Intercellular Adhesion Molecule-1 G241R Polymorphism Predicts Risk of Incident Ischemic Stroke
Atherosclerosis Risk in Communities Study
Background and Purpose— Intercellular adhesion molecule-1 levels are increased in pathological studies of atherosclerosis. We evaluated 13 491 participants from the Atherosclerosis Risk in Communities Study to determine the association of intercellular adhesion molecule-1 G241R and K469E polymorphisms with incident coronary heart disease and ischemic stroke.
Methods— Incidences of ischemic stroke (N=517) and coronary heart disease (N=1629) through 2003 were determined by annual telephone calls and hospital and death certificate surveillance. Risk factors were measured at the baseline examination. Cox proportional-hazards models were used to estimate hazard rate ratios.
Results— The intercellular adhesion molecule-1 G241RR genotype was associated with a significantly increased risk of ischemic stroke in both whites (hazard rate ratio=2.18; 95% CI, 1.01 to 4.68; P=0.05) and blacks (hazard rate ratio=7.04; 95% CI, 3.72 to 13.3; P<0.001).
Conclusions— The intercellular adhesion molecule-1 241RR genotype is associated with an increased risk of incident ischemic stroke in both whites and blacks.
Vascular inflammation, characterized by recruitment and adhesion of circulating leukocytes by cellular adhesion molecules on the endothelium, plays an important role in the pathogenesis of atherosclerosis.1,2 Intercellular adhesion molecule-1 (ICAM-1) plays an important role in the adhesion and subsequent transendothelial migration of circulating leukocytes into the vascular endothelium and is increased in pathological studies of atherosclerosis.1 Only a handful of studies have explored associations between ICAM-1 genetic variation and cardiovascular disease, with results proving contradictory.3–5 We evaluated participants from the large (N=15 792), biracial Atherosclerosis Risk in Communities (ARIC) Study to determine the association of the ICAM-1 G241R and K469E polymorphisms with incident coronary heart disease (CHD) and ischemic stroke.
Subjects and Methods
Participants were selected from the ARIC Study, a prospective investigation of atherosclerosis and its clinical sequelae involving 15 792 individuals age 45 to 64 years at recruitment (1987–1989). Institutional review boards approved the ARIC Study, and all participants provided written, informed consent. A detailed description of the ARIC Study design/methods and details on ascertainment/classification of CHD and stroke events have been published elsewhere.6–8 Participants were excluded from analyses (n=2301) if they had a positive or unknown history of prevalent stroke or CHD or a history of transient ischemic attack/stroke symptoms at baseline, prohibited use of their DNA, had an ethnic background other than white or black, or had missing information for both ICAM-1 genotypes or for the endpoints/covariates included in the analyses. Incident CHD cases (N=1629) were defined as a definite or probable myocardial infarction, a silent myocardial infarction detected by ECG, a definite CHD death, or a coronary revascularization. Incident ischemic stroke cases (N=517) were defined as validated definite/probable hospitalized embolic or thrombotic brain infarctions.
Seated blood pressure was measured 3 times with a random-zero sphygmomanometer, and the last 2 measurements were averaged. Hypertension was defined as systolic blood pressure ≥140 mm Hg, diastolic blood pressure ≥90 mm Hg, or current use of antihypertensive medications (assessed by questionnaires and in-person interviews). Diabetes was defined by a fasting glucose level ≥126 mg/dL, a nonfasting glucose level ≥200 mg/dL, and/or a history of or treatment for diabetes. Cigarette-smoking status compared current smokers with former/never smokers. Body mass index (in kg/m2) was calculated from height and weight measurements. Plasma total cholesterol was measured by an enzymatic method.9 HDL cholesterol was measured after dextran-magnesium precipitation of non-HDL lipoproteins.10 Plasma ICAM-1 levels were determined by ELISA (R&D Systems, Abingdon, UK).
Genotyping of the ICAM-1 G241R (rs1799969) and K469E (rs5498) polymorphisms was performed with the TaqMan assay (Applied Biosystems, Foster City, Calif). Primers and probes are available on request. The percentages of agreement for blind duplicate genotyping data were 98.7% for G241R and 98.2% for K469E.
