protective-effect-of-grape-seed-extract-andor-silymarin-against-thioacetamideinduced-hepatic-fibrosis-in-rats-2167-0889-1000178

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Protective Effect of Grape Seed Extract and/or Silymarin Against
Thioacetamide-induced Hepatic Fibrosis in Rats
Somaia A. Nada1*, Ayah M.H. Gowifel2, Ezz El-Din S. El-Denshary3, Abeer A. Salama1, Mona G Khalil2 and Kawkab A Ahmed4
1Pharmacology Department, National Research Center, Giza, Egypt
2Pharmacology and Toxicology Department, Faculty of Pharmacy, Modern University for Technology and information, Cairo, Egypt
3Pharmacology and Toxicology Department, Faculty of Pharmacy, Cairo University, Cairo, Egypt
4Pathology Department, Faculty of Veterinary Medicine, Cairo University, Giza, Egypt
*Corresponding author: Somaia A. Nada, Pharmacology Department, National Research Centre, Cairo, Egypt. E-mail: somaianada@yahoo.com
Received date: July 12, 2014; Accepted date: April 24, 2015; Published date: April 27, 2015
Copyright: © 2015 Nada SA, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted
use, distribution, and reproduction in any medium, provided the original author and source are credited.
Abstract
The aim of the present study was designed to evaluate the hepatoprotective and antioxidant potentials of GSE
(100 and 200 mg/kg) and/or silymarin against TAA-induced liver fibrosis in rats.
This study was designed to investigate the protective effect of grape seed extract (GSE) and/or silymarin against
thioacetamide (TAA)-induced hepatic fibrosis in Sprague-Dawley rats. Mature male Sprague-Dawley rats were
divided into 7 equal groups (8 rats each) and treated as follows: Group 1, kept as control group and orally given
saline; groups 2-7 were injected intraperitoneally (i.p.) with TAA (100 mg/Kg) twice weekly for 6 weeks to induce
hepatic fibrosis. Group 2, kept as control positive; groups 3-5 were administered daily oral doses of silymarin (50
mg/kg), GSE (100 mg/kg) and GSE (200 mg/kg), respectively. While groups 6-7 were administered combined
treatments of silymarin and GSE (100 mg/kg) or GSE (200 mg/kg), respectively. Our results indicated that TAA
caused significant elevation of hydroxyproline (Hyp), malondialdehyde (MDA) and nitric oxide (NO) contents in liver
homogenate and increased serum levels of: aminotransferases (AST and ALT), alkaline phosphatase (ALP) and
total bilirubin. While, TAA-treatment alone significantly decreased serum total protein and reduced glutathione (GSH)
content in liver homogenate. Administration of GSE (100 and 200 mg/kg) and/or silymarin attenuated TAA-induced
hepatic fibrosis, improved enzymes and reduced the oxidative stress in dose dependant manner Histopathological
study showed disruption of the hepatic architecture and collagen fibers deposition in the portal tract of TAA-injected
group. Concomitant treatment with GSE (100 and 200 mg/kg) and/or silymarin significantly improved
histopathological structure of liver tissue in variable degrees. In conclusion, the combined effect of GSE (200 mg/kg)
with silymarin (50 mg/kg) had powerful hepatoprotective effect than any other studied doses.
Keywords: Grape seed extract; Liver fibrosis; Rat
Introduction
Hepatic fibrosis, defined by excessive accumulation of extracellular
matrix (ECM) and resultant loss of pliability and liver function, is the
result of wound-healing responses triggered by either acute or chronic
liver injury [1]. Hepatic stellate cells (HSCs) are the main ECM-
producing cells in the injured liver [2].
Thioacetamide (TAA) is a thiono-sulfur containing compound. It
has been used as a fungicide, organic solvent, accelerator in the
vulcanization of rubber, and as a stabilizer of motor oil [3]. Treatment
of rodents with TAA is known to lead to liver fibrosis and eventually
cirrhosis, with the morphological and biochemical changes resembling
that of the human disease [4]. The toxicity of TAA results from its
bioactivation by a mixed-function oxidase system, particularly by
CYP2E1 and flavin adenine dinucleotide (FAD) monooxygenases [5].
