J. Nutr.
Effects
of Dexamethasone
Sci.
Vitaminol.,
on Experimental
in Cholesterol-fed
FUNAKI,
1992
Atherosclerosis
Rabbits
Michitaka NAITO, Masahiro YASUE, Kanichi Kazuyoshi YAMADA, Toshio HAYASHI, Masafumi Chiaki
38, 255-264,
Noboru
and Fumio
ASAI, KUZUYA,
YOSHIMINE,
KUZUYA
Department of Geriatrics; Nagoya University School of Medicine, Nagoya 466, Japan (Received December 26, 1991) Summary
We
studied
dexamethasone,
on
cholesterol-fed
rabbits.
(0.125 feeding
mg/day) a 1%
diet-induced was
in
as
normal
chow
the
diet no
injection to
inhibit
of
blood
into
structure
through
of
the
Key
Words
rich
diet,
aortic
1%
the
lipoprotein
intima,
or
experimental foam
involve
the
an of
involve
resulting
monocyte,
were
in
fed
with
the
drug
again
The
anti
inhibition
atherogenic in
a change
their
a
diet
observed
(ƒÀ-VLDL)
it may
the
was
insudation
density
into
rabbits
atherosclerosis. may
by
accumulation
cholesterol-rich
however,
of
and
lipoproteins,
endothelium
dexamethasone, hyperlipidemia,
progression
induced aggravated
it
dexamethasone-treated
lesions
dexamethasone
low
the
aortic
the
in
dexamethasone
cell
When
receiving
atherosclerotic
monocytes
the
the
(0.125mg/day);
of
mainly ƒÀ-very
experiments,
protein,
of
of
foam
animals.
after
of
aortic atherosclerosis 8 weeks, although less
steroid,
atherosclerosis
injection
lesions control
10 weeks
further
recruitment proteins,
the
dexamethasone
mechanism
and
intramuscular the for
adrenocortical
experimental
Histologically,
regression
atherogenic
a synthetic of
inhibited diet
with for
of
of
atherosclerotic
compared
8 weeks,
tended
Daily
remarkably cholesterol-rich
observed
daily
effects
development
hyperlipidemia.
rabbits
for
the
the
of lipo
the in
decreased
present the
size
passage
intima. atherosclerosis, cell, ƒÀ-very
cholesterol low
density
lipo
rabbit
Hypercholesterolemia is one of the most important risk factors for athero sclerosis (1, 2). Although cholesterol feedings are sufficient to produce atheroscl erotic changes in susceptible animal species such as rabbits and monkeys, in atherosclerosis-resistant species such as dogs and rats it is necessary to use other approaches, among them the production of mechanical injury to the endothelium, 255
256
M. NAITOet al.
the induction of hypertension, and the administration of anti-thyroid drugs, vitamin D, or foreign proteins (3). Although rabbits are relatively resistant to spontaneous atherosclerosis, the addition of 1g of cholesterol to the daily diet produces extensive atherosclerotic lesions in the aorta within a few months (4). In 1952, Oppenheimer and Bruger (5) showed that daily injections of cortisone inhibited the development of atherosclerosis in cholesterol-fed rabbits. An anti atherosclerotic effect was evident despite the exacerbation by cortisone of the diet-induced hyperlipidemia. Bailey and Butler (3) then reported that all of the steroids they examined, including fluorohydrocortisone, dexamethasone, methylp rednisolone, triamcinolone, prednisone and cortisone acetate, were similarly eff ective. This protective effect could be partially duplicated by various non-steroidal anti-inflammatory drugs including flufenamic acid, phenylbutazone, oxyphenyl butazone and mefenamic acid, although aminopyrine and aspirin were ineffective. Makheja et al. (6) recently reported that cortisone acetate (5mg daily for 3 months) reduced the formation of atherosclerotic plaques in Watanabe heritable hyperlipidemic (WHHL) rabbits, an animal model for familial hypercholesterole mia, although its administration increased plasma levels of cholesterol and trigl ycerides. In this study, we confirmed the inhibitory effect of dexamethasone on exper imental atherosclerosis in cholesterol-fed rabbits and examined the effect of this steroid on the regression of cholesterol-induced atherosclerosis. We also discuss the anti-atherosclerotic mechanism of steroids. MATERIALS ANDMETHODS Study design. Experiment 1 (Fig. 1): A total of 20 male New Zealand White (NZW) rabbits were selected according to body weight and serum lipid levels to form a homogenous group, and were then randomly allocated into study groups D and C. Both groups received an atherogenic diet (1% cholesterol-rich diet, 100g/ day) for 8 weeks. Group D received 0.125mg of dexamethasone (in 0.1ml vehicle) injected intramuscularly every day except Sunday for 8 weeks. The dose of dexamethasone was determined according to Bailey and Butler (3). Group C, the control group, received the same volume of vehicle (0.5mg sodium bisulfite, 1.5mg methyl p-hydroxybenzoate, 0.2mg propyl p-hydroxybenzoate, and 8.0mg creatin ine in 1ml of distilled water) during the 8-week period. One rabbit in group C died during the study and was excluded from analysis. The animals were sacrificed and autopsied after 8 weeks. Experiment 2 (Fig. 1): A total of 26 male NZW rabbits were fed a 1% cholesterol-rich diet (100g/day) for 8 weeks to induce atherosclerosis. The 26 rabbits were then divided into 3 groups so there would be no significant difference in serum total cholesterol levels. Group 1 (N=8) was sacrificed at 8 weeks. Groups 2 and 3 (N=9, respectively) were fed normal chow (100g/day) for an additional 10 weeks, the normal chow period, and were then sacrificed. During the J. Nutr.Sci. Vitaminol.
