(Circulation. 2001;103:1778.)
© 2001 American Heart Association, Inc.
Basic Science Reports |
Acyl-CoA:Cholesterol Acyltransferase Inhibitor Avasimibe Reduces Atherosclerosis in Addition to Its Cholesterol-Lowering Effect in ApoE*3-Leiden Mice
From Gaubius Laboratory, TNO-PG (D.J.M.D., E.H.O., W.v.D., H.v.d.B., E.C.M.d.W., L.M.H., H.M.G.P.); the Department of Human Genetics, Leiden University Medical Centre (M.J.J.G.); and the Department of Cardiology, Leiden University Medical Centre (A.v.d.L., J.W.J.), Leiden, the Netherlands.
Correspondence to Dr H.M.G. Princen, Gaubius Laboratory, TNO-PG, PO Box 2215, 2301 CE Leiden, the Netherlands. E-mail jmg.princen{at}pg.tno.nl
 |
Abstract |
|---|
 Top Abstract IntroductionMethodsResultsDiscussionReferences |
|---|
Background—The present study investigated whether the ACAT inhibitor avasimibe can reduce atherogenesis independently of its cholesterol-lowering effect in ApoE*3-Leiden mice.
Methods and Results—Two groups of 15 female ApoE*3-Leiden mice were put on a high-cholesterol (HC) diet; 1 group received 0.01% (wt/wt) avasimibe mixed into the diet. The HC diet resulted in a plasma cholesterol concentration of 18.7±2.6 mmol/L. Addition of avasimibe lowered plasma cholesterol by 56% to 8.1±1.2 mmol/L, caused mainly by a reduction of and composition change in VLDL and LDL. In a separate low-cholesterol (LC) control group, plasma cholesterol was titrated to a level comparable to that of the avasimibe group (10.3±1.4 mmol/L) by lowering the amount of dietary cholesterol. After 22 weeks of intervention, atherosclerosis in the aortic root area was quantified. Treatment with avasimibe resulted in a 92% reduction of lesion area compared with the HC control group. Compared with the LC control, avasimibe reduced lesion area by 78%. After correction for the slight difference in cholesterol exposure between the LC control and avasimibe groups, the effect of avasimibe on lesion area (73% reduction) remained highly significant. In addition, monocyte adherence to the endothelium, free cholesterol accumulation, and lesion severity were reduced by avasimibe treatment.
Conclusions—Treatment with avasimibe potently lowered plasma cholesterol levels in ApoE*3-Leiden mice and considerably reduced atherosclerotic lesion area in addition to its cholesterol-lowering effect. Because monocyte adherence to the endothelium and lesion severity were also reduced by avasimibe, treatment with avasimibe may result in higher plaque stability and therefore a reduced risk of plaque rupture.
Key Words: atherosclerosis • inhibitors • lipoproteins • cell adhesion molecules
|
Introduction |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
The enzyme acyl-CoA:cholesterol-O-acyltransferase (ACAT) catalyzes the intracellular formation of cholesteryl esters. It is involved in the absorption of cholesterol in the intestine and the secretion of VLDL from the liver. The enzyme has also been implicated in esterification and thereby storage of cholesteryl ester in monocyte-derived macrophages in the arterial wall.1 Because the accumulation of cholesteryl esters in the latter cells is one of the earliest steps in the development of atherosclerosis,2 it has been suggested that inhibition of the enzyme ACAT has a direct antiatherogenic potential.3 4 Moreover, because the stability of the atherosclerotic plaque is increased when fewer lipid-laden macrophages are present,5 a decrease of the number of lipid-laden macrophages in the arterial wall on ACAT inhibition could lead to a higher plaque stability and consequently a reduced risk of plaque rupture.
Several animal studies with various ACAT inhibitors have shown cholesterol-lowering effects of these drugs.6 7 8 Data on direct antiatherosclerotic effects of ACAT inhibitors, however, remain disputed. ACAT inhibition has been shown to decrease atherosclerotic lesion areas or cholesteryl ester enrichment in the arterial wall of animal models for atherosclerosis, but in most of these studies, substantial lipid-lowering was involved.9 10 11 12 Others found less atherosclerosis at a dose that showed some systemic availability but did not lower plasma total cholesterol levels.13 14 Thus, a number of models have been used to test the direct antiatherosclerotic activity of ACAT inhibitors, but so far, conclusions drawn from these experiments are based on circumstantial evidence.15
The present study was designed to determine the direct antiatherosclerotic effects of the ACAT inhibitor avasimibe (CI-1011; [[2,4,6-tris-(1-ethylethyl)phenyl]acetyl]sulfamic acid, 2,6-bis(1-methyl-ethyl)phenyl ester) in ApoE*3-Leiden transgenic mice by ruling out the confounding effects of cholesterol lowering. ApoE*3-Leiden mice were used because they exhibit elevated plasma cholesterol and triglyceride levels, mainly confined to the VLDL/LDL-size lipoprotein fraction.16 Because they are highly responsive in their plasma lipid levels to dietary treatment,17 their plasma cholesterol levels can easily be titrated to a constant level by the amount of cholesterol in the diet. In addition, on high-fat, high-cholesterol feeding, these mice develop atherosclerotic lesions resembling those found in humans, depending on their plasma cholesterol levels.18 19 Because ApoE*3-Leiden mice have been shown to be responsive to a number of cholesterol-lowering therapies, such as lovastatin, gemfibrozil, and fish oil,20 21 these mice were considered a suitable model for the testing of the antiatherosclerotic properties of the ACAT inhibitor avasimibe.
