We aimed to identify genetic variants associated with cortical bone thickness (CBT) and bone mineral density (BMD) by performing two separate genome-wide association study (GWAS) meta-analyses for CBT in 3 cohorts comprising 5,878 European subjects and for BMD in 5 cohorts comprising 5,672 individuals. We then assessed selected single-nucleotide polymorphisms (SNPs) for osteoporotic fracture in 2,023 cases and 3,740 controls. Association with CBT and forearm BMD was tested for ∼2.5 million SNPs in each cohort separately, and results were meta-analyzed using fixed effect meta-analysis. We identified a missense SNP (Thr>Ile; rs2707466) located in the WNT16 gene (7q31), associated with CBT (effect size of −0.11 standard deviations [SD] per C allele, P = 6.2×10−9). This SNP, as well as another nonsynonymous SNP rs2908004 (Gly>Arg), also had genome-wide significant association with forearm BMD (−0.14 SD per C allele, P = 2.3×10−12, and −0.16 SD per G allele, P = 1.2×10−15, respectively). Four genome-wide significant SNPs arising from BMD meta-analysis were tested for association with forearm fracture. SNP rs7776725 in FAM3C, a gene adjacent to WNT16, was associated with a genome-wide significant increased risk of forearm fracture (OR = 1.33, P = 7.3×10−9), with genome-wide suggestive signals from the two missense variants in WNT16 (rs2908004: OR = 1.22, P = 4.9×10−6 and rs2707466: OR = 1.22, P = 7.2×10−6). We next generated a homozygous mouse with targeted disruption of Wnt16. Female Wnt16−/− mice had 27% (P<0.001) thinner cortical bones at the femur midshaft, and bone strength measures were reduced between 43%–61% (6.5×10−13

SUMMARYTo determine whether the calcium-sensing receptor (CaR) participates in tooth formation and dental alveolar bone development in mandibles in vivo, we examined these processes, as well as mineralization, in 2-week-old CaR-knockout (CaR -/-) mice. We also attempted to rescue the phenotype of CaR -/-mice by genetic means, in mice doubly homozygous for CaR and 25-hydroxyvitamin D 1a-hydroxylase [1a(OH)ase] or parathyroid hormone (Pth). In CaR -/-mice, which exhibited hypercalcemia, hypophosphatemia and increased serum PTH, the volumes of teeth and of dental alveolar bone were decreased dramatically, whereas the ratio of the area of predentin to total dentin and the number and surface of osteoblasts in dental alveolar bone were increased significantly, as compared with wild-type littermates. The normocalcemia present in CaR -/-

The calcium-sensing receptor complements parathyroid hormone-induced bone turnover in discrete skeletal compartments in mice. Am J Physiol Endocrinol Metab 302: E841-E851, 2012. First published January 24, 2012; doi:10.1152/ajpendo.00599.2011.-Although the calcium-sensing receptor (CaSR) and parathyroid hormone (PTH) may each exert skeletal effects, it is uncertain how CaSR and PTH interact at the level of bone in primary hyperparathyroidism (PHPT). Therefore, we simulated PHPT with 2 wk of continuous PTH infusion in adult mice with deletion of the PTH gene (Pth Ϫ/Ϫ mice) and with deletion of both PTH and CaSR genes (Pth Ϫ/Ϫ -Casr Ϫ/Ϫ mice) and compared skeletal phenotypes. PTH infusion in Pth Ϫ/Ϫ mice increased cortical bone turnover, augmented cortical porosity, and reduced cortical bone volume, femoral bone mineral density (BMD), and bone mineral content (BMC); these effects were markedly attenuated in PTH-infused Pth Ϫ/Ϫ -Casr Ϫ/Ϫ mice. In the absence of CaSR, the PTH-stimulated expression of receptor activator of nuclear factor-B ligand and tartrate-resistant acid phosphatase and PTH-stimulated osteoclastogenesis was also reduced. In trabecular bone, PTH-induced increases in bone turnover, trabecular bone volume, and trabecular number were lower in Pth Ϫ/Ϫ -Casr Ϫ/Ϫ mice than in Pth Ϫ/Ϫ mice. PTH-stimulated genetic markers of osteoblast activity were also lower. These results are consistent with a role for CaSR in modulating both PTH-induced bone resorption and PTH-induced bone formation in discrete skeletal compartments. (40) and their osteoblastic progeny (4, 34), resulting in increased bone turnover in both cortical and trabecular bone and producing either net bone loss or net bone gain. In the fetus (27) and neonate (52), endogenous PTH is required for normal trabecular bone formation, and in both neonates and adult animals, exogenous PTH, when administered intermittently, can have an anabolic effect predominantly by increasing trabecular bone formation, although it may also concomitantly produce a small reduction in cortical bone (9,29,48). Primary hyperparathyroidism (PHPT) and continuous rather than intermittent PTH treatment induce cortical bone loss at least in part by enhancing endosteal bone resorption and intracortical bone resorption (17,25,36). Although severe, persistent elevations of PTH levels may also lead to trabecular bone loss (17), both PHPT and continuous PTH treatment often produce a modest increase in trabecular bone (33,44,54). The catabolic effect of PTH has been shown to be mediated in part by enhanced production of the cytokine receptor activator of nuclear factor-B ligand (RANKL) and by decreased production of the RANKL decoy receptor osteoprotegerin (OPG) by BMSC and osteoblasts (26,49).CaSR has been reported to function in vitro in a variety of skeletal cells, including BMSC, osteoblasts, monocytes/macrophages, and osteoclasts (6). By in situ hybridization, CaSR transcripts have been found mainly in osteoblasts, osteocytes, and bone marrow cells but rarely in mature osteocl...

