Download Association of lipoprotein-associated phospholipase A2 levels with

European Heart Journal (2005) 26, 137–144
doi:10.1093/eurheartj/ehi010
Clinical research
Association of lipoprotein-associated phospholipase
A2 levels with coronary artery disease risk factors,
angiographic coronary artery disease, and major
adverse events at follow-up
Emmanouil S. Brilakis1{, Joseph P. McConnell2, Ryan J. Lennon3,
Ahmad A. Elesber1, Jeffrey G. Meyer2, and Peter B. Berger4*
1
Division of Cardiovascular Diseases, Department of Internal Medicine, Mayo Clinic, 200 First Street SW,
Rochester, MN 55905, USA
2
Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN, USA
3
Division of Biostatistics, Mayo Clinic, Rochester, MN, USA
4
Division of Cardiovascular Diseases, Duke Clinical Research Institute, Durham, NC 27715, USA
Received 20 April 2004; revised 1 August 2004; accepted 3 September 2004; online publish-ahead-of-print 29 November 2004
See page 107 for the editorial comment on this article (doi:10.1093/eurheartj/ehi042)
KEYWORDS
Lipoprotein-associated
phospholipase A2;
C-reactive protein;
Acute myocardial infarction;
Coronary disease
Aims We aimed to evaluate the association of lipoprotein-associated phospholipase
A2 (Lp-PLA2) with coronary artery disease (CAD) risk factors, with the severity of
angiographic CAD, and with the incidence of major adverse events.
Methods and results We measured Lp-PLA2 levels in 504 consecutive patients undergoing clinically indicated coronary angiography. Mean age was 60 + 11 years and 38%
were women. The mean (+SD) Lp-PLA2 level (ng/mL) was 245 + 91. Lp-PLA2 levels
correlated with male gender, LDL, HDL, and total cholesterol, fibrinogen, and creatinine. Lp-PLA2 levels correlated with the extent of angiographic CAD on univariate but
not on multivariable analysis. During a median follow-up of 4.0 years, 72 major
adverse events occurred in 61 of 466 (13%) contacted patients (20 deaths, 14 myocardial infarctions, 28 coronary revascularizations, and 10 strokes). Higher Lp-PLA2
levels were associated with a greater risk of events: the hazard ratio per SD was
1.28 (95% CI 1.06–1.54, P ¼ 0.009), and remained significant after adjusting for clinical and lipid variables and C-reactive protein.
Conclusion Higher Lp-PLA2 levels were associated with a higher incidence of
major adverse events at follow-up, independently of traditional CAD risk factors
and C-reactive protein.
Introduction
Lipoprotein-associated phospholipase A2 (Lp-PLA2) is a
50-kDalton enzyme that belongs to the A2 phospholipase
* Corresponding author. Tel: þ1 919 668 8355; fax: þ1 919 668 7058.
E-mail address: [email protected]
Present address: University of Texas Southwestern Medical School,
5323 Harry Hines Blvd, Dallas, TX 75216, USA
{
superfamily.1 Lp-PLA2 is produced by macrophages
and lymphocytes and 80% of it circulates bound, mainly
to LDL.2 Whether Lp-PLA2 is predominantly proatherogenic or anti-atherogenic is controversial. Most
evidence from animal3 and human2–5 studies suggests it is
pro-atherogenic.
The first aim of our study was to examine the association between Lp-PLA2 and: (i) traditional and emerging
coronary artery disease (CAD) risk factors: age, gender,
European Heart Journal vol. 26 no. 2 & The European Society of Cardiology 2004; all rights reserved.
138
smoking, hypertension, obesity, total, high-density lipoprotein (HDL), and low-density lipoprotein (LDL) cholesterol, triglycerides, and homocysteine; (ii) the presence
of an acute coronary syndrome; (iii) the presence and
extent of angiographic CAD; and (iv) the incidence of
major adverse events at follow-up. The second aim was
to examine the impact on those associations when
other risk factors, such as clinical and lipid parameters
and C-reactive protein (CRP) were included in multivariable analyses.
