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Pomegranate peel extract attenuates oxidative stress by decreasing coronary angiotensin-converting enzyme (ACE) activity in hypertensive female rats



Based on the antioxidant properties of pomegranate, this study was designed to investigate the effects of pomegranate peel extract on damage associated with hypertension and aging in a spontaneously hypertensive rat (SHR) model. The influence of pomegranate consumption was examined on systolic blood pressure (SBP), angiotensin-converting enzyme (ACE) coronary activity, oxidative stress, and vascular morphology. Four- or 28-wk-old SHR model rats were treated for 30 d, with terminal experimental animal age being 8 and 32 wk, respectively, with either pomegranate extract (SHR-PG) or filtered water (SHR). Data showed significant reduction in SBP and coronary ACE activity in both age groups. The levels of superoxide anion, a measure of oxidative stress, were significantly lower in animals in the SHR-PG group compared to SHR alone. Coronary morphology demonstrated total increases in vascular wall areas were in the SHR group, and pomegranate peel extract diminished this effect. Pomegranate peel extract consumption conferred protection against hypertension in the SHR model. This finding was demonstrated by marked reduction in coronary ACE activity, oxidative stress, and vascular remodelling. In addition, treatment was able to reduce SBP in both groups. Evidence indicates that the use of pomegranate peel extract may prove beneficial in alleviating coronary heart disease.
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Pomegranate peel extract attenuates oxidative
stress by decreasing coronary angiotensin-
converting enzyme (ACE) activity in hypertensive
female rats
Roger L. dos Santos, Lais O. Dellacqua, Nathalie T. B. Delgado, Wender N.
Rouver, Priscila L. Podratz, Leandro C. F. Lima, Mariela P. C. Piccin, Silvana S.
Meyrelles, Helder Mauad, Jones B. Graceli & Margareth R. Moyses
To cite this article: Roger L. dos Santos, Lais O. Dellacqua, Nathalie T. B. Delgado, Wender
N. Rouver, Priscila L. Podratz, Leandro C. F. Lima, Mariela P. C. Piccin, Silvana S. Meyrelles,
Helder Mauad, Jones B. Graceli & Margareth R. Moyses (2016): Pomegranate peel extract
attenuates oxidative stress by decreasing coronary angiotensin-converting enzyme (ACE)
activity in hypertensive female rats, Journal of Toxicology and Environmental Health, Part A,
DOI: 10.1080/15287394.2016.1213690
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Pomegranate peel extract attenuates oxidative stress by decreasing coronary
angiotensin-converting enzyme (ACE) activity in hypertensive female rats
Roger L. dos Santos
, Lais O. Dellacqua
, Nathalie T. B. Delgado
, Wender N. Rouver
, Priscila L. Podratz
Leandro C. F. Lima
, Mariela P. C. Piccin
, Silvana S. Meyrelles
, Helder Mauad
, Jones B. Graceli
and Margareth R. Moyses
Department of Physiological Sciences, Federal University of Espirito Santo, Vitoria, Espirito Santo, Brazil;
Department of Physiology and
Biophysics, Federal University of Minas Gerais, Belo Horizonte, Minas Gerais, Brazil;
Department of Biophysics, Federal University of Rio de
Janeiro, Rio de Janeiro, Brazil;
Department of Morphology, Federal University of Espirito Santo, Vitoria, Espirito Santo, Brazil
Based on the antioxidant properties of pomegranate, this study was designed to investigate the
effects of pomegranate peel extract on damage associated with hypertension and aging in a
spontaneously hypertensive rat (SHR) model. The influence of pomegranate consumption was
examined on systolic blood pressure (SBP), angiotensin-converting enzyme (ACE) coronary activ-
ity, oxidative stress, and vascular morphology. Four- or 28-wk-old SHR model rats were treated for
30 d, with terminal experimental animal age being 8 and 32 wk, respectively, with either
pomegranate extract (SHR-PG) or filtered water (SHR). Data showed significant reduction in SBP
and coronary ACE activity in both age groups. The levels of superoxide anion, a measure of
oxidative stress, were significantly lower in animals in the SHR-PG group compared to SHR alone.
Coronary morphology demonstrated total increases in vascular wall areas were in the SHR group,
and pomegranate peel extract diminished this effect. Pomegranate peel extract consumption
conferred protection against hypertension in the SHR model. This finding was demonstrated by
marked reduction in coronary ACE activity, oxidative stress, and vascular remodelling. In addition,
treatment was able to reduce SBP in both groups. Evidence indicates that the use of pomegranate
peel extract may prove beneficial in alleviating coronary heart disease.
Received 27 November 2015
Accepted 30 May 2016
Systemic arterial hypertension is considered a global
public health problem with 9.4 million deaths attrib-
uted to it annually (Lima et al., 2012). As this condi-
tion is a major cardiovascular manifestation (Kearney
et al.. 2005), prevention and treatment of systemic
arterial hypertension should receive high priority.
Age is a significant risk factor in development of
cardiovascular disease; however, striking gender dif-
ferences also exist in the chronological development
of heart disease (Rosamond et al., 2007).
In human and experimental hypertension mod-
els, such as spontaneously hypertensive rats (SHR),
endothelium-dependent relaxation may be attenu-
ated, and the resulting endothelial dysfunction
contributes to increased peripheral resistance.
Endothelial dysfunction has been linked to
decreases in nitric oxide (NO) bioavailability,
reflecting impaired generation of NO and/or
enhanced inactivation of NO by free radicals
(Púzserová et al., 2010).
Free radicals are any species capable of inde-
pendent existence with at least one unpaired elec-
tron, such as superoxide anions (O
) (Forman
et al., 2008). The relationship between free radi-
cals and hypertension was first suggested in the
1960s (Romanowskia et al., 1960), but only in the
1990s was this association investigated exten-
sively. The administration of heparin-bound
superoxide dismutase (SOD) to SHR animals
was associated with a reduction in blood pressure
(Nakazono et al., 1991). This decrease in systolic
blood pressure (SBP) following administration of
SOD may be due to a fall in reactive oxygen
species (ROS), which may contribute to elevation
in blood pressure, either directly, related to vaso-
constrictor effects, or indirectly, by reducing the
CONTACT Prof. Roger Lyrio dos Santos, PhD Department of Physiological Sciences, Biomedical Center, Federal University of
Espirito Santo, Marechal Campos Avenue, 1468, 29050-755, Vitoria, ES, Brazil.
Color versions of one or more of the figures in the article can be found online at
© 2016 Taylor & Francis
activity of vasodilators such as NO (Reckelhoff
and Romero, 2003).
Increased sympathetic nervous system activity
(Prodel et al., 2015), upregulation of the renin
angiotensinaldosterone system (Probstfield and
OBrien, 2010), and enhanced oxidative stress
(Hamilton et al., 2001) are important factors that
are modified in hypertension. These factors may
lead to changes in the vessel structure. Angiotensin
II was suggested to contribute to vascular hyper-
trophy and hypertension via stimulation of the
NADPH oxidase system to subsequently elevate
ROS levels in vascular cells (Nickenig and
Harrison, 2002).
Several epidemiological studies suggested that
regular consumption of foods and beverages that
are rich in polyphenols, such as red wine, berries,
cocoa, tea, soy, and pomegranate, is associated
with a reduction in risk of a range of pathological
conditions including hypertension, coronary heart
disease, stroke, and dementia (Ghosh and
Scheepens, 2009). Studies in rats demonstrated
that pomegranates (Mohan et al., 2010) are rich
in polyphenolic antioxidants, which include tan-
nins, anthocyanin, and flavonoids (Jurenka, 2008).
All parts of the fruit seem to possess considerable
amounts of polyphenols, but the husk appears to
contain the highest concentration of these antiox-
idants (Gil et al., 2000).
Antioxidants are well known to enhance the
biological actions of NO by protecting against
oxidative destruction mediated by ROS (Gil et al.,
2000). Pomegranate is a rich source of antioxi-
dants; however, little is known regarding the
action of pomegranate on the coronary vascular
bed. Therefore, the objective of this study was to
examine the potential of pomegranate peel extract
in protecting against damage mediated by hyper-
tension on angiotensin-converting enzyme (ACE)
activity, oxidative stress, and vascular remodelling.
Materials and Methods
Spontaneously hypertensive (SHR) Wistar female
rats (4 and 28 wk old) were randomly divided into
two groups: SHR and spontaneously hypertensive
pomegranate extract (SHR-PG). Animals were
obtained from the animal facilities at the Federal
University of Espirito Santo. Pomegranate extract
was dissolved in filtered water and administered
orally for 30 d by gavage. The control group
received filtered water. At the end of the treat-
ment, the animals were 8 and 32 wk old, respec-
tively. Rats were maintained in temperature-
controlled rooms (22°C) under a 12-h light/dark
cycle. All procedures were conducted in accor-
dance with the institutional guidelines for animal
research, and protocols were previously approved
by the Institutional Ethics Committee for Use of
Animals (CEUA 107/2011).
