European Heart Journal

European Heart Journal Advance Access originally published online on May 3, 2006
European Heart Journal 2006 27(11):1331-1337; doi:10.1093/eurheartj/ehl018

This Article Abstract FREE Full Text (PDF) Supplementary tables All Versions of this Article: 27/11/1331    most recent ehl018v1 E-letters: Submit a response Alert me when this article is cited Alert me when E-letters are posted Alert me if a correction is posted Services Email this article to a friend Related articles in EHJ Similar articles in this journal Similar articles in ISI Web of Science Similar articles in PubMed Alert me to new issues of the journal Add to My Personal Archive Download to citation manager Search for citing articles in: ISI Web of Science (18) Request Permissions Disclaimer Google Scholar Articles by Kardys, I. Articles by Witteman, J. C.M. Search for Related Content PubMed PubMed Citation Articles by Kardys, I. Articles by Witteman, J. C.M. Social Bookmarking          What's this?

© The European Society of Cardiology 2006. All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org

C-reactive protein gene haplotypes and risk of coronary heart disease: the Rotterdam Study

Isabella Kardys¹, Moniek P.M. de Maat², André G. Uitterlinden^1,3, Albert Hofman¹ and Jacqueline C.M. Witteman^1,*

¹ Department of Epidemiology and Biostatistics, Erasmus MC, PO Box 1738, 3000 DR Rotterdam, The Netherlands
² Department of Hematology, Erasmus MC, Rotterdam, The Netherlands
³ Department of Internal Medicine, Erasmus MC, Rotterdam, The Netherlands

Received 30 November 2005; revised 10 March 2006; accepted 30 March 2006; online publish-ahead-of-print 3 May 2006.

^* Corresponding author. Tel: +31 10 4087488; fax: +31 10 4089382. E-mail address: j.witteman{at}erasmusmc.nl

See page 1261 for the editorial comment on this article (doi:10.1093.eurheartj/ehi852)

	Abstract

Top Abstract Introduction Participants and methods Results Discussion Supplementary material Acknowledgements References

 
Aims C-reactive protein is associated with risk of cardiovascular disease. However, whether C-reactive protein is a marker of severity of cardiovascular disease or actually is involved in its pathogenesis remains unknown. We investigated the relation between C-reactive protein haplotypes, representing the comprehensive variation of the C-reactive protein gene, and coronary heart disease.

Methods and results The Rotterdam Study is a prospective population-based study among men and women aged 55 years and older. C-reactive protein was associated with risk of coronary heart disease, with a multivariable adjusted hazard ratio of 1.9 (95% CI 1.5–2.4) for the highest vs. the lowest quartile. Four C-reactive protein haplotypes were present with overall frequencies of 32.8, 31.7, 29.5, and 5.9%. C-reactive protein serum levels were significantly different according to C-reactive protein haplotypes. C-reactive protein haplotypes were not associated with coronary heart disease.

Conclusion Steady-state C-reactive protein serum level is influenced by C-reactive protein gene haplotypes. Although elevated C-reactive protein level has lately been found to be a consistent and relatively strong risk factor for cardiovascular disease, our study does not support that the common variation in the C-reactive protein gene has a large effect on the occurrence of coronary heart disease.

Key Words: Coronary heart disease • Inflammation • C-reactive protein • Genetics • Epidemiology

	Introduction

Top Abstract Introduction Participants and methods Results Discussion Supplementary material Acknowledgements References

 
C-reactive protein is associated with cardiovascular disease.¹ However, whether C-reactive protein is merely a marker of severity of cardiovascular disease or actually is involved in its pathogenesis remains unknown. Genetic markers offer a possibility to study this.

Evidence has emerged that C-reactive protein may play a pathogenic role in cardiovascular disease.² If this is true, genetic variants associated with high C-reactive protein level may be associated with greater risk of cardiovascular disease. The genes involved in this regulation remain ill-defined. The C-reactive protein gene is likely to play a part, as several studies have demonstrated associations between single nucleotide polymorphisms (SNPs) in the C-reactive protein gene and C-reactive protein level.^3–13 Two recent studies have identified comprehensive sets of common C-reactive protein gene haplotypes and found associations of these haplotypes with C-reactive protein level.^14,15 One of these studies has also examined the association between these haplotypes and myocardial infarction or ischaemic stroke in a nested case–control study within the Physicians' Health Study cohort,¹⁵ but the association between C-reactive protein variants and baseline C-reactive protein did not correlate with the effects of those variants on clinical cardiovascular events in this study.

Seattle SNPs (part of the National Heart Lung and Blood Institute's Programs for Genomic Applications) reports that four C-reactive protein gene haplotypes are present in populations of European descent. These haplotypes represent all common variation across the C-reactive protein gene in these populations. To further clarify the role of C-reactive protein in coronary heart disease, we set out to investigate the relation between these four C-reactive protein gene haplotypes, C-reactive protein serum level, and coronary heart disease prospectively in all participants of the large, population-based Rotterdam Study.

	Participants and methods

Top Abstract Introduction Participants and methods Results Discussion Supplementary material Acknowledgements References

 
Study population and baseline data collection
The present study is part of the Rotterdam Study, a population-based cohort study aimed at assessing the occurrence of and risk factors for chronic diseases in the elderly. Objectives and methods of the Rotterdam Study have been described in detail elsewhere.¹⁶ The Rotterdam Study cohort includes 7983 men and women aged 55 years and over (78% of the eligible population), living in a well-defined suburb of the city of Rotterdam, The Netherlands. The Medical Ethics Committee of Erasmus Medical Center, Rotterdam, approved the study. Participants gave written informed consent and permission to retrieve information from treating physicians. This study complies with the Declaration of Helsinki.