Methods
Patient population
The population included 504 patients (97% Caucasian), aged
26–76 years, undergoing clinically-indicated coronary angiography at our institution between June 1998 and January 1999.
Patients were excluded if they had diabetes mellitus, smoking
history .50 pack-years, history of organ transplantation, pregnancy, prior coronary revascularization, bleeding disorders,
blood transfusion within 30 days, HIV infection, renal failure,
or prior chest radiation therapy. The most frequent indications
for angiography were acute coronary syndrome (34%), an abnormal nuclear imaging study (25%), and dyspnoea upon exertion
(27%). The 504 patients represent .90% of patients eligible for
this study during the enrolment period (the remaining patients
refused to participate). The study was approved by our Institutional Review Board, and all subjects provided written
informed consent. Four hundred and sixty-six patients (92.5%)
were contacted by a follow-up questionnaire or by telephone
in September 2002. The remaining 38 patients either refused
to participate in the follow-up (18, 47%) or could not be
contacted (20, 53%). The medical records of the patients who
had an event were obtained and reviewed in order to ascertain
the type of the event or the cause of death.
Acute coronary syndromes and stroke
Patients were classified as having unstable angina (UA) if they
had new-onset chest pain or if they had a significant unexplained
change in the pattern of stable angina (such as increased frequency, intensity, or duration, or decreased response to nitrates)
in the previous 2 months. Patients were defined as having an
acute myocardial infarction (AMI) if they had cardiac marker
elevation [total creatinine kinase (CK) more than 3 the upper
limit of normal, or cardiac troponin (T) more than the upper
limit of normal] in association with chest pain or ischaemic
electrocardiographic changes. Stroke was defined as a new
neurological defect persisting for .24 h (CT or MRI documentation was available in 9 of 10 patients).
Angiographic analysis
Coronary angiograms were analysed according to the segmental
classification proposed by the Coronary Artery Surgery Study
(CASS) investigators.6 The maximum-diameter stenosis in each
of 27 coronary artery segments was assessed with hand-held
callipers or visual analysis. Angiograms were analysed blinded
to risk factors and biochemical analyses.
The extent of CAD was quantified as follows: normal coronaries (smooth arteries with no stenosis or stenosis ,10%),
mild disease (reduction in luminal diameter between 10% and
E.S. Brilakis et al.
50%), single-vessel disease (.50% luminal diameter stenosis in
one coronary artery or its major branches), two-vessel coronary
artery disease (.50% stenoses in two coronary arteries), and
three-vessel disease (.50% stenoses in three coronary arteries).
Parameter definitions
The body mass index (BMI) was calculated by dividing the
patient’s weight in kilograms by the square of the patient’s
height in metres. Patients were classified as normal weight
(BMI 18.5–24.9), overweight (BMI 25–29.9), and obese (BMI
30). Patients were considered to be hypertensive if their
blood pressure was .140/90 mmHg, or if they were being
treated with antihypertensive medications. Major adverse
events consisted of any of the following: death, AMI, coronary
revascularization, or stroke.
Blood collection and biochemical analyses
Blood was collected in EDTA-treated tubes and divided into
aliquots for measurement of plasma risk factors. Fibrinogen
was measured using immunoturbidimetric methods on a Roche
COBAS MIRA system. Lipids (total cholesterol, triglycerides,
and HDL cholesterol) were measured using standard automated
enzymatic methods on a Roche COBAS MIRA system. LDL cholesterol was calculated as total cholesterol minus HDL cholesterol
and 20% of the triglyceride level (all expressed in mg/dL).
Total plasma homocysteine was measured by high performance liquid chromatography following reduction of the
disulfide bonds with Tris (2-carboxyethyl) phosphine hydrochloride and derivatization with SBD-F (7-fluoro-2-oxa-1,
3-diazole-4-sulfonate). Cysteamine was used as an internal
standard.
