Professional Documents
Culture Documents
2010; 6(12)
1. Introduction
Almost all clinically used antitumor drugs
exhibit toxic side effects affecting heart function.
Because of cardiotoxicity during anticancer
chemotherapy, effective doses of cytostatics have to
be limited, which may worsen antitumor efficacy.
Doxorubicin (Dox) is an anthracyclin antibiotic that is
considered as one of the most effective antitumor
agents. Dox is an essential component of treatment of
breast cancer (1), soft tissue sarcomas (2) and many
other cancers (3). The immense value of Dox in
treating a variety of malignant conditions is
unquestioned. However, the clinical use of Dox soon
proved to be hampered by such serious problems as
the development of resistance in tumor cells (4) and
toxicity in healthy tissues, in the form of central
nervous system toxicity (5), nephrotoxicity (6) and
most notably in the form of cardiomyopathy and
congestive heart failure (7). These adverse effects of
the drug can preclude its use in some patients and
limit the duration of its use in many others (8).
Carvedilol is non cardioselective -blocker
which lacks intrinsic sympathomimetic activity. In
addition, it has blocking effects at vascular 1receptors, antioxidant, and calcium antagonist
properties (9). The antioxidant activity of carvedilol
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Biochemical parameters:
The remainder of heart tissue of each rat was
weighed and homogenized in ice-cold saline for 1
minute, using an ice-cold Potter Elvejhem glass
homogenizer, forming 10% w/v homogenate.
Estimation of cardiac glutathione (GSH) content
A portion of homogenate was mixed with
ice-cold 7.5% sulfosalicylic acid in a ratio 1:1 and
centrifuged at 3000 r.p.m/15 minutes. The resulted
supernatant was used for the determination of GSH
(22), depending on the reaction between GSH and 5,
5'- dithio-bis, 2-nitrobenzoic acid to yield a stable
yellow
colour
which
can
be
measured
colourimetrically.
C- Experimental design:
Animals were divided into a normal control
group (10 rats), receiving the appropriate volume of
saline i.p, and Dox-treated group (22 rats). Dox was
dissolved in saline and injected i.p. as total cumulative
dose of 15 mg/kg, divided into 6 equal doses, each of
2.5 mg/kg. They were injected twice weekly/ 3 weeks
(16). The Dox-treated animals were divided into two
groups, one was kept without further treatment termed
Dox-group(12 rats), and a second group (10 rats),
termed Dox + carvedilol group, received carvedilol as
an i.p dose of 1mg/kg / 7 times over a period of 4
weeks including a dose before the 1st Dox dose (17).
Histopathological study:
Samples were taken from hearts and livers of rats in
different groups and fixed in 10% formol saline for 24
hours. Washing was done in tap water, then serial
dilutions of alcohol (methyl, ethyl and absolute ethyl)
were used for dehydration. Specimens were cleared in
xylene, embedded in paraffin at 56O C in hot air oven
for 24 hours. Paraffin bees wax tissue blocks were
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Statistical analysis
Results were analyzed statistically by oneway analysis of variance (ANOVA test) with
subsequent multiple comparisons using Tukey test.
Differences were considered statistically significant at
p less than 0.05. Results are presented as the mean
standard error of the mean (SEM), with the number of
observations (n) given in parentheses. All data
obtained were submitted to a computerized statistical
treatment using SPSS statistical package, version 17.
Tables were represented by Microsoft Excel computer
program.
3. Results:
Effect of doxorubicin (Dox), separately or in
combination with carvedilol, on serum lactate
dehydrogenase (LDH) and creatine phosphokinase
(CPK) activities in rats:
Results of table (1) revealed that Dox caused
significant increase in serum levels of LDH and CPK
amounting to 182.4% and 183.6%, respectively, as
compared to the normal values. Carvedilol coadministration caused normalization of LDH as well
as significant decrease in CPK serum levels reaching
119.4% of the control value.
