Printer Friendly

Effect of two common polymorphisms in the ATP binding cassette transporter A1 Gene on HDL-cholesterol concentration.

HDL-cholesterol (HDL-C) has long been recognized as having an atheroprotective role (1). Epidemiologic studies have shown that decreased HDL-C concentrations are the most common lipid abnormality in patients with premature coronary artery disease (CAD). Thus, there has been increased recognition of the inverse relationship between HDL-C concentrations and risk for CAD (2,3). This heightened awareness is underscored by changes in the recommendations of the Adult Treatment Panel (ATP) of the National Cholesterol Education Program. For example, ATP 11, published in 1993, first introduced the concept of a low HDL-C [<0.91 mmol/L (<35 mg/dL)] as a risk factor for CAD (4). More recently, ATP III further revised the cutoff for HDL-C as a risk factor to <1.04 mmol/L (<40 mg/dL) (5).

HDL-C is the primary lipoprotein particle responsible for reverse cholesterol transport (RCT) (6). RCT involves the transport of cholesterol from nonhepatic cells to the liver and its subsequent elimination from the body as bile acid and free cholesterol. A major advance in the understanding of RCT was heralded by the discovery of the gene encoding for ATP binding cassette transporter A1 (ABCA1). ABCA1, a member of the ABC transporter family, facilitates the active transport of cholesterol and phospholipids from the intracellular compartments of peripheral cells to the lipid-poor nascent HDL particle (7), thus representing the first step of the RCT pathway.

Deleterious mutations, when present in both alleles in the ABCA1 gene, were identified as the molecular basis for patients with Tangier disease, a disorder characterized by an almost complete absence of plasma HDL-C and an increased risk for CAD (8-10). Studies from our own laboratory, however, demonstrated that these detrimental mutations of the ABCA1 gene are probably not highly represented in the general population, even in a group of individuals with low HDL (11). On the other hand, many common ABCA1 polymorphisms have been reported, and at least some have been shown to influence plasma HDL-C concentrations and/or CAD progression and severity (12,13).

Clee et al. (12) investigated the effect of several polymorphisms on plasma HDL-C concentrations and the associated risk of CAD. Of nine nonsynonymous ABCA1 polymorphisms, only the 1051G/A polymorphism (R219K) was significantly correlated with HDL-C concentrations and CAD risk. Moreover, carriers of the 1051A allele were found to have higher HDL-C concentrations and reduced severity of CAD. The association between the ABCA1 1051G/A polymorphism and HDL-C concentrations was confirmed by Kakko et al. (13), but only in the female study population. Lutucuta et al. (14) studied the -477C/T polymorphism located in the promoter region of ABCA1 and found that it was associated with a trend toward higher plasma HDL-C concentrations. Other groups have also investigated the effect of ABCA1 polymorphisms on HDL-C concentrations and found no significant association (15) or an association only in a particular ethnic group (16).

In the current investigation, we studied the prevalence of two frequently occurring variants of the ABCA1 gene, the 1051G/A and the -477C/T polymorphisms, and the effect of these two polymorphisms on plasma HDL-C concentrations. We studied 838 patients with premature CAD, as documented by angiographically confirmed atherosclerosis and/or one or more episodes of myocardial infarction or coronary artery bypass surgery before age 55, recruited over a time period of 4 years from the Minneapolis Heart Institute. We also studied a cohort of the Minnesota Heart Study that included 257 apparently healthy individuals with no family history of CAD. The CAD group had a mean (SE) age of 49.5 (0.2) years and mean HDL-C of 0.91 (0.01) mmol/L. The group without CAD had a mean age of 48.3 (0.5) years and mean HDL-C of 1.24 (0.02) mmol/L. DNA was obtained from all individuals for polymorphism studies. This study was approved by the Institutional Review Board: Human Subjects Committee of the University of Minnesota, and all participants gave informed consent.

HDL-C was measured in serum by precipitation of the non-HDL-C with magnetic Mr 50 000 dextran sulfate and magnesium chloride, followed by colorimetric reflectance spectrophotometry on a Vitros analyzer (Johnson & Johnson Clinical Diagnostic, Inc.). Genomic DNA was extracted from peripheral leukocytes isolated from acid-citrate-dextrose-anticoagulated blood with use of commercially available DNA isolation reagents (Puregene; Gentra Systems).

