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Journal of Clinical Endocrinology & Metabolism , doi:10.1210/jc.2006-2136
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The Journal of Clinical Endocrinology & Metabolism Vol. 92, No. 8 3206-3212
Copyright © 2007 by The Endocrine Society

The Catechol-O-Methyltransferase Met158 Low-Activity Allele and Association with Nonvertebral Fracture Risk in Elderly Men

Lisette Stolk, Joyce B. J. van Meurs, Mila Jhamai, Pascal P. Arp, Johannes P. T. van Leeuwen, Albert Hofman, Frank H. de Jong, Huibert A. P. Pols and André G. Uitterlinden

Departments of Internal Medicine (L.S., J.B.J.v.M., M.J., P.P.A., J.P.T.v.L., F.H.d.J., H.A.P.P., A.G.U.) and Epidemiology and Biostatistics (A.H., H.A.P.P., A.G.U.), Erasmus Medical Center, 3000 DR Rotterdam, The Netherlands

Address all correspondence and requests for reprints to: A. G. Uitterlinden, Room Ee575, Genetic Laboratory, Department of Internal Medicine, Erasmus MC, P.O. Box 1738, 3000 DR Rotterdam, The Netherlands. E-mail: a.g.uitterlinden{at}erasmusmc.nl.


   Abstract
Top
 Abstract
Introduction
 Subjects and Methods
 Results
 Discussion
 References
 
Context: Because sex steroids play an important role in bone development, variants in genes encoding proteins involved in estrogen synthesis and metabolism could contribute to interindividual variation in bone parameters and fracture risk. An example is catechol-O-methyltransferase (COMT), an estrogen-degrading enzyme involved in inactivation of catechol-estrogens. Its gene contains a functional valine to methionine substitution at codon 158.

Objective: The aim of our study was to determine whether this polymorphism is associated with bone parameters and fracture risk in elderly subjects.

Methods: COMT genotypes were determined using TaqMan allelic discrimination in 2515 men and 3554 women from the Rotterdam Study, a population-based cohort study of individuals aged 55 and older. Associations with bone mineral density (BMD) and bone loss were analyzed using ANOVA or analysis of covariance, whereas fracture risk was analyzed using Cox’s proportional hazard regression analysis. COMT mRNA expression in three osteoblastic cell lines (SaOS, MG63, and SVHFO) was analyzed by RT-PCR.

Results: Male carriers of the Met158 allele had an increased risk for osteoporotic fractures (hazard ratio = 1.6; 95% confidence interval, 1.0–2.4) and for fragility fractures (hazard ratio = 2.7; 95% confidence interval, 1.3–5.9), with evidence for a dominant effect. Adjustments for age, height, weight, and BMD did not change the risk estimates. In women, this association was weaker and not significant. BMD was not significantly associated with the variant in either men or women. COMT mRNA was expressed in all three osteoblastic cell lines tested.

Conclusion: The COMT Val158Met polymorphism is associated with fracture risk in elderly men, through a mechanism independent of BMD.


   Introduction
Top
 Abstract
 Introduction
Subjects and Methods
 Results
 Discussion
 References
 
OSTEOPOROSIS IS A skeletal disorder characterized by low bone mineral density (BMD) and microarchitectural deterioration of bone, resulting in an increase in bone fragility and fracture risk (1). Osteoporotic fractures are associated with substantial morbidity and mortality and are therefore a major health care problem both in postmenopausal women and elderly men (2). Twin studies have estimated a high heritability of BMD (up to 80%), bone turnover (63%), and bone geometry (62%) (3), whereas heritability of risk for fractures is lower (25–35%) (3). The genes involved in these traits, however, are largely unknown.