DEXAMETHASONE
Fig.
10-week
period,
0.125mg,
in
group 0.1ml
provided
by banyu
area
of
itizer of
the
Model
plaque
(%)
1000, as
analysis. from
hematoxylin-eosin lipid
Ins.
a
Corp., of
by
was
Japan). longitudinally,
their
plaques
computerized Japan).
plaque
injection.
(Decadron_??_)
atherosclerotic
with
proportion
Sudan
methods.
38, No.
The the and
staining,
area
internal
and
the
planimeter The
to
total (Dig
relative
deposition
aortic
area •~100
total
part
aortas
aortas elastica of
were
were
the
fixed
in
embedded
van aortas
in
10%
aqueous
paraffin,
formaldehyde
sectioned
and
stained
Gieson. were
fixed
in
95%
ethanol
and
stained
the
follow
III.
Laboratory
Vol.
calculated
dexamethasone,
only
injection
incised
of
of
vehicle
(Tokyo,
were
area
Watanabe the
injections
for
Co.
the
were
DT
Sections
For
ing
and
1 and 2.
2 received solution
aortas
257
(4).
solution.
with
The
expressed
Histological
with
group
Pharmaceutical
surfaces
No. was
of Experiments
phosphate
photographed, aortic
ATHEROSCLEROSIS
intramuscular
while
analysis.
were
Design
daily
vehicle, sodium
Morphological surfaces
1.
3 received of
Dexamethasone kindly
AND
determinations. Total
3, 1992
cholesterol,
Serum triglycerides,
lipid
levels and
were
phospholipid
measured were
by
determined
by
258
M. NAITO
enzymatic -in-Ca was
method method
kits
kit
isolated
by
Statistical
(Nippon
(Nippon
Statistical
Effects
of Dexamethasone
Sci.
Vitaminol.,
on Experimental
in Cholesterol-fed
FUNAKI,
1992
Atherosclerosis
Rabbits
Michitaka NAITO, Masahiro YASUE, Kanichi Kazuyoshi YAMADA, Toshio HAYASHI, Masafumi Chiaki
38, 255-264,
Noboru
and Fumio
ASAI, KUZUYA,
YOSHIMINE,
KUZUYA
Department of Geriatrics; Nagoya University School of Medicine, Nagoya 466, Japan (Received December 26, 1991) Summary
We
studied
dexamethasone,
on
cholesterol-fed
rabbits.
(0.125 feeding
mg/day) a 1%
diet-induced was
in
as
normal
chow
the
diet no
injection to
inhibit
of
blood
into
structure
through
of
the
Key
Words
rich
diet,
aortic
1%
the
lipoprotein
intima,
or
experimental foam
involve
the
an of
involve
resulting
monocyte,
were
in
fed
with
the
drug
again
The
anti
inhibition
atherogenic in
a change
their
a
diet
observed
(ƒÀ-VLDL)
it may
the
was
insudation
density
into
rabbits
atherosclerosis. may
by
accumulation
cholesterol-rich
however,
of
and
lipoproteins,
endothelium
dexamethasone, hyperlipidemia,
progression
induced aggravated
it
dexamethasone-treated
lesions
dexamethasone
low
the
aortic
the
in
dexamethasone
cell
When
receiving
atherosclerotic
monocytes
the
the
(0.125mg/day);
of
mainly ƒÀ-very
experiments,
protein,
of
of
foam
animals.
after
of
aortic atherosclerosis 8 weeks, although less
steroid,
atherosclerosis
injection
lesions control
10 weeks
further
recruitment proteins,
the
dexamethasone
mechanism
and
intramuscular the for
adrenocortical
experimental
Histologically,
regression
atherogenic
a synthetic of
inhibited diet
with for
of
of
atherosclerotic
compared
8 weeks,
tended
Daily
remarkably cholesterol-rich
observed
daily
effects
development
hyperlipidemia.