Metabolic activation of TAA then leads to the formation of reactive
metabolites that are represented by radicals derived from TAA-S-oxide
and by reactive oxygen species (ROS) generated as intermediates [6]. It
was documented that reactive metabolites can covalently bind to
cellular macromolecules and/or induce oxidative stress [7].
Grape seed extract (GSE) is a natural extract from the seed of Vitis
vinifera. It is a rich source of flavonoids, proanthocyanidins oligomers.
Proanthocyanidins are a class of phenolic compounds that take the
forms of oligomers or polymers of polyhydroxy falvan- 3-ol units, such
as (+)-catechin, (−)-epicatechin [8]. These flavonoids exert many
health promoting effects including the ability to increase intracellular
vitamin C levels, decrease capillary permeability and fragility and
scavenge oxidants and free radicals [9,10]. Although these bioactive
components of grape mainly exist in grape skin and seeds, grape skin
and seeds are usually discarded in regular dietary intake and the
winery and grape juice industry. These wastes contain bioactive
components with potent antioxidant and free radical scavenging
activity. Grape seed extract proanthocyanidins has antibacterial, and
anti-inflammatory effect, they have been reported to inhibit lipid
peroxidation [11-14].
Silymarin is a mixture of flavonolignans extracted from “milk
thistle” Silybum marianum [15]. It is a mixture of flavonoid isomers
such as silibinin, isosilibinin, silidianin, and silichristin. Silymarin is
used primarily to treat various liver diseases and dysfunctions
including alcoholic cirrhosis, hepatitis (due to viral infections or drug-
induced), as well as hepatic problems related to diabetes [16].
Silymarin has free radical scavenging properties and its ability to
enhance endogenous anti-oxidant defense systems in vivo [17].
Liver Nada SA, et al., J Liver 2015, 4:2
http://dx.doi.org/10.4172/2167-0889.1000178
Research Article Open Access
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ISSN:2167-0889 JLR, an open access journal Volume 4 • Issue 2 • 1000178

Silymarin has been shown to reduce liver fibrosis up to 30–35%, and in
few cases it has reversed the liver fibrosis [18]. Silymarin showed anti-
inflammatory and anti-metastatic activities; it has protective effect
against toxicity of chemotherapy and radiotherapy [19].
Materials and Methods
Chemicals
TAA was obtained from El-Gomhouria Company for drug and
chemicals, Egypt, silymarin was obtained from Sigma-Aldrich, USA;
GSE was obtained from Arab Gelatin Pharmaceutical Products
Company, Egypt. All other chemicals, used throughout the
experiment, were of the highest analytical grade available. Kits used to
measurement of serum aspartate transaminase (AST), alanine
transaminase (ALT); alkaline phosphatase (ALP); total protein; total
bilirubin levels; hepatic malondialdehyde (MDA); nitric oxide (NO)
and hepatic reduced glutathione (GSH) were purchased from
Biodiagnostic, Inc., (Egypt). Kit used for measurement of hepatic
hydroxyproline (Hyp) was purchased from Glory Science Co., Ltd.
Animals
Adult male Sprague- Dawley rats weighing 150-200 gm were
obtained from the animal house at the National Research Center
(Giza, Egypt), and fed a standard laboratory diet and tap water ad
libitum. Experimental animals were housed in an air-conditioned
room at 22–25 °C with a 12-h light/dark cycle. All animals received
humane care and the study protocols were in compliance with
institutional guidelines for the use of laboratory animals.
Experimental design
After an acclimatization period of one week, sixty-four healthy male
Sprague- Dawley rats were randomly assigned to seven groups of eight
rats per group and that were treated as follows:
Group (1): Received normal saline orally, daily and injected
intraperitoneally (i.p.) with sterile distilled water twice a week for 6
weeks (normal control group);
Groups (2-7): Rats injected with TAA (100 mg/kg, i.p.), according
to Hamed et al. [20], dissolved in distilled water, twice a week for 6
weeks; and the following treatments were given daily oral doses in
concomitant with TAA for six weeks: Group (2): Kept as positive
control, and received normal saline; Group (3) given silymarin (50
mg/kg) according to Wills and Asha [21]; Group (4) administered GSE
(100 mg/kg) according to Pallares et al. [22]; Group (5) received GSE
(200 mg/kg) according to Yousef et al. [23]; Group (6) treated daily
with GSE (100 mg/kg) and silymarin (50 mg/kg); and Group (7)
received GSE (200 mg/kg) and Silymarin (50 mg/kg).