|
Methods |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
Mice
Heterozygous female ApoE*3-Leiden mice of F14 generations, 31 to 41 weeks of age at the start, were used. Identification of transgenic mice was performed by an ELISA for human apoE, as previously described.17 All animal experiments were approved by the institutional committees on animal welfare of TNO Prevention and Health.
Diets
Before the start of the study, animals were kept on a
standard rat/mouse chow diet (Hope Farms). During a 3-week run-in
period, all animals received a semisynthetic high-fat/high-cholesterol
(HFC) diet as used before.17
After this period, the animals were divided into 3 groups of 15 mice
each on the basis of age and cholesterol level. The high-cholesterol
(HC) control group received an HFC diet with addition of 0.5% wt/wt
cholesterol and 0.1% wt/wt cholate, the latter added to facilitate
intestinal uptake of fat and cholesterol, thereby increasing plasma
cholesterol levels. The avasimibe group received the same diet as the
HC control group, with an extra addition of 0.01% wt/wt avasimibe
(kindly provided by Dr Krause, Parke Davis, Ann Arbor, Mich),
approximately equaling a daily dose of 10 mg/kg body wt. The diet of
the low-cholesterol (LC) control group was chosen to reach the same
plasma cholesterol level in these mice as in the avasimibe group and
contained 0.5% wt/wt cholesterol and no cholate. Animals had free
access to water and food.
Lipid and Lipoprotein Analysis
After a 4-hour fasting period from 9
AM to 1
PM, blood samples were
obtained from each individual mouse by tail incision. Total plasma
cholesterol and triglyceride levels were measured enzymatically, and
lipoprotein profiles were obtained by size fractioning, as
described.17 22
For the determination of the composition of the apoB-containing
lipoproteins, ultracentrifugation of total plasma was performed, VLDL
and IDL/LDL fractions were pooled, and lipids were analyzed as
described previously.23
Total plasma apoB and apoE were measured by gel electrophoresis
followed by Coomassie blue staining and calculation of the contribution
of apoB and apoE to total protein.
Histological Assessment of
Atherosclerosis
After 24 weeks of diet feeding, mice were killed
after anesthetization and blood collection as
described.17 The hearts were
dissected, stored overnight in phosphate-buffered 3.8% formalin
fixation, and embedded in paraffin. Serial cross sections (5 µm
thick) throughout the entire aortic valve area were used for
histological analysis. Sections were routinely stained with
hematoxylin-phloxine-saffron (HPS). Per mouse, 4 sections with
intervals of 30 µm were used for quantification and qualification of
atherosclerotic lesions. For determination of severity of
atherosclerosis, the lesions were classified into 5 categories (types I
through V) as described
before17 24 :
(I) early fatty streak, (II) regular fatty streak, (III) mild
plaque, (IV) moderate plaque, and (V) severe plaque. Mouse macrophages
were immunostained with AIA31240 (1:3000, Accurate Chemical and
Scientific), and smooth muscle cells were stained with 
-actin
(Roche).
To evaluate whether avasimibe has an effect on the accumulation of free cholesterol in atherosclerotic lesions, the sections containing free cholesterol clefts were counted in each group. To determine whether avasimibe has an effect on endothelial activation, the monocytes adhering to the endothelium were counted in the same 4 slides as used for the quantification of atherosclerosis. The average number of monocytes attached to the endothelium per section was used for comparison between groups.
ACAT Activity in J774 Cells
The murine macrophage cell line J774 was
cultured in DMEM supplemented with 10% (vol/vol) FCS, 100 IU/mL
penicillin, and 100 µg/mL streptomycin. For each experiment, cells
were plated in 12-well plates under culture conditions as described by
Bocan et al.25 Cells were
preincubated with avasimibe 1 hour before the addition of 200 µg
protein/mL ßVLDL or 50 µg protein/mL acetylated LDL. ACAT activity
was determined by measurement of incorporation of
[14C]oleate in cholesteryl oleate,
essentially as described by Post et
al.22
Statistics
Overall comparisons between groups were performed
with the Kruskal-Wallis test. Distribution-free multiple comparisons
were performed to find differences between groups. If only 2 groups
were compared, Mann-Whitney rank sum tests were used. Differences in
distributions of atherosclerotic lesions over the different lesion
categories and of lipids over different lipoproteins were tested with
Fisher’s exact test. Probability values of
P<0.05 were regarded as
significant. All data are presented as mean±SD, unless mentioned
otherwise.
|
Results |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
Effect of Avasimibe on Plasma Cholesterol Levels and Lipoprotein Composition
Avasimibe did not affect body weight or food intake. Average plasma cholesterol levels from 8 blood draws (at t=3, 6, 9, 10, 14, 19, 21, and 24 weeks) were 18.73±2.63 mmol/L for the HC group, 8.17±1.15 mmol/L for the avasimibe group, and 10.24±1.46 mmol/L in the LC control group. Compared with the HC control group, avasimibe lowered plasma total cholesterol concentrations by 56% (P<0.005). Compared with the LC group, the avasimibe group showed a slightly but significantly reduced plasma cholesterol level (P<0.005). Triglyceride levels did not differ significantly between the groups (1.29±0.60, 1.59±0.47, and 1.01±0.48 mmol/L for the HC, avasimibe, and LC groups, respectively).