Effect of doxorubicin (Dox), separately or in
combination with carvedilol,
on cardiac
malondialdehyde (MDA) and nitric oxide (NO)
levels in rats:
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Table (1): Effect of doxorubicin, separately or in combination with carvedilol, on serum lactate dehydrogenase
(LDH) and creatine phosphokinase (CPK) activities in rats:
Group
LDH( U/L)
CPK(U/L)
Control
581.1713 (9)
616.520.8 (9)
Doxorubicin
1060.229.4 a, c (10)
1131.932 a, c (10)
Doxorubicin + Carvedilol
598.523.6 b (9)
736.341.2 a, b (9)
Values are given as means SEM (No. of observations are given in parentheses)
a: Significant difference from control group at P<0.05
b: Significant difference from doxorubicin group at P<0.05
c: Significant difference from doxorubicin+
carvedilol group at P<0.05
Table (2): Effect of doxorubicin, separately or in combination with carvedilol, on cardiac malondialdehyde
(MDA) and nitric oxide (NO) levels in rats:
Group
MDA nmole /gm tissue
NO nmole / gm tissue
Control
57.61.9 (9)
17511 (9)
Doxorubicin
105.84.6 a, c (10)
31118 a, c (10)
Doxorubicin + Carvedilol
59.13.5 b (9)
1848 b (9)
Values are given as means SEM (No. of observations are given in parentheses)
a: Significant difference from control group at P<0.05
b: Significant difference from doxorubicin group at P<0.05
c: Significant difference from doxorubicin + carvedilol group at P<0.05
Table (3): Effect of doxorubicin, separately or in combination with carvedilol, on cardiac glutathione (GSH) content,
glutathione peroxidase (GPx), glutathione-S-tranferase (GST) and superoxide dismutase (SOD) activities in rats:
GSH
GPx
GST
SOD
Group
g/gm tissue
nmoles /min /mg protein
nmoles /min/mg protein U/mg protein
Control
64.31.5 (9)
33.61.9 (9)
29.61.0 (9)
21.31.4 (8)
Doxorubicin
41.21.1a,c (9)
137.87.1a,c (10)
54.72.9a,c (9)
48.12.2a,c (10)
Doxorubicin +Carvedilol
59.72.0a,b (9)
53.02.7a,b (8)
36.61.59 b (9)
32.31.8a,b (9)
Values are given as means SEM (No. of observations are given in parentheses)
a: Significant difference from control group at P<0.05
b: Significant difference from doxorubicin group at P<0.05
c: Significant difference from doxorubicin + carvedilol group at P<0.05
Table (4): Effect of doxorubicin, separately or in combination with carvedilol, on serum alanine aminotransferase
(ALT) activity in rats:
Group
ALT (U/L)
Control
52.41.8 (9)
Doxorubicin
60.21.5a,c (10)
Doxorubicin + Carvedilol
69.62.6a,b (9)
Values are given as means SEM (No. of observations are given in parentheses)
a: Significant difference from control group at P<0.05
b: Significant difference from doxorubicin group at P<0.05
c: Significant difference from doxorubicin + carvedilol group at P<0.05
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2010; 6(12)
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4. Discussion:
Doxorubicin (Dox) is a potent anticancer
drug that is used in treating both hematological and
solid tumors. However, severe cardiomyopathy and
heart failure have been observed in Dox-treated cancer
patients, which limit the clinical dosage of Dox in
cancer treatments (31). Oxidative stress is generally
held as the mediating mechanism in the multiple
biological processes leading to Dox cardiomyopathy,
e.g. redox mediated superoxide radical production
(32), tissue-specific mitochondrial DNA damage (33)
and disturbances of calcium (34) or iron (35). Results
of the present study revealed that 15 mg/kg total
cumulative dose of Dox induced cardiomyopathy and
hepatotoxicity manifested biochemically as significant
increase in serum levels of LDH; CPK and ALT. In
addition, Dox caused elevation in cardiac NO, MDA
levels, SOD, GPx, GST activities, and reduction in
GSH content. Histopathological examination of heart
and liver sections of Dox-treated animals supported
these biochemical results.
Results of table (1) revealed that Dox caused
significant increase in the serum levels of LDH and
CPK, considered as important markers of cardiac
injury. Many previous studies have demonstrated
similar elevations in cardiac enzymes activities in rats
following Dox administration (36, 37). Leakage of
cardiac enzymes directly correlates to ultrastructure
damage of heart tissues examined by electron
microscope. Dox-induced cardiomyopathy is mainly
attributed to increase oxidant production in heart.