To detect the 1051G/A polymorphism, we used a 433-bp fragment selectively amplified by PCR (primers: sense, 5'-CTCCAAAAGACTTCAAGGACCC-3'; anti-sense, 5'-GGCCCAAAAGTCTGAAAGAACAC-3'). The amplified fragment was digested with 4 U of StyI according to the manufacturer's instructions (New England Biolabs) and electrophoresed on 2% ultra PURE Agarose-1000 (Life Technologies) gel containing ethidium bromide. DNA from a patient homozygous for the G allele appeared as bands 189, 131, and 113 by in length relative to the size marker. Presence of the A allele abolishes a StyI cut site, and DNA from a patient homozygous for the A allele appears as bands of 320 and 113 bp. The -477C/T polymorphism was detected by selective PCR amplification of a 351-bp fragment (primers: sense, 5'-CTCGGGTCCTCTGAGGGACCT-3'; antisense, 5'-CCGCAGACTCTCTAGTCCAC-3'), followed by digestion of the amplified product with 3.5 U of AciI according to manufacturer's instructions (New England Biolabs). DNA from a patient homozygous for the C allele appears as bands of 148, 130, and 73 bp. Presence of the T allele abolishes an AciI cut site, and DNA from a patient homozygous for the T allele appears as bands of 278 and 73 bp.

Statistical analysis was performed by SPSS 10.0 for Windows. We used multivariate AMOVA to identify statistical differences at 95% confidence. The effect of individual genotypes and P values were determined by post hoc analysis using least-significant differences. Mean HDL-C concentrations were adjusted for sex and age. We used the Pearson [chi square] test to compare the genotype prevalence in the two populations. Significance was defined as P <0.05.

The prevalences of the ABCA1 1051G/A and -477C/T genotypes in 838 individuals with documented CAD and 257 control individuals are shown in Table 1. The prevalence of individuals homozygous for the 1051A allele was significantly higher (P <0.05) in the control group compared with the CAD group. We observed no significant difference in genotype frequency between the two populations for the -477C/T polymorphism.

The mean HDL-C concentrations in the CAD patients and controls classified according to ABCA1 1051G/A and -477C/T genotypes are also shown in Table 1. Individuals with the 1051AA genotype had a significantly higher (P <0.05) mean HDL-C concentration than did individuals with the 1051GG and 1051GA genotypes. This same trend was observed in the controls, but the difference did not reach statistical significance, in part because of the small number of individuals tested and, thus, the lack of statistical power of the control group. With respect to the -477C/T polymorphism, there were no significant differences in mean serum HDL-C concentrations in either group.

The results of the current study are the first reported for the 1051G/A polymorphism in a US population. The results confirm those of Clee et al. (12) and Kakko et al. (13), studies involving a French-Canadian and a Finnish cohort, respectively, that the 1051A allele is associated with a slightly higher HDL-C concentration. Additionally, our finding of a higher frequency of the 1051AA genotype in apparently healthy individuals vs individuals with documented CAD confirms the finding of Clee et al. (12) that the 1051A allele confers protection against CAD. Individuals with the 1051AA genotype had higher HDL-C concentrations than did individuals with the 1051GG and -GA genotypes, suggesting that the 1051AA genotype may confer protection against CAD, at least in part through its association with higher HDL-C concentrations. By confirming the findings of Clee et al. (12), our study strengthens the hypothesis that increased ABCA1 activity reduces the development of atherosclerosis by increasing the net efflux of cholesterol from peripheral cells.

With regard to the -477C/T polymorphism, we did not find a consistent association between the genotype and HDL-C or a significant difference in the allele frequency between CAD and control individuals. This suggests that the ABCA1 -477C/T polymorphism has little to no effect on plasma HDL concentrations. An earlier study (14), involving 429 members of a US cohort with coronary lesions, also showed that the -477C/T genotype did not affect serum HDL-C concentrations, but individuals with the T/T genotype had an increased number of lesions. Thus, additional studies are needed to determine whether this polymorphism is of significance in predisposing individuals to atherosclerosis.

In conclusion, we show that the 1051A allele of the ABCA1 gene is more prevalent in controls than in patients with CAD and that the apparent protective effect of the 1051A allele may be attributable in part to its association with moderately increased HDL-C concentrations. Thus, ABCA1 polymorphisms play a role in the polygenic regulation of HDL-C, as do variants in other genes, such as those encoding apolipoprotein A-1 and cholesteryl ester transfer protein. Because ABCA1 protein is the first step in RCT and is responsible for facilitating the transport of cholesterol across the membrane of extrahepatic cells, more studies are needed to determine whether increased ABCA1 protein plays an atheroprotective role beyond its association with serum HDL-C concentrations.


(1.) Miller GJ, Miller NE. Plasma high-density-lipoprotein concentration and ischaemic heart disease. Lancet 1975;1:16-9.

(2.) Gordon D, Rifkind BM. Current concepts: high density lipoproteins--the clinical implications of recent studies. N Engl J Med 1989;321:1311-5.

(3.) Wilson PWF, Abbott RD, Castelli WP. High density lipoprotein cholesterol and mortality: the Framingham Heart Study. Arteriosclerosis 1988;8:737-41.

(4.) Summary of the Second Report of the National Cholesterol Education Program (NCEP) Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults (Adult Treatment Panel II). JAMA 1993;269: 3015-23.

(5.) Executive Summary of the Third Report ofthe National Cholesterol Education Program (NCEP) Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults (Adult Treatment Panel III). JAMA 2001; 285:2486-97.