Lipoprotein profiles at 14 weeks of treatment showed that
avasimibe mainly reduced VLDL, IDL, and LDL cholesterol
(Figure 1A
).
Figure 1B shows the composition of VLDL and IDL/LDL in the
different groups. VLDL and IDL/LDL in the avasimibe group contained
relatively more triglyceride and less cholesteryl ester than in the LC
and HC controls. The IDL/LDL lipoproteins in all groups contained
relatively less triglyceride than their respective VLDL.
|
Total apoB was 44% decreased in the avasimibe group
compared with the HC control (210 and 378 µg/mL, respectively). In
the LC control group, apoB was 55% lower than in the avasimibe group
(93 µg/mL). ApoE was not different between the avasimibe group and
the LC control (443 and 436 µg/mL, respectively). Plasma apoE in
these latter groups was 39% lower than in the HC control (720
µg/mL). These data indicate that in the HC control group, the number
of VLDL/LDL particles and their total cholesteryl ester content are
largest. Treatment with avasimibe causes a reduction of these
particles, which become relatively enriched in triglycerides. Compared
with the LC control, the avasimibe-treated animals have 
2-fold more
VLDL/LDL particles in their plasma, which contain less cholesteryl
ester.
Effect of Avasimibe on Macrophage ACAT
Activity
Plasma concentrations of avasimibe were 50±24 nmol/L.
To investigate whether esterification of cholesterol is inhibited at
these concentrations, the effect of avasimibe on ACAT activity was
measured in the mouse macrophage cell line J774 in the presence of an
additional source of cholesterol to increase the intracellular
cholesterol pool. Incubation with avasimibe resulted in a
dose-dependent reduction in ACAT activity, showing a significant 25%
reduction at 50 nmol/L
(Figure 2, P<0.005).
|
Effect of Avasimibe on Atherosclerosis
Development
Atherosclerotic lesions were quantified in cross
sections of the aortic valve area in the heart of each individual
mouse.
The average lesion area per section for the individual
groups is shown in
Figure 3A. For the HC control group, the average lesion area
per section was 95.5±35.2 µm2x1000.
Treatment with avasimibe resulted in a 92% reduction of lesion area
(7.6±7.0 µm2x1000), related to the total
effect of avasimibe. Comparison of the avasimibe group and the LC
control (34.1±24.5 µm2x1000) showed a
78% reduction of atherosclerotic lesion area by avasimibe, suggesting
an effect independent of its cholesterol-lowering effect.
|
Although the avasimibe group and the LC control were almost
matched for their plasma total cholesterol levels, there was a slight
difference in cholesterol level between those groups. Because it has
been shown previously that there is a linear relationship between
atherosclerotic lesion area and exposure of the arterial wall to plasma
cholesterol in ApoE*3-Leiden
mice,18 total cholesterol
burden over time for the latter groups was calculated to correct the
lesion areas for differences in these cholesterol exposures. Mean
cholesterol exposure in mice treated with avasimibe was 184±26 mmol
· L-1 ·
wk-1 and in mice receiving the LC control
diet, 236±34 mmol · L-1 ·
wk-1
(P<0.005). Correction of the
lesion areas for cholesterol exposure by dividing the lesion area per
mouse by its individual cholesterol exposure showed that the lesion
area in the avasimibe group was 73% smaller than in the LC control
group (P=0.012 by Mann-Whitney
test,
Figure 3B
). These data indicate that avasimibe reduces the
development of atherosclerosis independently of its
cholesterol-lowering effect.
We also evaluated the effect of avasimibe treatment on the
severity of the lesions.
Figure 4 shows representative pictures. The percentages of
lesions classified in the respective lesion categories were calculated
(Figure 5). In the HC control group, the largest percentage
of all lesions is constituted of severe lesions. Lesions in the LC
control group are almost equally distributed over the fatty streaks and
severe lesions. In the avasimibe group, however, most lesions are mere
small fatty streaks, and only a few lesions belong to the severe types
of plaques. These results, in combination with the stainings for
macrophages
(Figure 4B, 4E, and 4H) and smooth muscle cells
(Figure 4C, 4F, and 4I) indicate that treatment with
avasimibe resulted in the development of only small numbers of foam
cells in the intima of the arterial wall containing small lipid pools
compared with the LC control group, in which foam cell accumulation
with larger lipid pools also occurred in the media. In the HC control
group, in contrast, large lipid pools were found in the intima and
media, the presence of smooth muscle cells in the media was diminished,
and severe necrosis and calcification were found.
|
); staining with AIA31240. C, F, and I, Immunohistochemistry of smooth muscle cells (SMC); staining with
|
Effect of Avasimibe on Free Cholesterol Content
of Atherosclerotic Lesions
Because avasimibe reduced cholesteryl ester
accumulation in the cell, it is conceivable that treatment with
avasimibe would result in the accumulation of free cholesterol in the
atherosclerotic lesions. Because free cholesterol above a certain
threshold concentration is toxic to the
cell,26 accumulations of
free cholesterol can be found only in the form of cholesterol clefts.