CRP was measured using a sensitive latex particle-enhanced
immunoturbidimetric assay on a Hitachi 912 automated analyser,
using reagents from Kamiya Biomedical Company. The CRP assay
was sensitive to 0.15 mg/L and was standardized against the
IFCC/BCR/CAP CRM 470 CRP references.
Lp-PLA2 measurement
Lp-PLA2 mass was measured in plasma aliquots that were collected at the time of enrolment and stored at 2708C using an
enzyme-linked immunoassay (PLACTM test, diaDexus, Inc., CA,
USA).2,4 Samples were incubated in microtitre plate wells with
immobilized monoclonal antibody (2C10) against Lp-PLA2. The
enzyme was identified by a second monoclonal anti-Lp-PLA2
antibody (4B4) labelled with horseradish peroxidase. The standard was recombinant Lp-PLA2. The range of detection was
50–1000 ng/mL and the interassay coefficients of variation
were 7.8% at 276 ng/mL, 6.1% at 257 ng/mL, and 13.5% at
105 ng/mL. There was no cross-reactivity with other A2
phospholipases.2 All analyses were performed blinded to risk
factors, biochemical, and clinical characteristics.
The mean Lp-PLA2 value in our study was 245 + 91 ng/mL,
which was lower than the values reported from previous
studies [mean Lp-PLA2 level was 2370 + 520 ng/mL in the West
of Scotland Coronary Prevention Study (WOSCOPS) patients4].
Although the antibodies used in our method were the same
antibodies as used in previous studies, the recombinant Lp-PLA2
that we used as calibrator in the Lp-PLA2 assay had greater than
five-fold more immunological action than the standard used
previously. This accounts for the differences in recovered
values in the patient samples (Robert Wolfert, diaDexus Inc.,
personal communication).
Lipoprotein-associated phospholipase A2 levels
139
Statistical analysis
Associations of Lp-PLA2
Most continuous variables are summarized as mean + standard
deviation. Variables with heavily skewed distributions (CRP,
homocysteine, triglycerides, and creatinine) are reported as
medians, with first and third quartiles in parentheses. Discrete
variables are presented as frequencies and group percentages.
The association of continuous variables with the extent of
CAD (none, mild, one-, two-, or three-vessel disease) was
tested using a linear contrast in association with one-way
analysis-of-variance. The Armitage trend test was used to assess
the association between categorical variables and the extent
of CAD. Differences in distribution between other groups were
tested using one-way analysis of variance or the Kruskal–Wallis
test. Spearman’s correlation coefficient was used to assess
linear relationships between continuous variables.
Multiple regression models were used to estimate conditional
relationships. The covariates used in the logistic models for CAD
and Cox proportional hazards models for incidence of major
adverse events were age, gender, smoking history, hypertension,
total and HDL cholesterol, triglycerides, and CRP. Heavily
skewed variables were logarithmically transformed for use in
these models. The proportional hazards assumption was satisfied
for both Lp-PLA2 and log(CRP). Linearity was assessed for the
continuous variables in logistic and Cox regression models by
the use of generalized additive models. Splines were fitted and
the fitted results, plus point-wise 95% confidence intervals,
were plotted. In this way the linearity was assessed visually,
by evaluating whether a straight line would fit through the
confidence limits.
All hypothesis tests were two-tailed with a 0.05 Type I error rate.
Lp-PLA2 was significantly higher in men and was positively associated with creatinine, total and LDL cholesterol, and fibrinogen, and was negatively associated
with HDL (Tables 3 and 4). It was not significantly
associated with age, BMI, current smoking, hypertension,
systolic or diastolic blood pressure, triglycerides, homocysteine, or CRP.
CRP correlated with age, gender (higher in women),
history of hypertension, BMI, systolic blood pressure,
total cholesterol, LDL cholesterol, triglycerides, and
fibrinogen, but not with homocysteine or HDL cholesterol
(Tables 3 and 4).