Baseline data were collected from 1990 until 1993, as described previously.¹⁷ A trained interviewer visited all participants at home and collected information on current health status, medical history, drug use, and smoking, using a computerized questionnaire. In addition, in 7129 participants, established cardiovascular risk factors were measured at the research center.

Measurement of C-reactive protein
At baseline, non-fasting blood was collected. All tubes were stored on ice before and after blood sampling. High-sensitivity C-reactive protein was determined in serum, which was stored at –20°C until performance of the C-reactive protein measurements in 2003–04. C-reactive protein was measured using Rate Near Infrared Particle Immunoassay (Immage^® Immunochemistry System, Beckman Coulter, USA). This system measures concentrations ranging from 0.2 to 1440 mg/L, with a within-run precision <5.0%, a total precision <7.5%, and a reliability coefficient of 0.995.

Genotyping
The Seattle SNPs Program for Genomic Applications has identified 31 SNPs in the C-reactive protein gene and has established that, based on SNPs with overall frequencies above 5%, four common C-reactive protein gene haplotypes are present in 23 unrelated individuals of European descent from the CEPH pedigrees (http://pga.gs.washington.edu/data/crp, ‘visual haplotype’ option). These four haplotypes can be identified by ‘haplotype tagging’ SNPs. By genotyping three haplotype tagging SNPs we were able to infer all four haplotypes and consequently to describe the common variation across the C-reactive protein gene (Figure 1). These three tagging SNPs were chosen partly based on their presence in existing literature and on their proximity to the C-reactive protein gene. Other SNPs were also eligible, as a range of SNP trios across the C-reactive protein gene captures the four most common haplotypes among European participants.

View larger version (11K): [in this window] [in a new window]   Figure 1 The C-reactive protein gene, C-reactive protein gene polymorphisms determined in this study, and common C-reactive protein gene haplotypes.

 
All participants were genotyped for the 1184 C>T, 2042 C>T, and 2911 C>G SNPs of the C-reactive protein gene. The polymorphisms are described in relation to the start of the coding sequence of exon 1 using the Human May 2004 (hg 17) assembly (http://genome.ucsc.edu). These polymorphisms have also been described at http://www.ncbi.nlm.nih.gov/SNP under identification numbers rs1130864 (1184 C>T), rs1205 (2042 C>T), and rs3093068 (2911 C>G).

DNA was extracted according to standard procedures. DNA was solubilized in double-distilled water and stored at –20°C until used for DNA amplification. Genotypes were determined in 2 ng genomic DNA with the Taqman allelic discrimination assay (Applied Biosystems, Foster City, CA, USA). Primer and probe sequences were optimized using the SNP assay-by-design service of Applied Biosystems (http://store.appliedbiosystems.com). Reactions were performed with the Taqman Prism 7900HT 384 wells format. Haplotype alleles present in the population were inferred by means of the haplo.em function of the program Haplo Stats (http://cran.r-project.org/src/contrib/Descriptions/haplo.stats.html), which computes maximum likelihood estimates of haplotype probabilities.^18,19 Haplotype reconstruction resulted in seven haplotypes, but the fifth, sixth, and seventh haplotypes were present in <0.001% of the alleles and were therefore not used in the analyses. Haplotype alleles were coded as haplotype numbers 1 through 4 in the order of decreasing frequency in the population: coding from 1184 C>T, 2042 C>T, and 2911 C>G, haplotype 1=C-T-C, 2=T-C-C, 3=C-C-C, and 4=C-C-G (Figure 1).

Follow-up procedure
Follow-up started at the baseline examination and for the present study lasted until January 1, 2002. Information on fatal and non-fatal cardiovascular endpoints was obtained from general practitioners (GPs) and letters and discharge reports from medical specialists.¹⁷ Two research physicians independently coded all reported events according to the International Classification of Diseases, 10th edition (ICD-10).²⁰ In case of disagreement, consensus was reached. A medical expert in cardiovascular disease, whose judgment was considered final, reviewed all events.

We defined incident coronary heart disease as myocardial infarction, coronary artery bypass grafting (CABG), percutaneous transluminal coronary angioplasty (PTCA), and cardiac death. In identifying myocardial infarctions, all available information, which included ECG, cardiac enzyme levels, and the clinical judgment of the treating specialist, was used. We defined cardiac death as death from myocardial infarction or other ischaemic heart disease (ICD-10: I20–I25), sudden cardiac death (I46), sudden death undefined (R96), or death from heart failure (I50).

Population for analysis
C-reactive protein serum levels were available for 6658 participants. C-reactive protein measurements were lacking for participants who did not visit the research center (854) and for participants of whom no blood was available due to logistic reasons (471). After excluding participants with coronary heart disease at baseline (870), defined as a history of myocardial infarction, PTCA, or CABG, 5788 participants were left for the analysis of the association between C-reactive protein serum levels and coronary events.

DNA was available for 6571 participants. Genotyping of all three polymorphisms was successful in 6007 participants. For 5584 of these participants, C-reactive protein serum levels were available. After excluding participants with coronary heart disease at baseline, 5231 participants were left for the analysis of the association between C-reactive protein haplotypes and coronary events.