Noninvasive Arterial Blood Pressure Assessment
Noninvasive measurement of tail-cuff pressure as
an estimate of systolic arterial pressure was carried
out 1 d before treatment started and on the last
day (d 30) of administration. Rats were warmed in
a restraining chamber, and occluding cuffs and
pneumatic pulse transducers were placed on their
tails. A sphygmomanometer was inflated and
deflated automatically, and tail-cuff signals from
the transducer were automatically recorded using
an IITC apparatus (IITC, Inc., Woodland Hills,
CA) connected to a computer. For each blood
pressure measurement session, the mean of three
arterial blood pressure readings was recorded for
each rat.
Plant Material
The plant material of choice (Punica granatum L.)
popularly known as pomegranate, belonging to the
family Lythraceae, was collected in the city of
Vitoria, state of Espirito Santo, Brazil. Plant sam-
ples were authenticated by Dra. Valquíria Ferreira
Dutra at the Department of Biological Sciences,
Federal University of Espirito Santo, where a sam-
ple (voucher specimen number 37631) was depos-
ited in the herbarium of the VIES/UFES in botany
Preparation of Pomegranate Peel Extract
The peel of Punica granatum L. was removed and
dried in shade for 10 d before grinding. Extract
was prepared according to Lapornik et al. (2005)
with modification. Briefly, pomegranate was col-
lected; peel was removed, dried for 5 d, and then
ground. The ground material (85.71 g) was mixed
in 1000 ml ethanol (95°GL) in an amber bottle
until the complete extraction of peel compounds.
Subsequently, the sample was vacuum filtered,
supernatant was collected, and alcohol was evapo-
rated in a rotary evaporator at 60°C. The resultant
crude extract (hydroalcoholic extract) (68%, w/w)
was kept at 4°C in an amber bottle. Because the
hydroalcoholic extract undergoes a certain degree
of hydration, a dry weight determination was
made. The hydroalcoholic extract was diluted in
filtered water and administered for 30 d orally by
gavage at a concentration of 25 mg/100 g rat.
Determining Estrous Cycle Phase
Daily vaginal smears were taken from each female
rat as previously described (Marcondes et al.,
2002) to confirm that estrous cycles were proceed-
ing normally. The vaginal epithelial cells were
examined by microscopy for at least 7 consecutive
days before the experiment. The swabs were per-
formed between 8:00 and 10:00 a.m. to maintain
consistency. The females exhibiting normal estrous
cycles were killed at proestrus between 9:00 a.m.
and 1:00 p.m.
Isolation of Coronary Arteries
At the end of treatment, animals were anesthetized
with sodium thiopental (50 mg/kg, ip) and eutha-
nized via decapitation. The thorax cavity was
opened, and the heart was removed and placed
in a buffer solution of Tris-HCl, pH 7, with 50
mMNaCl (Carmona et al., 2006). The left anterior
descending branch and septal branch coronary
were isolated using a dissection microscope (D.F.
Vasconcelos M900, São Paulo, Brazil).
Subsequently, samples were stored at 80°C until
protein quantification (Furieri et al., 2011).
Measurement of Angiotensin-Converting Enzyme
(ACE) Coronary Activity
Angiotensin-converting enzyme (ACE) coronary
activity was determined using the fluorescence
resonance energy transfer (FRET) peptide Abz-
FRK(Dnp)P-OH (Aminotech Pesquisa and
Desenvolvimento, SP, Brazil) as a substrate
(Alves et al., 2005). Coronary samples were homo-
genized in 0.1 MTris-HCl buffer, pH 7, containing
50 mMNaCl and centrifuged at 1000 × g for 10
min. The hydrolysis rate of the Abz-FRK(Dnp)P-
OH substrate (10 μM) after incubation for 30 min
at 37°C in coronary homogenate aliquots was
assessed to obtain ACE enzymatic activity. The
assay methodology was adapted for a 96-well
plate reader. Fluorescence was measured at 320
nm excitation and 420 nm emission wavelengths
(Synergy2 BiotekR USA). Assays were performed
in triplicate, and results were averaged. Coronary
ACE activity was expressed in arbitrary fluores-
cence units (AFU/μg protein). The protein content
was determined by the Lowry et al. method (1951)
Histological Analysis of Anterior Septal Coronary
At the end of treatments, each rat was anesthetized
with sodium thiopental (50 mg/kg, intraperitoneal
injection) and the heart was removed. For each
animal, the medial portion from the septal coron-
ary artery was used. Subsequently, 10 sections
every 100 µm were obtained. The morphometric
analyses of total vascular and wall areas corre-
sponded to average values obtained from 10 cross
sections. Subsequently, tissues were embedded in
optical cutting temperature (OCT) compound and
cross-sectioned on a cryostat (Jung CM1850; Leica,
Wetzlar, Germany) at a thickness of 8 μm. For
each animal, the coronary cross sections were
mounted on gelatin-coated slides and stained
with hematoxylineosin for morphometric analy-
sis as shown previously (Borgo et al., 2016).
Images of the coronary arteries were captured
using a 20× objective with a color video camera
(VKC150, Hitachi, Tokyo, Japan) connected to a
microscope (Olympus AX70, Olympus, Center
Valley, PA) and analyzed employing a National
Institutes of Health (NIH) imaging program. An
examiner blind to the experimental groups per-
formed the image analysis to prevent any bias in
the interpretation of the results.
Detection of Superoxide Production
Dihydroethidium (DHE) staining was used to
determine ROS generation. Unfixed frozen coron-
ary sections (8 μm) after dehydration with a 30%
sucrose solution were incubated with 2 μmol DHE
(Molecular Probes, Sigma, D7008) in modified
Krebs solution containing 20 mmol HEPES for
30 min in a light-protected chamber at 37ºC.
Subsequently, the samples were subjected to three
washes with phosphate-buffered saline (PBS), air
dried (light-protected), and mounted with neutral
glycerine. The levels of ROS were determined
using microscopy, and coronary fluorescence was
quantified with microscope software (NIS-
Elements BR 7.0, Nikon Instruments, Inc.,
Champigny-sur-Marne, France). An enhanced
red fluorescence suggested elevated levels of super-
oxide anion. The fluorescence intensity of the cor-
onaries was quantified in 10 arbitrarily selected
coronaries, and the mean value for each islet was
Statistical Analysis
All data are expressed as the mean ± SEM. To
identify possible outlier data, a two-sided Grubbs
test was used to identify whether at least one out-
lier was present in each dataset. When Grubbstest
identified one outlier, an adapted ROUT method
was used to detect any outliers from that column
data and remove them according to the Q setting
at 1% (alpha = .01). For each data set, the
DAgostinoPearson omnibus normality test was
also performed. If data passed the normality test,
then one-way analysis of variance (ANOVA)
followed by Tukeys post hoc test for multiple
comparisons was applied. The significance was
set at p< .05.
Figure 1 shows the systolic blood pressure (SBP)
on d 30 of treatment with water or pomegranate
peel extract in the 8-wk-old animals (A) and 32-
wk-old animals (B). A significant decrease was
noted in SBP in the SHR-PG group compared
with the SHR alone after 30 d in both 8-wk-old
and 32-wk-old animals. Data in Figure 2 demon-
strate the effect of treatment with pomegranate
peel extract on coronary ACE activity. The activity
of ACE was significantly reduced in SHR-PG com-
pared with SHR in both age groups.
Figure 3 summarizes the data obtained from
coronary morphology analyses of 8-wk-old (left
panels) and 32-wk-old animals (right panels). An
increase in total vascular (1.86 ± 0.19 vs. 1.13 ± 0.1
,Figure 3D) and wall areas (1.06 ± 0.11 vs.
0.56 ± 0.04 µm
,Figure 3E) was noted in SHR with
8-wk-old animals compared to age-matched con-
trols. Similarly, an elevation in total vascular (3.75
± 0.16 vs. 2.27 ± 0.16 µm
,Figure 3J) and wall
areas (1.82 ± 0.13 vs. 0.78 ± 0.06 µm
,Figure 3K)
was found in SHR at 32 wk compared to age-
matched controls. However, treatment of SHR
with pomegranate peel extract resulted in a return
to control values in total vascular area in SHR at
both 8 wk (1.86 ± 0.19 to 0.89 ± 0.07, Figure 3D)
and 32 wk (3.75 ± 0.16 to 2.25 ± 0.29 µm
Figure 3J) respectively. Further, treatment of SHR
with pomegranate peel extract also produced a
return to control levels in wall areas in SHR at 8
Figure 1. Effect of pomegranate extract on systolic blood pressure (SBP) on d 0 and on d 30 in 8-wk-old (A, n= 18) and 32-wk-old
animals (B, n=1618). Values are expressed as means ± SEM. Asterisk indicates significant at p< .05 compared with SHR alone
(unpaired Studentst-test).
wk (1.06 ± 0.11 to 0.64 ± 0.03 µm
,Figure 3E) and
32 wk (1.82 ± 0.13 to 1.13 ± 0.09 µm
,Figure 3K).
