular
hypertension. Urol Clin North Am 1975; 2: 227-236.
6. Boissiere J, Eder V, Machet MC, Courteix D, Bonnet P.
Moderate exercise training does not worsen left ventricle
remodeling and function in untreated severe hypertensive
rats. J Appl Physiol (1985) 2008; 104: 321-327.
7. Ersahin M, Sehirli O, Toklu HZ, et al. Melatonin improves
cardiovascular function and ameliorates renal, cardiac and
cerebral damage in rats with renovascular hypertension. 
J Pineal Res 2009; 47: 97-106.
8. Montezano AC, Touyz RM. Oxidative stress, Noxs, and
hypertension: experimental evidence and clinical
controversies. Ann Med 2012; 44 (Suppl. 1): S2-S16.
9. Rodrigo R, Gonzalez J, Paoletto F. The role of oxidative
stress in the pathophysiology of hypertension. Hypertens
Res 2011; 34: 431-440.
10. Toklu HZ, Sehirli O, Ersahin M, et al. Resveratrol improves
cardiovascular function and reduces oxidative organ damage
in the renal, cardiovascular and cerebral tissues of two-
kidney, one-clip hypertensive rats. J Pharm Pharmacol
2010; 62: 1784-1793.
11. Hooper WC, Catravas JD, Heistad DD, Sessa WC, Mensah
GA. Vascular endothelium summary statement I: Health
promotion and chronic disease prevention. Vascul
Pharmacol 2007; 46: 315-317.
12. Reckelhoff JF, Romero JC. Role of oxidative stress in
angiotensin-induced hypertension. Am J Physiol Regul
Integr Comp Physiol 2003; 284: R893-R912.
13. Yetik-Anacak G, Catravas JD. Nitric oxide and the
endothelium: history and impact on cardiovascular disease.
Vascul Pharmacol 2006; 45: 268-276.
14. Rodrigues MC, Campagnole-Santos MJ, Machado RP, et al.
Evidence for a role of AT(2) receptors at the CVLM in the
cardiovascular changes induced by low-intensity physical
activity in renovascular hypertensive rats. Peptides 2007;
28: 1375-1382.
15. Semlitsch T, Jeitler K, Hemkens LG, et al. Increasing physical
activity for the treatment of hypertension: a systematic review
and meta-analysis. Sports Med 2013; 43: 1009-1023.
16. Soares ER, Lima WG, Machado RP, et al. Cardiac and renal
effects induced by different exercise workloads in
renovascular hypertensive rats. Braz J Med Biol Res 2011;
44: 573-582.
17. Parker L, McGuckin TA, Leicht AS. Influence of exercise
intensity on systemic oxidative stress and antioxidant
capacity. Clin Physiol Funct Imaging 2014; 34: 377-383.
18. Blair SN, Kohl HW, III, Barlow CE, Gibbons LW. Physical
fitness and all-cause mortality in hypertensive men. Ann
Med 1991; 23: 307-312.
19. Kokkinos P, Manolis A, Pittaras A, et al. Exercise capacity
and mortality in hypertensive men with and without
additional risk factors. Hypertension 2009; 53: 494-499.
20. Myers J, Prakash M, Froelicher V, Do D, Partington S, Atwood
JE. Exercise capacity and mortality among men referred for
exercise testing. N Engl J Med 2002; 346: 793-801.
21. Rossoni LV, Oliveira RA, Caffaro RR, et al. Cardiac benefits
of exercise training in aging spontaneously hypertensive
rats. J Hypertens 2011; 29: 2349-2358.
22. Evenwel R, Struyker-Boudier H. Effect of physical training
on the development of hypertension in the spontaneously
hypertensive rat. Pflugers Arch 1979; 381: 19-24.
23. Hayashi A, Kobayashi A, Takahashi R, Suzuki F, Nakagawa
T, Kimotro K. Effects of voluntary running exercise on
blood pressure and renin-angiotensin system in
spontaneously hypertensive rats and normotensive Wistar-
Kyoto rats. J Nutr Sci Vitaminol (Tokyo) 2000; 46: 165-170.