Collection of blood samples
At the end of the experimental period; blood samples were
withdrawn from the retro-orbital vein of each animal, under light
anesthesia by diethyl ether, according to the method described by
Cocchetto and Bjornsson [24]. Blood was allowed to coagulate and
then centrifuged at 3000 rpm for 15 min. The obtained serum was
used to estimate the levels of AST; ALT activities according to the
method of Reitman and Frankel [25]; ALP was determined according
to the method of Belfield and Goldberg [26]; total bilirubin was
analyzed by the method of Walter and Gerade [27]; and total protein
was estimated by the method of Gornal et al. [28].
Preparation of liver homogenate
Immediately after blood sampling, animals were sacrificed by
cervical dislocation under ether anesthesia and livers were collected for
biochemical and histopathological examinations. Liver tissues were
rapidly removed, washed in ice-cooled saline, plotted dry and weighed.
A weighed part of each liver was homogenized, using a homogenizer
(Medical instruments, MPW-120, Poland), with ice-cooled saline
(0.9% NaCl) to prepare 20% w/v homogenate. The homogenate was
then centrifuged at 4000 rpm for 5 min. at 4°C using a cooling
centrifuge to remove cell debris (Laborzentrifugen, 2k15, Sigma,
Germany). The aliquot was divided into four parts; the 1st part was
used for the assessment of lipid peroxidation (LPO) as MDA
according to the methods described by Ohkawa et al.; the 2nd part was
used for the determination of the level of NO by the method of
Montgomery and Dymock; the 3rd part was used to estimate GSH by
the method of Beutler et al.; and the 4th part was used to estimate Hyp
according to the instructions of the manufacturer, using glory science
ELISA kit [29-31].
Histopathological examinations
The specimens from the liver were taken and fixed immediately in
10% neutral buffered formalin, processed for light microscopy to get
(5μm) paraffin sections and stained with: Hematoxylin & Eosin (H &
E) to verify histological details and Masson’s trichrome staining (MTs)
to demonstrate the collagen fibers as described by Bancroft and
Gamble [32].
Statistical analysis
The degree in variability of results was expressed as means ±
standard error of means (SEM). Data were evaluated by one-way
analysis of variance (ANOVA) followed by Tukey’s multiple
comparisons test. The level of significance was accepted at P < 0.05.
Results
Effect of GSE (100 and 200 mg/kg) and/or silymarin (50
mg/kg) on serum liver function tests in TAA-induced hepatic
fibrosis in rats
Injection of TAA (100 mg/kg, i.p.) resulted in a considerable
hepatic injury as assessed by a significant elevations in serum AST,
ALT, ALP and total bilirubin by 636.39 %, 574.11%, 175.35%, and
438.67%, respectively as compared to normal control values (Table 1).
Treatment with silymarin (50 mg/kg) exerted a significant decrease in
serum AST, ALT, ALP, and total bilirubin levels by 53.49%, 34.9%,
37.48%, and 43.59%, respectively as compared to TAA control group.
Administration of GSE (100 and 200 mg/kg) significantly reduced
TAA-induced elevations by 42.09%, and 68.15% for AST; 25.45%, and
60.07% for ALT; 27.99%, and 49.68% for ALP; and 34.26%, and
61.33% for total bilirubin, respectively in dose-dependent manner,
compared to TAA control group. Meanwhile, administration of
silymarin (50 mg/kg) combined with either, GSE (100 mg/kg) or GSE
(200 mg/kg), significantly decreased elevated serum AST by 66%, and
84.02%; ALT by 49.62%, and 70.91%; ALP by 47.54%, and 57.10%, and
Citation: Nada SA, Gowifel AMH, El-Denshary EES, Salama AA, Khalil MG, et al. (2015) Protective Effect of Grape Seed Extract and/or
Silymarin Against Thioacetamide-induced Hepatic Fibrosis in Rats . J Liver 4: 178. doi:10.4172/2167-0889.1000178
Page 2 of 7
J Liver
ISSN:2167-0889 JLR, an open access journal Volume 4 • Issue 2 • 1000178

total bilirubin by 54.46%, and 69.13%, respectively, compared to TAA
control group.