An increase in wall/lumen ratios was found in
SHR at 8 wk (1.38 ± 0.10 vs. 0.89 ± 0.07 µm
Figure 3F) and 32 wk (1.28 ± 0.05 vs. 0.57 ± 0.05
,Figure 3L) compared to age-matched con-
trols. The treatment of SHR with pomegranate
peel extract were also able to lower these values
in SHR at 8 wk (1.38 ± 0.1 to 0.97 ± 0.09 µm
Figure 3F) and 32 wk (1.28 ± 0.05 to 0.8 ± 0.13
,Figure 3L).
The antioxidant potential effect of the pomegra-
nate peel extract treatment was observed in the cor-
onary arteries using DHE staining, as illustrated in
the microphotographs in Figure 4.ADHEoxidative
assay revealed intense fluorescence in SHR animals
but none in SHR-PG. Pomegranate peel extract
administration reduced vascular oxidative stress in
coronary arteries in the SHR at ages 8 wk (11.3 ± 0.9
AU vs. 3.66 ± 0.9 AU) and 32 wk (55.7 ± 4.9 AU vs.
25.96 ± 5.0 AU) compared with respective controls.
On average, coronary arteries from SHR-PG animals
exhibited approximately 50% less ethidium fluores-
cence than SHR.
Our main finding was that treatment with hydroal-
coholic extracts of Punica granatum peel was able to
reduce oxidative stress and coronary ACE activity in
SHR rats. Further, the treatments lowered SBP and
prevented vascular remodeling in coronary arteries
in this hypertension model. In the present study, 30
d of treatment with pomegranate peel extract
significantly decreased SBP in 8- and 32-wk-old ani-
mals. Among the possible causes of hypertension, an
overproduction of ROS is implicated. Oxidative
stress is defined as an imbalance between ROS levels
and antioxidant defenses, which worsens with aging
and hypertension (Wind et al., 2010; Ghio et al.,
2012). Further, oxidative stress was shown to be
intimately related with endothelial dysfunction
(Hamilton et al., 2001). Wind et al. (2010)found
that ROS production increased in the aortas of
aged SHR compared with aged Wistar-matched con-
trol aortas. This augmented ROS production may
lead to elevated SBP directly by vasoconstrictor
effects or indirectly by reduced activities of vasodila-
tors (Reckelhoff and Romero, 2003). Although the
animal model used in this study was characterized by
enhanced generation of ROS, oxidative stress
appeared to increase in conditions of hypertension
and aging. Thus, greater oxidative stress was
expected in 32-wk-old versus 8-wk-old animals.
Treatment with pomegranate peel extract in both
8-wk and 32-wk SHR may have led to lower ROS
production, which may have been sufficient to lower
SBP. Further, as ROS contribute to endothelium-
dependent contraction and increase in vascular resis-
tance, antioxidant substances found in pomegranate
extracts may be associated with the observed
decrease in SBP (Kitiyakara and Wilcox, 1998).
This finding is in agreement with other studies of
advanced aging, which demonstrated a 5% decline in
SBP with a daily consumption of pomegranate juice
for 2 wk (Aviram and Dornfeld, 2001).A decrease in
coronary ACE activity was noted in the SHR-PG
group compared with SHR alone at 8 and 32 wk.
Figure 2. Influence of pomegranate extract on coronary ACE activity on d 30. Eight-week-old (A) SHR rats and SHR-PG, and 32-wk-old
(B) SHR rats and SHR-PG. Values are expressed as means ± SEM; n= 5 animals in each group. Asterisk indicates significant at p< .05
compared with SHR alone.
Angiotensin-converting enzyme, a zinc (Zn) metal-
lopeptidase, possesses two large homologous
domains, the N and C domains. These domains are
often the targets of ACE inhibitors that act by bind-
ing to the Zn (Comini et al., 2007). Polyphenols have
chemical structures that favor chelation of redox-
active metals (Fraga, 2007) which may have favored
ACE inhibition observed in this study. In addition,
Mohan et al. (2010) demonstrated that consumption
of pomegranate juice for 4 wk in diabetic hyperten-
sive rats decreased serum ACE levels. In hyperten-
sion, angiotensin II is known to play a role in
vascular remodeling. This effect seems to be
mediated by a rise in production of free radicals
(Ushio-Fukai et al., 1996; Zalba et al., 2000). Thus,
coronary ACE inhibition produced by pomegranate
peel extract may have influenced the morphology of
the cells of the coronary arteries as noted in this
investigation. Althunibat et al. (2010) reported that
administration of Punica granatum peel methanol
extract improved antioxidant enzyme activities in
diabetic rats. Aviram et al. (2000) demonstrated
marked antioxidant capacity of pomegranate juice
to scavenge free radicals. Based on the antioxidant
properties of Punica granatum described in the lit-
erature (Gil et al., 2000) and its scavenging capacity,
it was postulated that treatment with pomegranate
peel extract might decrease oxidative stress in this
experimental model. Data demonstrated that coron-
ary arteries of SHR-PG exhibited a marked fall in
generation of O
. These results support the notion
that one of the mechanisms involved in attenuation
of damage derived from hypertension may be related
to reduced oxidative stress. Cardiovascular morpho-
physiological abnormalities have been associated
with increased oxidative stress (Lima et al., 2012).
Consequently, interventions that are able to potenti-
ate tissue antioxidant capacity, including pharmaco-
logical therapies, have been used to provide vascular
and cellular benefits (Bazargani-gilani et al., 2014;
Danesi et al., 2014). Evidence indicates that pome-
granate juice extract may also exert beneficial actions
in hypertensive individuals.
Our morphometric analyses showed that
pomegranate extract treatment prevented altera-
tions induced by hypertension and/or aging on
total vascular and coronary wall area. The
reninangiotensinaldosterone system has trophic
actions on the components of the arterial wall, and
angiotensin II might initiate hypertrophic pro-
cesses on vascular smooth muscles (Touyz et al.,
2003). Not surprisingly, ACE inhibition was found
to affect hypertrophied arterial walls in SHR ani-
mal models (Dedkov et al., 2006). A novel finding
Figure 3. Effect of pomegranate extract on morphometric
parameters of coronary arteries. Top panel, microphotographs
are typical cross sections of coronary arteries in SHR and SHR-
PG rats at 8 (left panel) and 32 (right panel) wk, respectively.
Bar = 100 μm. Bottom bar graphs show influence of pome-
granate peel extract on total vascular and wall area at 8 (left
panel) and 32 (right panel) wk old, respectively. Values are
means ± SEM, n=47 per group. Asterisk indicates significant
at p< .05, and double asterisk at p< .01 compared with
control, and
p< .05 compared with SHR alone.
of this study revealed that in SHR that received
pomegranate extract, the morphological alterations
induced by hypertension and/or aging were dimin-
ished. This finding may be attributed to a reduction
in oxidative stresses mediated by coronary ACE
inhibition and the antioxidant actions of polyphenol
compounds. These effects may be correlated with the
potent antioxidant activity of pomegranate asso-
ciated with high polyphenol content and to the spe-
cific type of polyphenols present in pomegranate,
specifically, hydrolyzable tannins, which display a
high scavenging capacity for free radicals (Aviram
and Rosenblat, 2012).
Our study corroborated the observations of Touyz
et al. (2003), which indicated that ROS might induce
morphological alterations. Indeed, ROS promoted
these alterations by (i) modifying the activity of
tyrosine kinases, metalloproteinases, and mitogen-
activated protein kinases (Baas and Berk, 1995;
Intengan and Schiffrin, 2001); (ii) acting on gene
and protein expression mechanisms by activating
transcription factors, such as nuclear factor (NF)-
κB and AP-1 (Touyz and Schiffrin, 2000); and (iii)
stimulating ion channels, such as plasma membrane
and K
channels, leading to changes in cation
concentrations (Lounsbury et al., 2000).
Data demonstrated for the first time the influ-
ence of pomegranate peel extract administration
on the vascular remodeling process of coronary
arteries for young and elderly hypertensive female
Figure 4. Influence of pomegranate peel extract treatment on superoxide anion production in coronary arteries. The top panels
show higher fluorescence intensity (red) in SHR rats when compared to SHR-PG rats using dihydroethidium (DHE) staining. The bar
graph shows the average DHE fluorescence (AU: arbitrary units) comparing all groups (n= 6). Left panel, 8-wk-old animals and right
panel, 32-wk-old animals. Values are means ± SEM. Asterisk indicates significant at p< .01 compared with controls, and
p< .01
compared with SHR-PG group. Scale bar: 100 μm.
rats. The coronary vascular remodeling was char-
acterized by increased wall and cross-sectional
vascular areas. Thus, the coronary artery remodel-
ing in SHR might reflect an adaptive response to
elevated arterial pressures to normalize increased
wall tension (Dedkov et al., 2006; Irwin et al.,
In conclusion, this study showed that treatment
with pomegranate peel extracts was able to prevent
morphological alterations in coronary arteries
induced by hypertension and/or aging, which
likely occurred through antihypertensive actions
such as antioxidant effects and decreasing coron-
ary ACE activity. Therefore, the beneficial effects
of pomegranate peel extracts in coronary arteries
may be considered in the development of better
therapies for hypertension.
