In two independent populations of predominantly Caucasian men and women, we related common variation in the CRP gene and estimated common haplotypes with plasma CRP levels and risk of incident CHD. CRP SNPs and associated haplotypes were strongly associated with plasma CRP levels. However, neither the individual SNPs nor common haplotypes were associated with risk of CHD in the direction that would be predicted by their association with CRP levels.
Inflammation is thought to contribute to increased cardiovascular risk, and C-reactive protein, an important component of the innate immune system, is the most extensively studied marker of inflammation.
Many studies have reported individual single nucleotide polymorphisms (SNPs) to be significantly associated with baseline CRP levels ().
Though some of the individual SNP results have been conflicting, our results are consistent with the majority of published findings.
Brief summary of literature examining single CRP SNPs with CRP levels, and CHD where available
On the other hand, the studies that assessed common variation and haplotypes in the CRP gene with plasma CRP levels differed in SNP selection methodology and estimated haplotypes.
For example, Carlson et al.
resequenced 47 samples from panels with African American and European descent individuals to select seven tagging SNPs for eight haplotypes in the CARDIA study. On the other hand, Miller et al.
resequenced 192 individuals in the Women's Health Study with extreme discordant baseline CRP levels and selected seven tagging SNPs for six common haplotypes among Caucasians participants in three sub-cohorts. In the Rotterdam Study, Kardys et al. genotyped 3 tagging SNPs (3014, 3872, and 4741) selected from SeattleSNPs which were sufficient to infer four common haplotypes.
Though overall haplotypes were not exactly replicated, the more recent studies shared much overlap in selected SNPs. Based on LD patterns and similar reference groups, our results were similar to what has been previously reported. The haplotypes associated with lower CRP levels were similar, as were the haplotypes associated with higher CRP levels.
Several studies have shown H4 as associated with significantly lower CRP levels.
However, specifically of importance, few studies have assessed haplotypes, and H4 in particular, with risk of incident CHD. Thus far, the results have been unclear and conflicting. Though the Rotterdam Study did not genotype 2667G>C, they reported four haplotypes significantly associated with CRP levels, and reported no associations with CHD.
A recent study in the Cardiovascular Health Study of older adults reported no association between five CRP haplotypes and risk of MI among white participants, and the relative risk of MI for our H4 equivalent was 0.93 (95% CI 0.72–1.21).
On the other hand, though H4 was not associated with CRP levels in the Physicians' Health Study, 3872C>T individually was associated with lower CRP levels, and with an increased risk of MI (odds ratio: 1.29, 95% CI 0.99–1.67).
Furthermore, the odds ratio for 2667G>C was suggestive of a 50% increase as well, but the confidence interval was much wider. Taken together, these results suggest that 2667G>C and 3872C>T, though associated with lower CRP levels, may be associated with an elevated risk of CHD.
A number of SNPs in the promoter region of the CRP gene have been shown to affect changes in transcription factor binding and gene promoter activity.
The triallelic SNP has been associated with increased promoter activity and higher CRP levels,
and haplotypes of the promoter triallelic and rs3093032 SNPs affect transcription factor binding, transcriptional activity, and CRP levels.
The major alleles of 3872C>T and 5237A>G together serve as a proxy for the triallelic,
and the 2-SNP haplotype was associated with higher CRP levels in our studies as well as in others.
Otherwise, more functionality data are needed for CRP SNPs.
There is emerging consistency in the literature that polymorphisms in the CRP gene are associated with plasma CRP levels, but whether there is also an association between polymorphisms and risk of coronary events remains unclear. While CRP polymorphisms are strongly associated with plasma levels, these polymorphisms contribute only 1.4–5.0% of the phenotypic variation, a level comparable to (or less than) that from environmental and lifestyle factors.
In addition to environmental stimuli, CRP is induced by other inflammatory cytokines, and additional genes in the inflammatory pathway may regulate CRP levels as well. Thus, our results are consistent with CRP as a good indicator of the inflammatory state and as a risk marker. However, the apparent influence in baseline CRP levels due to CRP polymorphisms may not be large enough to alter CHD risk, which may explain the lack of association between the majority of common haplotypes in the CRP gene and risk of CHD in either the NHS or the HPFS. The remaining exception would be haplotype 4, and it is possible that CRP is an epiphenomenon of vascular inflammation and not directly involved in vascular pathobiology, or possibly protective. Alternatively, the CRP polymorphisms and haplotypes may be in LD with pro-atherogenic variants in nearby gene regions which would not be captured in this study.
Several potential limitations should be discussed regarding the current two studies. In genetic association studies, population stratification is often a concern. However, population stratification should be minimized in the NHS and HPFS because both cohorts are predominantly Caucasian, and similar results were observed when restricted to Caucasians only. Additionally, we recognize that the relative socioeconomic homogeneity of the cohorts does not represent random samples of U.S. men and women and may not be generalizeable to other populations. Though the homogeneity is unlikely to influence genetic predisposition, it may be a strength in reducing residual confounding from unmeasured factors related to socioeconomic status. Also, the distribution of cardiovascular risk factors such as smoking status and history of diabetes were different between the men and women, and may have mediated influences through CRP. Finally, chance may be an explanation for the statistically significant findings. Nevertheless, we included a replication study by examining both the NHS and HPFS, and the overall results were identical between the two independent populations.
Our study provides a comprehensive assessment of the common genetic variation in the CRP gene with plasma CRP levels and risk of incident CHD among two independent populations. Among predominantly Caucasian men and women, CRP genotypes and haplotypes were consistently and significantly associated with plasma CRP levels. However, the relationship between genotype and CRP levels did not translate largely to an associated change in CHD risk, and Haplotype 4 in particular may be important in predicting other clinical events. These data suggest that the underlying inflammatory processes which predict coronary events cannot be captured solely by variation in the CRP gene, and other regulatory genes should be examined.