Results showed that TAA injection significantly decreased serum
total protein level by 38.95%, compared to normal control group
(Table 1). A significant elevation in serum total protein level was
observed after treatment with silymarin (50 mg/kg) by 17.49%, as
compared to TAA control group. Meanwhile, GSE (100 and 200
mg/kg) showed a significant elevation in serum total protein level as
compared to TAA control group by 20.37%, and 37.14%, respectively.
Administration of silymarin (50 mg/kg) combined with either, GSE
(100 mg/kg) or GSE (200 mg/kg), significantly increased serum total
protein level by 35.06%, and 51.83%, respectively, compared to TAA
control group (Figure 1).
Figure 1: The effect of oral administration of GSE (100 and 200 mg/kg) and/or silymarin (50 mg/kg) on liver MDA (A), NO (B),
hydroxyproline (Hyp) (C), and reduced GSH (D) contents in TAA-induced hepatic fibrosis in rats: (Means ± SEM, n = 8 rats/group),
ANOVA – one way (groups with different letters are significantly different at P < 0.05). MDA, Malondialdehyde; NO, Nitric oxide; Hyp,
Hydroxyproline; GSH, Reduced glutathione; TAA, Thioacetamide; GSE, Grape seed extract.
Citation: Nada SA, Gowifel AMH, El-Denshary EES, Salama AA, Khalil MG, et al. (2015) Protective Effect of Grape Seed Extract and/or
Silymarin Against Thioacetamide-induced Hepatic Fibrosis in Rats . J Liver 4: 178. doi:10.4172/2167-0889.1000178
Page 3 of 7
J Liver
ISSN:2167-0889 JLR, an open access journal Volume 4 • Issue 2 • 1000178

Groups\Parameters AST(U/ml) ALT(U/ml) ALP(IU/L) Total bilirubin(mg/dl) Total Protein(gm/dl)
Normal control (Saline) 34 ± 0.74a28 ± 0.82a77.47 ± 1.09a0.32 ± 0.01a7.04 ± 0.17a
TAA (100 mg/Kg), i.p. 250.38 ± 2.11b188.75 ± 2.13b213.32 ± 2.03b1.71 ± 0.03b4.29 ± 0.14b
Silymarin (50 mg/Kg) 116.44 ± 1.62c122.88 ± 1.59c133.38 ± 1.74c0.96 ± 0.02c5.05 ± 0.07c
GSE (100 mg/Kg) 145 ± 1.73d140.71 ± 2.07d153.62 ± 1.85d1.12 ± 0.02d5.17 ± 0.06c
GSE (200 mg/Kg) 79.75 ± 1.05e75.38 ± 1.07e107.34 ± 1.41e0.66 ± 0.02e5.89 ± 0.11d
GSE (100 mg/Kg) + Silymarin 85.13 ± 1.14e95.08 ± 1.38f111.92 ± 1.63e0.78 ± 0.02f5.8 ± 0.08d
GSE (200 mg/Kg) + Silymarin 40 ± 0.74a54.92 ± 0.99g91.39 ± 1.24f0.53 ± 0.01g6.52 ± 0.15e
ANOVA one way, within each raw, means the different superscript letters are significantly different between groups at P< 0.05; Data are expressed as Mean ± SEM
(n=8 rats).
Table 1: Effect of GSE (100 and 200 mg/kg) and/or silymarin (50 mg/kg) on serum liver function tests in TAA-induced hepatic fibrosis in rats.
Effect of GSE (100 and 200 mg/kg) and/or silymarin (50
mg/kg) on liver MDA, NO, Hyp, and GSH contents in TAA-
induced hepatic fibrosis in rats
TAA injection caused a significant elevation in MDA, NO, and Hyp
values as well as significant depletion in GSH value (Figure 1).
Injection of TAA significantly elevated liver MDA, NO, and Hyp
contents by 325.65%, 150.57%, and 481.96%, respectively, as compared
to normal control values. Treatment with silymarin (50 mg/kg),
significantly decreased liver MDA, NO, and Hyp contents by 16.22%,
14.12%, and 24.89%, respectively, compared to TAA control group.