Lp-PLA2 and angiographic CAD
Lp-PLA2 levels were higher in patients with more extensive angiographic CAD, even when AMI patients were
excluded (Table 1). However, after adjusting for clinical
and lipid variables (age, gender, smoking, hypertension,
total and HDL cholesterol, triglycerides, and CRP),
Lp-PLA2 was not independently predictive of angiographic CAD. CRP levels did not have a statistically
significant association with angiographic CAD on either
univariate (Table 1) or multivariable analysis (OR ¼ 1.13
per standard deviation, P ¼ 0.16).
Lp-PLA2 and major adverse events
Results
Patient characteristics
Table 1 summarizes the study population characteristics.
Mean age was 60.1 + 10.9 years and 38% were women.
Coronary angiography revealed normal coronaries in 122
patients (24%), mild disease in 111 patients (22%), and
one-, two-, and three-vessel disease in 85 (17%), 80
(16%), and 106 (21%) patients, respectively.
As expected, patients with significant CAD were more
likely to be male, older, and to have a history of hyperlipidaemia, hypertension, or myocardial infarction
(Table 1). Patients with significant CAD also had a
higher mean creatinine, LDL cholesterol, fibrinogen, and
Lp-PLA2, and lower HDL cholesterol. Mean Lp-PLA2 level
was 245 + 91 ng/mL, and its distribution is shown in
Figure 1.
Lp-PLA2 in acute coronary syndromes
Of the 504 patients, 169 had an acute coronary syndrome: 41 had an AMI and 128 had UA. Lp-PLA2 levels
were similar in patients with or without an acute coronary syndrome (Table 2). In contrast, median CRP was significantly higher in AMI patients (Table 2). Therefore,
analyses of the correlation between CRP and the extent
of angiographic CAD were performed in only the 463
patients who did not have an AMI at the time of study
enrolment (Tables 1, 3, and 4).
During a median follow-up of 4.0 years (interquartile
range 3.9–4.2 years), 72 major adverse events occurred
in 61 of 466 patients (the Kaplan–Meier estimated
event rate was 3.2% at 1 year and 10% at 4 years): 20
patients died (6 cardiac deaths), 14 had a myocardial
infarction, 26 underwent coronary revascularization
(15 percutaneous intervention only, 9 coronary artery
bypass surgery only, and 2 with both), and 10 had a
stroke. Seven patients had two events and two had
three (one patient had a myocardial infarction, percutaneous coronary intervention and coronary artery bypass
grafting surgery, and one had a myocardial infarction,
a stroke, and eventually died).
On univariate analysis, higher levels of both Lp-PLA2
and CRP (log-transformed) were associated with the
higher incidence of events: the hazard ratio (HR)
per standard deviation was 1.28 and 1.40, respectively
(Table 5). Figure 2 depicts the incidence of major
adverse events in the study population over time, classified according to Lp-PLA2 levels (tertiles), suggesting
that individuals in the lowest tertile had fewer events
during the follow-up period than those in the upper two
tertiles.
Cox proportional hazard models were developed to
examine the association of Lp-PLA2 with events. Those
models included Lp-PLA2, age, gender, smoking history,
hypertension, total and HDL cholesterol, triglycerides,
and log-CRP. Both Lp-PLA2 and log-CRP were independent
predictors of major adverse events (HR per standard
deviation 1.30 and 1.34, respectively) (Table 5).
