Significant association between common polymorphisms in the aromatase gene CYP19A1 and bone mineral density in postmenopausal women.

17ß-Estradiol is important in maintaining bone structure, and regulation of its synthesis plays an important role in the development of postmenopausal osteoporosis. We and others have demonstrated associations between variation in the CYP19A1 gene (encoding aromatase) and areal bone mineral density (aBMD) phenotypes in women. In the present study 33 tag polymorphisms were genotyped across the CYP19A1 gene in a population of 1,185 Caucasian postmenopausal women to test the association between sequence variations, total DXA hip aBMD, and circulating 17ß-estradiol levels. An in silico bioinformatics analysis was performed for single nucleotide polymorphisms (SNPs) associated with aBMD to identify putative functional effects, while linkage disequilibrium analysis of these SNPs was undertaken with previously published sequence variants. Five SNPs located in the central third of the gene were strongly associated with total-hip aBMD after adjustment for age (P = 0.006-0.013). A haplotype analysis of these five SNPs revealed an association between the haplotype C-G-G-G-C and increased aBMD (P = 0.008) and the haplotype A-A-A-A-A and a decreased aBMD (P = 0.021). The haplotype frequency was 9.0% for C-G-G-G-C and 15.4% for A-A-A-A-A, with the variation in mean total-hip aBMD explained by the haplotype analyses being 5% and 7%, respectively. None of these polymorphisms was significantly associated with circulating 17ß-estradiol levels. In conclusion, common genetic variations within the CYP19A1 gene are significantly associated with aBMD in postmenopausal Caucasian women.

Homozygous deletion of the UGT2B17 gene is not associated with osteoporosis risk in elderly Caucasian women.

UNLABELLED: Previously, homozygous deletion of the UGT2B17 gene has shown association with hip fracture. Using a high-throughput qRT-PCR assay, we genotyped UGT2B17 copy number variation (CNV) in 1,347 elderly Caucasian women and examined for effects on bone phenotypes. We found no evidence of association between UGT2B17 CNV and osteoporosis risk in this population. INTRODUCTION: Genetic studies of osteoporosis commonly examine SNPs in candidate genes or whole genome analyses, but insertions and deletions of DNA, collectively called CNV, also comprise a large amount of the genetic variability between individuals. Previously, homozygous deletion of the UGT2B17 gene in CNV 4q13.2, which encodes an enzyme that mediates the glucuronidation of steroid hormones, has shown association with the risk of hip fracture. METHODS: We used a quantitative real-time PCR assay for genotyping the UGT2B17 CNV in a well-characterized population study of 1,347 Caucasian women aged 75.2 ± 2.7 years (mean ± SD), to assess the effect of the CNV on bone mass density (BMD) at the total hip site and osteoporosis risk. RESULTS: The UGT2B17 CNV distribution was consistent with the expected Hardy-Weinberg distribution and not different from frequencies previously reported in a Caucasian population. Data from ANCOVA of age- and weight-adjusted BMD for UGT2B17 CNV genotype showed no significant difference between genotype groups. Individuals with homozygous or heterozygous deletion of the UGT2B17 gene showed no increased risk of incident fragility fracture. CONCLUSIONS: These data suggest that quantitative real-time PCR is a rapid and efficient technique for determination of candidate CNVs, including the UGT2B17 CNV; however, we found no evidence of an effect of UGT2B17 CNV on osteoporosis risk in elderly Caucasian women.

Bone mineral density, osteoporosis, and osteoporotic fractures: a genome-wide association study.