(6.) Fielding CJ, Fielding PE. Molecular physiology of reverse cholesterol trans- port. J Lipid Res 1995;36:211-28.

(7.) Oram JF, Lawn RM. ABCA1: the gatekeeper for eliminating excess tissue cholesterol. J Lipid Res 2001;42:1173-9.

(8.) Bodzioch M, Orsb E, Klucken J, Langmann T, Bbttcher A, Diederich W, et al. The gene encoding ATP-binding cassette transporter 1 is mutated in Tangier disease. Nat Genet 1999;22:347-51.

(9.) Rust S, Rosier M, Funke H, Real J, Amoura Z, Piette JC, et al. Tangier disease is caused by mutations in the gene encoding ATP-binding cassette transporter 1. Nat Genet 1999;22:352-5.

(10.) Oram JF. Tangier disease and ABCA1. Biochim Biophys Acta 2000;1529: 321-30.

(11.) Woll PS, Hanson NQ, Tsai MY. Absence of ABCA1 mutations in individuals with low serum HDL-cholesterol. Clin Chem 2003;49:521-2.

(12.) Clee SM, Zwinderman AH, Engert JC, Zwarts KY, Molhuizen HOF, Roomp K, et al. Common genetic variation in ABCA1 is associated with altered lipoprotein levels and a modified risk for coronary artery disease. Circulation 2001;103:1198-205.

(13.) Kakko S, Kelloniemi J, von Rohr P, Hoeschele I, Tamminen M, Brousseau ME, et al. ATP-binding cassette transporter A1 locus is not a major determinant of HDL-C levels in a population at high risk for coronary heart disease. Atherosclerosis 2003;166:285-90.

(14.) Lutucuta S, Ballantyne CM, Elghannam H, Gotto AM, Marian AJ. Novel polymorphisms in promoter region of ATP binding cassette transporter gene and plasma lipids, severity, progression, and regression of coronary atherosclerosis and response to therapy. Circ Res 2001;88:969-73.

(15.) Brousseau ME, Bodzioch M, Schaefer EJ, Goldkamp AL, Kielar D, Probst M, et al. Common variants in the gene encoding ATP-binding cassette transporter 1 in men with low HDL cholesterol and coronary heart disease. Atherosclerosis 2001;154:607-11.

(16.) Wang J, Burnett JR, Near S, Young K, Zinman B, Hanley AJ, et al. Common and rare variants affecting plasma HDL cholesterol. Arterioscler Thromb Vasc Biol 2000;20:1983-9.

DOI: 10.1373/clinchem.2004.047126

Petter S. Woll, Naomi Q. Hanson, Valerie L. Arends, and Michael Y. Tsai* (Department of Laboratory Medicine and Pathology, University of Minnesota Medical School, Minneapolis, MN 55455; * address correspondence to this author at: 420 Delaware Street SE, Mayo Mail Code 609, Minneapolis, MN 55455-0392; fax 612-625-5622, e-mail
Table 1. Prevalence of the ABCA1 1051G/A and 477C/T
genotypes in 838 CAD patients and 257 controls and
effect on HDL-C concentration.

 CAD patients Controls

 HDL-C,(a) HDL-C, (a)
Genotype n (%) mmol/L n (%) mmol/L

1051GG 450 (53.7) 0.90 (0.01)(b) 115 (44.7) 1.23 (0.03)
1051GA 327 (39.0) 0.91 (0.01)(b) 112 (43.6) 1.24 (0.03)
1051AA 61 (7.3)(c) 0.99 (0.03)(b) 30 (11.7)(c) 1.31 (0.06)
-- 477CC 237 (28.3) 0.90 (0.02) 71 (27.6) 1.29 (0.04)
-- 477CT 426 (50.8) 0.90 (0.01) 133 (51.8) 1.23 (0.03)
-- 477TT 175 (20.9) 0.94 (0.020) 53 (20.6) 1.21 (0.04)

(a) Values are the mean (SE) and are adjusted for age and sex.
(b)P< 0.05 for 1051AA vs 1051GG and 1051GA in CAD group.
(c)P< 0.05 for CAD patients vs controls.
COPYRIGHT 2005 American Association for Clinical Chemistry, Inc.
No portion of this article can be reproduced without the express written permission from the copyright holder.
Copyright 2005 Gale, Cengage Learning. All rights reserved.

Article Details
Printer friendly Cite/link Email Feedback
Title Annotation:Technical Briefs
Author:Woll, Petter S.; Hanson, Naomi Q.; Arends, Valerie L.; Tsai, Michael Y.
Publication:Clinical Chemistry
Date:May 1, 2005
Previous Article:Screening for dysbetalipoproteinemia by plasma cholesterol and apolipoprotein B concentrations.
Next Article:Experimental studies on capturing human leukocytes with cell immuno-chip.

Terms of use | Privacy policy | Copyright © 2019 Farlex, Inc. | Feedback | For webmasters