Accumulation of Apolipoprotein C-I–Rich and
Cholesterol-Rich VLDL Remnants During Exaggerated
Postprandial Triglyceridemia in Normolipidemic Patients
With Coronary Artery Disease
Johan Björkegren, MD, PhD; Susanna Boquist, MD; Ann Samnegård, MD; Pia Lundman, MD;
Per Tornvall, MD, PhD; Carl-Göran Ericsson, MD, PhD; Anders Hamsten, MD, PhD
Downloaded from http://circ.ahajournals.org/ by guest on July 31, 2017
Background—Exaggerated postprandial triglyceridemia is common in normolipidemic patients with coronary artery
disease (CAD). Alterations in the composition of triglyceride-rich lipoproteins (TRLs) are likely to underlie this
metabolic disturbance. However, the composition of very-low-density lipoproteins (VLDLs), which are the most
abundant postprandial TRLs, has never been defined in CAD patients.
Methods and Results—We examined postprandial changes in the number and composition of VLDLs in middle-aged,
normolipidemic CAD patients and control subjects. TRLs from 14 patients and 14 control subjects aged 45 to 55 years
were subfractionated by density gradient ultracentrifugation into Svedberg flotation rate (Sf) fractions ⬎400, 60 to 400,
and 20 to 60. The VLDLs were separated from chylomicron remnants by immunoaffinity chromatography. In CAD
patients, the postprandial concentrations of triglycerides and large (Sf 60 to 400) VLDL particles were elevated. In
addition, their postprandial large VLDLs were enriched in apolipoprotein (apo) C-I and their postprandial small (Sf 20
to 60) VLDL remnants were enriched with apo C-I and cholesterol.
Conclusions—Perturbed handling of postprandial triglycerides in normolipidemic CAD patients involves the accumulation
of apo C-I–rich large VLDL particles and the generation of small, apo C-I– and cholesterol-rich VLDL remnants.
(Circulation. 2000;101:227-230.)
Key Words: lipids 䡲 lipoproteins 䡲 atherosclerosis 䡲 myocardial infarction
E
Perturbations in the numbers and composition of postprandial
VLDL subfractions in CAD patients have never been carefully
defined. In the present study, we sought to test the hypothesis
that exaggerated postprandial triglyceridemia in normolipidemic
CAD patients might be due to an elevated number of postprandial VLDL particles from the liver. We also sought to define
whether there were abnormalities in the composition of postprandial VLDLs from CAD patients. Our studies revealed
abnormalities in the numbers and composition of postprandial
VLDLs in CAD patients. These abnormalities are likely to be
relevant to the pathogenesis of postprandial hyperlipidemia and
to the development of atherosclerotic disease.
xaggerated postprandial triglyceridemia has been reported to be common in humans with coronary artery
disease (CAD).1 A significant fraction of the postprandial
triglycerides are carried on intestinally derived chylomicrons.
However, most of the triglyceride-rich lipoproteins (TRLs)
that accumulate in the plasma after a meal are actually
VLDLs.2 The composition of postprandial VLDL is likely to
be important, both for understanding the mechanisms of the
accumulation of TRLs and for understanding how these
lipoproteins might contribute to the pathogenesis of atherosclerosis. An accumulation of VLDL after a meal might be
quite relevant to the pathogenesis of atherosclerotic disease,
because several clinical studies have suggested a link between the fasting levels of VLDLs and coronary atherosclerosis.3,4 In addition, the VLDLs that accumulate postprandially have an altered apolipoprotein (apo) and lipid
composition.5 In recent studies,6 specific families of TRLs, as
defined by their apolipoprotein content, appeared to be better
predictors of CAD progression than the total plasma concentrations of apolipoproteins or lipids.
Methods
Human Subjects
The subjects were 45- to 55-year-old male myocardial infarction
survivors (n⫽14) without congestive heart failure who had normal
fasting lipid and lipoprotein levels (LDL cholesterol ⬍4.5 mmol/L
and plasma triglycerides ⬍2.0 mmol/L; ie, ⬍90th percentile). The
control subjects were 14 healthy normolipidemic men matched for
Received July 16, 1999; revision received November 9, 1999; accepted November 18, 1999.
From the Atherosclerosis Research Unit, King Gustaf V Research Institute (J.B., S.B., A.H.), Division of Cardiology, Department of Medicine,
Karolinska Hospital (S.B., P.T., A.H.), and Division of Cardiology, Department of Medicine, Danderyd Hospital, Karolinska Institute (A.S., P.L.,
C.-G.E.), Stockholm, Sweden.
Correspondence to Dr Johan Björkegren, Gladstone Institute of Cardiovascular Disease, San Francisco General Hospital, UCSF, PO Box 419100, San
Francisco, CA 94141-9100. E-mail [email protected]
© 2000 American Heart Association, Inc.
Circulation is available at http://www.circulationaha.org
227
228
Circulation
TABLE 1.
