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Cholesterol vehicle in experimental atherosclerosis Part II. Peanut oil
‘4 thevosclerosis Elsevier Publishing
Company,
CHOLESTEROL
Amsterdam
VEHICLE
PART 11. PEANUT
IN
EXPERIMENTAL
ATHEROSCLEROSIS
OIL
D. KRITCHEVSKY*, R. W. \VISSLEK
SHIRLEY
A. TEPPER,
The Wistar Institute of Anatomy and PathologJf, Pvitzkev School of Medicine, (Received
53
- Printed in The Netherlands
November
DR,%GOSLX\‘,1
VESSELINO\-ITCH
AND
Biology, Philadelphia, Penn. 19104, and Departwaerzt University of Chicago, Chicago, Ill. iU..S..-l.)
of
13th, 1970)
SUMMARY In 4 consecutive fed diets, containing
experiments,
young
adult,
male, Dutch
belted
6% of fats of varying fatty acid composition.
rabbits
coconut oil (CNO), corn oil (CO), peanut oil (PNO), and a special fat (PGF) to resemble cholesterol findings
peanut
oil minus arachidic
were then
acids. All diets contained
for 8 weeks.
the following
fed corn oil showed a slightly
with the group fed coconut the group fed coconut of this group;
simulated
Biochemical
2”d
and histological
compared.
In the 4 experiments, rabbits
and behenic
and were fed to the rabbits
were
These fats included
oil exhibiting
the liver weight
were generally
seen. The
the least weight gain. The liver weight of
oil was consistently
groups were comparable.
characteristics
greater weight gain than did the other groups, lowest, as was the liver cholesterol
and liver cholesterol
Serum cholesterol
levels observed
levels were highest
aortas showed the
groups fed the special fat or corn oil to have lesions of comparable or peanut
in the other 3
in the group fed the
special fat and those fed corn oil. Visual grading of the prestained severe than the lesions of the groups fed coconut
content
severity,
but less
oil, which were similar
to each other. The gross visual and microscopic indicated
that the rabbits
as the most frequent fat consistently
fed coconut
and severe,
showed
much
evaluations or peanut
lesions,
less aortic
of the severity
of aortic
oil had the most extensive,
In contrast, disease,
the animals
no matter
fed the
lesions as well special
how the disease
was
This work was supported in part by U.S. Public Health Service Research Grants HE-03299 and HE-12487 and by a Research Career Award, HE-0734, from the National Institute of Health; and a grant-in-aid from the National Dairy Council. * Wistar Professor of Biochemistry, Division of Animal Biology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Penn. (U.S.A.) . Atherosclerosis,
1971, 14: 53-64
54
D. KRITCHEVSKY,
evaluated;
generally
in the rabbits peanut
D. VESSELINOVITCH,
in these animals the aortic involvement
fed corn oil. The main characteristic
oil was a prominent
whereas
S. A. TEPPER,
in animals
intimal
fed coconut
proliferation
oil intimal
W. WISSLER
was similar to that seen
of the lesions seen in rabbits with a high proportion
proliferation
fed
of collagen,
was over the area of lipid
deposition. The trend in severity
and frequency
to that seen in the aorta. Intimal tic finding, regardless These
of the coronary
proliferation
artery
lesion was similar
with lipid deposition
was a characteris-
of the lipid fed.
findings
support
the concept
that
arachidic
and behenic
acids, which
are present in peanut oil, but not in the special fat, may be responsible, the unexpectedly
greater
atherogenic
suggest that other factors,
effect of peanut oil. However,
such as triglyceride
structure,
in part,
for
our observations
may also play an important
role in atherogenesis.
Key words:
Arachidic Peavmt
acid - Atherosclerosis
- Rehenic
acid - Cholesterol-fed
rabbit
-
oil
INTRODUCTION
Experimental
data from several
that the type of fat used in atherogenic scopic
character
atherogenicity
of the experimental
Peanut atherosclerosis peanut
and
his
that peanut
they
saturated properly peanut
coworkers
reported
fatty
acids,
by blending
and behenic
Atherosclerosis.
names
(lo%),
cottonseed
this material
arachidic
the composition
“PGF”.
were fed 2% cholesterol
* The trivial
and 25%
cholic
judging
rats
acid,
by
developed
thiouracil
and
fat was butter. have
reported monkeys
et al.5
WISSLER
fat for 52 weeks. it contains
5-6:/,
long-chain,
and behenic
(22:0)
acids*.
acids towards
the atherogenicity
of the fat that would result if peanut
To of oil
acids and found that such an oil could be safflower
A series
of eicosanoic
(35%),
of experiments
plus 6% of different
are used instead
1971, 14: 53-64
(20:0)
The
or corn oil when Rhesus
findingss.
of these fatty
found that
when the dietary
than butter
2% cholesterol
principally
oil, we estimated
designated
than would be expected
cholesterol,
similar
specie+8.
to its degree of unsaturation.
HOWARD~
oil differs from many other fats in that
assess the contribution
obtained
AND
thrombosis
oil was more atherogenic
were freed of its arachidic
rabbits
GRESHAM
exhibited
were fed a diet containing Peanut
correspond
when fed a diet containing
oil, whereas
SCOTT
diets may affect the gross visual and micro-
oil appears to be more atherogenic 3).
ours, have suggested
lesions in at least 4 different
of the fat did not necessarily
its iodine value alone (95 f
including
laboratories,
(20:0)
and olive (55%) was designed
oils. We in which
fats. The fats used were coconut
and docosanoic
(22:0)
acid.
CHOLESTEROL
T;\RLlT
VEHICLE
IN EXPERIMENTAL
ATHEROSCLEROSIS,
PART 11
1
FATTY ACID COMPOSITION OF COCONUT
8:O IO:0
10.0 6.1
I”:0 1J:O 16:O 16:l 18:O 18:l 18:2 18:3 20:o 22:o
16.7 18.5 9.0
OIL,
PEASUT
OIL,
CORN
OIL,
AK,)
l’(;F
-
9.7 0.2 1.9 46.1 35.6 1.2 1.6 3.8
11.1 0.2 0.8 29.5 56.0 0.8 0.9
2.6 5.6 0.5
a Calc’d for PNO minus 20:0 and 22:0. I) IO”,, cottonseed oil, 5536 olive oil, 350/, saftlowcr
0. 1 9.4 0.7 2.5 19.1 36.3 1.1
10.2 0.2 2.0 48.7 37.5 1.3
oil.
with these oil (CXO), corn oil (CO), peanut oil (PNO) and PGF. The experiments fats are the basis of this report. The goal was to correlate the presence or absence of lesions
with various
3IATERLtLS
biochemical
measurements
these studies.
ANT) 3fETHODS
One hundred-eighty
young
adult
(17OOG2000 g) were used in these studies. comparable
made during
groups
in all the experiments.
male
rabbits
The rabbits
of the
Dutch
were divided
In each experiment
belted
at random
the average
weight of the 4 groups was identical (+ 3 g). The diets were prepared commercial cholesterol in the appropriate oil and thoroughly mixing
strain into 4 starting
by suspending the suspension
with rabbit chow (Purina). The fatty acid composition of these oils is given in Table 1. The final mix contained 2% cholesterol and Sq/, of the added fat. The rabbits were fed the diets for 8 weeks; venous blood samples were then taken and the rabbits were killed. Complete post-mortem examinations were performed. Livers were removed and weighed, and a sample was taken for cholesterol analysis. The total cholesterol in liver and serum was determined by the method of M4Ny9. Tile aorta, heart and liver were examined microscopically. The aorta was removed frown the animal, cleaned of all adventitial fat, and graded visually for atherosclerotic lcsiona on a O-4 scale according to the scheme of DUFF AND Mc~IILLAs~~J. The parts of each aorta which were to be used for microscopic examination were pinned on wax blocks and fixed for 24 hours in 10% neutral formalin and then stained with Sudan IV according to the method of EGGEN et al.11. The areas of gross cardiovascular sudanophilia were estimated with the aid of a dissecting microscope and an evaluation of each area was made. The degree of aortic surface area involved was expressed as a percent of the total surface of the aorta. After grading, two blocks of aorta, one frcjm the ascending and the other from the middle part of thoracic aorta, were obtained A therosrzerosis,
1971,
14. 53 6J.