Sex steroids play a major role in bone development (4), and therefore, genes regulating sex steroid production and metabolism are good candidates to study for involvement in the development of osteoporosis. There are several candidate genes in the estrogen pathway. We previously reported on genetic variations in the estrogen receptor {alpha} (ER{alpha}) gene to be associated with BMD, height, and fracture risk (5, 6, 7). Variations in genes involved in estrogen synthesis, like aromatase (CYP19), are also associated with bone parameters (8). Until now, not much attention has been given to genetic variation of genes involved in estrogen degradation. Estrogens are degraded in several steps; the two major steps are shown in Fig. 1Go. The first step is oxidation by cytochrome P450s (e.g. CYP1A1, -1B1, and -3A4), resulting in catechol-estrogens. One study examining CYP1A1 observed a nonsynonymous single-nucleotide polymorphism (SNP) in the gene to be associated with BMD in postmenopausal Caucasian women (9). This suggests that there might also be a link between genetic variation in estrogen-degrading enzymes and differences in BMD.


Figure 1
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  		FIG. 1. Pathway for oxidative metabolism of estradiol (modified from Refs. 10 and 34 ).

 
COMT is the gene coding for catechol-O-methyltransferase (COMT), which inactivates circulating catechol-estrogens by catalyzing O-methylation of 2-hydroxylated and 4-hydroxylated estrogens to their methoxy derivatives 2-OH-methoxy-estrogen and 4-OH-methoxy-estrogen (10).

The best-known polymorphism in the COMT gene is a functional G to A substitution, leading to a valine (G) to methionine (A) substitution at codon 158. The methionine variant results in thermolability of the enzyme (11) and a 3- to 4-fold lower enzyme activity compared with the valine variant (12). The alleles are therefore known as a low-activity (COMTL) and a high-activity (COMTH) allele. The frequency of the methionine variant is around 50% in Caucasian populations.

In middle-aged men, the low-activity allele was shown to be associated with increased serum estradiol levels (13). This finding indicates a link between COMT polymorphisms and estrogens. In postmenopausal women, however, there was no association found between the COMTL genotype and normal estradiol levels (14).

With regard to bone metabolism, not many studies have been published on COMT. In 458 young adult Swedish men, the COMTL allele was associated with a 4% lower peak BMD in the femur (15). Yet, in a cohort of 1795 postmenopausal women, no association of the COMTL allele with femoral neck and spine BMD was found (16).

The expression of COMT in bone cells could indicate a more local effect of COMT on bone than a systemic effect on estrogen levels; however, this has not been studied as far as we know.

Here we investigated the effect of the COMT Val158Met variant on BMD, fracture risk, and estrogen levels in elderly men and women of the Rotterdam Study. To study the local expression of COMT in bone cells, we looked at mRNA expression in three different osteoblast-like cell lines.


   Subjects and Methods
Top
 Abstract
 Introduction
 Subjects and Methods
Results
 Discussion
 References
 
Study populations

Subjects were participants of the Rotterdam Study, a large prospective population-based cohort study of Caucasian subjects aged 55 yr and over, living in the Ommoord district of Rotterdam, The Netherlands. The study was designed to investigate the incidence and determinants of chronic disabling diseases in the elderly. Rationale and design have been described previously (17). All 10,275 inhabitants aged 55 yr and over were invited for baseline examination between August 1990 and June 1993. Of those, 7983 participated. Among the subjects living independently, the overall response rate was 77% for home interview and 71% for examination in the research center, where anthropometric characteristics and BMD were measured and blood samples were taken. The Rotterdam Study was approved by the medical ethics committee of the Erasmus University Medical School, and written informed consent was obtained from each subject. The current study is based on 6298 subjects (2570 male) for whom genotype data were available for the COMT Val158Met polymorphism. Baseline measurements of BMD were available for 5156 subjects (82% of the genotyped cohort, 2217 males) and both height and weight data for 6069 subjects (96% of the genotyped cohort, 2515 males).

In addition, we determined allele frequencies in a panel of subjects of different ethnic background from the National Institute of General Medical Sciences Human Diversity Panel (Coriell Institute, Camden, NJ). The panel consisted of 60 African-Americans (HD04 and HD50) and 110 Han-Chinese (HD02 and HD100).

