To our knowledge, this study is the first to evaluate the genetic contribution to variation in serum MMP-1 levels in a large human population. Our analyses revealed that variation in MMP-1 levels is highly heritable (h2 = 81%) in the Amish, with strong and compelling evidence for association of serum MMP-1 levels with a large cluster of SNPs in an 11.4 Mb region on chromosome 11q. Of note, the strongest evidence for association was to a group of SNPs located near a cluster of MMP genes, particularly between MMP-1 and MMP-3. The estimated effect sizes of the associated SNPs were very high, with rs495366, the most strongly associated SNP, accounting by itself for 17.5% of the variance in MMP-1 levels. Two other SNPs in this region, rs12289128 and rs11226373, were also strongly associated with serum MMP-1 levels, and the three SNPs collectively accounted for 31% of the trait variance.
The location of these associated SNPs near the MMP
gene cluster lends strong biological plausibility to the observed associations. Several in vitro20, 25
and epidemiologic13, 20–24
studies have shown genetic variants in the MMP-1
promoter to be associated with transcriptional level of MMP-1
and/or CVD endpoints (Data Supplement Table S3
). The peak associations observed in our study (i.e., rs495366, rs603050, rs11225434 and rs7926920) were also located in the upstream region of MMP-1
gene, consistent with the existence of functional variants in the MMP-1
promoter region affecting expression. However, further studies are needed to verify this speculation given that the previously reported promoter polymorphisms were not highly correlated with our most strongly associated SNP (i.e. rs495366) based on the HAPMAP Caucasian population or with other associated SNPs in the MMP-1
upstream region. The absence of an association between intragenic MMP-1
SNPs and serum MMP-1 levels is possibly due to suboptimal SNP coverage of the MMP-1
gene by the Affymetrix 500K panel given that this panel includes only 3 SNPs within MMP-1
despite the fact that this gene spans a region of 8,243 bp.
Our analyses do not permit us to resolve whether the observed associations are attributable to a single functional SNP or to multiple functional SNPs residing in the MMP gene cluster. However, our analysis revealed that the rs12289128-rs495366 haplotype provides an even stronger association with MMP-1 serum levels than either SNP alone, suggesting that this haplotype is tagging a single (unmeasured) functional polymorphism better than either single SNP alone. Including rs11226373, which is partially correlated with the other two SNPs, in the haplotype analysis did not provide a clear dose-response relationship with MMP-1 serum levels. Therefore, it remains unclear whether rs11226373 is independently associated with MMP-1 levels or simply tags the same (unmeasured) causal polymorphism as do rs12289128 and rs495366. The latter is possible despite the >1.5 MB distance of rs11226373 from rs12289128 and rs495366 given the long-range LD present in this region in the Amish.
Previous studies suggest that MMP-1
may play an important role in atherosclerotic plaque disruption by influencing stability of the plaque fibrous cap6, 7, 29
. For example, the expression of MMP-1 is elevated in human atherosclerotic plaques, particularly in shoulder regions and areas of foam cell accumulation in the plaques 7
, and is higher in plaques with thin cap compared to those with a thick fibrous cap29
. Foam cells isolated from rabbit aortic lesions are capable of synthesizing and releasing MMP-1 constitutively in vitro 30
. Furthermore, treating patients with carotid endarterectomy with MMP inhibitor reduces the MMP-1 concentration in carotid plaques 31
. These findings suggest a critical role of MMP-1 in the matrix degradation that predisposes plaque rupture and acute coronary syndromes. Furthermore, MMP-1 can induce tyrosine phosphorylation of intracellular proteins in platelets and the movement of β3
integrins to areas of cell contact and thus prime platelets for aggregation 9
, indicating a potential role of MMP-1 in thrombotic events.
MMP-1 is found to be highly expressed in smooth muscle, pancreatic islets, cardiac myocytes and bronchial epithelial cells and normal vascular tissues (data sources: UCSC Genome Bioinformatics Site: http://genome.ucsc.edu
; GeneHub-GEPIS database: http://www.cgl.ucsf.edu/Research/genentech/genehub-gepis/index.html
). However, there has been limited data, if any, on how well MMP-1 expression levels in tissues correlate with their concentrations in the circulation, and the clinical significance of elevated MMP-1 levels in humans remains unclear. MMP-1 serum levels increase with increasing age as shown in our data and reported previously in Chinese women 32
. In two relatively small studies, higher plasma MMP-1 serum levels were correlated with increasing carotid-femoral pulse-wave velocity (PWV), a marker of arterial stiffness 10
and with increasing pulse pressure and mean blood pressure 11
while in our large study, we observed significant, albeit modest, correlations of MMP-1 levels with fasting triglycerides and pulse pressure.
Only a few studies have reported correlations of circulating MMP-1 levels with CHD outcomes. Several studies have reported higher circulating MMP-1 levels to be associated with acute MI and/or unstable angina 14–16
and ruptured plaques12
, suggesting that MMP-1 levels may be a marker of plaque instability, although others have failed to find such an association13
. MMP-1 levels have also been associated carotid stenosis 17
and left ventricular remodeling among patients with cardiovascular disease 15, 18, 19
. Given the limitations and inconsistency of existing studies, the utility of MMP-1 serum levels as a marker for screening, diagnosis or identifying individuals at risk of various disease outcomes remains unproven. Studies assessing the effects of polymorphisms in the MMP-1
on cardiovascular events have found inconsistent associations13, 20–24
. The lack of association of MMP-1
SNPs with cardiovascular risk factors in our data is possibly due either subjects in our study being relatively healthy with few individuals having symptomatic CHD or an absence of a true relationship effect between MMP-1
-associated genetic variants and cardiovascular risk.
We conclude that serum MMP-1 levels are highly heritable and that genetic variation at a locus on chromosome 11q that includes MMP-1 is a major determinant of MMP-1 levels. Given the relatively high frequency of the associated SNPs in this Amish population (15–36%), we expect that the strong associations detected will be present in other populations as well (especially European Caucasians). However, the large number of associated SNPs spanning across a relatively large region will make it difficult to localize the causative SNP(s) in this population based on statistical criteria alone. Conducting additional genetic association studies and functional characterization of the most likely causative variants in other populations will be needed to better define the genetic architecture of MMP-1 levels and to determine the relevance of genetic variation at this locus to CHD.
Extracellular matrix (ECM) remodeling is an essential process in the pathogenesis of atherosclerosis and coronary heart diseases (CHD). Matrix metalloproteinases (MMPs) mediate this process by participating in the degradation of the ECM, resulting in weakening of the fibrous cap and predisposing the atherosclerotic plaques to disruption and embolic events. One of the key MMPs is MMP-1, although the relation of circulating MMP-1 levels to cardiovascular risk and its potential value as a biomarker for screening for disease is unknown. To identify genetic determinants of MMP-1 serum levels, we carried out a genome-wide association study in 778 healthy Old Order Amish individuals. We found serum MMP-1 levels to be highly heritable in this population (h2 = 81%). We identified a large cluster of SNPs spanning an 11.5 Mb region on chromosome 11q to be strongly associated with serum MMP-1 levels, with the peak association observed with rs495366 (p = 5.73 × 10−34; minor allele frequency = 0.36). This region covers a cluster of MMP genes, including MMP-1. However, there was little evidence that the MMP-1-associated SNPs were associated with any other cardiovascular risk factors in this relatively healthy population. These results suggest that genetic variation on chromosome 11q is a major determinant of circulating MMP-1 levels in the Amish; additional studies in other populations are needed to determine the relevance of genetic variation at this region to CHD.