Treatment with GSE (100 and 200 mg/kg) showed a significant
decrease in elevated liver MDA by 24.56%, and 59.98% (Figure 1A);
NO by 15.45%, and 44.82% (Figure 1B), and Hyp by22.97%, and
64.84% (Figure 1C), compared to TAA control group, respectively.
Administration of silymarin (50 mg/kg) combined with either, GSE
(100 mg/kg) or GSE (200 mg/kg), significantly decreased elevated liver
MDA, NO, and Hyp contents by 44.63%, and 68.18% for MDA (Figure
1A); 29.68%, and 58.02% for NO (Figure 1B); and 48.56%, and 81.3%
for Hyp (Figure 1C), respectively, compared to TAA control group.
Results showed that TAA significantly decreased liver GSH content
by 56.13%, as compared to normal control group. Treatment with
silymarin (50 mg/kg), significantly increased the hepatic concentration
of GSH by 27.93%, compared to TAA control group. Moreover,
treatment with GSE (100 and 200 mg/kg) caused a significant increase
in GSH level by 54.83%, and 82.41%, respectively, compared to TAA
control value. Administration of silymarin (50 mg/kg) combined with
either, GSE (100 mg/kg) or GSE (200 mg/kg), significantly increased
liver GSH content by 89.31%, and 108.97%, respectively, compared to
TAA control group (Figure 1D).
Histopathological studies
The histological examination of the liver of control rats revealed
normal architecture of hepatic lobules with hepatocytes radiating from
the central veins, with narrow sinusoids and prominent nucleus
(Figure 2A).
Light microscopic examination of the TAA control rats showed
disruption of normal architecture of hepatic lobules and collagen
fibers deposition in portal tract, and pseudolobulation of hepatocytes
with fibroblasts (Figure 2B).
Figure 2: photomicrographs of liver sections from (A) normal
control group showing normal histological structure of hepatic
parenchyma. Note central vein (CV) and hepatocytes (H). (B) Rat
injected with TAA showing disarrangement of normal hepatic cells
disruption of normal architecture of hepatic lobules and collagen
fibers deposition in portal tract, and pseudolobulation of
hepatocytes with fibroblasts. (C) Rat treated with silymarin (50
mg/kg) showing scanty collagen deposition in portal tract, marked
activation of kupffer cells and necrosis of sporadic hepatocytes. (D)
Rat treated with GSE (100 mg/kg) showing scanty collagen fibers
deposition in the portal tract and marked activation of kupffer cells.
(E) Rat treated with GSE (200 mg/kg) showing improvement in the
liver tissue with necrosis of sporadic hepatocytes. (F) Rat treated
with combination of GSE (100 mg/kg) and silymarin (50 mg/kg)
showing mild improvement in the liver tissue with thin strands of
fibroblasts in portal tract. (G): Rat treated with combination of GSE
(200 mg/kg) and silymarin (50 mg/kg) showing apparent normal
hepatic parenchyma. (H & E X 100).
Treatment with silymarin (50 mg/kg), showed scanty collagen
deposition in portal tract, marked activation of kupffer cells and
necrosis of sporadic hepatocytes (Figure 2C).
Citation: Nada SA, Gowifel AMH, El-Denshary EES, Salama AA, Khalil MG, et al. (2015) Protective Effect of Grape Seed Extract and/or
Silymarin Against Thioacetamide-induced Hepatic Fibrosis in Rats . J Liver 4: 178. doi:10.4172/2167-0889.1000178
Page 4 of 7
J Liver
ISSN:2167-0889 JLR, an open access journal Volume 4 • Issue 2 • 1000178

Treatment with GSE (100 mg/kg) showed scanty collagen fibers
deposition in the portal tract and marked activation of kupffer cells
(Figure 2D).
Treatment with GSE, (200 mg/kg), showed necrosis of sporadic
hepatocytes (Figure 2E). Administration of combination of GSE (100
mg/kg) and silymarin (50 mg/kg) showed thin strands of fibroblasts in
portal tract (Figure 2F).
In group treated with GSE (200 mg/kg) and silymarin 50 mg/kg)
combination, the liver sections exhibited apparent normal hepatic
parenchyma (Figure 2G).