24. Melo RM, Martinho E, Jr, Michelini LC. Training-induced,
pressure-lowering effect in SHR: wide effects on circulatory
profile of exercised and nonexercised muscles. Hypertension
2003; 42: 851-857.
25. Hagg U, Andersson I, Naylor AS, et al. Voluntary physical
exercise-induced vascular effects in spontaneously
hypertensive rats. Clin Sci (Lond) 2004; 107: 571-581.
53
26. Dorffel Y, Latsch C, Stuhlmuller B, et al. Preactivated
peripheral blood monocytes in patients with essential
hypertension. Hypertension 1999; 34: 113-117.
27. Peeters AC, Netea MG, Janssen MC, Kullberg BJ, Van der
Meer JW, Thien T. Pro-inflammatory cytokines in patients
with essential hypertension. Eur J Clin Invest 2001; 31:
31-36.
28. Navar LG, Zou L, Von Thun A, Tarng Wang C, Imig JD,
Mitchell KD. Unraveling the mystery of goldblatt
hypertension. News Physiol Sci 1998; 13:170-176.
29. Cakir B, Kasimay O, Kolgazi M, Ersoy Y, Ercan F, Yegen
BC. Stress-induced multiple organ damage in rats is
ameliorated by the antioxidant and anxiolytic effects of
regular exercise. Cell Biochem Funct 2010; 28: 469-479.
30. Schiller NB, Shah PM, Crawford M, et al. Recommendations
for quantitation of the left ventricle by two-dimensional
echocardiography. American Society of Echocardiography
Committee on Standards, Subcommittee on Quantitation of
Two-Dimensional Echocardiograms. J Am Soc Echocardiogr
1989; 2: 358-367.
31. Bradley PP, Priebat DA, Christensen RD, Rothstein G.
Measurement of cutaneous inflammation: estimation of
neutrophil content with an enzyme marker. J Invest
Dermatol 1982; 78: 206-209.
32. Casini AF, Ferrali M, Pompella A, Maellaro E, Comporti M.
Lipid peroxidation and cellular damage in extrahepatic
tissues of bromobenzene-intoxicated mice. Am J Pathol
1986; 123: 520-531.
33. Aykac G, Uysal M, Yalcin AS, Kocak-Toker N, Sivas A,
Oz H. The effect of chronic ethanol ingestion on hepatic
lipid peroxide, glutathione, glutathione peroxidase and
glutathione transferase in rats. Toxicology 1985; 36: 
71-76.
34. Aebi H. Catalase in vitro. Methods Enzymol 1984; 105: 121-
126.
35. Frohlich ED. State of the Art lecture. Risk mechanisms in
hypertensive heart disease. Hypertension 1999; 34: 782-9.
36. Frohlich ED. Ischemia and fibrosis: the risk mechanisms of
hypertensive heart disease. Braz J Med Biol Res 2000; 33:
693-700.
37. Higashi Y, Sasaki S, Nakagawa K, Matsuura H, Oshima T,
Chayama K. Endothelial function and oxidative stress in
renovascular hypertension. N Engl J Med 2002; 346: 1954-
1962.
38. Hamilton CA, Brosnan MJ, McIntyre M, Graham D,
Dominiczak AF. Superoxide excess in hypertension and
aging: a common cause of endothelial dysfunction.
Hypertension 2001; 37: 529-534.
39. Hilgers RH, Xing D, Gong K, Chen YF, Chatham JC, Oparil
S. Acute O-GlcNAcylation prevents inflammation-induced
vascular dysfunction. Am J Physiol Heart Circ Physiol 2012;
303: H513-H522.
40. Kerr S, Brosnan MJ, McIntyre M, et al. Superoxide anion
production is increased in a model of genetic hypertension:
role of the endothelium. Hypertension 1999; 33:1353-8.
41. Patzak A, Kleinmann F, Lai EY, Kupsch E, Skelweit A,
Mrowka R. Nitric oxide counteracts angiotensin II induced
contraction in efferent arterioles in mice. Acta Physiol Scand
2004; 181: 439-444.