140
Table 1 Clinical characteristics of the study population, classified according to the extent of CAD at angiography
Age, years
Female (%)
Hypertension (%)
Current smoking (%)
History of AMI (%)
History of CHF (%)
Family history of CAD (%)
History of hyperlipidaemia (%)
BMI, kg/m2
Creatinine, mmol/La
Total cholesterol, mmol/L
Triglycerides, mmol/La
HDL cholesterol, mmol/L
LDL cholesterol, mmol/L
Homocysteine, mmol/La
Fibrinogen, mg/dL
CRP, mg/La
Lp-PLA2, ng/mL
All patients
(n ¼ 504)
No CAD
(n ¼ 122)
Mild CAD
(n ¼ 111)
One-vessel CAD
(n ¼ 85)
Two-vessel CAD
(n ¼ 80)
Three-vessel CAD
(n ¼ 106)
P-value
60.1 + 10.9
192 (38)
232 (46)
40 (8)
77 (15)
59 (12)
128 (25)
286 (57)
29.3 + 5.6
97 (88–115)
5.4 + 1.2
1.7 (1.3–2.3)
1.2 + 0.4
3.2 + 0.9
8.8 (7.5–10.7)
455 + 125
2.9 (1.2–6.7)
245 + 91
53.7 + 11.3
76 (62)
39 (32)
8 (7)
3 (2)
25 (20)
27 (22)
44 (36)
28.8 + 6.6
97 (88–106)
5.3 + 1.1
1.5 (1.1–2.2)
1.4 + 0.5
3.0 + 0.9
8.6 (7.2–10.9)
422 + 113
3.2 (1.1–6.7)
223 + 85
61.9 + 9.5
55 (50)
50 (45)
9 (8)
6 (5)
19 (17)
24 (22)
52 (47)
29.2 + 6.0
97 (80–106)
5.3 + 1.1
1.8 (1.3–2.5)
1.3 + 0.4
3.1 + 0.9
8.6 (7.5–10.3)
442 + 121
2.5 (1.1–6.0)
248 + 80
59.9 + 11.1
22 (26)
42 (49)
12 (14)
16 (19)
6 (7)
20 (24)
45 (53)
29.1 + 4.3
106 (88–115)
5.2 + 1.0
1.8 (1.3–2.5)
1.2 + 0.3
3.2 + 0.8
8.7 (7.4–10.7)
466 + 144
2.9 (1.4–8.0)
243 + 73
61.2 + 10.4
18 (23)
46 (58)
7 (9)
20 (25)
3 (4)
28 (35)
66 (83)
30.8 + 5.8
106 (88–115)
5.4 + 1.0
1.7 (1.3–2.4)
1.1 + 0.3
3.4 + 1.0
8.8 (7.4–10.4)
472 + 126
2.9 (1.4–7.7)
251 + 97
64.7 + 8.7
21 (20)
55 (52)
4 (4)
32 (30)
6 (6)
29 (27)
79 (75)
29.1 + 4.3
106 (97–115)
5.6 + 1.5
1.8 (1.3–2.5)
1.2 + 0.3
3.5 + 1.1
9.6 (7.8–11.1)
487 + 118
3.0 (1.2–7.6)
263 + 114
,0.001
,0.001
,0.001
0.58
,0.001
,0.001
0.10
,0.001
0.21
,0.001
0.07
0.010
,0.001
0.001
0.026
,0.001
0.27
0.003
Plus–minus values are mean + standard deviation.
a
Median (interquartile range).
CHF, congestive heart failure.
E.S. Brilakis et al.
Lipoprotein-associated phospholipase A2 levels
141
and oxidatively modified non-esterified fatty acid, which
may promote atherogenesis by functioning as monocyte
chemoattractants and by inducing endothelial leukocyte
adhesion molecules.7,8
Lp-PLA2 and angiographic CAD
Figure 1
Distribution of Lp-PLA2 levels in the study population.
When LDL cholesterol was substituted for total cholesterol and fibrinogen was added to the model, the effect
of Lp-PLA2 remained statistically significant (HR per
standard deviation 1.25, 95% CI 1.02–1.52, P ¼ 0.03),
whereas the effect of log-CRP became non-statistically
significant (HR per standard deviation 1.10, 95% CI
0.79–1.51, P ¼ 0.58).
Discussion
Our study demonstrates that Lp-PLA2: (i) is associated
with gender, total LDL, and HDL cholesterol, fibrinogen,
and creatinine but not with other traditional or emerging
CAD risk factors such as age, hypertension, obesity, and
CRP; (ii) is not increased in AMI patients, in contrast to
acute-phase reactants such as CRP and fibrinogen; (iii)
is associated with angiographic CAD on univariate but
not multivariable analysis; and (iv) is associated with
the incidence of major adverse events at follow-up independently of clinical and lipid risk factors and CRP.