56
D.
TABLE
KRITCHEVSKY,
S. A. TEPPER,
D. VESSELINOVITCH,
W. WISSLER
2
INFLUENCE INDUCED
OF
COCONUT
OIL
ATHEROSCLEROSIS
Survival ratio
GrouP
PEANUT
(CNO), IN
OIL
(PNO),
Weight gain (g)
Liver weight (d
OIL
(CO)
AND
Cholesterol
PGF
ON
CHOLESTEROL
Atheromata
liver
sewm
(g/f00 Expt. 1 CNO
CORN
RABBITS&
(mg/lOO
d
RAEb
arch
thoracic
ml)
14114
335
113.3
3.57 f
1743 + 164
2.9
2.3
PNO PGF co
14/14 13/14 14/14
214 213 261
134.4 138.8 131.3
5.52 $= 0.42 4.75 + 0.37 5.90 * 0.36
1844 5 223 2193 & 207 1833 * 207
2.4 2.5 2.5
1.4 1.7 1.7
1.49 1.03 0.96 1.09
2 CNO PNO PGF co
lo/lo lo/lo lo/lo lo/lo
81 161 117 187
112.1 132.4 124.9 132.5
7.38 f 9.35 f
0.56 0.44 9.19 + 0.68 8.75 h 0.48
1055 961 1424 1324
& 99 h 135 f 182 & 137
1.6 1.6 1.3 1.4
1.2 1.2 1.1 1.2
1.33 1.46 0.84 0.98
3 CNO PNO PGF co
lo/lo lo/lo S/l0 lo/lo
112 239 217 329
100.4 119.1 132.5 124.4
4.15 6.70 6.32 3.85
& f f &
0.54 1.08 0.53 0.53
696 688 725 675
& & h *
121 123 139 116
1.7 2.0 1.4 1.2
1.2 1.3 0.9 0.7
2.08 2.40 1.59 1.41
4 CNO PNO PGF co
11111 lljll ll/ll 9/l 1
7 53 122 141
98.5 105.3 119.6 118.8
7.70 7.69 8.47 8.65
& * & *
0.59 0.65 0.44 0.76
1787 1469 1676 2017
* + f f
258 226 218 242
2.2 1.9 1.3 1.5
1.5 1.3 1.0 1.1
1.04 1.09 0.69 0.64
0.29c
Expt.
Exfit.
Expt.
a 2% cholesterol in 6% fat fed for 8 weeks. b Relative atherogenic effect. c Standard error.
for microscopic sections
evaluation.
were stained
aldehyde fuchsin.
When
with
Sections
possible,
oil red 0,
abdominal
aorta was also sampled.
hematoxylin-eosin,
of the heart embedded
and Gomori
in carbowax
The
trichrome
were stained similarly.
RESULTS AND DISCUSSION
Results
of the 4 experiments
effect
(RAE)
serum
cholesterol
relating
was calculated
in Table 2. The relative
the average
level (g/100 ml). Th is method
the level of cholesterol
experiments
are detailed
by dividing
with a variety
In the first experiment,
was found to be effective
with the degree of atherosclerotic the CNO-fed group exhibited
it became
effect of PNO was similar
that the atherogenic
and that both were considerably 1971, 14: 53-64
lesions in previous
more severe atherosclero-
groups.
Atherosclerosis,
by the in cor-
of fat@.
tic lesions than did the other 3 dietary evident
atherogenic
grade of atheromata
In the other experiments,
more atherogenic
than CO or PGF.
to that
however, of CNO
CHOLESTEROL
TABLE
VEHICLE
ATHEROSCLEROSIS,
PART
57
11
3
DISTRIBUTION OIL
IN EXPERIMENTAL
OFATHEROMATAIN
(PXO),
SPECIAL
FAT
cxo
G vade
(44145)
0~
CORN
a
OIL
(co)
(AVERAGE
arch
(42145)
4.0
5
3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0
5 2 6 7 9 7 3
1 2 3 6 5 10 10 7
3 7 11 5 7 6
1 4 7 4 17 10 1
2 2 3 3 10 6 5 11 -
:\vcrage
2.15 & 0.16”
1.67 & 0.14
2.00 + 0.16
1.31 * 0.12
1.69 * 0.16
.\verage serum cholesterol (mg/lOO ml) RAEc a Survival IJ Standard r Relative
ratio. error. atherogenic
The
1 -
1360 1.40
of atherosclerotic experiments
There were no definite,
lesions
1 3 2 4 3 15 14 -
2 3 ‘2 5 13 5 9 -
4 1 2 5 13 17 -
1.2-t + 0.13
1.71 $I 0.16
1.13 f 0.12
1475 0.94
diet, is CNO > PNO
differences
fed cholesterol
liver cholesterol
levels in the CNO-fed
(7.08 g/100 g). The average
microscopic
Atherosclerotic
group (5.63 g/100 g) were lower
total
evaluation
lesions mainly
as compared
These are the combined separately,
(7.16 g/100 g), CO (6.82
liver cholesterol
levels for each
9.15 g.
to 21%
rabbits
lesions before
the aortic
and
and frequency arch.
Parts
of
of the
but to a less severe degree.
and 2776 of the intimal and 22%
results of all 4 experiments;
the peanut oil (PNO)-fed
in aortic
of the severity
affected
aorta were also involved,
In animals fed PNO and CNO, 29% ved, respectively,
(168 g)
(109.0 g) was slightly lower
fed PNO
by gross visualization
aortic
and abdominal
PGlT.
j&dings
findings
as well as a microscopic
lesions.
> CO =
plus PNO (123.4 g), PGF (129.3 g), or CO (130.4 g).
levels found in the rabbits
4 presents
the
of the 4
the average weight gain of the CO-fed group (240
after staining, thoracic
reflects
in weight gain or in liver weight.
group were: CNO, 6.19 g; PNO, 8.84 g; CO, 8.89 g; and PGF,
Table
--
higher than those of CNO- (152 g), PNO- (168 g), or PGF-fed
than liver cholesterol
Gross visual and
thovaclc
in all 4 experiments
groups. The average liver weight of the CNO-fed rabbits
g/100 g), or PGF
arch
(Table 3). The order of atherogenicity
consistent
over the 4 experiments,
The average
tikoracic
1564 0.94
fats, when added to a cholesterol-containing
than in the rabbits
(42145)
effect.
distribution
In general,
co
3
1300 1.27
results of the individual
g) was somewhat
4 EXPERIMENTS)
arch __~~
thoracic
____. 6
05
PGF
(45145)
PNO
fkoracic
arch
2 5; CHOLESYEROLhNDCOCONUTOIL(CNO),P&ANU'~
RABBITSFED
(PGF)
surface was invol-
in the CO- and PGF-fed if each experiment
rabbits.
was treated
still had the most severe atherosclerosis Athemclerosis,
1971,
14: 53 -6-l
58
n.