The authors thank Nazare S. Bissoli for critical review and
Polyana L. M. Dalpiaz for performing the measurement of
ACE activity.
Conflicts of Interest
All the authors disclose no financial and personal relation-
ships with other people or organizations that could inappro-
priately influence their work.
This work is supported by grants from the FAPES (54687578/
2011) and CNPq (55262312011-3).
Althunibat, O. Y., Al-Mustafa, A. H., Tarawneh, K., Khleifat,
K. M., Ridzwan, B. H., and Qaralleh, H. N. 2010. Protective
role of Punica granatum L. peel extract against oxidative
damage in experimental diabetic rats. Process Biochem. 45:
Alves, M. F., Araujo, M. C., Juliano, M. A., Oliveira, E. M.,
Krieger, J. E., Casarini, D. E., Juliano, L., and Carmona, A.
K. 2005. A continuous fluorescent assay for the determina-
tion of plasma and tissue angiotensin I-converting enzyme
activity. Braz. J. Med. Biol. Res. 38: 861868.
Aviram, M., and Dornfeld, L. 2001. Pomegranate juice con-
sumption inhibits serum angiotensin converting enzyme
activity and reduces systolic blood pressure. Atherosclerosis
158: 195198.
Aviram, M., and Rosenblat, R. 2012. Pomegranate protection
against cardiovascular diseases. J. Evidence-Based
Complement. Altern. Med. 2012: 382763
Aviram, M., Dornfeld, L., Rosenblat, M., Volkova, N.,
Kaplan, M., Coleman, R., Hayek, T., Presser, D., and
Fuhrman, B. 2000. Pomegranate juice consumption
reduces oxidative stress, atherogenic modifications to
LDL, and platelet aggregation: Studies in humans and in
atherosclerotic apolipoprotein Edeficient mice. Am. J.
Clin. Nutr. 71: 10621076.
Baas, A. S., and Berk, B. C. 1995. Differential activation of
mitogen-activated protein kinases by H
and O
vascular smooth muscle cells. Circ. Res. 77: 2936.
Bazargani-gilani, B., Tajik, H., and Aliakbarlu, J. 2014.
Physicochemical and antioxidative characteristics of
Iranian pomegranate (Punica granatum L. cv. Rabbab-e-
Neyriz) juice and comparison of its antioxidative activity
with Zataria multiflora Boiss essential oil. Vet. Res. Forum
5: 313318.
Borgo, M. V., Claudio, E. R., Silva, F. B., Romero, W. G.,
Gouvea, S. A., Moysés, M. R., Santos, R. L., Almeida, S. A.,
Podratz, P. L., Graceli, J. B., and Abreu, G. R. 2016.
Hormonal therapy with estradiol and drospirenone
improves endothelium-dependent vasodilation in the cor-
onary bed of ovariectomized spontaneously hypertensive
rats. Braz. J. Med. Biol. Res. 49: e4655.
Carmona, A. K., Schwager, S. L., Juliano, M. A., Juliano, L.,
and Sturrock, E. D. 2006. A continuous fluorescence reso-
nance energy transfer angiotensin I-converting enzyme
assay. Nat. Protocols 1: 19711976.
Comini, L., Bachetti, T., Cargnoni, A., Bastianon, D., Gitti, G.
L., Ceconi, C., and Ferrari, R. 2007. Therapeutic modula-
tion of the nitric oxide: All ACE inhibitors are not equiva-
lent. Pharmacol. Res. 56: 4248.
Danesi, F., Kroon, P. A., Saha, S., de Biase, D., DAntuono, L.
F., and Bordoni, A. 2014. Mixed pro- and anti-oxidative
effects of pomegranate polyphenols in cultured cells. Int. J.
Mol. Sci. 15: 1945819471.
Dedkov, E. I., Perloff, J. K., Tomanek, R. J., Fishbein, M. C.,
and Gutterman, D. D. 2006. The coronary microcircula-
tion in cyanotic congenital heart disease. Circulation 114:
Forman, H. J., Fukuto, J. M., Miller, T., Zhang, H., Rinna, A.,
and Levy, S. 2008. The chemistry of cell signaling by
reactive oxygen and nitrogen species and 4-hydroxynone-
nal. Arch. Biochem. Biophys. 477: 183195.
Fraga, C. G. 2007. Plant polyphenols: How to translate their
in vitro antioxidant actions to in vivo conditions. IUBMB
Life 59: 308315.
Furieri, L. B., Galán, M., Avendaño, M. S., García-Redondo,
A. B., Aguado, A., Martínez, S., Cachofeiro, V., Bartolomé,
M. V., Alonso, M. J., Vassallo, D. V., and Salaices, M. 2011.
Endothelial dysfunction of rat coronary arteries after expo-
sure to low concentrations of mercury is dependent on
reactive oxygen species. Br. J. Pharmacol. 162: 18191831.
Ghio, A. J., Carraway, M. S., and Madden, M. C. 2012.
Composition of air pollution particles and oxidative stress
in cells, tissues and living systems. J. Toxicol. Environ.
Health B 15: 121.
Ghosh, D., and Scheepens, A. 2009. Vascular action of poly-
phenols. Mol. Nutr. Food Res. 53: 322331.
Gil, M.I., Tomas-Barberan, F., Hess-Pierce, B., Holcroft, D.
M., and Kader, A. A. 2000. Antioxidant activity of pome-
granate juice and its relationship with phenolic composi-
tion and processing. J. Agric. Food Chem. 48: 45814589.
Hamilton, C. A., Brosnan, M. J., McIntyre, M., Graham, D.,
and Dominiczak, A. F. 2001. Superoxide excess in hyper-
tension and aging: A common cause of endothelial dys-
function. Hypertension 37: 529534.
Intengan, H. D., and Schiffrin, E. L. 2001. Vascular remodel-
ing in hypertension. Role of apoptosis, inflammation and
fibrosis. Hypertension 38: 581587.
Irwin, D. C., Garat, C. V., Crossno, J. T., Jr., MacLean, P. S.,
Sullivan, T. M., Erickson, P. F., Jackman, M. R., Harral, J.
W., Reusch, J. E., and Klemm, D. J. 2014. Obesity-related
pulmonary arterial hypertension in rats correlates with
increased circulating inflammatory cytokines and lipids
and with oxidant damage in the arterial wall but not with
hypoxia. Pulmon. Circ. 4: 638653.
Jurenka, J. 2008. Therapeutic applications of pomegranate
(Punica granatum L.): A review. Altern. Med. Rev. 13:
Kearney, P. M., Whelton, M., Reynolds, K., Muntner, P.,
Whelton, P. K., and He, J. 2005. Global burden of hyper-
tension: Analysis of worldwide data. Lancet 365: 217223.
Kitiyakara, C., and Wilcox, C.S. 1998. Antioxidants for
hypertension. Curr. Opin. Nephrol. Hypertens. 7: 531538.
Lapornik, B., Prosek, M., and Wondra, A.G. 2005. Comparison
of extracts prepared from plant by-products using different
solvents and extraction time. J. Food Eng. 71: 214222.
Lim, S. S., Vos, T., Flaxman, A. D., Danaei, G., Shibuya, K.,
Adair-Rohani, H., Amann, M., Anderson, H. R., Andrews,
K. G., Aryee, M., Atkinson, C., Bacchus, L. J., Bahalim, A.
N., Balakrishnan, K., Balmes, J., Barker-Collo, S., Baxter, A.,
Bell, M. L., Blore, J. D., Blyth, F., Bonner, C., Borges, G.,
Bourne, R., Boussinesq, M., Brauer, M., Brooks, P., Bruce,
N. G., Brunekreef, B., Bryan-Hancock, C., Bucello, C.,
Buchbinder, R., Bull, F., Burnett, R. T., Byers, T. E.,
Calabria, B., Carapetis, J., Carnahan, E., Chafe, Z.,
Charlson, F., Chen, H., Chen, J. S., Cheng, A. T., Child, J.
C., Cohen, A., Colson, K. E., Cowie, B. C., Darby, S.,
Darling, S., Davis, A., Degenhardt, L., Dentener, F., Des
Jarlais, D. C., Devries, K., Dherani, M., Ding, E. L.,
Dorsey, E. R., Driscoll, T., Edmond, K., Ali, S. E., Engell,
R. E., Erwin, P. J., Fahimi, S., Falder, G., Farzadfar, F.,
Ferrari, A., Finucane, M. M., Flaxman, S., Fowkes, F.G.,
Freedman, G., Freeman, M. K., Gakidou, E., Ghosh, S.,
Giovannucci, E., Gmel, G., Graham, K., Grainger, R.,
Grant, B., Gunnell, D., Gutierrez, H.R., Hall, W., Hoek, H.