42. Taddei S, Salvetti A. Endothelial dysfunction in essential
hypertension: clinical implications. J Hypertens 2002; 20:
1671-1674.
43. Albrecht EW, Stegeman CA, Heeringa P, Henning RH, van
Goor H. Protective role of endothelial nitric oxide synthase.
J Pathol 2003; 199: 8-17.
44. Huang PL. Endothelial nitric oxide synthase and endothelial
dysfunction. Curr Hypertens Rep 2003; 5: 473-480.
45. Lambeth JD. NOX enzymes and the biology of reactive
oxygen. Nat Rev Immunol 2004; 4: 181-189.
46. Martinez-Maldonado M. Pathophysiology of renovascular
hypertension. Hypertension 1991; 17: 707-719.
47. Ruiz-Ortega M, Ruperez M, Lorenzo O, et al. Angiotensin
II regulates the synthesis of proinflammatory cytokines
and chemokines in the kidney. Kidney Int Suppl 2002; 82:
S12-S22.
48. Minuz P, Patrignani P, Gaino S, et al. Increased oxidative
stress and platelet activation in patients with hypertension
and renovascular disease. Circulation 2002; 106: 2800-2805.49. Pauletto P, Rattazzi M. Inflammation and hypertension: the
search for a link. Nephrol Dial Transplant 2006; 21: 850-853.
50. Cohuet G, Struijker-Boudier H. Mechanisms of target organ
damage caused by hypertension: therapeutic potential.
Pharmacol Ther 2006; 111: 81-98.
51. Ceriello A. Possible role of oxidative stress in the
pathogenesis of hypertension. Diabetes Care 2008; 31
(Suppl. 2): S181-S184.
52. Sekiguchi F, Yanamoto A, Sunano S. Superoxide dismutase
reduces the impairment of endothelium-dependent relaxation
in the spontaneously hypertensive rat aorta. J Smooth Muscle
Res 2004; 40: 65-74.
53. Chen HH, Chiang IP, Jen CJ. Exercise training increases
acetylcholine-stimulated endothelium-derived nitric oxide
release in spontaneously hypertensive rats. J Biomed Sci
1996; 3: 454-460.
54. Cheng L, Yang C, Hsu L, Lin MT, Jen CJ, Chen H. Acute
exercise enhances receptor-mediated endothelium-
dependent vasodilation by receptor upregulation. J Biomed
Sci 1999; 6: 22-27.
55. Fagard RH. Exercise characteristics and the blood pressure
response to dynamic physical training. Med Sci Sports Exerc
2001; 33(Suppl. 6): S484-S492.
56. Graham DA, Rush JW. Exercise training improves aortic
endothelium-dependent vasorelaxation and determinants of
nitric oxide bioavailability in spontaneously hypertensive
rats. J Appl Physiol (1985) 2004; 96: 2088-2096.
57. Maiorana A, O’Driscoll G, Taylor R, Green D. Exercise and
the nitric oxide vasodilator system. Sports Med 2003; 33:
1013-1035.
58. Pescatello LS, Franklin BA, Fagard R, et al. American
College of Sports Medicine position stand. Exercise and
hypertension. Med Sci Sports Exerc 2004; 36: 533-553.
59. Yen MH, Yang JH, Sheu JR, Lee YM, Ding YA. Chronic
exercise enhances endothelium-mediated dilation in
spontaneously hypertensive rats. Life Sci 1995; 57: 2205-2213.
60. Locatelli J, Monteiro de Assis LV, Morais Araujo C, et al.
Swimming training promotes cardiac remodeling and alters
the expression of mRNA and protein levels involved in
calcium handling in hypertensive rats. Life Sci 2014; 117:
67-74.
61. Agarwal D, Elks CM, Reed SD, Mariappan N, Majid DS,
Francis J. Chronic exercise preserves renal structure and
hemodynamics in spontaneously hypertensive rats. Antioxid
Redox Signal 2012; 16: 139-152.