Clinical examination

Height and weight were measured at baseline examination with the subject in a standing position with indoor clothing without shoes. BMD (in grams per square centimeter) was determined by dual-energy x-ray absorptiometry (DPX-L densitometer; Lunar, Madison, WI) at the femoral neck and lumbar spine (vertebrae L2, L3, and L4), as described elsewhere (18). Age at menopause and smoking habits were assessed by a questionnaire.

Hormone assays

Estrone, estradiol, androstenedione, and testosterone levels were determined in a gender-stratified random sample (n = 1159) as described earlier (19).

Assessment of incident nonvertebral fractures and vertebral fractures

For nonvertebral fractures, follow-up started either January 1, 1991, or, if later, at the time of inclusion into the study. For this analysis, follow-up ended either at January 1, 2002, or, when earlier, at the participant’s death, comprising an average follow-up period of 7.4 (SD, 3.3) years for nonvertebral fractures. For approximately 80% of the study population, medical events were reported through computerized general practitioner diagnosis registers. For the remaining 20%, research physicians collected data from the general practitioners’ medical records of the study participants. All collected fractures were verified by reviewing discharge reports and letters from medical specialists. Fracture events were coded independently by two research physicians according to the International Classification of Diseases, 10th revision (ICD-10). Finally, an expert in osteoporosis reviewed all coded events for final classification. Any fracture was used as an outcome measure to have sufficient power. All fractures that were considered not osteoporotic (fractures caused by cancer and all hand, foot, skull, and face fractures) were excluded. In addition, we considered separately all fragility fractures that occur at older age, which included hip, proximal humerus, and pelvis fractures.

Both at baseline and at follow-up visits between 1997 and 2001, thoracolumbar radiographs of the spine were obtained. The follow-up radiographs were available for 3469 individuals (1498 men) who survived an average of 6.4 (SD 0.4) years after the baseline center visit and who were still able to come to our research center. All follow-up radiographs were scored for the presence of vertebral fracture by the McCloskey/Kanis method as described earlier (20). If a vertebral fracture was detected, the baseline radiograph was evaluated as well. If the vertebral fracture was already present at baseline, it was considered a baseline prevalent fracture. If it was not present at baseline, the fracture was defined to be incident.

Genotyping

Genomic DNA was extracted from samples of peripheral venous blood according to standard procedures. Genomic DNA (1–2 ng) was dispensed into 384-well plates using a Caliper Sciclone ALH3000 pipetting robot (Caliper LS, Mountain View, CA). Genotypes were determined using the TaqMan allelic discrimination assay. The Assay-by-Design service (www.appliedbio-systems.com) was used to set up a TaqMan allelic discrimination assay for the COMT Val158Met polymorphism [primers were CGAGATCAACCCCGACTGT (forward) and CAGGCATGCACACCTTGTC (reverse); probes were FAM, TCGCTGGCATGAAG, and VIC, TTTCGCTGGCGTGAAG]. The underlined letters indicate the SNP position. The PCR mixture included 1–2 ng genomic DNA in a 2-µl volume and the following reagents: FAM and VIC probes (200 nM), primers (0.9 µM), 2x TaqmAn PCR master mix (ABgene, Epsom, UK). Reagents were dispensed in a 384-well plate using the Deerac Equator NS808 (Deerac Fluidics, Dublin, Ireland). PCR cycling reaction were performed in 384-well PCR plates in an ABI 9700 PCR system (Applied Biosystems Inc., Foster City, CA) and consisted of initial denaturation for 15 min at 95 C and 40 cycles with denaturation of 15 sec at 95 C and annealing and extension for 60 sec at 60 C. Results were analyzed by the ABI TaqMan 7900HT using the sequence detection system 2.22 software (Applied Biosystems). To confirm the accuracy of genotyping results, 332 (5%) randomly selected samples were re-genotyped with the same method. No inconsistencies were observed.