Bone metastases represent a frequent complication of advanced breast cancer. As tumor growth-induced bone remodeling progresses, episodes of severe pain and fractures of weight-bearing limbs increase. All of these skeletal-related events influence the patient's quality of life and survival. In the present study, we sought to determine whether some of these pain-related behaviors could be directly correlated to tumor progression and bone remodeling. For this purpose, we used a rat model of bone cancer pain based on the implantation of mammary carcinoma cells in the medullary cavity of the femur. The bone content and tumor growth were monitored over time by magnetic resonance imaging (MRI) and micro X-ray computed tomography (μCT). The same animals were evaluated for changes in their reflexive withdrawal responses to mechanical stimuli (allodynia) and weight-bearing deficits. As assessed by MRI, we found a negative correlation between tumor volume and allodynia or postural deficits throughout the experiment. Using μCT, we found that the bone volume/total volume (BV/TV) ratios for trabecular and cortical bone correlated with both mechanical hypersensitivity and weight-bearing impairment. However, whereas trabecular BV/TV stabilized between days 7 and 10 post-tumor detection, the cortical bone loss reached its maximum at that time. Our imaging approach also allowed us to consistently detect the tumor before the onset of pain, paving the way for the preemptive identification of at-risk patients. Altogether, these results improve our understanding of the events leading to tumor-induced bone pain and could eventually help in the design of novel strategies for the management of bone diseases.

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The effects of melatonin implants were investigated on winter fur growth, monitored by counting growing and mature hairs per bundle and testicular recrudescence, judged by testis width, score count of spermatogenesis, and serum testosterone in the adult male raccoon dogs. Melatonin administration in July highly elevated melatonin concentrations in serum and urine and induced an earlier decrease in prolactin secretion (August in the treated group vs September in the control group), winter fur growth (July-beginning of November in the treated group vs. August-end of November in the control group) and testicular recrudescence (October in the treated group vs. November in the control group). In the control animals, urinary excretion of melatonin between 1500-0900 hr increased during autumn followed by a rapid fall in winter. The increase from July (1.8 +/- 0.4 ng) to August (3.9 +/- 0.5 ng) and the subsequent unchanged levels until October coincided with the period of winter fur growth. The further increase in November (6.5 +/- 1.2 ng) coincided with the significant elevation in both testis width and score count of spermatogenesis. These results suggest a role of the increase in endogenous melatonin secretion during autumn in the growth of winter fur and testicular recrudescence in this species under natural conditions. Relatively high serum concentrations of prolactin were shown in two animals, one in the control group and another in the treated group. However, the parameters for testis and winter fur growth in the two cases were similar to those in the remainder of the animals. Thereby, the role of prolactin in the winter fur growth and the initiation of testicular recrudescence, if it is truly involved, is manifested through its decreasing secretion rather than the actual blood concentrations.

Summary Temporary plugging during fracturing operations has become an efficient method to create a complex fracture network in tight reservoirs with natural fractures (NFs). Accurate prediction of the network-propagation process plays a critical role in plugging- and fracturing-parameter optimization. In this paper, the interaction between one single hydraulic fracture (HF) within a temporary plugging segment and multiple NFs was simulated using a complex fracture-development model. A new opening criterion for an NF penetrated by a nonorthogonal HF already was implemented to identify the dominant propagation direction of a HF under plugging conditions. Fracture displacements and the induced-stress field were determined by the 3D displacement-discontinuity method, and the Gauss-Jordan and Levenberg-Marquardt methods were combined to handle the coupling between rock mechanics and fluid flow numerically. Numerical results demonstrate that the opening and development of an NF are mainly dominated by its approaching angle and relative location. For a certain NF crossed by an HF within the plugging segment, the HF tends to propagate along the NF branch inclined in the main HF direction. The farther the interaction position is away from the HF tip, the easier the NF with an approaching angle less than 30° or larger than 150° can be open, and the outcome will be opposite if the approaching angle is larger than 45° or less than 135°. Higher approaching angle and plugging strength is necessary for expanding the position scope of NF that can be opened around HF. Under the effect of plugging, fluid pressure in the HF plummets at the beginning of the NF opening and keeps decreasing until the NF extends for a certain distance or encounters secondary NFs. Fluid-pressure drop occurs mainly in the unturned NF, together with the width of unturned NF being significantly lower than that of the turned NF and HF. Sensitivity analysis shows that the main factors, such as geometry, aperture profile, and fluid-pressure distribution, affecting the network progress under the temporary plugging conditions are the horizontal differential stress, NF position, approaching angle, plugging time, and plugging-segment length. The simulation results provide critical insight into complex fracture-propagation progress under temporary plugging conditions, which should serve as guidelines for well choosing and plugging optimization in temporary plugging fracturing.

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