Data analysis
Linear regression was used to investigate the association between C-reactive protein serum levels and established cardiovascular risk factors. After log-transformation of C-reactive protein, the residuals were normally distributed with a constant variance.

Subsequently, participants with coronary heart disease at baseline were excluded, and Cox proportional hazards analysis was used to determine the relative risks of coronary heart disease and myocardial infarction associated with increasing C-reactive protein quartiles (cut-points 0.9, 1.8, and 3.5 mg/L). The proportional hazards assumption was tested by drawing log minus log plots of the survival function. We adjusted for age and sex (model 1), and subsequently for age, sex, body mass index (BMI), systolic blood pressure, diastolic blood pressure, total cholesterol, HDL-cholesterol, smoking, and diabetes mellitus (model 2).

Hardy-Weinberg equilibrium of the three C-reactive protein gene polymorphisms was tested using a ² test. Differences in serum C-reactive protein levels (log-transformed) and established cardiovascular risk factors for the three polymorphisms were examined using analysis of covariance, adjusting for age and sex, categorizing the participants by their genotypes. We used the Bonferroni correction to account for multiple testing (three genotypes). All the above analyses were performed using SPSS 11.0 for Windows.

To test the associations of C-reactive protein gene haplotypes with cardiovascular risk factors, we used the program Haplo Stats http://cran.r-project.org/src/contrib/Descriptions/haplo.stats.html).^18,19,21 The probability for each haplotype pair in each individual was assigned and then an individual's phenotype was directly modelled as a function of each inferred haplotype pair, weighed by their estimated probability, to account for haplotype ambiguity. The haplo.score function of Haplo Stats was used to test the associations. Details on the background and theory of score statistics can be found in Schaid et al.²¹ We adjusted for age and sex and we computed global simulation P-values and simulation P-values for each haplotype. The number of simulations was set as 1000.

As haplo.score does not provide the magnitude of the effect of each haplotype, the association between C-reactive protein gene haplotypes and C-reactive protein serum level, coronary heart disease and myocardial infarction was investigated using the haplo.glm function of Haplo Stats.¹⁹ This approach is based on a generalized linear model, and computes the regression of a trait on haplotypes and other covariates. For the analysis regarding the disease outcomes, the haplotype that was found to be associated with the lowest serum C-reactive protein levels served as the reference category. First, we adjusted for age and sex, and second, we additionally adjusted for BMI, systolic blood pressure, diastolic blood pressure, total cholesterol, HDL-cholesterol, smoking, and diabetes mellitus. Haplo.em, haplo.score, and haplo.glm were all implemented in the Haplo Stats software using the R language.

Values for cardiovascular covariates were missing in <4% of participants. These missing values were handled by single imputation using the expectation–maximization algorithm in SPSS 11.0. All tests were two-sided.

	Results

Top Abstract Introduction Participants and methods Results Discussion Supplementary material Acknowledgements References

 
Table 1 shows baseline characteristics of the total cohort and their associations with C-reactive protein serum level. All studied characteristics, except for total cholesterol, were significantly associated with C-reactive protein serum level.

View this table: [in this window] [in a new window]   Table 1 Baseline characteristics of the population and age- and sex-adjusted regression coefficients for cardiovascular risk factors, describing the increase in log-C-reactive protein per unit increase in each risk factor

 
The mean follow-up time was 8.1 years (SD 3.0 years). Among participants without history of coronary heart disease, 584 (8.8%) participants experienced incident coronary heart disease during follow-up, including 224 myocardial infarctions. Hazard ratios for coronary heart disease and myocardial infarction increased significantly across quartiles of C-reactive protein (Table 2). When we repeated the analysis without excluding participants with coronary heart disease at baseline, the results did not change materially.

View this table: [in this window] [in a new window]   Table 2 Hazard ratios for coronary heart disease and myocardial infarction for quartiles of C-reactive protein in participants without history of coronary heart disease at baseline

 
Genotype distributions for the three haplotype tagging SNPs were in Hardy-Weinberg equilibrium. Both using the Seattle SNPs website and the HapMap website (http://www.hapmap.org), the SNPs were found to lie in one linkage disequilibrium block. The 1184 T allele was present in 31.6% of 12 014 chromosomes, the 2042 T allele in 32.7%, and the 2911 G allele in 6.0%. Figure 2 shows differences in serum C-reactive protein levels according to the genotype of the three C-reactive protein polymorphisms. For all three polymorphisms we observed an allele dose effect. No associations were present between genotypes and established cardiovascular risk factors (see online supplementary material, webtables 1, 2, and 3). Genotypes were not associated with coronary heart disease and myocardial infarction (see online supplementary material, webtable 4).

View larger version (43K): [in this window] [in a new window]   Figure 2 Geometric means and 95% CI of serum levels of C-reactive protein (mg/L) for three C-reactive protein gene polymorphisms. *P-value <10–3.

 
Haplotype alleles were present in the following frequencies: haplotype 1 (C-T-C) in 32.8%, haplotype 2 (T-C-C) in 31.7%, haplotype 3 (C-C-C) in 29.5%, haplotype 4 (C-C-G) in 5.9%, and remaining haplotypes (T-C-G, T-T-C and C-T-G) in less than 0.001%.