TABLE
KRITCHEVSKY,
S. A. TEPPER,
D. VESSELINOVITCH,
W. WISSLER
4
EFFECT
OF
DIETARY
FATS
(6%)
ON
CHOLESTEROL
(2°/o)-INDUCED
ATHEROMATA
(AVERAGE
4
OF
EXPERIMENTS)
Coconut (CNO) Peanut (PNO) PGF Corn (CO)
3.
Gross visual
No. of animals
Fat
arch
44 45 42 42
thoracic
2.15 2.00 1.69 1.71
+ * 5 +
0.16 0.16 0.16 0.16
1.67 1.31 1.24 1.13
& i_ + *
0.14 0.12 0.13 0.12
Gross visual stained aorta
Microscopic of lesions
(area 76)
zmolved
frequency
27 29 22 21
4.1 4.4 4.4 4.6
evaluation
severitya
( 76)
& & & *
50 56 38 35
36 37 20 20
Sum of severity Number
x
of animals
10.
whereas atherosclerosis visual differences trend
toward
animals.
greater
Even
was least apparent
in surface area involved involvement
visualized
lesions in PNO-fed
grossly,
in the CO-fed animals. Although between
was seen in the aorta the character
animals were raised and sometimes
had lesions, they were small, pale and only slightly Microscopic was carried
were observed monstrable
examination
out on standard in PNO-
of the severity
and CNO-fed
histochemically
the aortas of the rabbits In the rabbits high proportion
sections.
fed PNO a very prominent deposition
lesion in PNO-fed abundant
(Table
varied
with the type lesions in
intimal proliferation
by predominantly
Gomori rabbits
trichrome
as compared
with particularly to the rabbits
aorta of a PNO-fed
cellular proliferation,
fed
rabbit can be
composed
of a cap
of lipid giving the whole lesion a dense
aldehyde
fuchsin
stain
was used, the aortic
showed that the smooth muscle cells were separated
by the
collagen and elastin in which they were embedded.
The animals fed CNO had various sized, proliferative, on microscopic
examination,
revealed
remarkable
intimal
pale yellow lesions which, proliferation
characterized
by less collagen and elastin and more intracellular
lipid. Much of the stainable
in both types of lesions appeared
deposited
cells of innermost
media
(Fig.
to be diffusely
2). Lipid deposition
in or on smooth
inside pre-existing
in aorta was present in all of the groups, but was most frequent Except
4)
and severe lesions
of the atherosclerotic
was observed,
tissue with a small amount When
appearance.
of lesions
lesions showed lipid, de-
but the amount
other fats. In Fig. 1, a typical lesion in the thoracic of fibro-elastic
group but less severe.
All aortic
in the characteristics
CO-fed rabbits
Lesions of the PGF-fed
and frequency
rabbits.
The
fed PNO and CNO was seen consistently.
of collagen
seen. It is characterized
flat. Although
elevated.
The most frequent
with oil red 0,
and CNO-fed
very thick, while in the CNO-
to those in the PNO-fed
aortic
of fat in the diet. A difference
of PNO-
of the lesions seemed different.
fed animals the lesions were pale yellow and relatively group were similar in appearance
the gross
the groups were small, a definite
for these differences,
Atherosclerosis,
1971,
14: 53-64
there were histological
features
lipid muscle
medial cells
in CNO-fed animals. common
to both
the
CHOLI’STEROL
VEHICLE
IK
EXPERIMENTAL
ATHEROSCLEROSIS,
PART
11
I;ig. 1. Photomicrograph of a lesion in the thoracic aorta from a I’S@fed rabbit. The ksion ii charactc~riactl by pronounced proliferation with abundant intracellular mat&al which 15 pr,‘tlominrtntly collagen with some elastin. Oil red 0; x 195, rcduwd in rcprorluction L 0.75.
Fig. 2. Photomicrograph deposition was a frequent < 0.75.
of thoracic aortic lesion in a C?TO-fed rabbit. Proliferation over lipid finding in CSO-fed animals. Oil red 0; ’ 195, reduced in wprodwtlon
60
D. KRITCHEVSKY,
S. A. TEPPER, D. VESSELINOVITCH,
PNO- and CNO-fed groups. The lesions in PGF-fed
animals
W. WISSLER
also had some similarities
with those of the PNO-fed rabbits, but they were less severe (Fig. 3). The lesions in the CO-fed rabbits were characterized by less lipid deposition in pre-existing cells and by mild intimal A bar-graph
proliferation
(Fig. 5) summarizes
of the histopathological taken inside
animals.
pre-existing
the results
characteristics
from each aorta. A tendency
in PNO-fed
(Fig. 4). of microscopic
of the lesions
toward
collagen deposition
On the other hand, a more consistent medial
based
cells was seen in aortas
grading
of several
on standard
sections
was most pronounced
increase
of CNO-fed
in lipid deposition rabbits,
giving
the
appearance of a softer atheroma. Table 5 presents the frequency and severity evaluated microscopically of the lesions in the coronary arteries. The occurrence of lesions was highest in the group fed PNO and lowest in the group fed CO. The more advanced
coronary
artery
lesions
in PNO-fed rabbits were characterized by intimal proliferation and lipid deposition in both the intima and media. Lipid-rich proliferative intima sometimes resulted in severe
stenosis
coronary
of the intramural
coronary
vessels.
Qualitative
lesions were not great in spite of the different
Myocardial
infarction
cated on the endocardial third of the wall thickness. not completely
occluded,
was observed
in one rabbit
side of the left ventricular The coronary exhibited
differences
among
fats fed. fed PGF. The infarct wall and involved
artery in the vicinity
severe atherosclerotic
almost
of the infarct,
lesions.
was loone-
although
Most of the fibers
Fig. 3. Photomicrograph of thoracic aortic lesion of a PGF-fed animal. It is similar to the lesions produced by peanut oil, only less severe. Oil red 0; x 195, reduced in reproduction x 0.75. Atherosclerosis,
1971,
14: 53-64
CHOLESTEROL
VEHICLE
IGg. 1. l’hotomicrograph existing ~11s and mild duction i 0.75.
SMRITY
OF
IN EXPERIMEKTAL
of thoracic aortic intimal proliferation
AORTIC YICROSCOPIC
lizI COCONUT
fgj
PEANUT
ATHEROSCLEROSIS,
lesion in a CO-fed rabbit. wew seen. Oil ret1 0;
LESIONS
I!llI
FROM THREE
PGF
61
11
Deposition of lipid in prc~: 195, rcduccd in rcprc,-
STANDARD AREAS
CORN OIL
2.5 ??
PART
a
.
ASCENDINQ
??
THORACIC
A ABDOMINAL
1
k $
s
1.0
x
0.5
0
INT.
PROLIFER~ION
INT.
LIPIDS
LIPID
Fig. 5. Bar graph illustrates microscopic grading Imsvtl on standard sections taken in each aorta.
IN MEDIA
of several
FIBROUS CAP
characteristics
of the aortic
.4t/1erosc1erosis,
lesions,
1971, 14: 5x 6-i
62
D. KRITCHEVSKY,
TABLE
S. A. TEPPER,
D. VESSELINOVITCH,
W. WISSLER
5
MICROSCOPIC
LESIONS
IN
CORONARY
Treatwent
Group
1
Microscopic
coconut oil peanut oil PGF corn oil
II III
IV
were coagulated cation between a shadowy
ARTERIES
evaluation of lesions
frequency
severity
31 42 31 21
13 28 14 10
and their nuclei had disappeared. normal and ischemic
outline.
