Identification of non-synonymous polymorphisms in the WDSOF1 gene as novel susceptibility markers for low bone mineral density in Japanese postmenopausal women.

Genetic factors are important for the development of osteoporosis. During the search for novel markers of single-nucleotide polymorphisms (SNPs) associated with bone mineral density (BMD) by performing a large-scale SNP screen with 251 Japanese postmenopausal women utilizing 50K SNP array, we here focused on the rs1370005 in the WD repeats and SOF1 domain-containing (WDSOF1) gene because we could found common non-synonymous variants in this WDSOF1 gene. The analysis of linkage disequilibrium (LD) in the WDSOF1 gene revealed that rs1370005 and 3 other non-synonymous SNPs (Arg47Ser, Pro108Leu and Ile194Val) lie in a 30-kb region of high LD. Quantitative real-time PCR (qRT-PCR) analysis showed that WDSOF1 mRNA was expressed in mouse primary osteoblasts and osteoclasts, suggesting that WDSOF1 plays some roles in the bone metabolism. We examined the 3 non-synonymous SNPs in WDSOF1 gene in 750 Japanese postmenopausal women. A trend test showed that Arg47Ser, Pro108Leu, and Ile194Val genotypes were significant associated with total body BMD (Arg47Ser; P=0.021, Pro108Leu; P=0.022 and Ile194Val; P=0.009). We also compared Z scores for total body BMD between the subjects bearing at least one minor allele and those lacking the minor allele using unpaired t test. Subjects with the one or two minor alleles had significantly lower Z scores for total body BMD (Arg47Ser; P=0.010, Pro108Leu; P=0.019 and Ile194Val; P=0.003). The present study suggests that these non-synonymous WDSOF1 polymorphisms play a role in the genetic susceptibility to osteoporosis.

Association of a single nucleotide polymorphism in the constitutive androstane receptor gene with bone mineral density.

BACKGROUND: Nuclear receptors play an important role in bone metabolism. In bone cells, the vitamin D receptor (VDR) and the steroid and xenobiotic receptor (SXR) are activated by vitamin D and vitamin K2, respectively. VDR and SXR are the NR1I subfamily members of nuclear receptors. We speculated that the constitutive androstane receptor (CAR), the third member of the NR1I subfamily, also could be implicated in the regulation of bone metabolism. Therefore, we analyzed expression of CAR mRNA in osteoblasts and then examined association of a single nucleotide polymorphism (SNP) in the human CAR gene at intron 2 (IVS2-99C>T, rs2502815) with bone mineral density (BMD). METHODS: Expression levels of CAR mRNA were analyzed during the culture course of rat primary osteoblasts. Association of an SNP in the CAR gene with BMD was examined in 548 healthy Japanese postmenopausal women. RESULTS: CAR mRNA increased at day 16 and then increased during culture of rat primary osteoblasts. The increase of CAR mRNA was parallel with the increase of alkaline phosphatase expression, a differentiation marker of osteoblasts. As a result of association study of an SNP in the CAR gene at intron 2, subjects with the CC genotype (n = 208) had significantly higher BMD than subjects with the TT or CT genotype (n = 340) (lumbar spine BMD, P = 0.0185; total body BMD, P = 0.0416). CONCLUSION: CAR mRNA was expressed and regulated in primary osteoblasts. A genetic variation at the CAR gene locus is associated with BMD, suggesting an involvement of the CAR gene in bone metabolism.

A1330V variant of the low-density lipoprotein receptor-related protein 5 (LRP5) gene decreases Wnt signaling and affects the total body bone mineral density in Japanese women.