Figure 3: photomicrographs of liver sections from (A) normal
control group showing no histochemical reaction. (B) Rat injected
with TAA showing a strong positive reaction. (C) Rat treated with
silymarin (50 mg/kg) showing moderate fibrotic changes. (D) Rat
treated with GSE (100 mg/kg) showing moderate histochemical
reaction. (E) Rat treated with GSE (200 mg/kg) showing
improvement in the liver tissue with no histochemical reaction. (F)
Rat treated with combination of GSE (100 mg/kg) and silymarin
(50 mg/kg) showing mild fibrotic changes. (G): Rat treated with
combination of GSE (200 mg/kg) and silymarin (50 mg/kg)
showing no histochemical reaction. (MTs X100).
Histochemical studies
The histochemical examination of the liver sections of normal
control rats revealed no histochemical reaction (Figure 3A).
Examination of the TAA control rats showed a strong positive
reaction which indicate markedly increased accumulation of collagen
that was deposited (Figure 3B).
Silymarin (50 mg/kg) treatment showed moderate fibrotic changes
(Figure 3C).
Treatment with GSE (100 mg/kg) showed moderate histochemical
reaction (moderate fibrotic changes) (Figure 3D).
GSE (200 mg/kg) administration showed no histochemical reaction
(Figure 3E).
Administration of combination of GSE (100 mg/kg) and silymarin
(50 mg/kg) showed mild fibrotic changes which indicated ameliorated
fibrosis formation with only a few tiny, short bundles of collagen
(Figure 3F).
Treatment with combination of GSE (200 mg/kg) and silymarin (50
mg/kg) showed no histochemical reaction (Figure 3G).
Discussion
The present study shows that TAA-treatment alone had abundant
hepatotoxic effect, clearly demonstrated by the elevation in liver
enzyme markers, TB, Hyb., MDA and NO levels in hepatic
homogenate. As well as, significant decrease in serum TP and hepatic
–GSH content.
TAA bioactivation is mediated by microsomal cytochrome P450E1
and/or flavin-containing monooxygenase systems to form TAA
sulfoxide involving TAA-S,S-dioxide, which caused lipid peroxidation
at the plasma membrane level [33]. The S-oxide metabolite (TASO2)
covalently binds to liver macromolecules forming acetylimidolysine
derivatives that are responsible for hepatotoxic effects [34]. TAA
induces calcium (Ca 2+) mobilization from its intracellular stores [35].
Both ROS and Ca2+ were determined to activate multiple mechanisms
related to cell damage or proliferation [36]. The increased ROS
formation and disruption of calcium homeostasis increased the
permeability and disrupt the mitochondrial inner membrane and
inhibit its respiration [37].
In accordance with our results, total protein significantly decreased
as a result of TAA-induced liver toxicity this may be due to defect in
RNA synthesis [38] and disturbances in carbohydrate, protein, lipid
metabolisms due to acute liver injury induced by TAA [39].
Moreover, TAA injection significantly elevated liver MDA content
while, significantly decreased liver GSH content. This may be due to
chronic injection with TAA, a hepatotoxin containing thiono-sulfur
compound, which induce hepatic damage and fibrosis by generation of
ROS and suppressed antioxidant defense mechanism [40,41].
Meanwhile, TAA injection, showed a significant elevation in liver NO
content. This may be due to the formation of NO, which increases in
liver disease where L-arginine-NO pathway is activated [39,42].
Finally, TAA treatment caused a significant increase in hepatic Hyp
content, which supported by histopathological and histochemical
observations that showed the presence of fibrosis and numerous
connective tissue strands [43]. Hepatic fibrosis is the progressive
accumulation of ECM in hepatic tissue resulted from unbalance status
of ECM production and degradation [44]. The oxidative stress
activates HSCs, induces the secretion of growth factors and
profibrogenic cytokines that stimulates collagen synthesis [45].
The present study showed that treatment with GSE (100 and 200
mg/kg) significantly reduced serum AST, ALT, ALP, and TB levels
while, significantly increased serum TP level in a dose-dependent
manner, which attributed to GSE prevent the leakage of intracellular
enzymes [46].
Moreover, GSE (100 and 200 mg/kg) significantly reduced liver
MDA and NO contents and increased liver GSH content in a dose-
dependent manner [47].