All statistical analyses were conducted with STATA version 9.2 (College Station, Tex). Hardy-Weinberg equilibrium expectations were tested with a χ2 goodness-of-fit test. All analyses were done separately by race owing to ICAM-1 allele-frequency differences between whites and blacks. Cox proportional-hazards models were used to estimate the hazard rate ratios (HRRs) of incident CHD and ischemic stroke. For incident cases, follow-up time was defined as the time between the initial clinical visit and the date of the first event. For noncases, follow-up continued until December 31, 2003, the date of death, or the date of last contact if lost to follow-up, whichever came first. For CHD analyses, covariates included age, sex, field center, and traditional CHD risk factors (body mass index, HDL and total cholesterol, smoking, diabetes, hypertension). For stroke analyses, covariates included age, sex, field center, smoking, diabetes, and hypertension (identified by the National Institute of Neurological Disorders and Stroke; available at www.ninds.nih.gov). Covariates were assessed for statistical significance by the Wald χ2 statistic.
No significant differences in traditional cardiovascular/cerebrovascular risk factors (ie, body mass index, HDL or total cholesterol, smoking, diabetes, hypertension) were observed in whites or blacks for the ICAM-1 G241R or K469E polymorphisms (data not shown). Genotype frequencies for the ICAM-1 K469E polymorphism did not differ between CHD cases and noncases or between ischemic stroke cases and noncases (Table 1). Genotype frequencies for the ICAM-1 G241R polymorphism did not differ between CHD cases and noncases, but ICAM-1 241GR and 241RR genotypes were observed at a higher frequency in ischemic stroke cases compared with noncases for both racial groups (Table 1).
Genotypes containing the ICAM-1 241R allele (GR and RR) were a significant predictor of ischemic stroke in both whites (HRR=1.80; 95% CI, 1.34 to 2.43; P<0.001) and blacks (HRR=2.77; 95% CI, 1.68 to 4.57; P<0.001) (Table 2). No significant findings were observed for the ICAM-1 K469E polymorphism and incident ischemic stroke. Additionally, no significant findings were observed for analyses of incident CHD for either ICAM-1 polymorphism in whites or blacks (data not shown).
ICAM-1 levels were available only on a stratified random sample of the ARIC cohort (558 whites, 244 blacks). ICAM-1 G241R genotype distributions in this stratified random sample were similar to those observed in the entire ARIC cohort [f(R)=0.11 in whites, f(R)=0.04 in blacks]. The ICAM-1 G241R polymorphism was associated with higher ICAM-1 levels in both whites and blacks, but this finding was not statistically significant (Table 3).
The current study evaluated 2 polymorphisms located within exons 4 and 6 of the ICAM-1 gene. The ICAM-1 G241R polymorphism results in a glycine-to-arginine change within immunoglobulin-like domain 3, whereas the ICAM-1 K469E polymorphism results in a lysine-to-glutamine change in immunoglobulin-like domain 5. Immunoglobulin-like domain 3 mediates binding to macrophage antigen-1 (Mac-1) and may also affect accessibility of leukocyte function–associated antigen-1 (LFA-1) binding to the immunoglobulin-like domain 1.11–13 ICAM-1 mediates adhesive interactions of circulating leukocytes to the blood vessel wall by binding to Mac-1 and LAF-1, and increased expression of ICAM-1 has been found during all phases of atherogenesis.11–13 Additionally, molecular investigation of ICAM-1 variation has shown the G241R polymorphism to be a functional variant.14 Although our study did not investigate the mechanism(s) by which the ICAM-1 G241R polymorphism increases the risk of stroke, the observed association with stroke risk is potentially due to the functional impact of this variant on the binding affinity of ICAM-1 to both Mac-1 and LAF-1, thus possibly affecting adhesion and further interactions between circulating leukocytes and the vascular wall. On the basis of these results and those of previous studies, we hypothesize that the 241R allele enhances Mac-1 and LAF-1 binding to ICAM-1 while increasing the positive feedback on ICAM-1 synthesis. Further investigation of the ICAM-1 G241R polymorphism and its functional role in ischemic disease is warranted.
The authors thank the staff and participants of the ARIC study for their important contributions.
Source of Funding
The ARIC Study is carried out as a collaborative study supported by National Heart, Lung, and Blood Institute contracts N01-HC-55015, N01-HC-55016, N01-HC-55018, N01-HC-55019, N01-HC-55020, N01-HC-55021, and N01-HC-55022.
- Received December 8, 2009.
- Revision received January 19, 2010.
- Accepted January 25, 2010.
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