Wnt signaling is an important regulator of bone homeostasis. The Wnt co-receptor, namely, low-density lipoprotein receptor-related protein 5 (LRP5), initiates Wnt signal transduction. Recently, we and several other groups have shown that there is a single nucleotide polymorphism (SNP) located in the exon 18 of the LRP5 gene that leads to an amino acid change (3989C > T, A1330V), and is associated with lumbar spine, femoral neck, and radial bone mineral density (BMD), and incidence of fracture. These data suggest that the A1330V variation in the LRP5 gene may affect the pathogenesis of osteoporosis. However, the functional basis of the A1330V variation remains unclear. In the present study, we analyzed the effect of the A1330V variation on Wnt activity. We also investigated the association between this LRP5 SNP and total body BMD using 739 postmenopausal women. LRP5 with the A1330V SNP were transiently coexpressed with Wnt3a in 293T cells and their activity was evaluated by the TCF-Lef reporter assay. In vitro, the TCF-Lef activity in presence of Wnt3a in cells expressing LRP5 and carrying the T allele (Valine at 1330 (V1330)) of exon 18 was significantly reduced as compared to the wild-type allele. The association between the A1330V SNP and total body BMD were replicated in 739 postmenopausal Japanese women (AA vs. VV; P = 0.0026). These data suggest that the V1330 variant in the LRP5 gene decreases Wnt activity, which in turn decreases the BMD.

TRIM44 interacts with and stabilizes terf, a TRIM ubiquitin E3 ligase.

Terf/TRIM17 is a member of the TRIM family of proteins, which is characterized by the RING finger, B-box, and coiled-coil domains. In the present study, we found that terf interacts with TRIM44. Terf underwent ubiquitination in vitro in the presence of the E2 enzyme UbcH6; this suggests that terf exhibits E3 ubiquitin ligase activity. It was also found that terf was conjugated with polyubiquitin chains and stabilized by the proteasome inhibitor in mammalian cells; this suggested that terf rendered itself susceptible to proteasomal degradation through polyubiquitination. We also found that TRIM44 inhibited ubiquitination of terf, and thus stabilized the protein. The N-terminal region of TRIM44 contains a zinc-finger domain found in ubiquitin hydrolases (ZF UBP) and ubiquitin specific proteases (USPs). Thus, we proposed that TRIM44 may function as a new class of the "USP-like-TRIM" which regulates the activity of associated TRIM proteins.

Bone mass effects of a Smad6 gene polymorphism in Japanese postmenopausal women.

Smad6 plays pivotal roles in the negative regulation of transforming growth factor beta (TGFbeta) family signaling as one of the feedback molecules. Here, we analyzed whether the human Smad6 gene is involved in the regulation of bone mass, using association analysis between bone mineral density (BMD) and single-nucleotide polymorphism (SNP) in the Smad6 gene. Association of an SNP at IVS3+26115A>C (intron 3, rs755451) in the Smad6 gene with BMD was examined in 721 Japanese postmenopausal Japanese women (age 65.2 +/- 9.6 years; mean +/- SD). The subjects bearing at least one variant C allele (CC +/- AC; n = 387) had significantly lower Z-scores for total body and lumbar BMD than the subjects with no C allele (AA; n = 334) (total body, 0.23 +/- 0.98 versus 0.50 +/- 1.07; P = 0.0004; lumbar spine, -0.20 +/- 1.38 versus 0.10 +/- 1.48; P = 0.0050). These findings suggest that the Smad6 gene is a candidate for the genetic determinants of BMD in postmenopausal women, and this SNP could be useful as a genetic marker for predicting the risk of osteoporosis.

Association of a single nucleotide polymorphism in the insulin-like growth factor-1 receptor gene with spinal disc degeneration in postmenopausal Japanese women.

STUDY DESIGN: An association study investigating the genetic etiology for spinal disc degeneration. OBJECTIVE: To determine the association of single-nucleotide polymorphism (SNP) in the insulin-like growth factor-1 receptor (IGF1R) with spinal disc degeneration. SUMMARY OF BACKGROUND DATA: Insulin-like growth factor-1 (IGF-1) signaling pathway is involved in cartilage development and homeostasis, suggesting that genetic variations of genes involved in this pathway may affect the pathogenesis of cartilage-related diseases, such as disc degeneration. METHODS: We evaluated the presence of endplate sclerosis, osteophytes, and narrowing of disc spaces in 434 Japanese postmenopausal women. A SNP in the IGF1R gene at intron 1 was determined using TaqMan polymerase chain reaction method. RESULTS: We compared those who carried the G allele (GG or GC, n = 290) with those who did not (CC, n = 144). We found that the subjects with the G allele (GG or GC) were significantly over-represented in the subjects having higher disc narrowing score (P = 0.0033; odds ratio, 2.04; 95% confidence interval, 1.27-3.29 by logistic regression analysis). CONCLUSION: We suggest that a genetic variation at the IGF1R gene locus is associated with spinal disc degeneration, in line with the involvement of the IGF1R gene in the cartilage metabolism.