Associations of Lp-PLA2
Lp-PLA2 levels were measured in both men and women in
this study; two previous such studies included either men
(WOSCOPS sub-study),4 or women,5 but not both. In our
study, Lp-PLA2 levels were higher in men, but the difference was no longer significant after adjusting for HDL,
which was higher in women. The correlation between
Lp-PLA2 and LDL observed in both the current and the
WOSCOPS study was expected, since 80% of Lp-PLA2 is
associated with LDL.2 Similar to the WOSCOPS study,
Lp-PLA2 was not associated with age or BMI.
Both Lp-PLA2 and CRP were associated with lipid parameters (total cholesterol and LDL cholesterol; Tables 3
and 4). In contrast to what was reported in the
WOSCOPS study where there was a very weak correlation,
Lp-PLA2 had a significant negative association with HDL
cholesterol in our study (Table 4). CRP was associated
with obesity and AMI, whereas Lp-PLA2 was not. The
difference in those associations suggests that Lp-PLA2
may act on the atherosclerotic process through different
pathophysiological mechanisms than CRP. Lp-PLA2 cleaves
oxidized phosphatidylcholine on LDL in the vessel wall
to produce the bioactive lipids lysophosphatidylcholine
Caslake et al. 2 demonstrated that Lp-PLA2 levels were
higher in 94 patients with CAD than in 54 controls. The
association persisted after adjusting for LDL and HDL
cholesterol, smoking, and systolic blood pressure.
However, this study had several important limitations:
it was relatively small, only men were included, and
coronary angiography was performed in only one-third
of the patients, so that a possible association between
Lp-PLA2 levels and the severity of CAD could not be
explored.
In our population, Lp-PLA2 was higher in patients with
CAD than those without CAD, as also found by Caslake
et al. However, in our study the association between
Lp-PLA2 and CAD was not independent of other CAD risk
factors. Compared with the study by Caslake et al.,2
our study differed in the following three ways: (i) a
larger number of patients (504 vs. 148); (ii) data on the
severity and the extent of CAD, rather than just the presence of CAD; and (iii) inclusion of a significant proportion of women (192 of 504, 38%).
In the current study CRP did not correlate with the
angiographic extent of atherosclerosis. In the largest
published study correlating CRP with the extent of angiographic CAD, the correlation was weak (Pearson’s correlation coefficients 0.02–0.08), but reached statistical
significance because of the large sample size (n ¼ 2554).9
Lp-PLA2 and major adverse events
In our study, higher Lp-PLA2 levels were associated with a
higher incidence of major adverse events independently
of traditional CAD risk factors and CRP, a finding that is
consistent with the recently reported results from three
other studies.4,10
In the WOSCOPS population (exclusively men), Lp-PLA2
was a predictor of coronary events independent of CRP,
fibrinogen level, white cell count, age, systolic blood
pressure, plasma triglycerides, HDL cholesterol, and
LDL cholesterol levels (relative risk 1.18 per 1 standard
deviation increase in Lp-PLA2, P ¼ 0.005).4
In the Atherosclerosis Risk in Communities (ARIC)
study10 12 819 apparently healthy middle-aged men and
women were followed for 6 years: 609 individuals
developed CAD and were compared with 741 randomly
selected controls. In subjects with LDL cholesterol
below the median (130 mg/dL), Lp-PLA2 and CRP were
both significantly and independently associated with
CHD in fully adjusted models.
Similarly, in a post hoc analysis of the MONICA (MONItoring trends and determinants of CArdiovascular
disease) Augsburg cohort, Lp-PLA2 levels were significantly higher in the 97 of the 934 apparently healthy
men aged 45–64 who had suffered a coronary event
during 14 years of follow-up (data presented at the
142
E.S. Brilakis et al.
