J Peeters

Polymorphisms of the VDR, ER and COLIA1 Genes and Osteoporotic Hip Fracture in Elderly Postmenopausal Women

Abstract: In view of the reported associations between osteoporosis and polymorphisms of the vitamin D receptor (VDR), collagen Iα1 (COLIA1) and estrogen receptor (ER) genes, an association study was performed between VDR, COLIA1, and ER genotypes and bone mineral density, biochemical markers of bone turnover and hip fracture occurrence in Belgian older postmenopausal women. The gene polymorphisms were evaluated by restriction fragment length polymorphism analyses, using the restriction enzymes BsmI (VDR), AccB7I (COLIA1), and PvuII and XbaI (ER), respectively. As expected, bone mineral density and biochemical analyses demonstrated significant differences between hip fracture patients and elderly controls. However, no significant differences in genotype distributions or allele frequencies were observed between the cases (n= 135, age 78 ± 9 years) and controls (n= 239, age 76 ± 4 years) for any of the gene polymorphisms. Stratification of both study populations according to VDR, COLIA1 or ER genotype did not reveal any statistically significant difference in bone density or bone turnover between subgroups with different genotypes. In conclusion, despite its limited statistical power the outcome of this study does not support the hypothesis of a major contribution of the VDR, COLIA1 or ER polymorphisms to explain variations in bone mineral density or bone turnover, or to identify elderly women at risk of osteoporotic hip fracture.

Lack of association between osteoarthritis of the hip and gene polymorphisms of VDR, COL1A1, and COL2A1 in postmenopausal women

OBJECTIVE To evaluate whether DNA polymorphisms of the VDR (vitamin D receptor), COL1A1 (alpha1 type I collagen), and COL2A1 (alpha1 type II collagen) genes, which have previously been linked to bone mineral density (BMD) and/or osteoarthritis (OA), are also associated with OA of the hip (OAH). METHODS Genotype frequency distributions were compared between a group of 239 elderly healthy female controls and a group of 75 female OAH patients who underwent total hip replacement. All participants underwent BMD measurements of the total body, lumbar spine, and femoral neck. Mutation detection was done using polymerase chain reaction followed by restriction enzyme digestion. Polymorphisms were detected by digestion with Bsm I for VDR, Acc B7I for COL1A1, and Pvu II for COL2A1. RESULTS None of the genotype frequency distributions of any of the examined polymorphisms was significantly different between the OAH group and the control group. No significant differences in BMD variables were observed after stratification of the subjects according to the examined genotypes within each study group. CONCLUSION In Belgian postmenopausal women, the examined polymorphisms of the candidate genes VDR, COL1A1, and COL2A1 do not significantly contribute to an increased prevalence of OAH or to differences in BMD.

Alterations of the mineralization profile and osteocalcin concentrations in osteoarthritic cortical iliac crest bone

SummaryThe relation between bone mineralization and osteocalcin content was investigated in iliac crest cortical bone obtained at necropsy in young females and in two groups of elderly women with and without osteoarthritis of the hands evaluated by X-ray. Using density fractionation technique, the bone was separated into fractions of increasing density from 1.72 to 2.30 g/ml. The mineralization profile revealed a significant shift to higher densities in the osteoarthritis cases compared with young adults (P<0.005) and age-sexmatched controls (P<0.001). The ash, calcium, and phosphorus content of the bone increased with increasing density of the fractions whereas collagen content, measured as hydroxyproline, decreased. The osteocalcin concentration of each fraction was determined in the supernatants obtained after EDTA-extraction in the presence of protease inhibitors. In the young control and osteoarthritis group, the osteocalcin content in the lowest density fractions was higher compared with the older non-osteoarthritic group. Osteocalcin content of the high density fractions, representing highly mineralized osteons, was the same in the three groups studied. These findings support the hypothesis that quality differences in bone may explain the inverse relationship between osteoarthritis and osteoporosis.