Transmission of the parasite Ichthyophonus hoferi in cultured rainbow trout and comparison of epidemic models.

The epidemic process of the parasite Ichthyophonus hoferi in cultured rainbow trout Oncorhynchus mykiss was quantitatively estimated by both the cohabitation experiment and two standard models (the Kermarck-McKendrick model and the Reed-Frost model). For analysis of the parasite transmission by cohabitation, fish in two replicate tanks were exposed to 1, 5, or 10 infected fish, and daily mortality was counted for 102 d. Despite simple experiments for artificial exposure to the pathogen, the daily estimate of dead fish in the Kermarck-McKendrick model did not fit the observed number of dead fish in the experiment. In contrast, when the longest possible incubation period (generation time) was assumed to be 51 d in the Reed-Frost model, the estimated number of dead fish in discrete generations was close to the observed number of dead fish. If the time unit was 51 d, the estimated mortalities in the generation-based Kermarck-McKendrick model were significantly correlated with observed mortalities. These results suggest that the deterministic aspects of the epidemic process of the parasite can be quantitatively demonstrated on a 51-d timescale or longer, whereas transmission on a daily timescale is uncertain.

Association of a single nucleotide polymorphism in the WISP1 gene with spinal osteoarthritis in postmenopausal Japanese women.

The Wnt-beta-catenin signaling pathway that regulates bone density is also involved in cartilage development and homeostasis in vivo. Here, we assumed that genetic variation in Wnt-beta-catenin signaling genes can affect the pathogenesis of cartilage related diseases, such as osteoarthritis. Wnt-1-induced secreted protein 1 (WISP1) is a target of the Wnt pathway and directly regulated by beta-catenin. In the present study, we analyzed the association of a single nucleotide polymorphism (SNP) in the WISP1 3'-UTR region with the development of radiographically observable osteoarthritis of the spine. For this purpose, we evaluated the presence of osteophytes, endplate sclerosis, and narrowing of disc spaces in 304 postmenopausal Japanese women. We compared those who carried the G allele (GG or GA, n = 184) with those who did not (AA, n = 120). We found that the subjects without the G allele (AA) were significantly over-represented in the subjects having higher endplate sclerosis score (P = 0.0069; odds ratio, 2.91; 95% confidence interval, 1.34-6.30 by logistic regression analysis). On the other hand, the occurrence of disc narrowing and osteophyte formation did not significantly differ between those with and without at least one G allele. Thus, we suggest that a genetic variation in the WISP1 gene locus is associated with spinal osteoarthritis, in line with the involvement of the Wnt-beta-catenin-regulated gene in bone and cartilage metabolism.

Q89R polymorphism in the LDL receptor-related protein 5 gene is associated with spinal osteoarthritis in postmenopausal Japanese women.

STUDY DESIGN: An association study investigating the genetic etiology for spinal osteoarthritis. OBJECTIVE: To determine the association of single-nucleotide polymorphism (SNP) causing an amino-acid change (Q89R) in the low-density lipoprotein receptor-related protein 5 (LRP5) coding region with spinal osteoarthritis. SUMMARY OF BACKGROUND DATA: Wnt/beta-catenin signaling pathway regulates bone density through a Wnt coreceptor LRP5. This pathway is also involved in cartilage development and homeostasis, suggesting that genetic variation in LRP5 gene may affect the pathogenesis of cartilage-related diseases, such as osteoarthritis. METHODS: We evaluated the presence of osteophytes, endplate sclerosis, and narrowing of disc spaces in 357 Japanese postmenopausal women. Missense coding SNP for Q89R of LRP5 gene was determined using TaqMan polymerase chain reaction (PCR) method. RESULTS: We found that subjects without the R allele (QQ; n = 321) had a significantly lower osteophyte formation score than did subjects bearing at least one R allele (QR + RR; n = 36) (7.80 vs. 10.89, P = 0.0019 by analysis of covariance). CONCLUSIONS: We suggest that a genetic variation at the LRP5 gene locus is associated with spinal osteoarthritis, in line with the involvement of the LRP5 gene in the bone and cartilage metabolism.