rabbits
for
the
the
of lipo
the in
decreased
present the
size
passage
intima. atherosclerosis, cell, ƒÀ-very
cholesterol low
density
lipo
rabbit
Hypercholesterolemia is one of the most important risk factors for athero sclerosis (1, 2). Although cholesterol feedings are sufficient to produce atheroscl erotic changes in susceptible animal species such as rabbits and monkeys, in atherosclerosis-resistant species such as dogs and rats it is necessary to use other approaches, among them the production of mechanical injury to the endothelium, 255
256
M. NAITOet al.
the induction of hypertension, and the administration of anti-thyroid drugs, vitamin D, or foreign proteins (3). Although rabbits are relatively resistant to spontaneous atherosclerosis, the addition of 1g of cholesterol to the daily diet produces extensive atherosclerotic lesions in the aorta within a few months (4). In 1952, Oppenheimer and Bruger (5) showed that daily injections of cortisone inhibited the development of atherosclerosis in cholesterol-fed rabbits. An anti atherosclerotic effect was evident despite the exacerbation by cortisone of the diet-induced hyperlipidemia. Bailey and Butler (3) then reported that all of the steroids they examined, including fluorohydrocortisone, dexamethasone, methylp rednisolone, triamcinolone, prednisone and cortisone acetate, were similarly eff ective. This protective effect could be partially duplicated by various non-steroidal anti-inflammatory drugs including flufenamic acid, phenylbutazone, oxyphenyl butazone and mefenamic acid, although aminopyrine and aspirin were ineffective. Makheja et al. (6) recently reported that cortisone acetate (5mg daily for 3 months) reduced the formation of atherosclerotic plaques in Watanabe heritable hyperlipidemic (WHHL) rabbits, an animal model for familial hypercholesterole mia, although its administration increased plasma levels of cholesterol and trigl ycerides. In this study, we confirmed the inhibitory effect of dexamethasone on exper imental atherosclerosis in cholesterol-fed rabbits and examined the effect of this steroid on the regression of cholesterol-induced atherosclerosis. We also discuss the anti-atherosclerotic mechanism of steroids. MATERIALS ANDMETHODS Study design. Experiment 1 (Fig. 1): A total of 20 male New Zealand White (NZW) rabbits were selected according to body weight and serum lipid levels to form a homogenous group, and were then randomly allocated into study groups D and C. Both groups received an atherogenic diet (1% cholesterol-rich diet, 100g/ day) for 8 weeks. Group D received 0.125mg of dexamethasone (in 0.1ml vehicle) injected intramuscularly every day except Sunday for 8 weeks. The dose of dexamethasone was determined according to Bailey and Butler (3). Group C, the control group, received the same volume of vehicle (0.5mg sodium bisulfite, 1.5mg methyl p-hydroxybenzoate, 0.2mg propyl p-hydroxybenzoate, and 8.0mg creatin ine in 1ml of distilled water) during the 8-week period. One rabbit in group C died during the study and was excluded from analysis. The animals were sacrificed and autopsied after 8 weeks. Experiment 2 (Fig. 1): A total of 26 male NZW rabbits were fed a 1% cholesterol-rich diet (100g/day) for 8 weeks to induce atherosclerosis. The 26 rabbits were then divided into 3 groups so there would be no significant difference in serum total cholesterol levels. Group 1 (N=8) was sacrificed at 8 weeks. Groups 2 and 3 (N=9, respectively) were fed normal chow (100g/day) for an additional 10 weeks, the normal chow period, and were then sacrificed. During the J. Nutr.Sci. Vitaminol.
DEXAMETHASONE
Fig.
10-week
period,
0.125mg,
in
group 0.1ml
provided
by banyu
area
of
itizer of
the
Model
plaque
(%)
1000, as
analysis. from
hematoxylin-eosin lipid
Ins.
a
Corp., of
by
was
Japan). longitudinally,
their
plaques
computerized Japan).
plaque
injection.
(Decadron_??_)
atherosclerotic
with
proportion
Sudan
methods.
38, No.
The the and
staining,
area
internal
and
the
planimeter The
to
total (Dig
relative
deposition
aortic
area •~100
total
part
aortas
aortas elastica of
were
were
the
fixed
in
embedded
van aortas
in
10%
aqueous
paraffin,
formaldehyde
sectioned
and
stained
Gieson. were
fixed
in
95%
ethanol
and
stained
the
follow
III.
Laboratory
Vol.
calculated
dexamethasone,
only
injection
incised
of
of
vehicle
(Tokyo,
were
area
Watanabe the
injections
for
Co.
the
were
DT
Sections
For
ing
and
1 and 2.
2 received solution
aortas
257
(4).
solution.
with
The
expressed
Histological
with
group
Pharmaceutical
surfaces
No. was
of Experiments
phosphate
photographed, aortic
ATHEROSCLEROSIS
intramuscular
while
analysis.
were
Design
daily
vehicle, sodium
Morphological surfaces
1.
3 received of
Dexamethasone kindly
AND
determinations. Total
3, 1992
cholesterol,
Serum triglycerides,
lipid
levels and
were
phospholipid
measured were
by
determined
by
258
M. NAITO
enzymatic -in-Ca was
method method
kits
kit
isolated
by
Statistical
(Nippon
(Nippon
Statistical