Haplotype 4 was associated with lower BMI (P=0.03), haplotype 1 with higher systolic blood pressure (P=0.04), and haplotype 3 with higher percentage of prevalent myocardial infarction (P=0.04). No other associations with cardiovascular risk factors were present (see online supplementary material, webtable 5). All haplotypes provided significantly higher C-reactive protein levels than haplotype 1 (see online supplementary material, webtable 6). The effect of haplotype on C-reactive protein level, calculated from the regression coefficients, is displayed in Figure 3. Additional adjustment for BMI, systolic blood pressure, diastolic blood pressure, total cholesterol, HDL-cholesterol, smoking, and diabetes mellitus did not materially change the results.

View larger version (9K): [in this window] [in a new window]   Figure 3 Relative effects of C-reactive protein gene haplotypes on C-reactive protein serum levels (mg/L). *P-value<10–3. Regression coefficients were estimated with haplo.glm for each haplotype, adjusted for age and sex. Coefficients reflect difference in mean ln(C-reactive protein) per copy relative to haplotype 1, the most frequent haplotype. Values were transformed back to normal scale. Haplotypes 2–4 were all significantly higher than haplotype 1.

 
In Table 3, age- and sex-adjusted odds ratios (OR) for coronary heart disease and for myocardial infarction are displayed for different C-reactive protein haplotypes. As haplotype 1 was associated with the lowest serum C-reactive protein levels, it served as the reference. For both outcomes, the OR for all haplotypes were around one. Additional adjustment for BMI, systolic blood pressure, diastolic blood pressure, total cholesterol, HDL-cholesterol, smoking, and diabetes mellitus did not materially change the point estimates, and neither did repeating the analysis without excluding participants with coronary heart disease at baseline.

View this table: [in this window] [in a new window]   Table 3 Age- and sex-adjusted OR for coronary heart disease and myocardial infarction for C-reactive protein haplotypes in participants without history of coronary heart disease

 

	Discussion

Top Abstract Introduction Participants and methods Results Discussion Supplementary material Acknowledgements References

 
In this population-based study, elevated C-reactive protein serum level was a strong and independent marker of increased risk of coronary heart disease and myocardial infarction in participants without a history of coronary heart disease. C-reactive protein haplotypes were associated with C-reactive protein serum levels. However, C-reactive protein haplotypes were not associated with coronary heart disease and myocardial infarction.

The approach we used in this study has also been termed ‘Mendelian randomization’. This approach has been used recently in studies of C-reactive protein, hypertension, and metabolic syndrome.^8,22 It deals with residual confounding, as alleles of the C-reactive protein gene that influence C-reactive protein level are transmitted from parent to offspring at random, and factors that could confound associations of C-reactive protein level with cardiovascular disease should be evenly distributed in those who do, and those who do not, have alleles that cause high C-reactive protein level. Furthermore, it deals with reverse causation, because genotype is determined before onset of disease.²³

The present study uses haplotypes describing the total common variation of the C-reactive protein gene. So far, two other studies have used this approach. First, Carlson et al.¹⁴ defined all common genetic variation across the C-reactive protein gene region by resequencing the region in a multiethnic variation discovery panel (24 African–Americans and 23 European–Americans), and selected SNPs for genotyping in a larger panel (CARDIA study), in which associations between common haplotypes and C-reactive protein levels were investigated. Carlson et al. used a population that was partly of European descent and partly of African descent, and used all haplotypes that occur in these populations. We used a population of European descent; therefore, the studies are not strictly comparable. In approximation, Carlson et al.'s haplotypes 4, 5, and 7 concur with our haplotypes 3, 2, and 4, respectively, and Carlson et al.'s haplotype 1 and 2 taken together concur with our haplotype 1. Remaining haplotypes in the Carlson et al. study were only present in African–Americans. Carlson et al. found that their haplotypes 5 and 7 were associated with high C-reactive protein levels, haplotype 1 and 2 with the lowest levels, and haplotype 4 with intermediate levels. These results are in agreement with ours. Furthermore, Carlson et al. did a promoter transcriptional analysis of the C-reactive protein gene, which suggested that the C-reactive protein haplotype–phenotype associations are at least partially attributable to functional changes at promoter sites rs3093062 (SNP 1421 in their paper) and rs3091244 (SNP 1440 in their paper).

Secondly, Miller et al.¹⁵ resequenced 192 individuals to ascertain a comprehensive set of common variants in the C-reactive protein gene, studied their association with C-reactive protein level in (subsets of) three cohorts, and also studied their association with myocardial infarction or ischaemic stroke in a nested case–control study within the Physicians' Health Study cohort. Interestingly, after resequencing this large number of individuals, Miller et al. found a haplotype pattern similar to the pattern of Seattle SNPs. Miller et al.'s haplotypes 1, 2, and 5 concur with haplotypes 3, 2, and 4 in our study, respectively. There were only two minor differences: Miller et al.'s haplotypes 3 and 4 together constitute our haplotype 1, and we did not determine Miller et al.'s haplotype 6, but the mean frequency of this haplotype was only 2.1%.¹⁵ Miller et al.'s haplotypes 2 and 5 were associated with higher C-reactive protein levels and haplotypes 3 and 4 with lower C-reactive protein levels; these results are again in agreement with ours. Also, Miller et al. found that the minor allele of SNP rs2794521 was associated with reduced risk of atherothrombotic events. However, this SNP was associated with higher C-reactive protein level, so the association between the C-reactive protein variant and baseline C-reactive protein did not correlate with the effect of this variant on clinical cardiovascular events. We did not determine this SNP in our study. Our study has the advantage that we had data available on all cases and non-cases in a large, population-based cohort.