Quantitative real-time PCR

Cell nuclear extracts from SaOS and MG-63 (human osteoblast-like sarcoma cells) and SV-HFO (human preosteoblast cells) at different differentiation stages were pooled per cell line and were prepared as described elsewhere (21). Quantitative real-time PCR was carried out using an ABI 7700 sequence detection system (Applied Biosystems). Reactions were performed in 25-µl volumes using a qPCR core kit (Eurogentec, Seraing, Belgium). Reaction mixtures contained 20 ng cDNA, 5 mM MgCl2, 200 µM dNTPs, and 0.025 U/µl Hot GoldStar enzyme (QIAGEN, Valencia, CA). Primer and probe sets were designed using the Primer Express software (version 2.0; Applied Biosystems). The amount of human glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA was used as internal control to normalize for possible differences in RNA extraction and degradation as well as efficiency of the cDNA synthesis. The cDNA primers COMT-For (5'-AGGAGTGGGCCATGAACGT-3'), COMT-Rev (5'-GGCTGGTGCTCCTGAATCAC-3'), GAPDH-For (5'-ATGGGGAAGGTGAAGGTCG-3'), and GAPDH-Rev (5'-TAAAAGCAGCCCTGGTGACC-3') were used to amplify the COMT and GAPDH cDNA, respectively. The probe sequences for COMT and GAPDH were COMT-FAM (5'-FAM-ACAAGAAAGGCAAGATCGTGGACGAA-3') and GAPDH-FAM (5'-FAM-CGCCCAATACGACCAAATCCGTTGAC-3'). Cycling conditions were 50 C for 2 min and 95 C for 10 min followed by 40 cycles of 95 C for 15 sec and 60 C for 1 min. Data are presented as relative mRNA levels calculated by the equation: 2–{Delta}Ct ({Delta}Ct = Ct of COMT– Ct of GAPDH, where Ct is cycle threshold).

Statistical analysis

Hardy-Weinberg Equilibrium was calculated according to standard procedures using {chi}2 analysis.

For each allele, subjects were grouped according to genotype by allele copy number (0, 1, and 2, corresponding to noncarriers, heterozygote carriers, and homozygote carriers, respectively). We allowed for three possible genetic models to explain differences between groups, i.e. an allele-dose effect, a dominant effect, or a recessive effect. Allele dose was defined as the number of copies of a certain allele in the genotype. In case of a consistent trend reflected as an allele-dose effect, we performed a linear regression analysis to quantify the association. In case of a dominant or a recessive effect of the test allele, ANOVA or analysis of covariance was performed to test for differences between two genotype groups. For dominant alleles, we compared test-allele carriers vs. noncarriers.

Odds ratios with 95% confidence intervals (95% CI) were calculated by logistic regression analyses to estimate the relative risk of fractures by genotype for the risk allele, with no copies of the risk allele as the reference group. We first calculated crude odds ratios and then adjusted for potentially confounding factors (e.g. age, height, weight, and BMD). To estimate nonvertebral fracture risk by genotype, we used Cox proportional hazard models, thereby taking potential differences in time-to-event into account. To estimate the risk of vertebral fractures, odds ratios with 95% CI were calculated using logistic regression models. We were not able to use Cox proportional hazard models for vertebral fractures because the exact time of the event was not known. All statistical analyses were performed using SPSS version 11.0.1 (SPSS Inc., Chicago, IL). P values are two-sided, and ≤0.05 was considered significant.