Dox may undergo a one-electron reduction through a
metabolic activation by NADPH-cytochrome P-450
reductase. This reduction leads to the formation of the
free radical semiquinone, which in turn can produce a
variety of active ROS/RNS, including H2O2, OH and
ONOO (38). These species can attack the
cardiomyocyte
membrane,
damage
several
macromolecular cellular components, cause protein
and lipid peroxidation and consequently lead to
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5. References:
1
. Bo
na
d
o
nna G, Va
l
a
g
u
s
s
a P: Se
q
u
e
nt
i
a
lo
r
a
l
t
e
r
na
t
i
ng d
o
x
o
r
u
b
i
c
i
na
nd CMF r
e
g
i
me
nsi
n
b
r
e
a
s
tc
a
nc
e
rwi
t
hmo
r
et
ha
nt
hr
e
ep
o
s
i
t
i
v
eno
de
s
.
JAMA.1
9
9
6
;2
3
7
:
5
4
2
5
4
7
.
2
. Ba
c
c
iG,Fe
r
r
a
r
iS,De
l
e
p
i
neN:Pr
e
d
i
c
t
i
v
ef
a
c
t
o
r
s
o
fhi
s
t
o
l
o
g
i
cr
e
s
p
o
ns
et
op
r
i
ma
r
yc
he
mo
t
he
r
a
py
i
n o
s
t
e
o
s
a
r
c
o
ma o
fe
x
t
r
e
mi
t
y
. J.Clin.Oncol.
1
9
9
8
;
1
6
:6
5
8
6
6
3
.
3
. Ku
f
eDW,Ho
l
l
a
ndJ
F,Fr
e
iE:Ame
r
i
c
a
nCa
nc
e
r
So
c
i
e
t
y
:Ca
nc
e
rme
d
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c
i
ne6e
d
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t
i
o
n.Ed
i
t
e
db
yBC
De
c
k
e
r
,Ha
mi
l
t
o
n,2
0
0
3
:
3
4
4
3
6
7
.
4
. La
g
eM,He
l
b
a
c
hH,Di
e
t
e
lM,Sc
ha
d
e
ndo
r
fD:
o
p
o
i
s
o
me
r
a
s
eI
Ia
c
t
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v
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t
y
Mo
d
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l
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t
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no
fDNA t
a
n
de
x
p
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s
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o
n i
nme
l
a
no
ma c
e
l
l
s wi
t
h
r
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gr
e
s
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s
t
a
n
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e
.
Br J Cancer 2000;
a
c
q
u
i
r
e
d d
8
2
:
4
8
84
9
1
.
5
. Ta
ng
p
o
ngJ
,Co
l
eM,Su
l
t
a
naR:Ad
r
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a
my
c
i
nme
d
i
a
t
e
d ni
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t
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o
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ng
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ne
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s
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0
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5. Conclusion:
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.
Competing Interests:
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u
t
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r
s de
c
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a
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y ha
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e no
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s
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s
.
Authors' Contributions:
SSI
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HSH: c
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Acknowledgements:
Wea
r
et
ha
nkf
ult
o Dr
.Mo
ha
me
d A Al
i
,
As
s
i
s
t
a
ntp
r
o
f
e
s
s
o
ro
fHi
s
t
o
l
o
gy
,Fa
c
ul
t
yo
fMe
d
i
c
i
ne
,
Ca
i
r
oUni
v
e
r
s
i
t
y
,f
o
rhi
sp
r
o
f
e
s
s
i
o
na
lhe
l
pi
nc
a
r
r
y
i
ng
o
u
tt
heh
i
s
t
o
p
a
t
ho
l
o
gi
c
a
le
xa
mi
na
t
i
o
n.
Corresponding author
Sa
f
i
na
zS.I
b
r
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m
Bi
o
c
he
mi
s
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r
yDe
pa
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Ca
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Uni
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s
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,Ca
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,Egy
pt
h
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:
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u
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yB:I
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t
i
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n o
f
ma
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a
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de
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c
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