Remaining studies on C-reactive protein gene haplotypes and risk of cardiovascular events have mostly been conducted in smaller numbers of high-risk patients.^24–26 The haplotypes used in these studies have been constructed without consideration of the patterns of variation across the locus as a whole. Remaining studies that have examined the association of C-reactive protein gene haplotypes with C-reactive protein levels^6,7,12,26,27 are not comparable to our study, because of different polymorphisms used to reconstruct the haplotypes and different populations used in terms of health status, ethnicity, or age. The results of these studies are diverse, some finding associations with C-reactive protein levels, and some not. Interesting to note is the finding of Szalai et al.,²⁷ that haplotypes reconstructed from the –409G/A (rs3093032) and –390C/T/A (rs3091244) C-reactive protein gene promoter polymorphisms affect transcription factor binding, alter transcriptional activity, and associate with differences in baseline serum C-reactive protein level. According to Seattle SNPs, the latter polymorphism is present in all participants with haplotypes 2 and 4 in our study, and therefore it may result in functional differences between the haplotypes in our study, leading to different serum C-reactive protein levels. Several, mostly smaller, studies have demonstrated associations between various C-reactive protein SNPs and C-reactive protein levels.^3–13 These studies were different in design and were conducted in various populations, and are therefore not similar to ours.

In this study, we found an independent association between serum C-reactive protein levels and coronary heart disease. Since our earlier report on the role of C-reactive protein in the prediction of myocardial infarction in the Rotterdam Study, which was then investigated by means of a nested case–control study,²⁸ data from four more years of follow-up have become available and C-reactive protein has been determined in the total cohort. This may in part explain the difference with the previous results that showed a lack of association after multivariable adjustment.

An issue that warrants consideration in this study is that C-reactive protein measurements were lacking for 854 participants who did not visit the research center at baseline. These participants generally had a higher age and worse general health. However, the association that we found between C-reactive protein serum level and coronary heart disease is in line with the results from previous studies,¹ and this suggests that although we cannot entirely exclude the presence of selection bias, it is not likely that it has substantially influenced the results.

Although serum C-reactive protein levels were found to influence risk of coronary heart disease and C-reactive protein gene haplotypes were found to influence steady-state serum C-reactive protein levels, no association could be demonstrated between C-reactive protein haplotypes and coronary heart disease. Power calculation for the present study shows that, with a power of 80% and an alpha of 0.05, in reference to haplotype 1, (the most common haplotype, frequency 32.8%), we were able to demonstrate relative risks for coronary heart disease of at least 1.21 (for haplotype 2, frequency 31.7%).²⁹ Therefore, either there indeed is no association between C-reactive protein gene haplotypes and coronary heart disease, or, otherwise, the relative risk is of relatively small magnitude. Application of the instrumental variables approach²² to our data is in compliance with the latter; the expected relative risks of coronary heart disease for haplotypes 2, 3, and 4, as estimated from the association between C-reactive protein serum level with coronary heart disease and the association between haplotypes and C-reactive protein serum level were 1.03 (95% CI 0.90–1.20), 1.02 (95% CI 0.88–1.18), and 1.05 (95% CI 0.82–1.36), respectively, as compared with haplotype 1. We were not able to demonstrate estimates of such small magnitude in the present study. Therefore, the door may still be open for a pathophysiological role of C-reactive protein in the development of cardiovascular disease.

Another explanation of the absence of an association in the present study is that baseline C-reactive protein levels are not solely determined by the variation in the C-reactive protein gene, but also by its interaction with several transcription factors induced by regulatory cytokines such as IL-6 and IL-1ß³⁰ which may have a larger influence on serum C-reactive protein levels. Furthermore, high C-reactive protein levels may exert their harmful effects in the acute phase of a coronary event, with high peak C-reactive protein levels leading to enhanced infarct size and more complications such as arrhythmias.³¹ As high C-reactive protein responders may not necessarily have high baseline serum C-reactive protein levels, this aspect needs to be studied by means of a study design different from ours.

In conclusion, this study confirms that steady-state C-reactive protein serum level is predictive of coronary heart disease. Furthermore, it demonstrates that steady-state C-reactive protein serum level is influenced by C-reactive protein haplotypes. Although elevated C-reactive protein level has lately been found to be a consistent and relatively strong risk factor for cardiovascular disease, our study does not support that the common variation in the C-reactive protein gene has a large effect on the occurrence of coronary heart disease.

	Supplementary material

Top Abstract Introduction Participants and methods Results Discussion Supplementary material Acknowledgements References

 
Supplementary material is available at European Heart Journal online.

	Acknowledgements

Top Abstract Introduction Participants and methods Results Discussion Supplementary material Acknowledgements References

 
The authors thank Theo Stijnen and Aaron Isaacs for their advice on statistical analyses and Lamberto Felida for his help in genotyping.

Conflict of interest: none declared for all authors.