January 25, 2000
Characteristics of Patients and Control Subjects
Patients
(n⫽14)
Controls
(n⫽14)
51⫾3.0
51⫾2.0
27⫾3.8
27⫾2.9
Apo E-3/E-3
10
9
Apo E-3/E-4
4
5
Age, y
2
BMI, kg/m
Genotype
Cholesterol, mmol/L
Plasma
5.10⫾0.63
5.15⫾0.68
VLDL
0.49⫾0.22
0.39⫾0.23
LDL
3.36⫾0.53
3.49⫾0.62
HDL
0.97⫾0.31
1.08⫾0.25
Plasma
1.42⫾0.43
1.33⫾0.41
VLDL
1.02⫾0.43
0.91⫾0.36
LDL
0.34⫾0.06
0.33⫾0.09
HDL
0.13⫾0.02
0.12⫾0.02
Figure 1. Changes in plasma triglycerides after ingestion of a
mixed meal in middle-aged healthy controls (n⫽14) and patients
with recent myocardial infarction (n⫽14). Blood samples were
drawn before and every hour until 6 hours after intake of test
meal. Values are mean⫾SEM. *P⬍0.05 vs controls. F and E
denote mean plasma triglyceride concentrations for patients and
controls, respectively.
Triglycerides, mmol/L
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Values are mean⫾SD. VLDL, d⬍1.006 g/mL lipoproteins; LDL, d⫽1.006 –
1.063 g/mL lipoproteins; HDL, d⬎1.063 g/mL lipoproteins. There were no
statistically significant differences between patients and controls.
age, smoking habits, alcohol consumption, waist-hip circumference
ratio, and blood pressure (Table 1). All subjects gave informed
consent to the study, which was approved by the ethics committee of
the Karolinska Hospital.
n⫽8). The lipids were determined enzymatically.5 Apo B and apo
E were quantified by SDS-PAGE, and apo Cs were determined by
urea gel electrophoresis.5
Statistical Analysis
Differences between CAD patients and the control group were
assessed by Mann-Whitney tests. Within-group comparisons of
measurements from the fasting state to various time points during the
fat-tolerance test were assessed by Wilcoxon signed rank test.
Associations between lipoprotein parameters were assessed by calculation of Spearman correlation coefficients.
Results
Plasma Triglycerides and Apo B-48 and B-100
Concentrations in TRL Fractions
TRL Separation and Lipid and
Apolipoprotein Determination
Participants underwent a mixed meal-type oral fat-tolerance test.5
TRLs were subfractionated by cumulative flotation in a density
gradient into Svedberg flotation rate fractions (Sf) ⬎400, 60 to
400, and 20 to 60. Within these fractions, VLDLs and VLDL
remnants were purified from chylomicron remnants by immunoaffinity chromatography with apo B-100 –specific monoclonal
antibodies 4G3 and 5E11.5 The recovery of TRLs from the
immunoaffinity chromatography was 90⫾5.3% (mean⫾SD,
Although postprandial plasma triglyceride levels were elevated in the CAD patients (Figure 1), they returned toward
fasting levels at 6 hours. The large (Sf 60 to 400) VLDL and
chylomicron remnants remained elevated in the patients 6
hours after the meal compared with fasting levels (Table 2;
P⬍0.05 for apo B-48, P⬍0.005 for apo B-100). The CAD
patients had significantly more large VLDL particles in their
TABLE 2. Apo B-48 and Apo B-100 Concentrations in Sf >400, Sf 60 – 400, and
Sf 20 – 60 Fractions of TRLs in Fasting and Postprandial Plasma Samples From
Healthy Controls and Patients With Previous Myocardial Infarction
Time After Test Meal, h
0
3
6
Patients
Controls
Patients
Controls
Patients
Controls
Apo B-48
ND
ND
0.5⫾0.3
0.4⫾0.3
ND
ND
Apo B-100
ND
ND
0.6⫾0.6*
0.2⫾0.1
ND
ND
Apo B-48
0.8⫾0.9
0.5⫾0.5
2.0⫾0.8‡
2.0⫾1.3‡
1.4⫾0.9†
0.9⫾0.7
Apo B-100
28⫾14
22⫾15
42⫾21‡
37⫾21‡
42⫾23‡*
28⫾23
Apo B-48
1.2⫾0.9
0.8⫾0.7
1.3⫾0.8
1.4⫾1.0‡
0.9⫾0.5
1.0⫾0.6
Apo B-100
50⫾21
37⫾23
40⫾17†
34⫾11
46⫾23
43⫾31
Sf ⬎400
Sf 60–400
Sf 20–60
ND indicates not detectable in all subjects. Values are mean⫾SD in mg/L; n⫽14.
*P⬍0.05 compared with controls.
†P⬍0.05, ‡P⬍0.005 compared with baseline values.
Björkegren et al
Postprandial VLDL Composition and CAD
229
plasma 6 hours after the test meal than the control subjects
(P⬍0.05).
Composition of Fasting and Postprandial VLDL
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The most striking aspect of VLDL composition was the
postprandial increase in apo C-I in VLDLs from CAD
patients (Figure 2). At 6 hours, large and small (Sf 20 to 60)
VLDLs from CAD patients had 50% to 100% more apo C-I
than VLDLs from controls (P⬍0.005, P⬍0.05).