W., Hogan, A., Hosgood, H. D. 3rd., Hoy, D., Hu, H.,
Hubbell, B. J., Hutchings, S. J., Ibeanusi, S. E., Jacklyn, G.
L., Jasrasaria, R., Jonas, J. B., Kan, H., Kanis, J.A.,
Kassebaum, N., Kawakami, N., Khang, Y. H., Khatibzadeh,
S., Khoo, J. P., Kok, C., Laden, F., Lalloo, R., Lan, Q.,
Lathlean, T., Leasher, J. L., Leigh, J., Li, Y., Lin, J. K.,
Lipshultz, S. E., London, S., Lozano, R., Lu, Y., Mak, J.,
Malekzadeh, R., Mallinger, L., Marcenes, W., March, L.,
Marks, R., Martin, R., McGale, P., McGrath, J., Mehta, S.,
Mensah, G. A., Merriman, T. R., Micha, R., Michaud, C.,
Mishra, V., Mohd Hanafiah, K., Mokdad, A. A., Morawska,
L., Mozaffarian, D., Murphy, T., Naghavi, M., Neal, B.,
Nelson, P. K., Nolla, J.M., Norman, R., Olives, C., Omer,
S. B., Orchard, J., Osborne, R., Ostro, B., Page, A., Pandey,
K. D., Parry, C. D., Passmore, E., Patra, J., Pearce, N.,
Pelizzari, P. M., Petzold, M., Phillips, M. R., Pope, D.,
Pope, C. A. 3rd, Powles, J., Rao, M., Razavi, H., Rehfuess,
E.A., Rehm, J. T., Ritz, B., Rivara, F. P., Roberts, T.,
Robinson, C., Rodriguez-Portales, J. A., Romieu, I., Room,
R., Rosenfeld, L. C., Roy, A., Rushton, L., Salomon, J. A.,
Sampson, U., Sanchez-Riera, L., Sanman, E., Sapkota, A.,
Seedat, S., Shi, P., Shield, K., Shivakoti, R., Singh, G. M.,
Sleet, D. A., Smith, E., Smith, K. R., Stapelberg, N. J.,
Steenland, K., Stöckl, H., Stovner, L. J., Straif, K., Straney,
L., Thurston, G. D., Tran, J. H., Van Dingenen, R., van
Donkelaar, A., Veerman, J. L., Vijayakumar, L.,
Weintraub, R., Weissman, M. M., White, R. A.,
Whiteford, H., Wiersma, S. T., Wilkinson, J. D., Williams,
H. C., Williams, W., Wilson, N., Woolf, A. D., Yip, P.,
Zielinski, J. M., Lopez, A. D., Murray, C. J., Ezzati, M.,
AlMazroa, M. A., and Memish, Z. A. 2012. A comparative
risk assessment of burden of disease and injury attributable
to 67 risk factors and risk factor clusters in 21 regions,
19902010: A systematic analysis for the Global Burden of
Disease Study 2010. Lancet 380: 22242260
Lima, L. C. F., Porto, M. L., Campagnaro, B. P., Tonini, C. L.,
Nogueira, B. V, Pereira, T. M., Vasquez, E. C., and
Meyrelles, S. S. 2012. Mononuclear cell therapy reverts
cuff-induced thrombosis in apolipoprotein E-deficient
mice. Lipids Health Dis. 11: 96.
Lounsbury, K. M., Hu, Q., and Ziegelstein, R. C. 2000.
Calcium signaling and oxidant stress in the vasculature.
Free Radical Biol. Med. 28: 13621369.
Lowry, O. H., Rosebrough, N. J., Farr, A. L., and Randall, R. J.
1951. Protein measurement with the Folin phenol reagent.
J. Biol. Chem. 193: 265275.
Marcondes, F. K., Bianchi, F. J., and Tanno, A. P. 2002.
Determination of the estrous cycle phases of rats: Some
helpful considerations. Braz. J. Biol. 62: 609614.
Mohan, M., Waghulde, H., and Kasture, S. 2010. Effect of
pomegranate juice on angiotensin II-induced hypertension
in diabetic Wistar rats. Phytother. Res. 24: 196203.
Nakazono, K., Watanabe, N., Matsuno, K., Sasaki, J., Sato, T.,
and Inoue, M. 1991. Does superoxide underlie the patho-
genesis of hypertension? Proc. Natl. Acad. Sci. USA 88:
Nickenig, G., and Harrison, D. G. 2002. The AT1-type angio-
tensin receptor in oxidative stress and atherogenesis: Part
II: AT1 receptor regulation. Circulation 105: 530536.
Probstfield, J. L., and OBrien, K. D. 2010. Progression of
cardiovascular damage: The role of renin-angiotensin sys-
tem blockade. Am. J. Cardiol. 105: 1020.
Prodel, E., Barbosa, T., Machado, A., Mansur, D., Nobrega,
A. C., and Vianna L. 2015. Arterial stiffening in human
hypertension: Is there a contribution of the sympathetic
nervous system? FASEB J. 29(1 Suppl): 649613.
Púzserová, A., Kopincová, J., and Bernátová, I. 2010.
Endothelial (dys)function in the experimental model of
primary hypertension. Cesk. Fysiol. 59: 414.
Reckelhoff, J. F., and Romero, J. C. 2003. Role of oxidative
stress in angiotensin induced hypertension. Am. J. Physiol.
Reg. Integr. Comp. Physiol. 284: 893912.
Romanowskia, A., Murray, J. R., and Huston, M. J. 1960.
Effects of hydrogen peroxide on normal and hypertensive
rats. Pharm. Acta Helv. 35: 354357.
C.,Nichol,G.,ODonnell, C. J., Roger, V., Rumsfeld, J., Sorlie,
P., Steinberger, J., Thom, T., Wasserthiel-Smoller, S., and
Hong, Y. 2007. Heart disease and stroke statistics: 2007
Update: A report from the American Heart Association
Statistics Committee and Stroke Statistics Subcommittee.
Circulation 115: 69171.
Touyz, R. M., and Schiffrin, E. L. 2000. Signal transduction
mechanisms mediating the physiological and pathophysiolo-
gical actions of angiotensin II in vascular smooth muscle cells.
Pharmacol. Rev. 52: 639672.
Touyz, R. M., Tabet, F., and Schiffrin, E. L. 2003. Redox-depen-
dent signalling by angiotensin II and vascular remodelling in
hypertension. Clin. Exp. Pharmacol. Physiol. 30: 860866.
Ushio-Fukai, M., Zafari, A. M., Fukui, T., Ishizaka, N., and
Griendling, K. K. 1996. p22phox is a critical component of
the superoxide-generating NADH/NADPH oxidase system
and regulates angiotensin II-induced hypertrophy in vascu-
lar smooth muscle cells. J. Biol. Chem. 271: 2331723321.
Wind, S., Beuerlein, K., Armitage, M. E., Taye, A., Kumar, A.
H. S., Janowitz, D., Neff, C., Shah, A. M., Wingler, K., and
Schmidt, H.H. 2010. Oxidative stress and endothelial dys-
function in aortas of aged spontaneously hypertensive rats
by NOX1/2 is reversed by NADPH oxidase inhibition.
Hypertension 56: 490497.
A., Etayo, J. C., and Díez, J. 2000. Vascular NADH/NADPH
spontaneously hypertensive rats. Hypertension 35: 10551061.
... It was observed that consumption of pomegranate juice reduces the level of serum ACE and SBP by 36% and 5%, respectively, as well as providing protection against cardiovascular diseases by the inhibition of oxidative stress and serum ACE activity. An in vivo study was conducted by dos Santos et al. [43], which detected the effect of pomegranate peel on the damaging effect of hypertension and aging on spontaneously hypertensive rat (SHR) model and also examined their systolic blood pressure (SBP), angiotensin-converting enzyme (ACE) coronary activity, oxidative stress, and vascular morphology. It was concluded that in a dose dependent manner pomegranate peel significantly lowered the SBP, coronary ACE activity; oxidative stress and vascular remodeling provide beneficial effects for the treatment of coronary heart disease and hypertension. ...
... Anticancer and antimutagenic activity Flavonoids, phenols, phytates, alkaloids, glycosidic cyanide, tannins, saponins, and steroids, ellagic acid, gallic acid, punicalagin, ellagitannin,. [35,36,37,38,39] Cardioprotective and antihypertensive activity Flavonoids, anthocyanins (such as cyanidin-3glucoside, cyanidin-3,5-diglucoside and delphindin-3glucoside), catechins, ellagic tannins, gallic acid, ellagic acids, tocopherols, sterols, terpenoids, alkaloids [40,42,43] Antidiabetic activity Flavonoids, terpenoids, glycosides, saponins, alkaloids, condensed tannins, catechins, gallocatechins and prodelphinidins [44,46] Hepatoprotective activity Flavonoids, epicatechin, epigallocatechingallate, quercetin, luteolin and naringenin, phenolic acids, chlorogenic acid, caffeic acids ellagitannin, tannic acid and corilagin. [47] Nephroprotective activity Ellagitannins, ellagic acid and gallic acid [48] Antimicrobial activity Ellagitannin, punicalagin, catechin, rutin and epicatechin [50,52] Antioxidant activity Tannins, flavonoids, ellagic acid, gallic acids, protocatchoic acid, P-cumaric acid, chlorogenic acid, catechin, epicatechin, vanillic acid, Caffeic acid and ferulic acid. ...