62. Finkel T, Holbrook NJ. Oxidants, oxidative stress and the
biology of ageing. Nature 2000; 408: 239-247.
63. McArdle A, Jackson MJ. Exercise, oxidative stress and
ageing. J Anat 2000; 197: 539-541.
64. McArdle A, Pattwell D, Vasilaki A, Griffiths RD, Jackson
MJ. Contractile activity-induced oxidative stress: cellular
origin and adaptive responses. Am J Physiol Cell Physiol
2001; 280: C621-C627.
65. Ji LL. Oxidative stress during exercise: implication of
antioxidant nutrients. Free Radic Biol Med 1995; 18:
1079-1086.
54
66. Sen CK. Oxidants and antioxidants in exercise. J Appl
Physiol (1985) 1995; 79: 675-686.
67. Moraes-Teixeira JA, Felix A, Fernandes-Santos C, Moura
AS, Mandarim-de-Lacerda CA, de Carvalho JJ. Exercise
training enhances elastin, fibrillin and nitric oxide in the
aorta wall of spontaneously hypertensive rats. Exp Mol
Pathol 2010; 89: 351-357.
68. Bahner U, Geiger H, Palkovits M, Ganten D, Klotz B,
Heidland A. Changes in the central ANF-system of
renovascular hypertensive rats. Kidney Int 1991; 39: 33-38.
69. Wrzosek M, Sokal M, Sawicka A, et al. Impact of obesity
and nitric oxide synthase gene G894T polymorphism on
essential hypertension. J Physiol Pharmacol 2015; 66:
681-689.
70. Goligorsky MS, Brodsky SV, Noiri E. Nitric oxide in acute
renal failure: NOS versus NOS. Kidney Int 2002; 61: 
855-861.
71. Bertagnolli M, Schenkel PC, Campos C, et al. Exercise
training reduces sympathetic modulation on cardiovascular
system and cardiac oxidative stress in spontaneously
hypertensive rats. Am J Hypertens 2008; 21: 1188-1193.
72. Fernandes T, Nakamuta JS, Magalhaes FC, et al. Exercise
training restores the endothelial progenitor cells number and
function in hypertension: implications for angiogenesis. 
J Hypertens 2012; 30: 2133-2143.
73. Husain K, Hazelrigg SR. Oxidative injury due to chronic
nitric oxide synthase inhibition in rat: effect of regular
exercise on the heart. Biochim Biophys Acta 2002; 1587:
75-82.
74. Kimura H, Kon N, Furukawa S, et al. Effect of endurance
exercise training on oxidative stress in spontaneously
hypertensive rats (SHR) after emergence of hypertension.
Clin Exp Hypertens 2010; 32: 407-415.
75. Roque FR, Hernanz R, Salaices M, Briones AM. Exercise
training and cardiometabolic diseases: focus on the vascular
system. Curr Hypertens Rep 2013; 15: 204-214.
76. Mason RP, Corbalan JJ, Jacob RF, Dawoud H, Malinski T.
Atorvastatin enhanced nitric oxide release and reduced
blood pressure, nitroxidative stress and rantes levels in
hypertensive rats with diabetes. J Physiol Pharmacol 2015;
66: 65-72.
77. Skurk T, van Harmelen V, Hauner H. Angiotensin II
stimulates the release of interleukin-6 and interleukin-8 from
cultured human adipocytes by activation of NF-kappaB.
Arterioscler Thromb Vasc Biol 2004; 24: 1199-1203.
78. Cai H, Harrison DG. Endothelial dysfunction in
cardiovascular diseases: the role of oxidant stress. Circ Res
2000; 87: 840-844.
79. Munoz-Canoves P, Scheele C, Pedersen BK, Serrano AL.
Interleukin-6 myokine signaling in skeletal muscle: a
double-edged sword? FEBS J 2013; 280: 4131-4148.
80. Toblli JE, DiGennaro F, Giani JF, Dominici FP. Nebivolol:
impact on cardiac and endothelial function and clinical
utility. Vasc Health Risk Manag 2012; 8: 151-160.