In contrast to the apo C-I content, the apo C-II and C-III
contents of VLDL were unaffected by the oral fat load. The
apo E content of large VLDLs increased after the fatty meal
in both groups (Figure 2) but remained elevated only in the
CAD group at 6 hours (P⬍0.005 versus fasting values).
The cholesterol content of small VLDLs in CAD patients
had increased significantly at the end of the postprandial
period and was then significantly higher in patients than in
controls (P⬍0.05). Interestingly, the cholesterol content correlated positively to the apo C-I content of small postprandial
VLDL particles in the CAD patients (r⫽0.73, n⫽14,
P⬍0.005 at 6 hours).
Discussion
Figure 2. Changes in VLDL apolipoprotein and lipid content (number
of molecules per particle) in Sf 60 to 400 and Sf 20 to 60 lipoprotein
fractions after ingestion of a mixed meal in middle-aged healthy controls and CAD patients with recent myocardial infarction. Blood samples were drawn before and 3 and 6 hours after intake of test meal.
Values are mean⫾SEM (n⫽14). *P⬍0.05 and **P⬍0.005 vs controls;
†P⬍0.05 and ‡P⬍0.005 vs fasting levels. F and E denote VLDL from
patients and controls, respectively.
Twenty years ago, Zilversmit7 suggested that postprandial
lipoproteins might be particularly atherogenic. Since then,
much of the research in this area has focused on intestinally
derived chylomicrons and chylomicron remnants. However,
most of the TRLs that accumulate in postprandial plasma are
not chylomicrons but VLDLs, which originate in the liver. In
the present study, we sought to define compositional differences in the postprandial VLDL subfractions in normolipidemic CAD patients and controls. Using a combination of
ultracentrifugation techniques and immunoaffinity chromatography, we were able to purify and analyze VLDL subfractions after an oral fat-tolerance test in both CAD patients and
matched controls. In our study, the CAD patients had an
expected exaggerated postprandial triglyceridemia compared
with the controls. In addition, the CAD patients manifested a
postprandial accumulation of large, apo C-I–rich VLDLs, and
their small VLDL remnants were enriched with apo C-I and
cholesterol.
Interestingly, we found no significant differences in the
apo C-II and apo C-III concentrations in the VLDLs of CAD
patients and controls, even though both of those apolipoproteins are known to influence the metabolism of TRLs. The
fact that apo C-I was uniquely elevated in the postprandial
large VLDL of CAD patients suggests that it could be pivotal
in causing the postprandial accumulation of those lipoproteins. We suspect that the enrichment in apo C-I causes a
delay in VLDL clearance, even though these lipoproteins
contained large amounts of apo E, the key ligand that
mediates the uptake of hepatic lipoproteins. This idea is
supported by in vitro studies, which have shown that apo C-I
enrichment of TRLs inhibits apo E-mediated uptake by both
the LDL receptor8 and the LDL receptor–related protein
pathways.9 Interestingly, in vivo studies have suggested that
apo C-I enrichment of VLDLs has no effect on the binding
capacity of these particles to proteoglycans in the arterial wall
and does not interfere with TRL hydrolysis by lipoprotein
230
Circulation
January 25, 2000
lipase.10 In agreement with those studies, it is likely that the
small postprandial VLDLs rich in apo C-I were generated in
part from lipoprotein lipase–mediated hydrolysis of apo
C-I–rich large VLDL particles.
The small postprandial VLDLs from CAD patients were, in
addition to apo C-I, enriched in cholesterol. On the basis of
the strong positive correlation between the number of apo C-I
and cholesterol molecules on these particles, we strongly
suspect that an apo C-I–mediated delay in VLDL clearance
contributes to the cholesterol enrichment of smaller VLDL
particles, simply because it prolongs the VLDL circulation
time and extends the time during which cholesterol transfer
from HDL could occur. The cholesterol enrichment of small
VLDLs could be very relevant to the development of premature coronary atherosclerosis in CAD patients, because small
VLDL remnants, like LDLs, are thought to be very susceptible to retention within the arterial intima.11
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Acknowledgments
This study was supported by the Swedish Medical Research Council
(8691), the Swedish Heart-Lung Foundation, and the Marianne and
Marcus Wallenberg Foundation. We thank Dr Stephen G. Young for
editorial assistance and Dr Peter Bacchetti for statistical advice.
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Accumulation of Apolipoprotein C-I−Rich and Cholesterol-Rich VLDL Remnants During
Exaggerated Postprandial Triglyceridemia in Normolipidemic Patients With Coronary
Artery Disease
Johan Björkegren, Susanna Boquist, Ann Samnegård, Pia Lundman, Per Tornvall, Carl-Göran
Ericsson and Anders Hamsten
Downloaded from http://circ.ahajournals.org/ by guest on July 31, 2017
Circulation. 2000;101:227-230
doi: 10.1161/01.CIR.101.3.227
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