Inflammatory
There
tissue. Architecture
exudation
infiltrated
was a fairly good demar-
was visible in some parts as only partly into the necrotic
focus (Fig. 6), Fatty Although
liver changes
were observed
there was no remarkable
was a slight tendency and slightly
in most animals
difference
regardless
among the dietary
of the diet fed.
groups,
still there
for the fatty changes to be more severe in the PGF-fed
rabbits
milder in the CO-fed animals.
Pathologists of atherosclerosis.
have been puzzled about the factors responsible It appears
aortic wall may, at least partly,
that the chemical determine
lesions, as well as the rate of progression
nature
the quantity
for the progression
of the lipid deposited and quality
in the
of atherosclerotic
from fatty streak to fibrous plaque.
Fig. 6. Photomicrograph of the heart of a PGF-fed animal. Most of the fibers are coagulated their nuclei have disappeared. H.-E.; x 195, reduced in reproduction X 0.75. Atherosclerosis,
1971, 14: 53-64
and
63
CHOLESTEROL VEHICLE IS EXPERIMENTAL ATHEROSCLEROSIS, PART 11 There are several possible ways to explain the pathogenetic sclerosis
caused by the various food fats. One possibility
may be metabolized There
more effectively
may be either a greater
or a relatively consistent position
retarded
accumulation of stainable
of CSO-fed
of the exogenous
of stainable
suggests
bolism of lipid, particularly A further
absorption
aortic
that
that brought
lesions.
Only further
The results the rabbits
responsible
experimentation
the obvious suggestion
reaction
plus the 4 different
of coconut
of the
examinations
of the aortas
of
fats would, at first, appear to uphold
from its level of unsaturation
First is that the lesions of PGF-fed
is the pre-
must also be important
105; CO, 126. While PGF has
similar to that of PNO, the structure different
of this fat. Hence,
to cholesterol
some aspect
absorption,
varies;
however,
transport
the percentage
was calculated
of the component
from PNO since 3 different
ANI) VOLPENHEIN14,15 have shown that the percentage the oils used in these experiments tion of the triglycerides,
shown, in a
corn oil and corn oil, that
in fact, it is not. The iodine values of the 3 fats
must be considerably
were used for preparation
morphologic-
we have previously
hydrogenated
were: CNO, 8; PNO, 95; PGF,
a gross fatty acid composition
rabbits
This might be explained
with iodine value 7.8. On this basis, PGF should be
than CO, whereas,
of PGF
However,
oil, lard, partially
may be correlated
used in this experiment triglycerides
This reaction
morphology
and behenic acids. However, several factors suggest a more complex
on the basis of the level of unsaturation.
more atherogenic
within cells of the
that the factor that causes peanut oil to exert an atherogenic
reason for our observations.
atherogenicity
in the form
will resolve this question.
ally resemble the lesions of PNO- rather than CO-fed rabbits. comparison
in the meta-
than others.
for the characteristic
effect more severe than would be expected sence of arachidic
This de-
in the smooth muscle cells
cells may be critical
of the gross visual and microscopic
fed cholesterol
with the
rabbits.
to them by the blood stream
aortic wall may lead to a more severe metabolic partly
cholesterol
associated
is that some types of lipids when deposited
at least
wall than others.
before by GEER et al.12 and WISSLER tit &.I”.
This has been suggested
possibility
is probably
cholesterol
lipid in the CNO-fed
these arterial
of athero-
of the dietary
lipid in the aortic walls, particularly
animals,
of lipoproteins.
by the liver and the artery
gastrointestinal
catabolism
patterns
is that some types of lipid
of triglyceride and deposition.
of component
oils
structure MATTSON
fatty
acids in
of fatty acids in the Z-posi-
to be comparable
for PNO and
PGF. The primary
fatty
fats used were oleic (18:l) 95q;, of all the fatty position
acids present in the 2-position and linoleic (18:2).
acids present
in this position.
of CO and PNO triglycerides
to be 0.88. Thus there is little difference
glycerides
which arise during digestion
the structure
of the intact
fatty
acids that arise on lipolysis
may exert
respectively;
in the structure
and absorption
triglyceride
of all the
acids represented
The ratio of 18:1/18:2
is 0.36 and 0.59,
calculated then,
of the triglycerides
These 2 fatty
in the 2-
for PGF
it is
of the P-mono-
of PNO and PGF.
may be important
about
Possibly,
or the specific
free
an effect.
‘ltherosczerosis, 1971, 14: 53 ~6.l
64
D. KRITCHEVSKY,
S. A. TEPPER,
D. VESSELINOVITCH,
W. WISSLER
ACKNOWLEDGEMENTS
We are deeply indebted to Doctor Fred H. Mattson of the Proctor and Gamble Company for his assistance in providing one of the oils used (PGF) and for mrny helpful discussions,
REFERENCES 1
2
3
KRITCHEVSKY, D., A. W. MOYER, W. C. TESAR, R. F. J. MCCANDLESS, J. B. LOGAN, R. A. BROWN AND M. E. ENGLERT, Cholesterol vehicle in experimental atherosclerosis, Part 2 (Influence of unsaturation), Amer. J. Physiol., 1956, 185: 279-280. GRESHAM, G. A. AND A. N. HOWARD, The independent production of atherosclerosis and thrombosis in the rat, &it. J. Exp. Pathol., 1960, 41: 395-402. SCOTT, R. F., E. S. MORRISON, W. A. THOMAS, R. JONES AND S. C. NAM, Short-term feeding of unsaturated versus saturated fat in the production of atherosclerosis in the rat, Exp. Mol. Pathol.,
4
5
0
7 8
9 IO
11
12
13
14
15
1964,
3: 421-443.
IMAI, H., K. T. LEE, S. PASTORI, E. PANLILIO, R. FLORENTIN AND W. A. THOMAS, Atherosclerosis in rabbits: architectural and subcellular alteration of smooth muscle cells of aortas in response to hyperlipemia, Exp. Mol. Pathol., 1966, 5: 273-310. WISSLER, R. W., D. VESSELINOVITCH, G. S. GETZ AND R. H. HUGHES, Aortic lesions and blood lipids in Rhesus monkeys fed three food fats, Fed. Proc., 1967, 26: 371. FLORENTIN, R. A. AND S. C. NAM, Dietary-induced atherosclerosis in miniature swine, Part 1 (Gross and light microscopy observations. Time of development and morphologic characteristics of lesions), Exp. Mol. Pathol., 1968, 8: 263-277. KRITCHEVSKY, D. AND S. A. TEPPER, Cholesterol vehicle in experimental atherosclerosis, Part 7 (Influences of naturally occurring saturated fats), Med. Pharmacol. Exp., 1965, 12: 315320. KRITCHEVSKY, D. AND S. A. TEPPER, Cholesterol vehicle in experimental atherosclerosis, Part 8 (Effect of a medium chain triglyceride (MCT)), Exp. Mol. Pathol., 1965, 4: 489-499. MANN, G. V., A method for measurement of cholesterol in blood serum, Cl&. Chem., 1961, 7 : 275-284. DUFF, G. L. AND G. C. MCMILLAN, The effect of alloxan diabetes on experimental cholesterol atherosclerosis in the rabbit, J. Exp. Med., 1949, 89: 611-630. EGGEN, D. A., J. P. STRONG AND H. C. MCGILL, JR., An objective method for grading atherosclerotic lesions, Lab. Invest., 1962, 11: 732-742. GEER, J. C., H. C. MCGILL, JR. AND J. P. STRONG, The fine structure of human atherosclerotic lesions, Amer. J. Pathol., 1961, 38: 263-287. WISSLER, R. W., L. E. FRAZIER, R. H. HUGHES AND R. A. RASMUSSEN, Atherogenesis in the cebus monkey, Arch. Pathol., 1962, 74: 312-322. MATTSON, F. H. AND R. A. VOLPENHEIN, The specific distribution of fatty acids in the glycerides of vegetable fats, J. Biol. Chem., 1961, 236: 1891-1894. MATTSON, F. H. AND R. A. VOLPENHEIN, The specific distribution of unsaturated fatty acids in the triglycerides of plants, J. Lipid Res., 1963, 4: 392-396.