Background Inclination of the world towards natural and super foods has intended researchers and nutritionists to develop foods that are highly nutritious, feasible, eco-friendly along with immuno-boosting potential. Massive amounts of agro-industrial waste are generated across the world which can be turned-up into a functional ingredient by utilization and transformation of these wastes into wealth. An Indian balanced diet consists of foods from all food categories, with fruits and vegetables constituting one-fourth of the diet. Since ancient times, pomegranate (Punica granatum) and its various parts have been well-known for their therapeutic properties and delicious flavor. Worldwide approximately million tons of pomegranate peels are produced per annum which are a considerable source of vitamins, minerals, and numerous bioactive compounds such as phenolics, flavonoids (anthocyanins, flavonols, flavones) and tannins. Consequently, it has many health benefits, including antioxidative, anticancer, antidiabetic, anti-inflammatory, anti-microbial, antimutagenic, cardio-protective, hepato-protective, nephro-protective properties as well as it is also helpful in the prevention of many other chronic diseases. Its safety and efficacy for consumption have been proved by incorporating it in food products and industrial applications. The main aim of this review is to focus on the vital bioactive compounds present in pomegranate peel and their positive health benefits on human health
... The metabolic consequences also comprise leptin-induced ROS production in heart muscle, leading to lipotoxicity [68]. The results from various studies indicated that Punica granatum L. peel extract reduced vascular remodeling and heart tissue damage [67,69]. Our observations did not indicate a marked amendment of the GAL-3 concentration in ethanolic-peel-treated groups, both with MetS and diet-driven obesity. ...
... The proposed molecular mechanism of heart failure is pyroptosis, marked by inflammatory-regulated cell death triggered by metabolic alternations [73]. Evidence has shown that pomegranate peel extract can reduce the hypertrophied coronary arterial walls induced via the downregulation of angiotensin-converting enzyme activity (ACE) [69]. Of note, referring to cardiac muscle, phenolic extract from peels improved myofibril composition via the reduction in separated and disorientated muscle fibers with pyknotic nuclei, inhibition in cytoplasmatic lipid deposition, and vacuolar degeneration of cardiomyocytes [63,67,74]. ...
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Citation: Niewiadomska, J.; Kasztura, M.; Janus, I.; Chełmecka, E.; Stygar, D.M.; Frydrychowski, P.; Wojdyło, A.; Noszczyk-Nowak, A. Abstract: Metabolic syndrome (MetS) significantly increases the risk of cardiovascular diseases (CVD), a leading cause of death globally. The presented study investigated the cardioprotective role of dietary polyphenols found in pomegranate peels in an animal model of metabolic syndrome. Zucker diabetic fatty rats (ZDF, MetS rats, fa/fa) were supplemented with polyphenol-rich pomegranate peel extract (EPP) at two dosages: 100 mg/kg BW and 200 mg/kg BW. The extract was administered for 8 weeks. The effect of ethanolic peel extract on the concentration of oxidative stress markers (CAT, SOD, MnSOD, GR, GST, GPx, TOS, SH, and MDA), biomarkers of heart failure (cTnI, GAL-3), and alternations in tissue architecture was assessed. The results showed a significant increase in SH concentration mediated via EPP supplementation (p < 0.001). Treatment with a 100 mg/kg BW dosage reduced the TOS level more efficiently than the higher dose. Interestingly, the CAT and GST activities were relevantly higher in the MetS 100 group (p < 0.001) compared to the MetS control. The rats administered EPP at a dose of 200 mg/kg BW did not follow a similar trend. No differences in the GR (p = 0.063), SOD (p = 0.455), MnSOD (p = 0.155), and MDA (p = 0.790) concentration were observed after exposure to the pomegranate peel extract. The administration of EPP did not influence the cTnI and GAL-3 levels. Histology analysis of the heart and aorta sections revealed no toxic changes in phenolic-treated rats. The findings of this study prove that the extract from pomegranate peels possesses free radical scavenging properties in the myocardium. The effect on alleviating ventricular remodeling and cardiomyocyte necrosis was not confirmed and requires further investigation.
... However, studies investigating the health benefits of pomegranate peel showed that polyphenols in this material possess even higher antioxidant activity than the pulp itself (14,24). Its nutritional value has been explored in many in vivo studies on animal models, and these proved that pomegranate peel extract reduces obesity (38), dyslipidaemia (34), hypertension (11), and hyperinsulinaemia and insulin resistance (49), and also ameliorates chronic inflammatory status (30). In the presented study, we aimed to evaluate the health benefits of pomegranate peel extract supplementation in a model of MetS and healthy control rats maintained on a high-calorie diet. ...
... Cardiovascular protection is also provided with polyphenols from pomegranate peel. A study in a spontaneously hypertensive rat model found the consumption of EPP to reduce systolic blood pressure, coronary angiotensinconverting enzyme activity and oxidative stress level, and prevent vascular remodelling (11). Evidence suggests that phenolics from peel may find application in mitigating coronary heart disease by attenuation of electrocardiographic changes, myocardial tissue damage and heart weight increase. ...
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Introduction Metabolic syndrome (MetS) is a cluster of pathological conditions well described in humans but still investigated insufficiently in animals. A novel approach in its management is the utilisation of nutrients from natural sources. Recent studies suggested that phenolic compounds from pomegranate peel could be a promising dietary intervention for MetS. This study evaluated the potency of polyphenol-rich pomegranate peel extract (EPP) in mitigating some MetS components in an animal model. Material and Methods Zucker diabetic fatty rats (with an fa/fa missense mutation in the Lepr leptin receptor gene) and their healthy counterparts (fa/+) as controls were fed a high-calorie diet to induce MetS and supplemented with EPP at two doses: 100 mg/kg body weight (b.w.) and 200 mg/kg b.w. The extract was administered for eight weeks. The rats’ body weights were monitored twice per week, and blood samples were taken before EPP administration after four weeks and eight weeks of study. Echocardiography measurement was performed at the beginning and at the end of the study. Results The extract restrained the dynamic of weight gain. A cardioprotective effect of the highest dose of EPP supplementation was manifested in a relative decrease in heart rate and improved mid-fractional shortening, representing myocardial contractility. No improvement in fasting blood glucose or lipid profile was observed. Conclusion Pomegranate peel extract possesses beneficial health properties that could be useful in dietary intervention in MetS. However, its bioavailability still requires further investigation in clinical trials in humans and animals suffering from endocrine and metabolic disorders.
... Studies on pomegranate juice, which contains some similar polyphenols to pomegranate peel, have also confirmed the antihypertensive effects of pomegranate on systolic and diastolic blood pressure [19,24,25]. The probable mechanism is related to the polyphenols present in pomegranate peel and fruit, which lead to a reduction in the level of angiotensin-converting enzyme and improvement of vasodilation through the effect on nitric oxide [60][61][62]. ...
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Abstract Introduction Non-alcoholic fatty liver disease (NAFLD) is a metabolic syndrome (MS)-related liver disorder that has an increasing prevalence. Thus, the aim of our study is to evaluate the effects of pomegranate peel extract (PP) supplementation on hepatic status and metabolic syndrome risk factors. Methods In phase one, the hydro-alcoholic extraction of the peel of 750 kg of pomegranate (Punica granatum L.) was performed by the soaking method. Then, in phase two, NAFLD patients received 1500 mg of placebo (n = 37) or pomegranate peel capsules (n = 39) with a 500-kcal deficit diet for 8 weeks. Gastrointestinal intolerance, dietary intake, lipid and glycemic profiles, systolic and diastolic blood pressure, body composition, insulin resistance indexes, and elastography-evaluated NAFLD changes were followed. Results The mean age of participants was 43.1 ± 8.6 years (51.3% female). Following the intervention, the mean body weight (mean changes: -5.10 ± 2.30 kg), waist circumference (-7.57 ± 2.97 cm), body mass index (-1.82 ± 0.85 kg/m2), body fat index (-1.49 ± 0.86), and trunk fat (− 3.93 ± 3.07%), systolic (-0.63 ± 0.29 cmHg) and diastolic (-0.39 ± 0.19 cmHg) blood pressure, total cholesterol (-10.51 ± 0.77 mg/dl), triglyceride (-16.02 ± 1.7 mg/dl), low-density lipoprotein cholesterol (-9.33 ± 6.66 mg/dl; all P
Trichinella spiralis (T. spiralis) is a prevalent foodborne intestinal parasite in many developing countries. Albendazole (ABZ) is the drug of choice for treating trichinosis despite its several drawbacks as its week effect against encapsulated larvae, low bioavailability, and emerging drug resistance. As a result, new anthelmintic agents are required. This study aims to investigate the in vivo and in vitro effects of Punica granatum peels extract (PGPE) on intestinal and muscle phases of T. spiralis. The adult worms and larvae were isolated and cultured with different concentrations of PGPE ranging from 6.75 to 100 µg/ml and measuring the survival rate was done after 1, 3, 18, 24 and 48 h of incubation, followed by scanning electron microscopic (SEM) examination of isolated parasites. For the in vivo experiment, the infected animals were divided into two main groups: intestinal phase group and muscular phase group, each group was subdivided into; infected not treated, infected treated with PGPE, ABZ and combined PGPE and ABZ (6 mice in each). The drug effect was assessed by adults and larvae load. A significant increase in the percentage of dead adult parasite and muscle larvae cultured with PGPE with severe destruction and deformity of the tegument were observed with SEM. Also, a significant reduction of adult parasite number in the intestine and muscle larva number in the diaphragm of infected treated mice in comparison to the control group. This study proved that PGPE has a potential activity against trichinosis, particularly when combined with ABZ, and this could serve as a new agent in trichinosis therapy.