BACKGROUND: Osteoporosis is diagnosed by the measurement of bone mineral density, which is a highly heritable and multifactorial trait. We aimed to identify genetic loci that are associated with bone mineral density. METHODS: In this genome-wide association study, we identified the most promising of 314 075 single nucleotide polymorphisms (SNPs) in 2094 women in a UK study. We then tested these SNPs for replication in 6463 people from three other cohorts in western Europe. We also investigated allelic expression in lymphoblast cell lines. We tested the association between the replicated SNPs and osteoporotic fractures with data from two studies. FINDINGS: We identified genome-wide evidence for an association between bone mineral density and two SNPs (p<5x10(-8)). The SNPs were rs4355801, on chromosome 8, near to the TNFRSF11B (osteoprotegerin) gene, and rs3736228, on chromosome 11 in the LRP5 (lipoprotein-receptor-related protein) gene. A non-synonymous SNP in the LRP5 gene was associated with decreased bone mineral density (rs3736228, p=6.3x10(-12) for lumbar spine and p=1.9x10(-4) for femoral neck) and an increased risk of both osteoporotic fractures (odds ratio [OR] 1.3, 95% CI 1.09-1.52, p=0.002) and osteoporosis (OR 1.3, 1.08-1.63, p=0.008). Three SNPs near the TNFRSF11B gene were associated with decreased bone mineral density (top SNP, rs4355801: p=7.6x10(-10) for lumbar spine and p=3.3x10(-8) for femoral neck) and increased risk of osteoporosis (OR 1.2, 95% CI 1.01-1.42, p=0.038). For carriers of the risk allele at rs4355801, expression of TNFRSF11B in lymphoblast cell lines was halved (p=3.0x10(-6)). 1883 (22%) of 8557 people were at least heterozygous for these risk alleles, and these alleles had a cumulative association with bone mineral density (trend p=2.3x10(-17)). The presence of both risk alleles increased the risk of osteoporotic fractures (OR 1.3, 1.08-1.63, p=0.006) and this effect was independent of bone mineral density. INTERPRETATION: Two gene variants of key biological proteins increase the risk of osteoporosis and osteoporotic fracture. The combined effect of these risk alleles on fractures is similar to that of most well-replicated environmental risk factors, and they are present in more than one in five white people, suggesting a potential role in screening.

Bone structural effects of variation in the TNFRSF1B gene encoding the tumor necrosis factor receptor 2.

UNLABELLED: The 1p36 region of the human genome has been identified as containing a QTL for BMD in multiple studies. We analysed the TNFRSF1B gene from this region, which encodes the TNF receptor 2, in two large population-based cohorts. Our results suggest that variation in TNFRSF1B is associated with BMD. INTRODUCTION: The TNFRSF1B gene, encoding the TNF receptor 2, is a strong positional and functional candidate gene for impaired bone structure through the role that TNF has in bone cells. The aims of this study were to evaluate the role of variations in the TNFRSF1B gene on bone structure and osteoporotic fracture risk in postmenopausal women. METHODS: Six SNPs in TNFRSF1B were analysed in a cohort of 1,190 postmenopausal Australian women, three of which were also genotyped in an independent cohort of 811 UK postmenopausal women. Differences in phenotypic means for genotype groups were examined using one-way ANOVA and ANCOVA. RESULTS: Significant associations were seen for IVS1+5580A>G with BMD and QUS parameters in the Australian population (P = 0.008 - 0.034) and with hip BMD parameters in the UK population (P = 0.005 - 0.029). Significant associations were also observed between IVS1+6528G>A and hip BMD parameters in the UK cohort (P = 0.0002 - 0.003). We then combined the data from the two cohorts and observed significant associations between both IVS1+5580A>G and IVS1+6528G>A and hip BMD parameters (P = 0.002 - 0.033). CONCLUSIONS: Genetic variation in TNFRSF1B plays a role in the determination of bone structure in Caucasian postmenopausal women, possibly through effects on osteoblast and osteoclast differentiation.

Polymorphisms in ALOX12, but not ALOX15, are significantly associated with BMD in postmenopausal women.

The murine arachidonate 15-lipoxygenase gene (Alox15) has recently been identified as a negative regulator of peak bone mineral density (BMD). The human ALOX15 gene shares significant sequence homology with the murine Alox15 gene; however, the human arachidonate 12-lipoxygenase gene (ALOX12) is functionally more similar to the mouse gene. Multiple single-nucleotide polymorphisms (SNPs) in the human ALOX15 and ALOX12 genes have previously been reported to be significantly associated with BMD in humans. On the basis of these data, we carried out our own investigation of the human ALOX15 and ALOX12 genes and their relationship with hip and spine BMD parameters. The study population consisted of 779 postmenopausal women with a mean (+/- standard deviation) age of 62.5 +/- 5.9 years at BMD measurement and was recruited from a single large general practice in Chingford, northeast London. Three SNPs from ALOX15 and five from ALOX12 were analyzed. None of the SNPs that we analyzed in ALOX15 were significantly associated with any of the BMD parameters or fracture data. However, we found that three SNPs from ALOX12, all previously associated with spine BMD in women, were significantly associated with spine and various hip BMD parameters in our cohort (P = 0.029-0.049). In conclusion, we found no association between polymorphism in ALOX15 and BMD phenotypes but were able to replicate previous findings that genetic variation in ALOX12 seems to play a role in determining bone structure in Caucasian women.

Polymorphism of the follistatin gene in polycystic ovary syndrome.