Atherosclerosis,
1971, 14: 53-64
Company,
CHOLESTEROL
Amsterdam
VEHICLE
PART 11. PEANUT
IN
EXPERIMENTAL
ATHEROSCLEROSIS
OIL
D. KRITCHEVSKY*, R. W. \VISSLEK
SHIRLEY
A. TEPPER,
The Wistar Institute of Anatomy and PathologJf, Pvitzkev School of Medicine, (Received
53
- Printed in The Netherlands
November
DR,%GOSLX\‘,1
VESSELINO\-ITCH
AND
Biology, Philadelphia, Penn. 19104, and Departwaerzt University of Chicago, Chicago, Ill. iU..S..-l.)
of
13th, 1970)
SUMMARY In 4 consecutive fed diets, containing
experiments,
young
adult,
male, Dutch
belted
6% of fats of varying fatty acid composition.
rabbits
coconut oil (CNO), corn oil (CO), peanut oil (PNO), and a special fat (PGF) to resemble cholesterol findings
peanut
oil minus arachidic
were then
acids. All diets contained
for 8 weeks.
the following
fed corn oil showed a slightly
with the group fed coconut the group fed coconut of this group;
simulated
Biochemical
2”d
and histological
compared.
In the 4 experiments, rabbits
and behenic
and were fed to the rabbits
were
These fats included
oil exhibiting
the liver weight
were generally
seen. The
the least weight gain. The liver weight of
oil was consistently
groups were comparable.
characteristics
greater weight gain than did the other groups, lowest, as was the liver cholesterol
and liver cholesterol
Serum cholesterol
levels observed
levels were highest
aortas showed the
groups fed the special fat or corn oil to have lesions of comparable or peanut
in the other 3
in the group fed the
special fat and those fed corn oil. Visual grading of the prestained severe than the lesions of the groups fed coconut
content
severity,
but less
oil, which were similar
to each other. The gross visual and microscopic indicated
that the rabbits
as the most frequent fat consistently
fed coconut
and severe,
showed
much
evaluations or peanut
lesions,
less aortic
of the severity
of aortic
oil had the most extensive,
In contrast, disease,
the animals
no matter
fed the
lesions as well special
how the disease
was
This work was supported in part by U.S. Public Health Service Research Grants HE-03299 and HE-12487 and by a Research Career Award, HE-0734, from the National Institute of Health; and a grant-in-aid from the National Dairy Council. * Wistar Professor of Biochemistry, Division of Animal Biology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Penn. (U.S.A.) . Atherosclerosis,
1971, 14: 53-64
54
D. KRITCHEVSKY,
evaluated;
generally
in the rabbits peanut
D. VESSELINOVITCH,
in these animals the aortic involvement
fed corn oil. The main characteristic
oil was a prominent
whereas
S. A. TEPPER,
in animals
intimal
fed coconut
proliferation
oil intimal
W. WISSLER
was similar to that seen
of the lesions seen in rabbits with a high proportion
proliferation
fed
of collagen,
was over the area of lipid
deposition. The trend in severity
and frequency
to that seen in the aorta. Intimal tic finding, regardless These
of the coronary
proliferation
artery
lesion was similar
with lipid deposition
was a characteris-
of the lipid fed.
findings
support
the concept
that
arachidic
and behenic
acids, which
are present in peanut oil, but not in the special fat, may be responsible, the unexpectedly
greater
atherogenic
suggest that other factors,
effect of peanut oil. However,
such as triglyceride
structure,
in part,
for
our observations
may also play an important
role in atherogenesis.
Key words:
Arachidic Peavmt
acid - Atherosclerosis
- Rehenic
acid - Cholesterol-fed
rabbit
-
oil
INTRODUCTION
Experimental
data from several
that the type of fat used in atherogenic scopic
character
atherogenicity
of the experimental
Peanut atherosclerosis peanut
and
his
that peanut
they
saturated properly peanut
coworkers
reported
fatty
acids,
by blending
and behenic
Atherosclerosis.
names
(lo%),
cottonseed
this material
arachidic
the composition
“PGF”.
were fed 2% cholesterol
* The trivial
and 25%
cholic
judging
rats
acid,
by
developed
thiouracil
and
fat was butter. have
reported monkeys
et al.5
WISSLER
fat for 52 weeks. it contains
5-6:/,
long-chain,
and behenic
(22:0)
acids*.
acids towards
the atherogenicity
of the fat that would result if peanut
To of oil
acids and found that such an oil could be safflower
A series
of eicosanoic
(35%),
of experiments
plus 6% of different
are used instead
1971, 14: 53-64
(20:0)
The
or corn oil when Rhesus
findingss.
of these fatty
found that
when the dietary
than butter
2% cholesterol
principally
oil, we estimated
designated
than would be expected
cholesterol,
similar
specie+8.
to its degree of unsaturation.
HOWARD~
oil differs from many other fats in that
assess the contribution
obtained
AND
thrombosis
oil was more atherogenic
were freed of its arachidic
rabbits
GRESHAM
exhibited
were fed a diet containing Peanut
correspond
when fed a diet containing
oil, whereas
SCOTT
diets may affect the gross visual and micro-
oil appears to be more atherogenic 3).
ours, have suggested
lesions in at least 4 different
of the fat did not necessarily
its iodine value alone (95 f
including
laboratories,
(20:0)
and olive (55%) was designed
oils. We in which
fats. The fats used were coconut
and docosanoic
(22:0)
acid.
CHOLESTEROL
T;\RLlT
VEHICLE
IN EXPERIMENTAL
ATHEROSCLEROSIS,
PART 11
1
FATTY ACID COMPOSITION OF COCONUT
8:O IO:0
10.0 6.1
I”:0 1J:O 16:O 16:l 18:O 18:l 18:2 18:3 20:o 22:o
16.7 18.5 9.0
OIL,
PEASUT
OIL,
CORN
OIL,
AK,)
l’(;F
-
9.7 0.2 1.9 46.1 35.6 1.2 1.6 3.8
11.1 0.2 0.8 29.5 56.0 0.8 0.9
2.6 5.6 0.5
a Calc’d for PNO minus 20:0 and 22:0. I) IO”,, cottonseed oil, 5536 olive oil, 350/, saftlowcr
0. 1 9.4 0.7 2.5 19.1 36.3 1.1
10.2 0.2 2.0 48.7 37.5 1.3
oil.
with these oil (CXO), corn oil (CO), peanut oil (PNO) and PGF. The experiments fats are the basis of this report. The goal was to correlate the presence or absence of lesions
with various
3IATERLtLS
biochemical
measurements
these studies.