81. Versari D, Daghini E, Virdis A, Ghiadoni L, Taddei S.
Endothelium-dependent contractions and endothelial
dysfunction in human hypertension. Br J Pharmacol 2009;
157: 527-536.
82. Agarwal R, Campbell RC, Warnock DG. Oxidative stress in
hypertension and chronic kidney disease: role of angiotensin
II. Semin Nephrol 2004; 24: 101-114.
83. Noiri E, Peresleni T, Miller F, Goligorsky MS. In vivo
targeting of inducible NO synthase with
oligodeoxynucleotides protects rat kidney against ischemia.
J Clin Invest 1996; 97: 2377-2383.
84. Ruiz-Ortega M, Lorenzo O, Suzuki Y, Ruperez M, Egido J.
Proinflammatory actions of angiotensins. Curr Opin
Nephrol Hypertens 2001; 10: 321-329.
85. Linke A, Adams V, Schulze PC, et al. Antioxidative effects
of exercise training in patients with chronic heart failure:
increase in radical scavenger enzyme activity in skeletal
muscle. Circulation 2005; 111: 1763-1770.
86. Petersen AM, Pedersen BK. The anti-inflammatory effect of
exercise. J Appl Physiol (1985) 2005; 98: 1154-1162.
87. Pinho RA, Andrades ME, Oliveira MR, et al. Imbalance in
SOD/CAT activities in rat skeletal muscles submitted to
treadmill training exercise. Cell Biol Int 2006; 30: 848-853.
88. Radak Z, Kaneko T, Tahara S, et al. The effect of exercise
training on oxidative damage of lipids, proteins, and DNA in
rat skeletal muscle: evidence for beneficial outcomes. Free
Radic Biol Med 1999; 27: 69-74.
89. Radak Z, Sasvari M, Nyakas C, Pucsok J, Nakamoto H,
Goto S. Exercise preconditioning against hydrogen
peroxide-induced oxidative damage in proteins of rat
myocardium. Arch Biochem Biophys 2000; 376: 248-251.
90. Starnes JW, Barnes BD, Olsen ME. Exercise training
decreases rat heart mitochondria free radical generation but
does not prevent Ca2+-induced dysfunction. J Appl Physiol
(1985) 2007; 102: 1793-1798.
91. Walsh NP, Gleeson M, Shephard RJ, et al. Position
statement. Part one: Immune function and exercise. Exerc
Immunol Rev 2011; 17: 6-63.
92. Rossi NF, Chen H, Maliszewska-Scislo M. Paraventricular
nucleus control of blood pressure in two-kidney, one-clip
rats: effects of exercise training and resting blood pressure.
Am J Physiol Regul Integr Comp Physiol 2013; 305: R1390-
R1400.
93. Savoia C, Schiffrin EL. Vascular inflammation in
hypertensionand diabetes: molecular mechanisms and
therapeutic interventions. Clin Sci (Lond) 2007; 112: 
375-384.
94. Fagard RH, Bjornstad HH, Borjesson M, et al. ESC Study
Group of Sports Cardiology recommendations for
participation in leisure-time physical activities and
competitive sports for patients with hypertension. Eur J
Cardiovasc Prev Rehabil 2005; 12: 326-331.
95. O’Connor FG, Meyering CD, Patel R, Oriscello RP, Joint
National Committee on the Prevention, Detection,
Evaluation and Treatment of High Blood Pressure.
Hypertension, athletes, and the sports physician:
implications of JNC VII, the Fourth Report, and the 36th
Bethesda Conference Guidelines. Curr Sports Med Rep
2007; 6: 80-84.
R e c e i v e d : July 15, 2015
A c c e p t e d : January 19, 2016
Author’s address: Prof. Berrak C. Yegen, Marmara
University, School of Medicine, Department of Physiology,
Basibuyuk Mah. Maltepe Basibuyuk Yolu No. 9/1 , 34854
Maltepe Istanbul, Turkey
E-mail: byegen@marmara.edu.tr
55