Follistatin has been reported as a candidate gene for polycystic ovary syndrome (PCOS) from linkage and association studies. Acting to regulate the development of ovarian follicles and as an antagonist to aromatase activity, alterations in follistatin function or expression may result in key features of PCOS such as reduced serum FSH, impaired ovarian follicle development and augmented ovarian androgen production. We investigated polymorphisms in the FST gene to determine if genetic variation is associated with susceptibility to PCOS or key phenotypic features of PCOS patients in a case-control association study. One hundred and seventy-three PCOS patients of Caucasian descent (mean age 30.0 +/- 4.8 years), conforming to the NIH diagnostic criteria, were recruited from a clinical practice database and 107 normal ovulating women (mean age 38.8 +/- 13.4 years) were recruited from the general community as control subjects. Morphometric data, biochemistry and genomic DNA were collected from study subjects and genotyping was performed on seven Single nucleotide polymorphisms (SNPs) in the FST gene region. Allele frequencies of the SNPs were rs1423560 G/C (0.99/0.01), rs3797297 C/A (0.80/0.20), rs11745088 C/G (0.98/0.02), rs3203788 A/T (0.98/0.02) and rs1062809 G/C (1.00/-), rs1127760 A/T (0.98/0.02) and rs1127761 A/T (0.98/0.02), and these were not significantly different between the PCOS and control groups (P < 0.05). Statistical analysis revealed significant associations between the SNP rs3797297 and sex hormone-binding globulin (P = 0.04) and free androgen index (FAI) (P < 0.01). We conclude that FST is not a susceptibility locus for PCOS; however, the SNP rs3797297 from FST gene was associated with androgenic markers for PCOS and may be of importance in the hyperandrogenaemia of the disease.

No associations between OPG gene polymorphisms or serum levels and measures of osteoporosis in elderly Australian women.

Bone mass is the single most important risk factor for osteoporotic fractures in the elderly and is mainly influenced by genetic factors accounting for 40-75% of the inter-individual variation. Critical for the bone remodeling process is the balance between the newly discovered members of the tumor necrosis factor ligand and receptor superfamilies, osteoprotegerin (OPG) and receptor activator of nuclear factor-kappaB ligand, which mediate the effects of many upstream regulators of bone metabolism. In the present study, we evaluated the impact of sequence variations in the OPG gene on bone mass, bone-related biochemistry including serum OPG and fracture frequency in elderly Australian women. A total of 1101 women were genotyped for 3 different single nucleotide polymorphisms (SNP) within the OPG gene (G1181C, T950C and A163G). The effects of these SNPs and serum OPG on calcaneal quantitative ultrasound measurements, osteodensitometry of the hip and bone-related biochemistry were examined. We found no significant relationship between sequence variations in the OPG gene or serum OPG and bone mass, bone-related biochemistry or fracture frequency. Our findings confirm some recent publications investigating the same SNPs but diverge from others, indicating that generalization of the relationships found in this type of study must be done with caution and signify the importance of determining associations between polymorphisms and osteoporosis in different ethnic groups.

Klotho gene polymorphisms are associated with osteocalcin levels but not bone density of aged postmenopausal women.

Osteoporosis is known to have a strong genetic basis. It has been proposed that polymorphisms within the KL (klotho) gene have a significant effect on aging, in particular, the osteoblast defect of aging. The association between polymorphisms within this gene and biochemical markers of bone formation and resorption, bone structure, and fracture rates was studied in 1,190 postmenopausal women with a mean age of 75 years. Genotyping of these polymorphic sites was carried out using Matrix-Assisted Laser Desorption Ionization--Time of Flight (MALDI-ToF) mass spectrometry. The G allele of SNP c.1775G>A was associated with a lower osteocalcin level than the A allele (P = 0.004) in a codominant model. SNPs C-387T and IVS1+8262c>t both showed nonsignificant associations with osteocalcin (P values of 0.063 and 0.068, respectively), but a haplotype analysis of 2 of 5 haplotypes of the three SNPs with a frequency greater than 4% revealed a significant association with osteocalcin (P = 0.036). None of the individual polymorphisms or haplotypes analyzed showed any associations with a marker of bone resorption the deoxypyridinoline creatinine ratio, bone structure, or fracture data. Therefore, the G polymorphism within the c.1775G>A SNP site and a haplotype including this are associated with a reduced osteoblast product osteocalcin. These data suggest that variation in the KL gene product affects osteoblast activity independent of osteoclast activity but that this defect does not result in an effect on bone structure in this population, perhaps because of "rescue" by other genetic or environmental factors in this population.