ANT) 3fETHODS
One hundred-eighty
young
adult
(17OOG2000 g) were used in these studies. comparable
made during
groups
in all the experiments.
male
rabbits
The rabbits
of the
Dutch
were divided
In each experiment
belted
at random
the average
weight of the 4 groups was identical (+ 3 g). The diets were prepared commercial cholesterol in the appropriate oil and thoroughly mixing
strain into 4 starting
by suspending the suspension
with rabbit chow (Purina). The fatty acid composition of these oils is given in Table 1. The final mix contained 2% cholesterol and Sq/, of the added fat. The rabbits were fed the diets for 8 weeks; venous blood samples were then taken and the rabbits were killed. Complete post-mortem examinations were performed. Livers were removed and weighed, and a sample was taken for cholesterol analysis. The total cholesterol in liver and serum was determined by the method of M4Ny9. Tile aorta, heart and liver were examined microscopically. The aorta was removed frown the animal, cleaned of all adventitial fat, and graded visually for atherosclerotic lcsiona on a O-4 scale according to the scheme of DUFF AND Mc~IILLAs~~J. The parts of each aorta which were to be used for microscopic examination were pinned on wax blocks and fixed for 24 hours in 10% neutral formalin and then stained with Sudan IV according to the method of EGGEN et al.11. The areas of gross cardiovascular sudanophilia were estimated with the aid of a dissecting microscope and an evaluation of each area was made. The degree of aortic surface area involved was expressed as a percent of the total surface of the aorta. After grading, two blocks of aorta, one frcjm the ascending and the other from the middle part of thoracic aorta, were obtained A therosrzerosis,
1971,
14. 53 6J.
56
D.
TABLE
KRITCHEVSKY,
S. A. TEPPER,
D. VESSELINOVITCH,
W. WISSLER
2
INFLUENCE INDUCED
OF
COCONUT
OIL
ATHEROSCLEROSIS
Survival ratio
GrouP
PEANUT
(CNO), IN
OIL
(PNO),
Weight gain (g)
Liver weight (d
OIL
(CO)
AND
Cholesterol
PGF
ON
CHOLESTEROL
Atheromata
liver
sewm
(g/f00 Expt. 1 CNO
CORN
RABBITS&
(mg/lOO
d
RAEb
arch
thoracic
ml)
14114
335
113.3
3.57 f
1743 + 164
2.9
2.3
PNO PGF co
14/14 13/14 14/14
214 213 261
134.4 138.8 131.3
5.52 $= 0.42 4.75 + 0.37 5.90 * 0.36
1844 5 223 2193 & 207 1833 * 207
2.4 2.5 2.5
1.4 1.7 1.7
1.49 1.03 0.96 1.09
2 CNO PNO PGF co
lo/lo lo/lo lo/lo lo/lo
81 161 117 187
112.1 132.4 124.9 132.5
7.38 f 9.35 f
0.56 0.44 9.19 + 0.68 8.75 h 0.48
1055 961 1424 1324
& 99 h 135 f 182 & 137
1.6 1.6 1.3 1.4
1.2 1.2 1.1 1.2
1.33 1.46 0.84 0.98
3 CNO PNO PGF co
lo/lo lo/lo S/l0 lo/lo
112 239 217 329
100.4 119.1 132.5 124.4
4.15 6.70 6.32 3.85
& f f &
0.54 1.08 0.53 0.53
696 688 725 675
& & h *
121 123 139 116
1.7 2.0 1.4 1.2
1.2 1.3 0.9 0.7
2.08 2.40 1.59 1.41
4 CNO PNO PGF co
11111 lljll ll/ll 9/l 1
7 53 122 141
98.5 105.3 119.6 118.8
7.70 7.69 8.47 8.65
& * & *
0.59 0.65 0.44 0.76
1787 1469 1676 2017
* + f f
258 226 218 242
2.2 1.9 1.3 1.5
1.5 1.3 1.0 1.1
1.04 1.09 0.69 0.64
0.29c
Expt.
Exfit.
Expt.
a 2% cholesterol in 6% fat fed for 8 weeks. b Relative atherogenic effect. c Standard error.
for microscopic sections
evaluation.
were stained
aldehyde fuchsin.
When
with
Sections
possible,
oil red 0,
abdominal
aorta was also sampled.
hematoxylin-eosin,
of the heart embedded
and Gomori
in carbowax
The
trichrome
were stained similarly.
RESULTS AND DISCUSSION
Results
of the 4 experiments
effect
(RAE)
serum
cholesterol
relating
was calculated
in Table 2. The relative
the average
level (g/100 ml). Th is method
the level of cholesterol
experiments
are detailed
by dividing
with a variety
In the first experiment,
was found to be effective
with the degree of atherosclerotic the CNO-fed group exhibited
it became
effect of PNO was similar
that the atherogenic
and that both were considerably 1971, 14: 53-64
lesions in previous
more severe atherosclero-
groups.
Atherosclerosis,
by the in cor-
of fat@.
tic lesions than did the other 3 dietary evident
atherogenic
grade of atheromata
In the other experiments,
more atherogenic
than CO or PGF.
to that
however, of CNO
CHOLESTEROL
TABLE
VEHICLE
ATHEROSCLEROSIS,
PART
57
11
3
DISTRIBUTION OIL
IN EXPERIMENTAL
OFATHEROMATAIN
(PXO),
SPECIAL
FAT
cxo
G vade
(44145)
0~
CORN
a
OIL
(co)
(AVERAGE
arch
(42145)
4.0
5
3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0
5 2 6 7 9 7 3
1 2 3 6 5 10 10 7
3 7 11 5 7 6
1 4 7 4 17 10 1
2 2 3 3 10 6 5 11 -
:\vcrage
2.15 & 0.16”
1.67 & 0.14
2.00 + 0.16
1.31 * 0.12
1.69 * 0.16
.\verage serum cholesterol (mg/lOO ml) RAEc a Survival IJ Standard r Relative
ratio. error. atherogenic
The
1 -
1360 1.40
of atherosclerotic experiments
There were no definite,
lesions
1 3 2 4 3 15 14 -
2 3 ‘2 5 13 5 9 -
4 1 2 5 13 17 -
1.2-t + 0.13
1.71 $I 0.16
1.13 f 0.12
1475 0.94
diet, is CNO > PNO
differences
fed cholesterol
liver cholesterol
levels in the CNO-fed
(7.08 g/100 g). The average
microscopic
Atherosclerotic
group (5.63 g/100 g) were lower
total
evaluation
lesions mainly
as compared
These are the combined separately,
(7.16 g/100 g), CO (6.82
liver cholesterol
levels for each
9.15 g.
to 21%
rabbits
lesions before
the aortic
and
and frequency arch.
Parts
of
of the
but to a less severe degree.
and 2776 of the intimal and 22%
results of all 4 experiments;
the peanut oil (PNO)-fed
in aortic
of the severity
affected
aorta were also involved,
In animals fed PNO and CNO, 29% ved, respectively,
(168 g)
(109.0 g) was slightly lower
fed PNO
by gross visualization
aortic
and abdominal
PGlT.
j&dings
findings
as well as a microscopic
lesions.
> CO =
plus PNO (123.4 g), PGF (129.3 g), or CO (130.4 g).
levels found in the rabbits
4 presents
the
of the 4
the average weight gain of the CO-fed group (240
after staining, thoracic
reflects
in weight gain or in liver weight.
group were: CNO, 6.19 g; PNO, 8.84 g; CO, 8.89 g; and PGF,
Table
--
higher than those of CNO- (152 g), PNO- (168 g), or PGF-fed
than liver cholesterol
Gross visual and
thovaclc
in all 4 experiments
groups. The average liver weight of the CNO-fed rabbits
g/100 g), or PGF
arch
(Table 3). The order of atherogenicity
consistent
over the 4 experiments,
The average
tikoracic
1564 0.94
fats, when added to a cholesterol-containing
than in the rabbits
(42145)
effect.
distribution
In general,
co
3
1300 1.27
results of the individual
g) was somewhat
4 EXPERIMENTS)
arch __~~
thoracic
____. 6
05
PGF
(45145)
PNO
fkoracic
arch
2 5; CHOLESYEROLhNDCOCONUTOIL(CNO),P&ANU'~
RABBITSFED
(PGF)
surface was invol-
in the CO- and PGF-fed if each experiment
rabbits.
was treated
still had the most severe atherosclerosis Athemclerosis,
1971,
14: 53 -6-l
58
n.