Table 2 Lp-PLA2, CRP, and fibrinogen levels in patients with and without an acute coronary syndrome
Lp-PLA2, ng/mL
CRP, mg/La
Fibrinogen, mg/dL
Total cholesterol, mmol/L
Triglycerides, mmol/La
HDL cholesterol, mmol/L
LDL cholesterol, mmol/L
AMI (n ¼ 41)
UA (n ¼ 128)
Non-ACS (n ¼ 335)
P-value
254 + 75
13.4 (5.9–32.8)
549 + 164
5.3 + 1.1
1.7 (1.3–2.10)
1.1 + 0.3
3.4 + 1.0
245 + 83
3.2 (1.3–6.6)
468 + 123
5.4 + 1.0
1.7 (1.3–2.6)
1.2 + 0.4
3.3 + 1.0
243 + 97
2.2 (1.0–5.6)
438 + 115
5.4 + 1.2
1.7 (1.3–2.3)
1.3 + 0.4
3.2 + 0.9
0.77
,0.001
,0.001
0.79
0.74
,0.001
0.39
Plus–minus values are mean + standard deviation.
a
Median (interquartile range).
ACS, acute coronary syndrome. Other abbreviations as in text and Table 1.
Table 3 Relationship between Lp-PLA2 and CRP levels with gender, current smoking, hypertension, and BMI
Lp-PLA2a
(n ¼ 504)
Lp-PLA2a
(n ¼ 463 without AMI)
CRPb
(n ¼ 463 without AMI)
Men
Women
P
254 + 87
229 + 97
0.003
254 + 88
227 + 97
0.002
2.0 (0.9–4.5)
3.6 (1.4–7.3)
,0.001
Current smoker
Current non-smoker
P
264 + 73
243 + 93
0.16
270 + 76
242 + 94
0.089
2.4 (1.1–6.0)
3.7 (1.7–4.7)
0.37
Hypertension
No hypertension
P
240 + 99
249 + 85
0.28
240 + 99
248 + 85
0.26
3.1 (1.4–6.2)
2.0 (0.9–5.5)
0.015
BMI , 24.9
BMI 25–29.9
BMI 30
P
242 + 89
243 + 80
248 + 103
0.84
240 + 89
243 + 81
247 + 106
0.82
1.7 (0.7–4.9)
2.1 (1.0–4.8)
3.5 (1.6–7.7)
,0.001
a
ng/mL.
mg/L, median, IQR.
Abbreviations as in text and Table 1.
b
Table 4 Relationship between Lp-PLA2, CRP, and other CAD risk factors
Lp-PLA2 (n ¼ 504)
Age
BMI
Systolic blood pressure
Diastolic blood pressure
Creatinine
Total cholesterol
Triglycerides
HDL cholesterol
LDL cholesterol
Homocysteine
Fibrinogen
CRP
Lp-PLA2
Abbreviations as in text.
Lp-PLA2 (n ¼ 463 without AMI)
CRP (n ¼ 463 without AMI)
Spearman’s rho
P
Spearman’s rho
P
Spearman’s rho
P
0.004
20.005
20.08
0.04
0.13
0.25
0.08
20.26
0.32
0.08
0.12
0.01
—
0.92
0.91
0.07
0.37
0.004
,0.001
0.06
,0.001
,0.001
0.09
0.007
0.83
—
0.02
20.007
20.08
0.05
0.13
0.24
0.09
20.26
0.32
0.08
0.12
20.02
—
0.71
0.88
0.11
0.33
0.007
,0.001
0.07
,0.001
,0.001
0.10
0.01
0.62
—
0.10
0.22
0.11
0.02
20.10
0.11
0.12
20.01
0.11
0.08
0.49
—
20.02
0.04
,0.001
0.01
0.60
0.03
0.02
0.01
0.76
0.02
0.07
,0.001
—
0.62
Lipoprotein-associated phospholipase A2 levels
143
Table 5 Univariate and multivariable association between different baseline parameters and the incidence of major adverse
events at follow-up
Parameter (SD)
Univariate HR (95% CI)
P
Multivariable HR (95% CI)a
P
Age (10.8 years)
Male gender
Smoking history
Hypertension
Total cholesterol (1.17 mmol/L)
HDL cholesterol (0.37 mmol/L)
log triglycerides (0.01 mmol/L)
log-CRP (1.32 mg/L)
Lp-PLA2 (92.8 ng/mL)
1.59
1.18
1.22
1.49
0.92
0.82
1.17
1.40
1.28
0.002
0.53
0.45
0.12
0.52
0.15
0.22
0.006
0.009
1.55
1.33
1.21
1.40
0.75
1.05
1.32
1.34
1.30
0.003
0.38
0.49
0.20
0.08
0.78
0.11
0.02
0.01
(1.19–2.13)
(0.70–2.01)
(0.73–2.03)
(0.90–2.48)
(0.70–1.19)
(0.62–1.07)
(0.91–1.51)
(1.10–1.79)
(1.06–1.54)
(1.16–2.08)
(0.70–2.54)
(0.70–2.09)
(0.84–2.35)
(0.55–1.04)
(0.74–1.50)
(0.94–1.87)
(1.05–1.72)
(1.06–1.59)
a