Therefore, we determined whether treatment with avasimibe increased the
number of sections containing cholesterol clefts. However, the opposite
was found
(Figure 6). Treatment with avasimibe significantly reduced
the number of atherosclerotic lesions containing cholesterol clefts,
indicating that avasimibe does not increase free cholesterol
accumulation.
|
Effect of Avasimibe on Monocyte Adhesion to
the Endothelium
The number of monocytes adhering to the endothelium, as
shown in
Figure 7A through 7D, was counted in the same slides as used
for quantification of atherosclerosis.
Figure 7E shows that monocyte attachment to the endothelium
was not different between the HC and LC control groups. Avasimibe,
however, showed a significant 80% reduction of the number of monocytes
adhering to the endothelium
(P<0.001), suggesting that
avasimibe reduces endothelial activation independently of its
cholesterol-lowering effect.
|
|
Discussion |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
In the present study, the ACAT inhibitor avasimibe decreases plasma cholesterol levels by >50% in cholesterol-fed ApoE*3-Leiden mice by reduction of VLDL/LDL cholesterol and induces a relative enrichment of triglycerides and decrease of cholesteryl esters in these lipoproteins, making them potentially less atherogenic. In addition, treatment with avasimibe results in reduced progression and severity of atherosclerosis, in addition to its cholesterol-lowering effect.
The present study was designed to demonstrate a direct effect of avasimibe on the development of atherosclerosis, in addition to its cholesterol-lowering effect. Therefore, an HC control group, an HC group receiving avasimibe, and a matched LC group were used to compare the development of atherosclerosis in the avasimibe group with that in the groups without ACAT inhibitor at high or at equally low cholesterol levels.
Our results on the lipid-lowering effects of the ACAT inhibitor avasimibe are in agreement with previous studies showing that avasimibe can potently reduce plasma lipid concentrations in rats, rabbits, and hamsters. At the dosage used (10 mg · kg body wt-1 · d-1), however, the plasma cholesterol reduction observed in the present study (-56%) exceeds that in previous studies in rats (-32%)27 and hamsters (-37%).9 The reduction of plasma cholesterol levels by ACAT inhibitors can be attributed primarily to a decreased intestinal cholesterol absorption.1 For inhibitors with increased bioavailability compared with the early nonabsorbable hypocholesterolemic agents,27 however, a reduction of the secretion of VLDL apoB from the liver28 29 30 and the subsequent proper disposal of cholesterol into the bile acid synthetic pathway22 may also contribute to the cholesterol-lowering effect. The present study shows that treatment with avasimibe also changed lipoprotein composition. Compared with the HC and LC controls, avasimibe treatment resulted in a relative enrichment of VLDL/LDL with triglycerides, probably as a consequence of the decrease in cholesterol esterification in the liver. Compared with the HC control group, the avasimibe group showed 44% lower total plasma apoB levels, in agreement with data obtained in miniature pigs.30 Compared with the LC control, however, apoB levels were more than twice as high in the avasimibe group. This indicates that plasma from the avasimibe group contains VLDL/LDL particles that are less atherogenic than those in the LC control, but also that more of these particles are present in plasma, which again adversely contributes to atherogenicity in the avasimibe group.
Because the enzyme ACAT is also responsible for the esterification and storage of cholesterol in the macrophage, it has been hypothesized that ACAT inhibition may prevent the accumulation of lipid-laden macrophages in the arterial wall.3 4 Bocan et al25 recently showed that avasimibe reduced the intracellular cholesteryl ester concentration in human monocyte–derived macrophages in a dose-dependent manner. To investigate whether esterification of cholesterol is inhibited in mouse macrophages at concentrations found in plasma of the avasimibe group (50±24 nmol/L), the effect of avasimibe on ACAT activity was measured in the mouse macrophage cell line J774. At 50 nmol/L avasimibe, cholesterol esterification was significantly inhibited by 25%, suggesting that in our animal study, macrophage ACAT in the vessel wall was also inhibited by avasimibe.