   Results
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
Discussion
 References
 
Genotyping

The allele frequency of the G allele (methionine, COMTL) was 55% in our study population, the genotype distribution was in Hardy-Weinberg equilibrium (P = 0.99). Figure 2Go shows the COMT gene with the exons numbered 1–6 and indicates SNPs found in several data sources: dbSNP, HapMap, and a resequencing study (22). The methionine to valine substitution is located in exon 4 and was found in both the dbSNP database, the HapMap genome browser (Phase II release 19), and the resequencing study (22). We analyzed the LD-block structure from publicly available HapMap data across the gene (Fig. 2Go). The Met158Val variant is situated in haploblock 2 in the gene, encompassing intron 3 to intron 5. Figure 2Go also shows the allele frequency of the Met158 variant in three different ethnic groups. The allele frequency of the Met allele in African-Americans (24%) and in Han-Chinese (21%) is lower compared with the frequency in Caucasians in our population (55%).


Figure 2
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  		FIG. 2. Schematic overview of the COMT gene showing the methionine to valine substitution at codon 158. In gray are the six exons (numbered 1–6), and the vertical lines represent SNPs found in this gene. We searched the NCBI database (http://www.ncbi.nlm.nih.-gov/), the HapMap database (Phase II release 19) (http://www.hapmap.org), and a study by Shield et al. (22 ) in which the COMT gene was resequenced to find SNPs. + indicates that the SNP is present in one of the data resources; – indicates not present. In the lower part of the figure, the allele frequencies of the low activity methionine allele are shown for three different ethnic populations [The Rotterdam Study (Caucasian), African-American, and Han-Chinese].

 
Baseline characteristics

None of the baseline characteristics were significantly different between the three genotype groups in men and women (Table 1Go). In Table 2Go, the bone density measures of the study population are shown. Both lumbar spine BMD and femoral neck BMD did not differ by COMT Val158Met genotype. No association of the COMT polymorphism with serum estrone and estradiol levels was found.


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  		TABLE 1. Baseline anthropometric measures according to COMT Met158Val genotype

 

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  		TABLE 2. BMD and hormone levels according to COMT Met158Val genotype

 
Fracture risks

Table 3Go shows the hazard ratios and 95% CI for osteoporotic and fragility fractures, and the odds ratios and 95% CI for vertebral fractures for both men and women. The COMT genotype was associated with osteoporotic fractures and fragility fractures. In men, the fracture risk for COMTLL [hazard ratio (HR) = 1.6; 95% CI, 1.0–2.5; P = 0.05] and COMTHL (HR = 1.6; 85% CI, 1.0–2.4; P = 0.04) was similar, suggesting a dominant effect, and therefore, we analyzed the fractures risks also in carriers vs. noncarriers of the COMTL allele. The risk for osteoporotic fractures in male COMTL-carriers was 1.6 (HR = 1.6; 95% CI, 1.0–2.4; P = 0.02) compared with noncarriers. For fragility fractures, the risk was 2.7 (HR = 2.7; 95% CI, 1.3–5.9; P = 0.005). Both HRs are adjusted for age, height, and weight. In women, a higher osteoporotic fracture risk was seen for the COMTLL genotype (HR = 1.2; 95% CI, 1.0–1.5; P = 0.04). No significant effect of the COMTL allele was seen on fragility fractures in women. We examined whether estradiollevels influenced the association between the polymorphism and fracture risk. We did not find any influence, probably due to low power. The COMT polymorphism did not show an association with vertebral fractures in either men or women.


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  		TABLE 3. Fracture risk according to COMT Met158Val genotype

 
Expression of COMT in osteoblasts

Figure 3Go shows mRNA expression levels of COMT mRNA relative to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA in three human bone cell lines and in the liver cell line HepG2. We observed a GAPDH Ct value of 16 in all three osteoblastic cell lines and the HepG2 cell line. The Ct value for COMT was around 20 for all three osteoblast-like cell lines and 19 for the HepG2 cell line. For SV-HFO cells, the relative mRNA expression level was 6.9% (SD 2.7), for MG-63 it was 3.5% (SD 0.6), for SaOS it was 4.3% (SD 0.3), and for HepG2 it was 9.3% (SD 1.4) compared with GAPDH. For SV-HFO, there was no significant difference in relative COMT mRNA expression compared with the expression in HepG2. For MG-63 and SaOS, the relative COMT mRNA expression was two times lower compared with HepG2 COMT mRNA expression.