Deletion of frizzled-related protein reduces voluntary running exercise performance in mice

OBJECTIVE To study the effect of frizzled-related protein (Frzb) deletion in mice on voluntary running wheel exercise performance and osteoarthritis. METHODS At the age of 7 weeks, Frzb(-/-) and wild-type mice were grouped and a running wheel was introduced into the cage. At week 8, all mice were caged solitarily with a running wheel available. Mice were allowed free exercise for 6-12 months and distances run were recorded daily. Non-running mice were used as additional control group. X-rays of knees and hips were taken at different time points. At the end of the experiment, mice were sacrificed and joints were processed for histological evaluation. Cartilage damage, synovitis and osteophyte formation were scored. Muscle fiber composition of the soleus and extensor digitorum longus was studied by immunofluorescence. RESULTS At the age of 6 months, both female and male wild-type mice showed a significantly greater exercise performance than the Frzb(-/-) mice (P<0.05). At 1 year, the difference was still significant for male mice, but not for females. Running exercise did not significantly affect severity of osteoarthritis. No statistical differences in osteoarthritis severity were seen between Frzb(-/-) mice and wild-type mice. No differences were seen in muscle composition between Frzb(-/-) mice and wild-type mice. CONCLUSION Absence of Frzb in mice reduced voluntary exercise performance in running wheels. These experiments demonstrate that the effects of genes in mice can also be evaluated using functional outcomes such as running wheel exercise performance, similar to evolving practice in human clinical trials.

Zfhx1a and Zfhx1b mRNAs have non-overlapping expression domains during chick and mouse midgestation limb development

Smad-interacting protein 1 (Zfhx1b, Sip1) and Zfhx1a (deltaEF1) are transcriptional repressors belonging to the family of two-handed zinc finger/homeodomain proteins. Both of the proteins bind to a bipartite CACCT/CACCTG DNA sequence, but only Sip 1 can interact with activated SMAD proteins in vitro. To gain insight into their developmental roles, we investigated the mRNA expression patterns of both genes in developing mouse and chick limbs by in situ hybridization. To improve the resolution of the expression mapping we have used a dual in situ hybridization (DISH) technique allowing for a detection of expression of two different mRNAs on the same tissue section. Using DISH we could demonstrate that both genes were expressed in distinct non-overlapping patterns in developing limbs of both species. Zfhx1a was expressed in a cell population immediately adjacent to cartilage anlagen as well as in developing tendons. Zfhx1b, in contrast, was present in a broad area around developing tendon and partially overlapping with the expression of genes associated with myogenic differentiation.

65 FRIZZLED RELATED PROTEIN (FRZB) IS INVOLVED IN DIFFERENT ASPECTS OF JOINT BIOLOGY AND PATHOLOGY

Conclusions: The knee cartilage of the Hartley guinea pig undergoes an increase in the expression of genes associated with hypertrophy by 12 months of age coincident with the development of severe osteoarthritic changes. Compared to the OA-prone Hartley strain, Strain 13 guinea pig knee cartilage showed much lower expression of genes associated with the hypertrophic cartilage phenotype. These findings are compatible with the lesser severity of OA in this model. Interestingly, Hartley and Strain 13 guinea pigs showed similar and high ADAMTS-5 mRNA expression, suggesting that ADAMTS-5 may be relevant in OA development in the guinea pig model, but may not account for the differential susceptibility of the two strains to OA.

Lack of In vitro evidence for storage of 1,25-dihydroxycholecalciferol (1,25(OH)2D3) and 1,25(OH)2D3 binding protein in skeletal matrix

A few studies have reported on the measurement of 1,25-dihydroxycholecalciferol (1,25(OH)2D3) in bone, using chloroform/methanol extraction and radioreceptor assay. As the significance of bone 1,25(OH)2D3 content was not defined in any of these reports, the objective of the current investigation was to determine whether 1,25(OH)2D3 may be stored in skeletal matrix. Bone powder samples from the iliac crest were extracted in ethylacetate/cyclohexane and 1,25(OH)2D3 isolated from the extract by means of Sephadex LH-20 and high pressure liquid chromatographic separation and subsequently measured by radioimmunoassay (RIA). Within the detection range of the RIA, no 1,25(OH)2D3 could be measured, suggesting that 1,25(OH)2D3 is not stored in skeletal matrix. Vitamin D bone concentrations previously measured may therefore have reflected plasma contamination. Consistent with this hypothesis, only traces of skeletal 1,25(OH)2D3 binding protein were measured when compared with serum values. Although 1,25(OH)2D3 may act as a potential local determinant of bone remodeling, there is no evidence supporting a delayed paracrine function by matrix-derived 1,25(OH)2D3.