Comparison of atherosclerotic lesion areas in the HC control and the avasimibe group shows that avasimibe reduced atherosclerosis by 92% in the ApoE*3-Leiden mice. This finding is in line with previous observations with avasimibe in hamsters9 and other ACAT inhibitors in rabbits10 11 showing an antiatherosclerotic activity of ACAT inhibitors concomitant with reductions in plasma cholesterol concentrations. After correction of the extent of atherosclerosis for the slight difference in cholesterol exposure between the LC control and the avasimibe groups, comparison of these corrected lesion areas showed that avasimibe significantly reduced the development of atherosclerosis by 73%. It has been suggested that an atherogenic diet containing cholate, such as that used in the avasimibe group, may induce an inflammatory response in the liver, resulting in adverse effects on atherogenesis.31 Others, however, have found no such effect in C57BL/6J32 and LDLr-/-33 mice, indicating that the effect of cholate on the liver has not been unequivocally established. The fact that we found that avasimibe in a cholate-containing diet significantly reduced atherosclerosis compared with the LC control group without cholate clearly favors the direct antiatherosclerotic effect of avasimibe.
We also demonstrate that avasimibe reduced lesion severity compared with the LC and the HC controls. In the HC control group, atherosclerotic lesions were characterized by large lipid depositions in the intima and media, with severe fibrosis, necrosis, calcification, and reduction of smooth muscle cells. Lesions found in the avasimibe group contained only a small number of foam cells and a small lipid pool. The media was hardly ever affected, and the number of smooth muscle cells was not reduced. This is in contrast to the mild plaques found in the LC control group, in which the lipid pools were larger and foam cells were also found among the smooth muscle cells in the media. It is currently understood that clinical manifestations of atherosclerosis are characterized by disruption of the plaque34 and that the most important features of a vulnerable and potentially unstable plaque are an abundant presence of T lymphocytes and macrophages, a large lipid pool in the plaque, a low density of smooth muscle cells, and a thin, collagen-poor fibrous cap.5 Therefore, our results indicate that treatment with avasimibe can contribute to a higher plaque stability and consequently reduce the risk of plaque rupture, as has recently also been suggested in studies with hypercholesterolemic rabbits.25 Also, the finding that treatment with avasimibe significantly reduced the number of atherosclerotic lesions containing cholesterol clefts indicates that avasimibe does not increase free cholesterol accumulation in the lesions. Thus, the additional effect of avasimibe is an attenuated accumulation of lipids in the arterial wall and inhibition of the infiltration of macrophages into the media. In addition, our results show that avasimibe significantly reduced the number of monocytes adhering to the endothelium, which is considered to be one of the first steps in the development of atherosclerosis.5 This is in agreement with in vitro studies that have shown that ACAT inhibitors can decrease monocyte adhesion to endothelial cells.35 It suggests that treatment with avasimibe reduces endothelial activation. Whether avasimibe indeed decreases the expression of adhesion and/or chemoattractant molecules in the endothelium remains to be investigated.
In conclusion, this study demonstrates that the ACAT inhibitor avasimibe potently lowers plasma cholesterol levels in transgenic ApoE*3-Leiden mice. Although it cannot be fully excluded that the change in lipoprotein composition in the avasimibe-treated animals contributes to its antiatherogenic effect, the dramatic reduction in atherosclerotic lesion area in the avasimibe group clearly appears to be attributable to an effect additional to that of cholesterol lowering. Because lesion severity is also reduced by avasimibe, treatment with avasimibe may result in higher plaque stability and therefore a reduced risk of plaque rupture.
|
Acknowledgments |
|---|
This study was supported in part by a grant from Parke Davis. The authors would like to thank Dr Krause (Parke Davis, Ann Arbor, Mich) for fruitful discussions and measurement of plasma avasimibe levels.
Received August 15, 2000; revision received October 9, 2000; accepted October 18, 2000.
|
References |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
1. Suckling KE, Stange EF. Role of acyl-CoA: cholesterol acyltransferase in cellular cholesterol metabolism. J Lipid Res. 1985;26:647–671.[Medline] [Order article via Infotrieve]
2. Ross R. The pathogenesis of atherosclerosis: a perspective for the 1990s. Nature. 1993;362:801–809.[Medline] [Order article via Infotrieve]
3. Krause BK, Sliskovic DR, Bocan TMA. Emerging therapies in atherosclerosis. Expert Opin Invest Drugs. 1995;4:353–387.
4. Sliskovic DR, White AD. Therapeutic potential of ACAT inhibitors as lipid lowering and anti-atherosclerotic agents. Trends Pharmacol Sci. 1991;12:194–199.[Medline] [Order article via Infotrieve]
5. Libby P, Geng YJ, Aikawa M, et al. Macrophages and atherosclerotic plaque stability. Curr Opin Lipidol. 1996;7:330–335.[Medline] [Order article via Infotrieve]
6. Krause BK, Bocan TMA. ACAT inhibitors: physiologic mechanisms for hypolipidemic and anti-atherosclerotic activities in experimental animals. In: Ruffolo RR, Hollinger MA, eds. Inflammation: Mediators and Pathways. Boca Raton, Fla: CRC Press; 1995:173–198.