HRs were adjusted for Lp-PLA2, age, gender, smoking history, hypertension, total and HDL cholesterol, triglycerides, and log-CRP.
CI, confidence intervals. Other abbreviations as in text.
Figure 2
Incidence of major adverse events in the study population (n ¼ 466) classified according to Lp-PLA2 levels (tertiles).
2003 American Heart Association Scientific Session in
November 2003).
In contrast, Blake et al. 5 did not find an independent
association between Lp-PLA2 and cardiac events in a
nested case–control study of apparently healthy women
from the Women’s Health study, with 123 cases, 40% of
which were stroke.
Similar to the WOSCOPS, ARIC, and MONICA studies, in
our study higher Lp-PLA2 levels were associated with a
higher incidence of major adverse events independently
of other CAD risk factors and CRP, suggesting that
Lp-PLA2 may help in risk stratification of those patients.
In contrast to the ARIC study, the predictive role of
Lp-PLA2 appeared to be similar in patients with high or
low LDL. The discordance between the weak association
of Lp-PLA2 with angiographic CAD and the stronger
association of Lp-PLA2 with major adverse events has
also been observed with CRP, which was only weakly
associated with coronary artery calcification but was
strongly associated with clinical events in one study.11
As specific inhibitors of Lp-PLA2 have been developed
and shown to be orally active in animal models,12
Lp-PLA2 has the potential to be a therapeutic target in
patients with cardiovascular disease.11
Limitations
Measurements of Lp-PLA2 were performed on frozen
rather than fresh plasma. We demonstrated that
Lp-PLA2 is stable in samples stored at 48C or 2708C for
at least 7 days and that repeated freeze thaw cycles
(three cycles) did not reduce Lp-PLA2 concentration.
The effect of long-term storage is yet to be addressed.
Patients in this study were all referred for cardiac catheterization; therefore, our control patients may not be
representative of the population-based controls believed
to be free of atherosclerosis. However, the use of
patients with normal coronary arteries on angiography
also has strengths over using population-based controls,
because the presence of subclinical coronary disease
144
E.S. Brilakis et al.
can be excluded. Follow-up was obtained in 92.5% of
patients and it is possible that some events were not
detected. The incidence of major adverse events was
low (13%), limiting the power of our study, but we were
still able to detect a significant association between
Lp-PLA2 and events.
4.
Conclusions
5.
Lp-PLA2 was associated with different risk factors for CAD
than CRP. Higher Lp-PLA2 levels were associated with
more severe angiographic CAD on univariate but not on
multivariable analysis. Higher Lp-PLA2 levels were associated with a higher incidence of major adverse events
during follow-up, independently of traditional CAD risk
factors and CRP.
3.
6.
7.
8.
Acknowledgements
Supported in part by research grants from diaDexus Inc.,
South San Francisco, CA, USA and Interleukin Genetics,
Waltham, MA, USA.
9.
10.
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