	References

Top Abstract Introduction Participants and methods Results Discussion Supplementary material Acknowledgements References

 

1. Danesh J, Wheeler JG, Hirschfield GM, Eda S, Eiriksdottir G, Rumley A, Lowe GD, Pepys MB, Gudnason V. (2004) C-reactive protein and other circulating markers of inflammation in the prediction of coronary heart disease. N Engl J Med 350:1387–1397.[Abstract/Free Full Text]
2. de Maat MP and Trion A. (2004) C-reactive protein as a risk factor versus risk marker. Curr Opin Lipidol 15:651–657.[CrossRef][Web of Science][Medline]
3. Brull DJ, Serrano N, Zito F, Jones L, Montgomery HE, Rumley A, Sharma P, Lowe GD, World MJ, Humphries SE, Hingorani AD. (2003) Human CRP gene polymorphism influences CRP levels: implications for the prediction and pathogenesis of coronary heart disease. Arterioscler Thromb Vasc Biol 23:2063–2069.[Abstract/Free Full Text]
4. Zee RY and Ridker PM. (2002) Polymorphism in the human C-reactive protein (CRP) gene, plasma concentrations of CRP, and the risk of future arterial thrombosis. Atherosclerosis 162:217–219.[CrossRef][Web of Science][Medline]
5. D'Aiuto F, Casas JP, Shah T, Humphries SE, Hingorani AD, Tonetti MS. (2005) C-reactive protein (+1444C>T) polymorphism influences CRP response following a moderate inflammatory stimulus. Atherosclerosis 179:413–417.[CrossRef][Web of Science][Medline]
6. Kovacs A, Green F, Hansson LO, Lundman P, Samnegard A, Boquist S, Ericsson CG, Watkins H, Hamsten A, Tornvall P. (2005) A novel common single nucleotide polymorphism in the promoter region of the C-reactive protein gene associated with the plasma concentration of C-reactive protein. Atherosclerosis 178:193–198.[CrossRef][Web of Science][Medline]
7. Russell AI, Cunninghame Graham DS, Shepherd C, Roberton CA, Whittaker J, Meeks J, Powell RJ, Isenberg DA, Walport MJ, Vyse TJ. (2004) Polymorphism at the C-reactive protein locus influences gene expression and predisposes to systemic lupus erythematosus. Hum Mol Genet 13:137–147.[Abstract/Free Full Text]
8. Davey Smith G, Lawlor DA, Harbord R, Timpson N, Rumley A, Lowe GD, Day IN, Ebrahim S. (2005) Association of C-reactive protein with blood pressure and hypertension: life course confounding and Mendelian randomization tests of causality. Arterioscler Thromb Vasc Biol 25:1051–1056.[Abstract/Free Full Text]
9. Suk HJ, Ridker PM, Cook NR, Zee RY. (2005) Relation of polymorphism within the C-reactive protein gene and plasma CRP levels. Atherosclerosis 178:139–145.[CrossRef][Web of Science][Medline]
10. Szalai AJ, McCrory MA, Cooper GS, Wu J, Kimberly RP. (2002) Association between baseline levels of C-reactive protein (CRP) and a dinucleotide repeat polymorphism in the intron of the CRP gene. Genes Immun 3:14–19.[CrossRef][Web of Science][Medline]
11. Szalai AJ, Alarcon GS, Calvo-Alen J, Toloza SM, McCrory MA, Edberg JC, McGwin G Jr, Bastian HM, Fessler BJ, Vila LM, Kimberly RP, Reveille JD. (2005) Systemic lupus erythematosus in a multiethnic US Cohort (LUMINA). XXX: association between C-reactive protein (CRP) gene polymorphisms and vascular events. Rheumatology 44:864–868.[Abstract/Free Full Text]
12. Obisesan TO, Leeuwenburgh C, Phillips T, Ferrell RE, Phares DA, Prior SJ, Hagberg JM. (2004) C-reactive protein genotypes affect baseline, but not exercise training-induced changes, in C-reactive protein levels. Arterioscler Thromb Vasc Biol 24:1874–1879.[Abstract/Free Full Text]
13. Eklund C, Lehtimaki T, Hurme M. (2005) Epistatic effect of C-reactive protein (CRP) single nucleotide polymorphism (SNP) +1059 and interleukin-1B SNP +3954 on CRP concentration in healthy male blood donors. Int J Immunogenet 32:229–232.[Web of Science][Medline]
14. Carlson CS, Aldred SF, Lee PK, Tracy RP, Schwartz SM, Rieder M, Liu K, Williams OD, Iribarren C, Lewis EC, Fornage M, Boerwinkle E, Gross M, Jaquish C, Nickerson DA, Myers RM, Siscovick DS, Reiner AP. (2005) Polymorphisms within the C-reactive protein (CRP) promoter region are associated with plasma CRP levels. Am J Hum Genet 77:64–77.[CrossRef][Web of Science][Medline]
15. Miller DT, Zee RY, Suk Danik J, Kozlowski P, Chasman DI, Lazarus R, Cook NR, Ridker PM, Kwiatkowski DJ. (2005) Association of common CRP gene variants with CRP levels and cardiovascular events. Ann Hum Genet 69:623–638.[CrossRef][Web of Science][Medline]
16. Hofman A, Grobbee DE, de Jong PT, van den Ouweland FA. (1991) Determinants of disease and disability in the elderly: the Rotterdam Elderly Study. Eur J Epidemiol 7:403–422.[CrossRef][Web of Science][Medline]
17. Kardys I, Kors JA, van der Meer IM, Hofman A, van der Kuip DA, Witteman JC. (2003) Spatial QRS-T angle predicts cardiac death in a general population. Eur Heart J 24:1357–1364.[Abstract/Free Full Text]
18. Epstein MP and Satten GA. (2003) Inference on haplotype effects in case-control studies using unphased genotype data. Am J Hum Genet 73:1316–1329.[CrossRef][Web of Science][Medline]
19. Lake SL, Lyon H, Tantisira K, Silverman EK, Weiss ST, Laird NM, Schaid DJ. (2003) Estimation and tests of haplotype-environment interaction when linkage phase is ambiguous. Hum Hered 55:56–65.[CrossRef][Web of Science][Medline]
20. WHO. (1992) International Statistical Classification of Diseases and Related Health Problems 10th revision (WHO, Geneva).
21. Schaid DJ, Rowland CM, Tines DE, Jacobson RM, Poland GA. (2002) Score tests for association between traits and haplotypes when linkage phase is ambiguous. Am J Hum Genet 70:425–434.[CrossRef][Web of Science][Medline]
22. Timpson NJ, Lawlor DA, Harbord RM, Gaunt TR, Day IN, Palmer LJ, Hattersley AT, Ebrahim S, Lowe GD, Rumley A, Davey Smith G. (2005) C-reactive protein and its role in metabolic syndrome: Mendelian randomisation study. Lancet 366:1954–1959.[CrossRef][Web of Science][Medline]
23. Hingorani A and Humphries S. (2005) Nature's randomised trials. Lancet 366:1906–1908.[CrossRef][Web of Science][Medline]
24. Zee RY, Hegener HH, Fernandez-Cruz A, Lindpaintner K. (2004) C-reactive protein gene polymorphisms and the incidence of post-angioplasty restenosis. Atherosclerosis 176:393–396.[CrossRef][Web of Science][Medline]
25. Zee RY, Hegener HH, Cook NR, Ridker PM. (2004) C-reactive protein gene polymorphisms and the risk of venous thromboembolism: a haplotype-based analysis. J Thromb Haemost 2:1240–1243.[CrossRef][Web of Science][Medline]
26. Chen J, Zhao J, Huang J, Su S, Qiang B, Gu D. (2005) –717A>G polymorphism of human C-reactive protein gene associated with coronary heart disease in ethnic Han Chinese: the Beijing atherosclerosis study. J Mol Med 83:72–78.[CrossRef][Web of Science][Medline]
27. Szalai AJ, Wu J, Lange EM, McCrory MA, Langefeld CD, Williams A, Zakharkin SO, George V, Allison DB, Cooper GS, Xie F, Fan Z, Edberg JC, Kimberly RP. (2005) Single-nucleotide polymorphisms in the C-reactive protein (CRP) gene promoter that affect transcription factor binding, alter transcriptional activity, and associate with differences in baseline serum CRP level. J Mol Med 83:440–447.[CrossRef][Web of Science][Medline]
28. van der Meer IM, de Maat MP, Kiliaan AJ, van der Kuip DA, Hofman A, Witteman JC. (2003) The value of C-reactive protein in cardiovascular risk prediction: the Rotterdam Study. Arch Intern Med 163:1323–1328.[Abstract/Free Full Text]
29. http://members.aol.com/krothman/episheet.xls (accessed on 10 March 2006).
30. Black S, Kushner I, Samols D. (2004) C-reactive Protein. J Biol Chem 279:48487–48490.[Abstract/Free Full Text]
31. Griselli M, Herbert J, Hutchinson WL, Taylor KM, Sohail M, Krausz T, Pepys MB. (1999) C-reactive protein and complement are important mediators of tissue damage in acute myocardial infarction. J Exp Med 190:1733–1740.[Abstract/Free Full Text]