7. Largis EE, Wang CH, DeVries VG, et al. CL 277,082: a novel inhibitor of ACAT-catalyzed cholesterol esterification and cholesterol absorption. J Lipid Res. 1989;30:681–690.[Abstract]
8. Burnett JR, Wilcox LJ, Telford DE, et al. Inhibition of cholesterol esterification by DuP 128 decreases hepatic apolipoprotein B secretion in vivo: effect of dietary fat and cholesterol. Biochim Biophys Acta. 1998;1393:63–79.[Medline] [Order article via Infotrieve]
9. Nicolosi RJ, Wilson TA, Krause BR. The ACAT inhibitor, CI-1011 is effective in the prevention and regression of aortic fatty streak area in hamsters. Atherosclerosis. 1998;137:77–85.[Medline] [Order article via Infotrieve]
10. White AD, Purchase CF II, Picard JA, et al. Heterocyclic amides: inhibitors of acyl-CoA:cholesterol O-acyl transferase with hypocholesterolemic activity in several species and antiatherosclerotic activity in the rabbit. J Med Chem. 1996;39:3908–3919.[Medline] [Order article via Infotrieve]
11. Ashton MJ, Bridge AW, Bush RJ, et al. RP-70676: a potent systemically available inhibitor of acyl-CoA:cholesterol o-acyl transferase (ACAT). Bioorg Med Chem Lett. 1992;2:375–380.
12. Schaffer SA, Bloom JD, De Vries VG, et al. CL 277,082, a novel inhibitor of cholesterol esterification and cholesterol absorption. In: Fidge NH, Nestel PJ, eds. Atherosclerosis VII. Amsterdam, Netherlands: Elsevier Science Publishers BV; 1986:633–636.
13. Kogushi M, Tanaka H, Ohtsuka I, et al. Anti-atherosclerotic effect of E5324, an inhibitor of acyl-CoA: cholesterol acyltransferase, in Watanabe heritable hyperlipidemic rabbits. Atherosclerosis. 1996;124:203–210.[Medline] [Order article via Infotrieve]
14.
Bocan TM, Mueller
SB, Uhlendorf PD, et al. Comparison of CI-976, an ACAT inhibitor, and
selected lipid-lowering agents for antiatherosclerotic activity in
iliac- femoral and thoracic aortic lesions: a biochemical,
morphological, and morphometric evaluation.
Arterioscler Thromb. 1991;11:1830–1843.
15. Bocan TM. Animal models of atherosclerosis and interpretation of drug intervention studies. Curr Pharm Des. 1998;4:37–52.[Medline] [Order article via Infotrieve]
16.
van den
Maagdenberg AM, Hofker MH, Krimpenfort PJ, et al. Transgenic mice
carrying the apolipoprotein E3-Leiden gene exhibit
hyperlipoproteinemia. J Biol
Chem. 1993;268:10540–10545.
17. van Vlijmen BJ, van den Maagdenberg AM, Gijbels MJ, et al. Diet-induced hyperlipoproteinemia and atherosclerosis in apolipoprotein E3-Leiden transgenic mice. J Clin Invest. 1994;93:1403–1410.
18.
Groot PH, van
Vlijmen BJ, Benson GM, et al. Quantitative assessment of aortic
atherosclerosis in APOE*3 Leiden transgenic mice and its relationship
to serum cholesterol exposure.
Arterioscler Thromb Vasc Biol. 1996;16:926–933.
19.
Lutgens E, Daemen
M, Kockx M, et al. Atherosclerosis in APOE*3-Leiden transgenic mice:
from proliferative to atheromatous stage.
Circulation. 1999;99:276–283.
20.
van Vlijmen BJ,
Mensink RP, et al. Effects of dietary fish oil on serum lipids and VLDL
kinetics in hyperlipidemic apolipoprotein E*3-Leiden transgenic mice.
J Lipid Res. 1998;39:1181–1188.
21. van Vlijmen BJ, Pearce NJ, Bergö M, et al. Apolipoprotein E*3-Leiden transgenic mice as a test model for hypolipidaemic drugs. Arzneimittelforschung. 1998;48:396–402.[Medline] [Order article via Infotrieve]
22.
Post SM,
Zoeteweij JP, Bos MH, et al. Acyl-coenzyme A:cholesterol
acyltransferase inhibitor, avasimibe, stimulates bile acid synthesis
and cholesterol 7-hydroxylase in cultured rat hepatocytes and in
vivo in the rat. Hepatology. 1999;30:491–500.[Medline]
[Order article via Infotrieve]
23. Havekes LM, de Wit EC, Princen HM. Cellular free cholesterol in Hep G2 cells is only partially available for down-regulation of low-density-lipoprotein receptor activity. Biochem J. 1987;247:739–746.[Medline] [Order article via Infotrieve]
24. Gijbels MJ, van der Cammen M, van der Laan LJ, et al. Progression and regression of atherosclerosis in APOE3-Leiden transgenic mice: an immunohistochemical study. Atherosclerosis. 1999;143:15–25.[Medline] [Order article via Infotrieve]
25.
Bocan TM, Krause
BR, Rosebury WS, et al. The ACAT inhibitor avasimibe reduces
macrophages and matrix metalloproteinase expression in atherosclerotic
lesions of hypercholesterolemic rabbits.
Arterioscler Thromb Vasc Biol. 2000;20:70–79.