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The yield of essential oils isolated from plants is not very high and the hydrodistilled residue by-products rich in polyphenols could be used to increase the profitability of such plant raw materials [1–4]. The aim of the study was to evaluate the compositions and contents of phenolic compounds in the dry extracts obtained from the by-product of American basil (Ocimum americanum L.) and wild bergamot (Monarda fistulosa L.) as well as to investigate their safety and pharmacological activities. The conducted chromatographic analyses of polyphenols revealed the domination of rosmarinic acid in both obtained dry extracts. Its amount analyzed by high-performance liquid chromatography method was 91.23 ± 1.62 mg/g in the Monarda fistulosa dry extract (ME) and 78.70 ± 1.13 mg/g in the Ocimum americanum dry extract (OE). Luteolin-7-O-glucoside was the second predominant polyphenol of both extracts, but its content differed significantly (76.30 ± 1.50 mg/g and 17.22 ±0.49 mg/g, respectively). Caffeic acid (21.62 ± 0.17 mg/g) followed by apigenin (15.12 ± 0.15 mg/g) were the other major compounds in the ME, whilst rutin (11.20 ± 0.26 mg/g) and ferulic acid (8.21 ± 0.09 mg/g) predominated in OE. The free radical scavenging activity against DPPH of ME and OE were IC50 = 0.285 mg/mL and IC50 = 0.298 mg/mL, respectively. Both tested extracts dose-dependently decreased the paw oedema in rats suggesting their anti-inflammatory properties. The administration of extracts at the doses of 500–5000 mg/kg to rats did not reveal any toxic reactions that indicates their safety. Consequently, the studied by-products are promising sources of bioactive compounds with antioxidant and anti-inflammatory effects.
Conference Paper
Proteolytic Enzymes Papain and Chymotrypsin Combined with Laser Techniques for the Management of Facial Hirsutism Aikaterini Liatsopoulou1, Athanasia Varvaresou1, Evangelia Protopapa1 1 Laboratory of Chemistry-Biochemistry-Cosmetic Science, Department of Biomedical Sciences, University of West Attica, 28 Ag. Spyridonos Str., 12243, Egaleo, Athens, Greece Background. Papain is a proteolytic enzyme, derived from the latex of Carica papaya. It is used for depilatory preparations and has been proposed as a safe treatment for hirsutism. Photo-epilation is the treatment of choice for hirsutism, but it does not offer complete and persistent results. Meanwhile, iontophoresis of proteolytic enzymes papain and chymotrypsin to skin of experimental animals, has shown long-lasting depilatory effects. Therefore, we think it would be of interest to combine photo-epilation with application of these enzymes, as a more natural approach, to enhance treatment efficacy. Methods. In this randomized controlled blinded clinical study, 30 adult Caucasian women with facial hirsutism were divided into two groups: Group I, treated with combination of laser Alexandrite 755nm and iontophoresis of aqueous solutions papain 0.48% and chymotrypsin 0.29% and Group II, treated with laser Alexandrite 755nm alone. Evaluations were based on digital hair counts. Comparison between groups of percentage change from baseline to 6 months was performed using the independent samples t-test and the statistical package SPSS vr 21.00 (IBM Corporation, Somers, NY, USA). The study has been approved by the Research Ethics Committee of the University of West Attica. Results. The decrease of percentage change from baseline to 6 months of the ‘Total Number of Hairs’ was statistically significantly higher in Group I compared to Group II (p=0.017). Conclusions. Iontophoretic delivery of papain and chymotrypsin provides a more natural adjuvant treatment, with a potential to increase laser-induced hair reduction in hirsute patients. Table: Analysis of Total Number of Hairs variable between groups, during the observation period. Groups Baseline After 6 months of treatment % Decrease I (Laser+ Enzymes) 73.53±13.93 37.60±11.55 -48.70%±13.77 II (Laser) 77.60±24.47 56.40±.27.0 -25.60%±32.39 p-value 0.017 All values are presented as mean±SD References [1] Mellou F, Varvaresou A, Papageorgiou S, Protopapa E, Liatsopoulou E. Papain: An Enzyme of Multiple Applications. Rev. Clin. Pharmacol. Pharmacokinet. Int. 2020; 34: 107–111 [2] Oliveira Pinto CAS, Lopes PS, Sarruf FD, Polakiewicz B, Kaneko TM, Baby AR, Maria Velasco MVR. Comparative study of the stability of free and modified papain incorporated in topical formulations. Brazilian Journal of Pharmaceutical Sciences. 2011; 47(4) [3] Mikiel D, Olszewska B, Polanska A, Adamski Z, Zaba R, Danczak-Pazdrowska A. Principles of management of women with hirsutism – a dermatologist’s perspective. Dermatol Rev/Przegl Dermatol. 2020; 107: 424–40 [4] Vissing AC, Taudorf EH, Haak CS, Philipsen PA, Hædersdal M. Adjuvant eflornithine to maintain IPL-induced hair reduction in women with facial hirsutism: A randomized controlled trial. J Eur Acad Dermatology Venereol. 2016; 30(2): 314–9 [5] Protopapa EE, Gaissert H, Xenakis A, Avramiotis S, Stavrianeas N, Sekeris CE, Schenkelf J, Alonso A. The effect of proteolytic enzymes on hair follicles of transgenic mice expressing the lac Z-protein in cells of the bulge region. J Eur Acad Dermatology Venereol. 1999; 13(1): 28–35
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Drospirenone (DRSP) is a progestin with anti-aldosterone properties and it reduces blood pressure in hypertensive women. However, the effects of DRSP on endothelium-dependent coronary vasodilation have not been evaluated. This study investigated the effects of combined therapy with estrogen (E2) and DRSP on endothelium-dependent vasodilation of the coronary bed of ovariectomized (OVX) spontaneously hypertensive rats. Female spontaneously hypertensive rats (n=87) at 12 weeks of age were randomly divided into sham operated (Sham), OVX, OVX treated with E2 (E2), and OVX treated with E2 and DRSP (E2+DRSP) groups. Hemodynamic parameters were directly evaluated by catheter insertion into the femoral artery. Endothelium-dependent vasodilation in response to bradykinin in the coronary arterial bed was assessed using isolated hearts according to a modified Langendorff method. Coronary protein expression of endothelial nitric oxide synthase and estrogen receptor alpha (ER-α) was assessed by Western blotting. Histological slices of coronary arteries were stained with hematoxylin and eosin, and morphometric parameters were analyzed. Oxidative stress was assessed in situ by dihydroethidium fluorescence. Ovariectomy increased systolic blood pressure, which was only prevented by E2+DRSP treatment. Estrogen deficiency caused endothelial dysfunction, which was prevented by both treatments. However, the vasodilator response in the E2+DRSP group was significantly higher at the three highest concentrations compared with the OVX group. Reduced ER-α expression in OVX rats was restored by both treatments. Morphometric parameters and oxidative stress were augmented by OVX and reduced by E2 and E2+DRSP treatments. Hormonal therapy with E2 and DRSP may be an important therapeutic option in the prevention of coronary heart disease in hypertensive post-menopausal women.