GSE contains polyphenolic compounds such as procyanidins and
proanthocyanidins that have powerful free radical scavenging effect
[48,49]. Previous study demonstrated that GSE prevent hepatic fibrosis
and dysfunction caused by chronic arsenic administration in rats [50].
Citation: Nada SA, Gowifel AMH, El-Denshary EES, Salama AA, Khalil MG, et al. (2015) Protective Effect of Grape Seed Extract and/or
Silymarin Against Thioacetamide-induced Hepatic Fibrosis in Rats . J Liver 4: 178. doi:10.4172/2167-0889.1000178
Page 5 of 7
J Liver
ISSN:2167-0889 JLR, an open access journal Volume 4 • Issue 2 • 1000178

Meanwhile, GSE significantly reduced liver Hyp content in a dose-
dependent manner, our finding was supported by significant
improvement in histopathological and histochemical examinations,
this may be attributed to GSE inhibited HSCs activation, which
subsequently resulted in suppressed mRNA level of the α_1-(I)-
collagen and decreased collagen accumulation [51].
In the present study, treatment with silymarin (50 mg/kg)
significantly decreased serum AST, ALT, ALP, and total bilirubin
levels while, significantly elevated serum total protein level. These may
attributed to protection of liver cells directly through stabilizing the
cell membrane by preventing liver glutathione depletion and
inhibiting lipid peroxidation [52]. The pharmacological properties of
silymarin involve regulating cell membrane permeability and integrity,
inhibiting leukotriene, scavenging reactive oxygen species [53]. Abdel-
Salam et al. reported that the silymarin (22 mg/kg) decreased leakage
of hepatocellular enzymes ALT and AST into the plasma, decreased
serum levels of ALP and lessened the development of liver necrosis
and fibrosis caused by carbon tetrachloride (CCl4) [54].
Moreover, silymarin significantly decreased liver MDA and NO
contents while, significantly increased liver GSH content, compared to
TAA control group. This due to silymarin has antioxidant and
scavenging free radicals (ROS) activities, thus protecting against
oxidative stress. It augments the non-enzymatic and enzymatic
antioxidant defense systems of cells involving GSH, superoxide
dismutase and catalase. It can protect the liver, brain, heart and other
vital organs from oxidative damage for its ability to prevent lipid
peroxidation and replenishing the reduced glutathione levels.
Furthermore, silibinin exhibits membrane protective properties and it
may protect blood constituents from oxidative damage [55].
Silymarin administration significantly declined liver Hyp content
induced by TAA –treatment and there were significant improvement
in the histopathological and histochemical hepatic architecture. It was
found that silymarin inhibit stellate hepatocytes transformation to
myofibroblasts, which is responsible for collagen deposition in CCl4 -
induced hepatic fibrosis [56,57].
Combined administration of GSE and silymarin effectively
attenuate TAA-induced hepatic fibrosis in dose dependant manner
and significantly improved the tested biochemical parameters,
decreased hepatic Hpy content, prevent the oxidative stress and
restoring GSH level in experimental hepatic fibrosis.
Our results indicated the potential and beneficial role of GSE and
silymarin combinations in preventing oxidative stress-mediated
damage and strengthening antioxidant defense mechanism increased
antioxidant status of animals.
Conclusion
It is suggested that GSE and silymarin combination had synergistic
effect as antifibrotic therapy than single treatment with silymarin or
GSE alone and GSE (200 mg/Kg) combination with silymarin had a
good resultant effect.
Conflict of interest
The authors declared no conflicts of interest.
Funding
This research received no specific grant from any funding agency in
the public, commercial, or not-for-profit sectors.
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Citation: Nada SA, Gowifel AMH, El-Denshary EES, Salama AA, Khalil MG, et al. (2015) Protective Effect of Grape Seed Extract and/or
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Citation: Nada SA, Gowifel AMH, El-Denshary EES, Salama AA, Khalil MG, et al. (2015) Protective Effect of Grape Seed Extract and/or
Silymarin Against Thioacetamide-induced Hepatic Fibrosis in Rats . J Liver 4: 178. doi:10.4172/2167-0889.1000178
Page 7 of 7
J Liver
ISSN:2167-0889 JLR, an open access journal Volume 4 • Issue 2 • 1000178