Figure 3
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  		FIG. 3. COMT mRNA expression relative to GAPDH mRNA expression in three osteoblast-like cell-lines and a HepG2 cell line. Results of two separate experiments are shown. n = 4 measurements for all cell lines. Mean relative mRNA expression for SV-HFO was 0.069 (SD 0.027); for MG-63, the relative COMT mRNA expression was 0.034 (SD 0.006); for SaOS, it was 0.043 (SD 0.003); and for HepG2, it was 0.093 (SD 0.014). #, Significant (P < 0.05) compared with HepG2 COMT mRNA expression.

 

   Discussion
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
 Discussion
References
 
In this study, we investigated whether the COMT Val158Met polymorphism is associated with parameters of bone health in a large prospective population-based cohort study, the Rotterdam Study. We found the Met158 variant to be associated with higher risk for osteoporotic and fragility fractures in elderly men.

In line with a previous study (16), we found no association with lumbar spine BMD and femoral neck BMD in postmenopausal women. Neither was there an association of the polymorphism with BMD measures in elderly men. This is not in concordance with a previous study reporting an association of the COMTL allele with lower femoral neck BMD in young adult Caucasian men (mean age 19 yr) (15). However, in our study population, the mean age for men was 68 yr. Because 19-yr-old males are not completely full-grown, and their BMD has not yet reached peak levels, the activity of COMT might influence bone mass during accrual, but this influence might stop once peak BMD is reached.

Interestingly, we observed that the COMTL allele increased fracture risk in elderly men, and with a similar trend for women. Male carriers of the COMTL allele had a 60% higher risk for osteoporotic fractures, whereas the risk for fragility fractures was even higher, 170%; however, confidence limits for this HR were wide, so the true effect of this polymorphism on fragility fractures is not clear yet. To be able to establish the true effect of the COMTL allele on fragility fractures, you need a larger study population with data on fragility fractures. The risk for vertebral fractures was not higher in men, suggesting a possible role for COMT in the strength of the appendicular skeleton more than in the axial skeleton. In women, there was a higher osteoporotic fracture risk in COMTLL genotype carriers, but there was no association with fragility fractures in women. It was shown previously that the activity of COMT in liver and red blood cells is 30% higher in men than in women (23). It could be that women, due to their already lower COMT activity, have developed an alternative pathway to compensate for this. In that case, the effect of the COMTL allele would be less dramatic in women than in men, which could explain the weaker and nonsignificant trend seen in women.

In line with a study by Worda et al. (14) of 159 postmenopausal women, we did not observe significant differences in serum estradiol levels between the three genotype groups (COMTHH, COMTHL, and COMTLL) in women. We also found no significant association with estradiol levels in men, which did not correspond with a recently published study by Eriksson et al. (13), who found increased serum estradiol levels in men with the COMTLL genotype. Our results indicate that COMT has no effect on systemic estradiol levels, which suggests that the COMT variant has more of a local effect than a systemic effect.

The fact that COMT mRNA is expressed in three different osteoblastic cell lines indicates that normally COMT is expressed in bone cells in vivo. We also found the expression levels in the osteoblastic cell lines to be quite similar to expression levels in HepG2 liver cells, where COMT is biologically active (24). This raises the possibility that the COMT variants influence estrogen levels locally, without affecting systemic levels.

The molecular mechanism by which lower activity of COMT could influence fracture risk is not yet known. A possible explanation might be that the estrogen metabolite 2-hydroxyestrone (2-OHE1), a catechol estrogen, which has already been shown to possess antiestrogenic activity in MCF-7 human breast cancer cells, also plays a role in bone. Binding of 2-OHE1 to the estrogen receptor {alpha} might prevent circulating estrogens from binding and interacting with this receptor (25). Lower activity of the COMTL variant would lead to higher concentrations of 2-OHE1 in target tissue, thereby inhibiting the function of estradiol in these tissues. In such a situation, one would not expect higher estrogen levels in both men and women carrying the COMTL allele.