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Obesity is causally linked to a number of comorbidities, including cardiovascular disease, diabetes, renal dysfunction, and cancer. Obesity has also been linked to pulmonary disorders, including pulmonary arterial hypertension (PAH). It was long believed that obesity-related PAH was the result of hypoventilation and hypoxia due to the increased mechanical load of excess body fat. However, in recent years it has been proposed that the metabolic and inflammatory disturbances of obesity may also play a role in the development of PAH. To determine whether PAH develops in obese rats in the absence of hypoxia, we assessed pulmonary hemodynamics and pulmonary artery (PA) structure in the diet-resistant/diet-induced obesity (DR/DIO) and Zucker lean/fatty rat models. We found that high-fat feeding (DR/DIO) or overfeeding (Zucker) elicited PA remodeling, neomuscularization of distal arterioles, and elevated PA pressure, accompanied by right ventricular (RV) hypertrophy. PA thickening and distal neomuscularization were also observed in DIO rats on a low-fat diet. No evidence of hypoventilation or chronic hypoxia was detected in either model, nor was there a correlation between blood glucose or insulin levels and PAH. However, circulating inflammatory cytokine levels were increased with high-fat feeding or calorie overload, and hyperlipidemia and oxidant damage in the PA wall correlated with PAH in the DR/DIO model. We conclude that hyperlipidemia and peripheral inflammation correlate with the development of PAH in obese subjects. Obesity-related inflammation may predispose to PAH even in the absence of hypoxia.
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In recent years, the number of scientific papers concerning pomegranate (Punica granatum L.) and its health properties has increased greatly, and there is great potential for the use of bioactive-rich pomegranate extracts as ingredients in functional foods and nutraceuticals. To translate this potential into effective strategies it is essential to further elucidate the mechanisms of the reported bioactivity. In this study HepG2 cells were supplemented with a pomegranate fruit extract or with the corresponding amount of pure punicalagin, and then subjected to an exogenous oxidative stress. Overall, upon the oxidative stress the gene expression and activity of the main antioxidant enzymes appeared reduced in supplemented cells, which were more prone to the detrimental effects than unsupplemented ones. No differences were detected between cells supplemented with the pomegranate juice or the pure punicalagin. Although further studies are needed due to the gaps existing between in vitro and in vivo studies, our results suggest caution in the administration of high concentrations of nutraceutical molecules, particularly when they are administered in concentrated form.
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Pomegranate juice (PJ) and its products are directly used in foods due to their pleasant taste and palatability as well as preservative effects. In spite of useful effects of essential oils such as zataria multiflora Boiss essential oil (ZEO) on prolonging shelf-life of foods, their application is restricted due to their vigorous taste and aroma. In the present study, physicochemical characteristics, chemical compositions and antioxidative activities of two Iranian native plants, PJ (Rabbab-e-Neyriz cultivar) and ZEO were investigated. 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging and reducing power tests were used for measuring antioxidant activity. The level of total phenolic of them were also determined. Total soluble solids content, pH value, titratable acidity content and total anthocyanins content of PJ were also measured. Chemical compositions of ZEO were determined using gas-chromatography, mass-spectrometry (GC-MS). The results of antioxidative tests indicated that the ZEO was significantly more potent (p < 0.05) than PJ. Also the phenolic content in ZEO (262.52 mg per g) was significantly higher (p < 0.05) than PJ (154.90 mg per 100g). Chemical compositions analysis of ZEO indicated that its major components were carvacrol (59.17%), linalool (23.67%), trans-caryophyllene (3.07%) and carvacrol methyl ether (2.44%). In the present study, physicochemical and antioxidative characteristics of Rabbab-e-Neyriz PJ were determined for first time. It was aslo found that ZEO in comparison with PJ had higher antioxidative activity and total phenolic content.
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Superoxide anion formation is vital to the microbicidal activity of phagocytes. Recently, however, there is accumulating evidence that it is also involved in cell growth in vascular smooth muscle cells (VSMCs). We have shown that the hypertrophic agent angiotensin II stimulates superoxide production by activating the membrane-bound NADH/NADPH oxidase and that inhibition of this oxidase attenuates vascular hypertrophy. However, the molecular identity of this oxidase in VSMCs is unknown. We have recently cloned the cytochrome b558 α-subunit, p22phox (one of the key electron transfer elements of the NADPH oxidase in phagocytes), from a rat VSMC cDNA library, but its role in VSMC oxidase activity remains unclarified. Here we report that the complete inhibition of p22phox mRNA expression by stable transfection of antisense p22phox cDNA into VSMCs results in a decrease in cytochrome b content, which is accompanied by a significant inhibition of angiotensin II-stimulated NADH/NADPH-dependent superoxide production, subsequent hydrogen peroxide production, and [³H]leucine incorporation. We provide the first evidence that p22phox is a critical component of superoxide-generating vascular NADH/NADPH oxidase and suggest a central role for this oxidase system in vascular hypertrophy.
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Quantification of the disease burden caused by different risks informs prevention by providing an account of health loss different to that provided by a disease-by-disease analysis. No complete revision of global disease burden caused by risk factors has been done since a comparative risk assessment in 2000, and no previous analysis has assessed changes in burden attributable to risk factors over time. METHODS We estimated deaths and disability-adjusted life years (DALYs; sum of years lived with disability [YLD] and years of life lost [YLL]) attributable to the independent effects of 67 risk factors and clusters of risk factors for 21 regions in 1990 and 2010. We estimated exposure distributions for each year, region, sex, and age group, and relative risks per unit of exposure by systematically reviewing and synthesising published and unpublished data. We used these estimates, together with estimates of cause-specific deaths and DALYs from the Global Burden of Disease Study 2010, to calculate the burden attributable to each risk factor exposure compared with the theoretical-minimum-risk exposure. We incorporated uncertainty in disease burden, relative risks, and exposures into our estimates of attributable burden. FINDINGS In 2010, the three leading risk factors for global disease burden were high blood pressure (7·0% [95% uncertainty interval 6·2-7·7] of global DALYs), tobacco smoking including second-hand smoke (6·3% [5·5-7·0]), and alcohol use (5·5% [5·0-5·9]). In 1990, the leading risks were childhood underweight (7·9% [6·8-9·4]), household air pollution from solid fuels (HAP; 7·0% [5·6-8·3]), and tobacco smoking including second-hand smoke (6·1% [5·4-6·8]). Dietary risk factors and physical inactivity collectively accounted for 10·0% (95% UI 9·2-10·8) of global DALYs in 2010, with the most prominent dietary risks being diets low in fruits and those high in sodium. Several risks that primarily affect childhood communicable diseases, including unimproved water and sanitation and childhood micronutrient deficiencies, fell in rank between 1990 and 2010, with unimproved water and sanitation accounting for 0·9% (0·4-1·6) of global DALYs in 2010. However, in most of sub-Saharan Africa childhood underweight, HAP, and non-exclusive and discontinued breastfeeding were the leading risks in 2010, while HAP was the leading risk in south Asia. The leading risk factor in Eastern Europe, most of Latin America, and southern sub-Saharan Africa in 2010 was alcohol use; in most of Asia, North Africa and Middle East, and central Europe it was high blood pressure. Despite declines, tobacco smoking including second-hand smoke remained the leading risk in high-income north America and western Europe. High body-mass index has increased globally and it is the leading risk in Australasia and southern Latin America, and also ranks high in other high-income regions, North Africa and Middle East, and Oceania. INTERPRETATION Worldwide, the contribution of different risk factors to disease burden has changed substantially, with a shift away from risks for communicable diseases in children towards those for non-communicable diseases in adults. These changes are related to the ageing population, decreased mortality among children younger than 5 years, changes in cause-of-death composition, and changes in risk factor exposures. New evidence has led to changes in the magnitude of key risks including unimproved water and sanitation, vitamin A and zinc deficiencies, and ambient particulate matter pollution. The extent to which the epidemiological shift has occurred and what the leading risks currently are varies greatly across regions. In much of sub-Saharan Africa, the leading risks are still those associated with poverty and those that affect children. FUNDING Bill & Melinda Gates Foundation.
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The current paper summarizes the antioxidative and antiatherogenic effects of pomegranate polyphenols on serum lipoproteins and on arterial macrophages (two major components of the atherosclerotic lesion), using both in vitro and in vivo humans and mice models. Pomegranate juice and its by-products substantially reduced macrophage cholesterol and oxidized lipids accumulation, and foam cell formation (the hallmark of early atherogenesis), leading to attenuation of atherosclerosis development, and its consequent cardiovascular events.
The antioxidant activity of pomegranate juices was evaluated by four different methods (ABTS, DPPH, DMPD, and FRAP) and compared to those of red wine and a green tea infusion. Commercial pomegranate juices showed an antioxidant activity (18−20 TEAC) three times higher than those of red wine and green tea (6−8 TEAC). The activity was higher in commercial juices extracted from whole pomegranates than in experimental juices obtained from the arils only (12−14 TEAC). HPLC-DAD and HPLC-MS analyses of the juices revealed that commercial juices contained the pomegranate tannin punicalagin (1500−1900 mg/L) while only traces of this compound were detected in the experimental juice obtained from arils in the laboratory. This shows that pomegranate industrial processing extracts some of the hydrolyzable tannins present in the fruit rind. This could account for the higher antioxidant activity of commercial juices compared to the experimental ones. In addition, anthocyanins, ellagic acid derivatives, and hydrolyzable tannins were detected and quantified in the pomegranate juices. Keywords: Pomegranate; Punica granatum; Punicaceae; juice; phenolics; anthocyanins; ellagic acid; punicalagin; tannins; antioxidant activity; ABTS; DPPH; DMPD; FRAP