CiteULike    Connotea    Del.icio.us    What's this?

Related articles in EHJ:

Linking observational and genetic approaches to determine the role of C-reactive protein in heart disease risk

Aroon D. Hingorani, Tina Shah, and Juan P. Casas
EHJ 2006 27: 1261-1263. [Extract] [FREE Full Text]  

This article has been cited by other articles:

				P. Elliott, J. C. Chambers, W. Zhang, R. Clarke, J. C. Hopewell, J. F. Peden, J. Erdmann, P. Braund, J. C. Engert, D. Bennett, et al. Genetic Loci Associated With C-Reactive Protein Levels and Risk of Coronary Heart Disease JAMA, July 1, 2009; 302(1): 37 - 48. [Abstract] [Full Text] [PDF]

				Y. M.T.A. van Durme, K. M.C. Verhamme, A.-J. L.H.J. Aarnoudse, G. R. Van Pottelberge, A. Hofman, J. C.M. Witteman, G. F. Joos, G. G. Brusselle, and B. H.C. Stricker C-Reactive Protein Levels, Haplotypes, and the Risk of Incident Chronic Obstructive Pulmonary Disease Am. J. Respir. Crit. Care Med., March 1, 2009; 179(5): 375 - 382. [Abstract] [Full Text] [PDF]

				J. Shen and J. M. Ordovas Impact of Genetic and Environmental Factors on hsCRP Concentrations and Response to Therapeutic Agents Clin. Chem., February 1, 2009; 55(2): 256 - 264. [Abstract] [Full Text] [PDF]

				T. B. Grammer, W. Marz, W. Renner, B. O. Bohm, and M. M. Hoffmann C-reactive protein genotypes associated with circulating C-reactive protein but not with angiographic coronary artery disease: the LURIC study Eur. Heart J., January 2, 2009; 30(2): 170 - 182. [Abstract] [Full Text] [PDF]

				J. Zacho, A. Tybjaerg-Hansen, J. S. Jensen, P. Grande, H. Sillesen, and B. G. Nordestgaard Genetically Elevated C-Reactive Protein and Ischemic Vascular Disease N. Engl. J. Med., October 30, 2008; 359(18): 1897 - 1908. [Abstract] [Full Text] [PDF]