We show an association of the COMTL allele with an increased risk for fractures in men but not with BMD. Such a genotype effect on fracture risk independent of BMD has already been seen for the estrogen receptor {alpha} XbaI polymorphism in the GENOMOS Study, a large prospective metaanalysis of almost 20,000 subjects (26) and also for polymorphisms in the vitamin D receptor and collagen 1A1 (27) genes. This observation suggests that COMT is involved in bone metabolic pathways other than those reflected in BMD but that still lead to higher fracture risk. It is known that other physiological factors, such as bone size (28), microarchitecture (bone structure) (29), and bone quality (30) can lead to an increased risk for fractures. On the basis of our data, we conclude that COMT does not have an effect directly on BMD but that it can affect one or more of these other factors, thereby leading to a higher fracture risk. Furthermore, in view of the pleiotropic effects of estradiol, we have to consider effects on phenotypes other than bone. For example, the COMTL allele might affect fall frequency or muscle strength, two predictors of fractures (31).

Our study has some limitations. First, genetic association studies can be influenced by population heterogeneity. However, all subjects in our study population were of Dutch-Caucasian ancestry and have a similar ethnic background. The COMT genotypes were also in Hardy-Weinberg equilibrium, and genotype frequencies were similar to those found in studies of other Caucasian populations (15, 32). Our study population can therefore be considered ethnically homogeneous and representative of the Dutch population. The second possible limitation is that we studied this association in only one Caucasian study sample. However, our study population is large, and we looked at well-defined endpoints. The third limitation might be that we examined a single SNP instead of haplotypes, whereas it has been suggested that studying haplotypes can increase power to detect rare causal alleles (33). Nevertheless, we studied a polymorphism with well-established functional effects.

Another limitation of genetic association studies are false-positive findings caused by multiple testing. Although we tested multiple endpoints, not all are independent. We have three main endpoints: BMD, fracture risk, and estradiol levels. Nevertheless, replication of our findings is needed.

In summary, this population-based study provides evidence that the COMTL allele is associated with an increased risk for osteoporotic and fragility fractures in elderly men. There was no association of this allele with bone parameters in women. The biological mechanism behind this relation remains to be elucidated.

Web resources

URLs for data presented herein are as follows: dbSNP, http://www.ncbi.nlm.nih.gov/SNP/; HapMap data, http://hapmap.jst.go.jp/cgi-perl/gbrowse/hapmap/.


   Acknowledgments
 
We are very grateful to the participants of the Rotterdam study and to the DXA and radiograph technicians, L. Buist and H. W. M. Mathot. Furthermore, we acknowledge all participating general practitioners and the many field workers in the research center in Ommoord, Rotterdam, The Netherlands.


   Footnotes
 
This study is supported by The Netherlands Organization of Scientific Research-Research Institute for Diseases in the Elderly (Grant 014-93-015; RIDE2) and the European Commission [Grants QLK6-CT-2002-02629 (GENOMOS) and QLK6–2002-00491 (NEMO)].

Disclosure: L.S., J.B.J.v.M., M.J., P.P.A., J.P.T.v.L., A.H., F.H.d.J., H.A.P.P., and A.G.U. have nothing to declare.

First Published Online May 15, 2007

Abbreviations: BMD, Bone mineral density; CI, confidence interval; COMT, catechol-O-methyltransferase; COMTH, high-activity COMT allele; COMTL, low-activity COMT allele; HR, hazard ratio; 2-OHE1, 2-hydroxyestrone; SNP, single-nucleotide polymorphism.

Received September 29, 2006.

Accepted May 4, 2007.


   References
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
 Discussion
 References
 

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