TABLE
KRITCHEVSKY,
S. A. TEPPER,
D. VESSELINOVITCH,
W. WISSLER
4
EFFECT
OF
DIETARY
FATS
(6%)
ON
CHOLESTEROL
(2°/o)-INDUCED
ATHEROMATA
(AVERAGE
4
OF
EXPERIMENTS)
Coconut (CNO) Peanut (PNO) PGF Corn (CO)
3.
Gross visual
No. of animals
Fat
arch
44 45 42 42
thoracic
2.15 2.00 1.69 1.71
+ * 5 +
0.16 0.16 0.16 0.16
1.67 1.31 1.24 1.13
& i_ + *
0.14 0.12 0.13 0.12
Gross visual stained aorta
Microscopic of lesions
(area 76)
zmolved
frequency
27 29 22 21
4.1 4.4 4.4 4.6
evaluation
severitya
( 76)
& & & *
50 56 38 35
36 37 20 20
Sum of severity Number
x
of animals
10.
whereas atherosclerosis visual differences trend
toward
animals.
greater
Even
was least apparent
in surface area involved involvement
visualized
lesions in PNO-fed
grossly,
in the CO-fed animals. Although between
was seen in the aorta the character
animals were raised and sometimes
had lesions, they were small, pale and only slightly Microscopic was carried
were observed monstrable
examination
out on standard in PNO-
of the severity
and CNO-fed
histochemically
the aortas of the rabbits In the rabbits high proportion
sections.
fed PNO a very prominent deposition
lesion in PNO-fed abundant
(Table
varied
with the type lesions in
intimal proliferation
by predominantly
Gomori rabbits
trichrome
as compared
with particularly to the rabbits
aorta of a PNO-fed
cellular proliferation,
fed
rabbit can be
composed
of a cap
of lipid giving the whole lesion a dense
aldehyde
fuchsin
stain
was used, the aortic
showed that the smooth muscle cells were separated
by the
collagen and elastin in which they were embedded.
The animals fed CNO had various sized, proliferative, on microscopic
examination,
revealed
remarkable
intimal
pale yellow lesions which, proliferation
characterized
by less collagen and elastin and more intracellular
lipid. Much of the stainable
in both types of lesions appeared
deposited
cells of innermost
media
(Fig.
to be diffusely
2). Lipid deposition
in or on smooth
inside pre-existing
in aorta was present in all of the groups, but was most frequent Except
4)
and severe lesions
of the atherosclerotic
was observed,
tissue with a small amount When
appearance.
of lesions
lesions showed lipid, de-
but the amount
other fats. In Fig. 1, a typical lesion in the thoracic of fibro-elastic
group but less severe.
All aortic
in the characteristics
CO-fed rabbits
Lesions of the PGF-fed
and frequency
rabbits.
The
fed PNO and CNO was seen consistently.
of collagen
seen. It is characterized
flat. Although
elevated.
The most frequent
with oil red 0,
and CNO-fed
very thick, while in the CNO-
to those in the PNO-fed
aortic
of fat in the diet. A difference
of PNO-
of the lesions seemed different.
fed animals the lesions were pale yellow and relatively group were similar in appearance
the gross
the groups were small, a definite
for these differences,
Atherosclerosis,
1971,
14: 53-64
there were histological
features
lipid muscle
medial cells
in CNO-fed animals. common
to both
the
CHOLI’STEROL
VEHICLE
IK
EXPERIMENTAL
ATHEROSCLEROSIS,
PART
11
I;ig. 1. Photomicrograph of a lesion in the thoracic aorta from a I’S@fed rabbit. The ksion ii charactc~riactl by pronounced proliferation with abundant intracellular mat&al which 15 pr,‘tlominrtntly collagen with some elastin. Oil red 0; x 195, rcduwd in rcprorluction L 0.75.
Fig. 2. Photomicrograph deposition was a frequent < 0.75.
of thoracic aortic lesion in a C?TO-fed rabbit. Proliferation over lipid finding in CSO-fed animals. Oil red 0; ’ 195, reduced in wprodwtlon
60
D. KRITCHEVSKY,
S. A. TEPPER, D. VESSELINOVITCH,
PNO- and CNO-fed groups. The lesions in PGF-fed
animals
W. WISSLER
also had some similarities
with those of the PNO-fed rabbits, but they were less severe (Fig. 3). The lesions in the CO-fed rabbits were characterized by less lipid deposition in pre-existing cells and by mild intimal A bar-graph
proliferation
(Fig. 5) summarizes
of the histopathological taken inside
animals.
pre-existing
the results
characteristics
from each aorta. A tendency
in PNO-fed
(Fig. 4). of microscopic
of the lesions
toward
collagen deposition
On the other hand, a more consistent medial
based
cells was seen in aortas
grading
of several
on standard
sections
was most pronounced
increase
of CNO-fed
in lipid deposition rabbits,
giving
the
appearance of a softer atheroma. Table 5 presents the frequency and severity evaluated microscopically of the lesions in the coronary arteries. The occurrence of lesions was highest in the group fed PNO and lowest in the group fed CO. The more advanced
coronary
artery
lesions
in PNO-fed rabbits were characterized by intimal proliferation and lipid deposition in both the intima and media. Lipid-rich proliferative intima sometimes resulted in severe
stenosis
coronary
of the intramural
coronary
vessels.
Qualitative
lesions were not great in spite of the different
Myocardial
infarction
cated on the endocardial third of the wall thickness. not completely
occluded,
was observed
in one rabbit
side of the left ventricular The coronary exhibited
differences
among
fats fed. fed PGF. The infarct wall and involved
artery in the vicinity
severe atherosclerotic
almost
of the infarct,
lesions.
was loone-
although
Most of the fibers
Fig. 3. Photomicrograph of thoracic aortic lesion of a PGF-fed animal. It is similar to the lesions produced by peanut oil, only less severe. Oil red 0; x 195, reduced in reproduction x 0.75. Atherosclerosis,
1971,
14: 53-64
CHOLESTEROL
VEHICLE
IGg. 1. l’hotomicrograph existing ~11s and mild duction i 0.75.
SMRITY
OF
IN EXPERIMEKTAL
of thoracic aortic intimal proliferation
AORTIC YICROSCOPIC
lizI COCONUT
fgj
PEANUT
ATHEROSCLEROSIS,
lesion in a CO-fed rabbit. wew seen. Oil ret1 0;
LESIONS
I!llI
FROM THREE
PGF
61
11
Deposition of lipid in prc~: 195, rcduccd in rcprc,-
STANDARD AREAS
CORN OIL
2.5 ??
PART
a
.
ASCENDINQ
??
THORACIC
A ABDOMINAL
1
k $
s
1.0
x
0.5
0
INT.
PROLIFER~ION
INT.
LIPIDS
LIPID
Fig. 5. Bar graph illustrates microscopic grading Imsvtl on standard sections taken in each aorta.
IN MEDIA
of several
FIBROUS CAP
characteristics
of the aortic
.4t/1erosc1erosis,
lesions,
1971, 14: 5x 6-i
62
D. KRITCHEVSKY,
TABLE
S. A. TEPPER,
D. VESSELINOVITCH,
W. WISSLER
5
MICROSCOPIC
LESIONS
IN
CORONARY
Treatwent
Group
1
Microscopic
coconut oil peanut oil PGF corn oil
II III
IV
were coagulated cation between a shadowy
ARTERIES
evaluation of lesions
frequency
severity
31 42 31 21
13 28 14 10
and their nuclei had disappeared. normal and ischemic
outline.