				J. Sunyer, R. Pistelli, E. Plana, M. Andreani, F. Baldari, M. Kolz, W. Koenig, J. Pekkanen, A. Peters, and F. Forastiere Systemic inflammation, genetic susceptibility and lung function Eur. Respir. J., July 1, 2008; 32(1): 92 - 97. [Abstract] [Full Text] [PDF]

				M. Kolz, W. Koenig, M. Muller, M. Andreani, S. Greven, T. Illig, N. Khuseyinova, D. Panagiotakos, G. Pershagen, V. Salomaa, et al. DNA variants, plasma levels and variability of C-reactive protein in myocardial infarction survivors: results from the AIRGENE study Eur. Heart J., May 2, 2008; 29(10): 1250 - 1258. [Abstract] [Full Text] [PDF]

				L. Qi, N. Rifai, and F. B. Hu Interleukin-6 Receptor Gene Variations, Plasma Interleukin-6 Levels, and Type 2 Diabetes in U.S. Women Diabetes, December 1, 2007; 56(12): 3075 - 3081. [Abstract] [Full Text] [PDF]

				M. Kivimaki, D. A. Lawlor, G. D. Smith, C. Eklund, M. Hurme, T. Lehtimaki, J. S. A. Viikari, and O. T. Raitakari Variants in the CRP Gene as a Measure of Lifelong Differences in Average C-Reactive Protein Levels: The Cardiovascular Risk in Young Finns Study, 1980 2001 Am. J. Epidemiol., October 1, 2007; 166(7): 760 - 764. [Abstract] [Full Text] [PDF]

				F. G. Hage and A. J. Szalai C-Reactive Protein Gene Polymorphisms, C-Reactive Protein Blood Levels, and Cardiovascular Disease Risk J. Am. Coll. Cardiol., September 18, 2007; 50(12): 1115 - 1122. [Abstract] [Full Text] [PDF]

				L. A. Lange, C. S. Carlson, L. A. Hindorff, E. M. Lange, J. Walston, J. P. Durda, M. Cushman, J. C. Bis, D. Zeng, D. Lin, et al. Association of Polymorphisms in the CRP Gene With Circulating C-Reactive Protein Levels and Cardiovascular Events JAMA, December 13, 2006; 296(22): 2703 - 2711. [Abstract] [Full Text] [PDF]

				D. C. Crawford, C. L. Sanders, X. Qin, J. D. Smith, C. Shephard, M. Wong, L. Witrak, M. J. Rieder, and D. A. Nickerson Genetic Variation Is Associated With C-Reactive Protein Levels in the Third National Health and Nutrition Examination Survey Circulation, December 5, 2006; 114(23): 2458 - 2465. [Abstract] [Full Text] [PDF]

				C. Siemes, L. E. Visser, J.-W. W. Coebergh, T. A.W. Splinter, J. C.M. Witteman, A. G. Uitterlinden, A. Hofman, H. A.P. Pols, and B. H.Ch. Stricker C-Reactive Protein Levels, Variation in the C-Reactive Protein Gene, and Cancer Risk: The Rotterdam Study J. Clin. Oncol., November 20, 2006; 24(33): 5216 - 5222. [Abstract] [Full Text] [PDF]

				P. Stenvinkel C-reactive protein--does it promote vascular disease? Nephrol. Dial. Transplant., October 1, 2006; 21(10): 2718 - 2720. [Full Text] [PDF]

				D. D. G. Despriet, C. C. W. Klaver, J. C. M. Witteman, A. A. B. Bergen, I. Kardys, M. P. M. de Maat, S. S. Boekhoorn, J. R. Vingerling, A. Hofman, B. A. Oostra, et al. Complement factor H polymorphism, complement activators, and risk of age-related macular degeneration. JAMA, July 19, 2006; 296(3): 301 - 309. [Abstract] [Full Text] [PDF]

				A. D. Hingorani, T. Shah, and J. P. Casas Linking observational and genetic approaches to determine the role of C-reactive protein in heart disease risk Eur. Heart J., June 1, 2006; 27(11): 1261 - 1263. [Full Text] [PDF]

This Article Abstract FREE Full Text (PDF) Supplementary tables All Versions of this Article: 27/11/1331    most recent ehl018v1 E-letters: Submit a response Alert me when this article is cited Alert me when E-letters are posted Alert me if a correction is posted Services Email this article to a friend Related articles in EHJ Similar articles in this journal Similar articles in ISI Web of Science Similar articles in PubMed Alert me to new issues of the journal Add to My Personal Archive Download to citation manager Search for citing articles in: ISI Web of Science (18) Request Permissions Disclaimer Google Scholar Articles by Kardys, I. Articles by Witteman, J. C.M. Search for Related Content PubMed PubMed Citation Articles by Kardys, I. Articles by Witteman, J. C.M. Social Bookmarking          What's this?

Online ISSN 1522-9645 - Print ISSN 0195-668x

Copyright © 2009 European Society of Cardiology

Oxford Journals Oxford University Press

• Site Map
• Privacy Policy
• Frequently Asked Questions

Other Oxford University Press sites: Oxford University Press Oxford Journals China Oxford Journals Japan American National Biography Booksellers' Information Service Children's Fiction and Poetry Children's Reference Corporate & Special Sales Dictionaries Dictionary of National Biography Digital Reference English Language Teaching Higher Education Textbooks Humanities International Education Unit Law Medicine Music Online Products Oxford English Dictionary Reference Rights and Permissions Science School Books Social Sciences Very Short Introductions World's Classics