26.
Kellner-Weibel G,
Jerome WG, Small DM, et al. Effects of intracellular free cholesterol
accumulation on macrophage viability: a model for foam cell death.
Arterioscler Thromb Vasc Biol. 1998;18:423–431.
27. Lee HT, Sliskovic DR, Picard JA, et al. Inhibitors of acyl-CoA: cholesterol O-acyl transferase (ACAT) as hypocholesterolemic agents: CI-1011: an acyl sulfamate with unique cholesterol-lowering activity in animals fed noncholesterol-supplemented diets. J Med Chem. 1996;39:5031–5034.[Medline] [Order article via Infotrieve]
28. Carr TP, Hamilton RLJ, Rudel LL. ACAT inhibitors decrease secretion of cholesteryl esters and apolipoprotein B by perfused livers of African green monkeys. J Lipid Res. 1995;36:25–36.[Abstract]
29.
Huff MW, Telford
DE, Barrett PH, et al. Inhibition of hepatic ACAT decreases apoB
secretion in miniature pigs fed a cholesterol-free diet.
Arterioscler Thromb. 1994;14:1498–1508.
30.
Burnett JR,
Wilcox LJ, Telford DE, et al. Inhibition of ACAT by avasimibe decreases
both VLDL and LDL apolipoprotein B production in miniature pigs.
J Lipid Res. 1999;40:1317–1328.
31. Liao F, Andalibi A, Qiao JH, et al. Genetic evidence for a common pathway mediating oxidative stress, inflammatory gene induction, and aortic fatty streak formation in mice. J Clin Invest. 1994;94:877–884.
32. Nishina PM, Verstuyft J, Paigen B. Synthetic low and high fat diets for the study of atherosclerosis in the mouse. J Lipid Res. 1990;31:859–869.[Abstract]
33.
Lichtman AH,
Clinton SK, Iiyama K, et al. Hyperlipidemia and atherosclerotic lesion
development in LDL receptor-deficient mice fed defined semipurified
diets with and without cholate.
Arterioscler Thromb Vasc Biol. 1999;19:1938–1944.
34.
Falk E, Shah PK,
Fuster V. Coronary plaque disruption.
Circulation. 1995;92:657–671.
35. Saxena U, Ferguson E, Newton RS. Acyl-coenzyme A:cholesterol-acyltransferase (ACAT) inhibitors modulate monocyte adhesion to aortic endothelial cells. Atherosclerosis. 1995;112:7–17. [Medline] [Order article via Infotrieve]
This article has been cited by other articles:
 |
|
 |
|
  M. C. Meuwese, E. de Groot, R. Duivenvoorden, M. D. Trip, L. Ose, F. J. Maritz, D. C. G. Basart, J. J. P. Kastelein, R. Habib, M. H. Davidson, et al. ACAT Inhibition and Progression of Carotid Atherosclerosis in Patients With Familial Hypercholesterolemia: The CAPTIVATE Randomized Trial JAMA, March 18, 2009; 301(11): 1131 - 1139. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. W.A. van der Hoorn, W. de Haan, J. F.P. Berbee, L. M. Havekes, J. W. Jukema, P. C.N. Rensen, and H. M.G. Princen Niacin Increases HDL by Reducing Hepatic Expression and Plasma Levels of Cholesteryl Ester Transfer Protein in APOE*3Leiden.CETP Mice Arterioscler. Thromb. Vasc. Biol., November 1, 2008; 28(11): 2016 - 2022. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Fujiwara, N. Kiyota, M. Hori, S. Matsushita, Y. Iijima, K. Aoki, D. Shibata, M. Takeya, T. Ikeda, T. Nohara, et al. Esculeogenin A, a New Tomato Sapogenol, Ameliorates Hyperlipidemia and Atherosclerosis in ApoE-Deficient Mice by Inhibiting ACAT Arterioscler. Thromb. Vasc. Biol., November 1, 2007; 27(11): 2400 - 2406. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Zadelaar, R. Kleemann, L. Verschuren, J. de Vries-Van der Weij, J. van der Hoorn, H. M. Princen, and T. Kooistra Mouse Models for Atherosclerosis and Pharmaceutical Modifiers Arterioscler. Thromb. Vasc. Biol., August 1, 2007; 27(8): 1706 - 1721. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. Cheng, D. Tempel, R. van Haperen, A. van der Baan, F. Grosveld, M. J.A.P. Daemen, R. Krams, and R. de Crom Atherosclerotic Lesion Size and Vulnerability Are Determined by Patterns of Fluid Shear Stress Circulation, June 13, 2006; 113(23): 2744 - 2753. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. T. Dalen, S. M. Ulven, B. M. Arntsen, K. Solaas, and H. I. Nebb PPAR{alpha} activators and fasting induce the expression of adipose differentiation-related protein in liver J. Lipid Res., May 1, 2006; 47(5): 931 - 943. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. E. Nissen, E. M. Tuzcu, H. B. Brewer, I. Sipahi, S. J. Nicholls, P. Ganz, P. Schoenhagen, D. D. Waters, C. J. Pepine, T. D. Crowe, et al. Effect of ACAT inhibition on the progression of coronary atherosclerosis. N. Engl. J. Med., March 23, 2006; 354(12): 1253 - 1263. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. P. Tam, J. B. Ancsin, R. Tan, and R. Kisilevsky Peptides derived from serum amyloid A prevent, and reverse, aortic lipid lesions in apoE-/- mice J. Lipid Res., October 1, 2005; 46(10): 2091 - 2101. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Moselewski, C. J. O'Donnell, S. Achenbach, M. Ferencik, J. Massaro, A. Nguyen, R. C. Cury, S. Abbara, I.-K. Jang, T. J. Brady, et al. Calcium Concentration of Individual Coronary Calcified Plaques as Measured by Multidetector Row Computed Tomography Circulation, June 21, 2005; 111(24): 3236 - 3241. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Vidal, S. Hernandez-Vallejo, T. Pauquai, O. Texier, M. Rousset, J. Chambaz, S. Demignot, and J.-M. Lacorte Apple procyanidins decrease cholesterol esterification and lipoprotein secretion in Caco-2/TC7 enterocytes J. Lipid Res., February 1, 2005; 46(2): 258 - 268. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Verschuren, R. Kleemann, E. H. Offerman, A. J. Szalai, S. J. Emeis, H. M. G. Princen, and T. Kooistra Effect of Low Dose Atorvastatin Versus Diet-Induced Cholesterol Lowering on Atherosclerotic Lesion Progression and Inflammation in Apolipoprotein E*3-Leiden Transgenic Mice Arterioscler. Thromb. Vasc. Biol., January 1, 2005; 25(1): 161 - 167. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J.-C. Tardif, J. Gregoire, P. L. L'Allier, T. J. Anderson, O. Bertrand, F. Reeves, L. M. Title, F. Alfonso, E. Schampaert, A. Hassan, et al. Effects of the Acyl Coenzyme A:Cholesterol Acyltransferase Inhibitor Avasimibe on Human Atherosclerotic Lesions Circulation, November 23, 2004; 110(21): 3372 - 3377. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. S. Meir and E. Leitersdorf Atherosclerosis in the Apolipoprotein E-Deficient Mouse: A Decade of Progress Arterioscler. Thromb. Vasc. Biol., June 1, 2004; 24(6): 1006 - 1014. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. M. S. Espirito Santo, N. M. M. Pires, L. S. M. Boesten, G. Gerritsen, N. Bovenschen, K. W. van Dijk, J. W. Jukema, H. M. G. Princen, A. Bensadoun, W.-P. Li, et al. Hepatic low-density lipoprotein receptor-related protein deficiency in mice increases atherosclerosis independent of plasma cholesterol Blood, May 15, 2004; 103(10): 3777 - 3782. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Lie, R. de Crom, T. van Gent, R. van Haperen, L. Scheek, F. Sadeghi-Niaraki, and A. van Tol Elevation of plasma phospholipid transfer protein increases the risk of atherosclerosis despite lower apolipoprotein B-containing lipoproteins J. Lipid Res., May 1, 2004; 45(5): 805 - 811. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. de Medina, B. L. Payre, J. Bernad, I. Bosser, B. Pipy, S. Silvente-Poirot, G. Favre, J.-C. Faye, and M. Poirot Tamoxifen Is a Potent Inhibitor of Cholesterol Esterification and Prevents the Formation of Foam Cells J. Pharmacol. Exp. Ther., March 1, 2004; 308(3): 1165 - 1173. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 I. Namatame, H. Tomoda, S. Ishibashi, and S. Omura Antiatherogenic activity of fungal beauveriolides, inhibitors of lipid droplet accumulation in macrophages PNAS, January 20, 2004; 101(3): 737 - 742. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. E. Sobel, D. J. Taatjes, and D. J. Schneider Intramural Plasminogen Activator Inhibitor Type-1 and Coronary Atherosclerosis Arterioscler. Thromb. Vasc. Biol., November 1, 2003; 23(11): 1979 - 1989. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Kleemann, H. M.G. Princen, J. J. Emeis, J. W. Jukema, R. D. Fontijn, A. J.G. Horrevoets, T. Kooistra, and L. M. Havekes Rosuvastatin Reduces Atherosclerosis Development Beyond and Independent of Its Plasma Cholesterol-Lowering Effect in APOE*3-Leiden Transgenic Mice: Evidence for Antiinflammatory Effects of Rosuvastatin Circulation, September 16, 2003; 108(11): 1368 - 1374. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. L. Willner, B. Tow, K. K. Buhman, M. Wilson, D. A. Sanan, L. L. Rudel, and R. V. Farese Jr. Deficiency of acyl CoA:cholesterol acyltransferase 2 prevents atherosclerosis in apolipoprotein E-deficient mice PNAS, February 4, 2003; 100(3): 1262 - 1267. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. K. Buhman, H. C. Chen, and R. V. Farese Jr. The Enzymes of Neutral Lipid Synthesis J. Biol. Chem., October 26, 2001; 276(44): 40369 - 40372. [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||