Inflammatory
There
tissue. Architecture
exudation
infiltrated
was a fairly good demar-
was visible in some parts as only partly into the necrotic
focus (Fig. 6), Fatty Although
liver changes
were observed
there was no remarkable
was a slight tendency and slightly
in most animals
difference
regardless
among the dietary
of the diet fed.
groups,
still there
for the fatty changes to be more severe in the PGF-fed
rabbits
milder in the CO-fed animals.
Pathologists of atherosclerosis.
have been puzzled about the factors responsible It appears
aortic wall may, at least partly,
that the chemical determine
lesions, as well as the rate of progression
nature
the quantity
for the progression
of the lipid deposited and quality
in the
of atherosclerotic
from fatty streak to fibrous plaque.
Fig. 6. Photomicrograph of the heart of a PGF-fed animal. Most of the fibers are coagulated their nuclei have disappeared. H.-E.; x 195, reduced in reproduction X 0.75. Atherosclerosis,
1971, 14: 53-64
and
63
CHOLESTEROL VEHICLE IS EXPERIMENTAL ATHEROSCLEROSIS, PART 11 There are several possible ways to explain the pathogenetic sclerosis
caused by the various food fats. One possibility
may be metabolized There
more effectively
may be either a greater
or a relatively consistent position
retarded
accumulation of stainable
of CSO-fed
of the exogenous
of stainable
suggests
bolism of lipid, particularly A further
absorption
aortic
that
that brought
lesions.
Only further
The results the rabbits
responsible
experimentation
the obvious suggestion
reaction
plus the 4 different
of coconut
of the
examinations
of the aortas
of
fats would, at first, appear to uphold
from its level of unsaturation
First is that the lesions of PGF-fed
is the pre-
must also be important
105; CO, 126. While PGF has
similar to that of PNO, the structure different
of this fat. Hence,
to cholesterol
some aspect
absorption,
varies;
however,
transport
the percentage
was calculated
of the component
from PNO since 3 different
ANI) VOLPENHEIN14,15 have shown that the percentage the oils used in these experiments tion of the triglycerides,
shown, in a
corn oil and corn oil, that
in fact, it is not. The iodine values of the 3 fats
must be considerably
were used for preparation
morphologic-
we have previously
hydrogenated
were: CNO, 8; PNO, 95; PGF,
a gross fatty acid composition
rabbits
This might be explained
with iodine value 7.8. On this basis, PGF should be
than CO, whereas,
of PGF
However,
oil, lard, partially
may be correlated
used in this experiment triglycerides
This reaction
morphology
and behenic acids. However, several factors suggest a more complex
on the basis of the level of unsaturation.
more atherogenic
within cells of the
that the factor that causes peanut oil to exert an atherogenic
reason for our observations.
atherogenicity
in the form
will resolve this question.
ally resemble the lesions of PNO- rather than CO-fed rabbits. comparison
in the meta-
than others.
for the characteristic
effect more severe than would be expected sence of arachidic
This de-
in the smooth muscle cells
cells may be critical
of the gross visual and microscopic
fed cholesterol
with the
rabbits.
to them by the blood stream
aortic wall may lead to a more severe metabolic partly
cholesterol
associated
is that some types of lipids when deposited
at least
wall than others.
before by GEER et al.12 and WISSLER tit &.I”.
This has been suggested
possibility
is probably
cholesterol
lipid in the CNO-fed
these arterial
of athero-
of the dietary
lipid in the aortic walls, particularly
animals,
of lipoproteins.
by the liver and the artery
gastrointestinal
catabolism
patterns
is that some types of lipid
of triglyceride and deposition.
of component
oils
structure MATTSON
fatty
acids in
of fatty acids in the Z-posi-
to be comparable
for PNO and
PGF. The primary
fatty
fats used were oleic (18:l) 95q;, of all the fatty position
acids present in the 2-position and linoleic (18:2).
acids present
in this position.
of CO and PNO triglycerides
to be 0.88. Thus there is little difference
glycerides
which arise during digestion
the structure
of the intact
fatty
acids that arise on lipolysis
may exert
respectively;
in the structure
and absorption
triglyceride
of all the
acids represented
The ratio of 18:1/18:2
is 0.36 and 0.59,
calculated then,
of the triglycerides
These 2 fatty
in the 2-
for PGF
it is
of the P-mono-
of PNO and PGF.
may be important
about
Possibly,
or the specific
free
an effect.
‘ltherosczerosis, 1971, 14: 53 ~6.l
64
D. KRITCHEVSKY,
S. A. TEPPER,
D. VESSELINOVITCH,
W. WISSLER
ACKNOWLEDGEMENTS
We are deeply indebted to Doctor Fred H. Mattson of the Proctor and Gamble Company for his assistance in providing one of the oils used (PGF) and for mrny helpful discussions,
REFERENCES 1
2
3
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IMAI, H., K. T. LEE, S. PASTORI, E. PANLILIO, R. FLORENTIN AND W. A. THOMAS, Atherosclerosis in rabbits: architectural and subcellular alteration of smooth muscle cells of aortas in response to hyperlipemia, Exp. Mol. Pathol., 1966, 5: 273-310. WISSLER, R. W., D. VESSELINOVITCH, G. S. GETZ AND R. H. HUGHES, Aortic lesions and blood lipids in Rhesus monkeys fed three food fats, Fed. Proc., 1967, 26: 371. FLORENTIN, R. A. AND S. C. NAM, Dietary-induced atherosclerosis in miniature swine, Part 1 (Gross and light microscopy observations. Time of development and morphologic characteristics of lesions), Exp. Mol. Pathol., 1968, 8: 263-277. KRITCHEVSKY, D. AND S. A. TEPPER, Cholesterol vehicle in experimental atherosclerosis, Part 7 (Influences of naturally occurring saturated fats), Med. Pharmacol. Exp., 1965, 12: 315320. KRITCHEVSKY, D. AND S. A. TEPPER, Cholesterol vehicle in experimental atherosclerosis, Part 8 (Effect of a medium chain triglyceride (MCT)), Exp. Mol. Pathol., 1965, 4: 489-499. MANN, G. V., A method for measurement of cholesterol in blood serum, Cl&. Chem., 1961, 7 : 275-284. DUFF, G. L. AND G. C. MCMILLAN, The effect of alloxan diabetes on experimental cholesterol atherosclerosis in the rabbit, J. Exp. Med., 1949, 89: 611-630. EGGEN, D. A., J. P. STRONG AND H. C. MCGILL, JR., An objective method for grading atherosclerotic lesions, Lab. Invest., 1962, 11: 732-742. GEER, J. C., H. C. MCGILL, JR. AND J. P. STRONG, The fine structure of human atherosclerotic lesions, Amer. J. Pathol., 1961, 38: 263-287. WISSLER, R. W., L. E. FRAZIER, R. H. HUGHES AND R. A. RASMUSSEN, Atherogenesis in the cebus monkey, Arch. Pathol., 1962, 74: 312-322. MATTSON, F. H. AND R. A. VOLPENHEIN, The specific distribution of fatty acids in the glycerides of vegetable fats, J. Biol. Chem., 1961, 236: 1891-1894. MATTSON, F. H. AND R. A. VOLPENHEIN, The specific distribution of unsaturated fatty acids in the triglycerides of plants, J. Lipid Res., 1963, 4: 392-396.
Atherosclerosis,
1971, 14: 53-64