Bone resorption starts at 14 days of treatment with gonadotropin-releasing hormone agonist in in vitro fertilization cycles

The effect of gonadotropin-releasing hormone agonist (GnRH-a) use on bone turnover was investigated in a prospective cohort study of female patients undergoing in vitro fertilization (IVF) treatment. In 46 couples diagnosed with male-factor infertility, the women underwent a long step-down ovulation induction protocol. Urinary cross-linked N-telopeptide (uNTx) level was used to demonstrate bone turnover rate and was measured at the first day of GnRH-a administration, the first day of gonadotropin administration, the day after human chorionic gonadotropin injection and 12 days after embryo transfer. Urinary NTx levels (mean?±?standard deviation (SD)) were 71?±?34, 81?±?40, 81?±?50 and 83?±?47?nmol BCE/mmol creatinine (BCE, bone collagen equivalents), respectively. There was no statistically significant difference between the four measurements (p?=?0.28). In 19 women GnRH-a was administered for ??14 days. Urinary NTx values of this group and the remaining 27 patients after GnRH-a treatment were 96.2?±?40.7 and 71.5?±?36.8?nmol BCE/mmol creatinine (mean?±?SD), respectively. The difference between these groups was statistically significant (p?=?0.038). These findings suggest that <?14 days' use of GnRH agonist in IVF patients has no effect on bone metabolism. To our knowledge, this is the first study demonstrating that the effect of agonists on bone metabolism starts as soon as estradiol suppression has started.     

An essential role for the association of CD47 to SHPS‐1 in skeletal remodeling

Integrin‐associated protein (IAP/CD47) has been implicated in macrophage‐macrophage fusion. To understand the actions of CD47 on skeletal remodeling, we compared Cd47^?/? mice with Cd47^+/+ controls. Cd47^?/? mice weighed less and had decreased areal bone mineral density compared with controls. Cd47^?/? femurs were shorter in length with thinner cortices and exhibited lower trabecular bone volume owing to decreased trabecular number and thickness. Histomorphometry revealed reduced bone‐formation and mineral apposition rates, accompanied by decreased osteoblast numbers. No differences in osteoclast number were observed despite a nonsignificant but 40% decrease in eroded surface/bone surface in Cd47^?/? mice. In vitro, the number of functional osteoclasts formed by differentiating Cd47^?/? bone marrow cells was significantly decreased compared with wild‐type cultures and was associated with a decrease in bone‐resorption capacity. Furthermore, by disrupting the CD47–SHPS‐1 association, we found that osteoclastogenesis was markedly impaired. Assays for markers of osteoclast maturation suggested that the defect was at the point of fusion and not differentiation and was associated with a lack of SHPS‐1 phosphorylation, SHP‐1 phosphatase recruitment, and subsequent dephosphorylation of non–muscle cell myosin IIA. We also demonstrated a significant decrease in osteoblastogenesis in bone marrow stromal cells derived from Cd47^?/? mice. Our finding of cell‐autonomous defects in Cd47^?/? osteoblast and osteoclast differentiation coupled with the pronounced skeletal phenotype of Cd47^?/? mice support the conclusion that CD47 plays an important role in regulating skeletal acquisition and maintenance through its actions on both bone formation and bone resorption. © 2011 American Society for Bone and Mineral Research     

Sodium/myo‐inositol cotransporter 1 and myo‐inositol are essential for osteogenesis and bone formation

myo‐Inositol (MI) plays an essential role in several important processes of cell physiology, is involved in the neural system, and provides an effective treatment for some psychiatric disorders. Its role in osteogenesis and bone formation nonetheless is unclear. Sodium/MI cotransporter 1 (SMIT1, the major cotransporter of MI) knockout (SMIT1^?/?) mice with markedly reduced tissue MI levels were used to characterize the essential roles of MI and SMIT1 in osteogenesis. SMIT1^?/? embryos had a dramatic delay in prenatal mineralization and died soon after birth owing to respiratory failure, but this could be rescued by maternal MI supplementation. The rescued SMIT1^?/? mice had shorter limbs, decreased bone density, and abnormal bone architecture in adulthood. Deletion of SMIT1 resulted in retarded postnatal osteoblastic differentiation and bone formation in vivo and in vitro. Continuous MI supplementation partially restored the abnormal bone phenotypes in adult SMIT1^?/? mice and strengthened bone structure in SMIT1^+/+ mice. Although MI content was much lower in SMIT1^?/? mesenchymal cells (MSCs), the I(1,4,5)P₃ signaling pathway was excluded as the means by which SMIT1 and MI affected osteogenesis. PCR expression array revealed Fgf4, leptin, Sele, Selp, and Nos2 as novel target genes of SMIT1 and MI. SMIT1 was constitutively expressed in multipotential C3H10T1/2 and preosteoblastic MC3T3‐E1 cells and could be upregulated during bone morphogenetic protein 2 (BMP‐2)–induced osteogenesis. Collectively, this study demonstrated that deficiency in SMIT1 and MI has a detrimental impact on prenatal skeletal development and postnatal bone remodeling and confirmed their essential roles in osteogenesis, bone formation, and bone mineral density (BMD) determination. © 2011 American Society for Bone and Mineral Research.     

Effects of bazedoxifene on bone mineral density,bone turnover,and safety in postmenopausal japanese women with osteoporosis

This randomized, double‐blind, placebo‐controlled, dose‐response late phase 2 study evaluated the efficacy and safety of bazedoxifene in postmenopausal Japanese women 85 years of age or younger with osteoporosis. Eligible subjects received daily treatment with oral doses of bazedoxifene 20 or 40 mg or placebo for 2 years. Efficacy assessments included bone mineral density (BMD) at the lumbar spine and other skeletal sites, bone turnover marker levels, lipid parameters, and incidence of new fractures. Of 429 randomized subjects, 387 were evaluable for efficacy, and 423 were included in the safety analyses (mean age, 64 years). At 2 years, the mean percent changes from baseline in lumbar spine BMD were significantly greater with bazedoxifene 20 and 40 mg (2.43% and 2.74%, respectively) than with placebo (?0.65%, p < .001 for both). Both bazedoxifene doses significantly improved BMD at the total hip, femoral neck, and greater trochanter compared with placebo (p < .001 for all). Decreases in bone turnover markers were observed with bazedoxifene 20 and 40 mg as early as 12 weeks (p < .05 for all) and were sustained throughout the study. Total and low‐density lipoprotein cholesterol levels were significantly decreased from baseline with both bazedoxifene doses compared with placebo (p < .05 for all). Incidences of new vertebral and nonvertebral fractures were similar among the bazedoxifene and placebo groups. Overall, the incidence of adverse events with bazedoxifene 20 and 40 mg was similar to that with placebo. Bazedoxifene significantly improved BMD, reduced bone turnover, and was well tolerated in postmenopausal Japanese women with osteoporosis. © 2011 American Society for Bone and Mineral Research.     

Bisphosphonate Withdrawal: Effects on Bone Formation and Bone Resorption in Maturing Male Mice

Bisphosphonates are being increasingly used to treat pediatric patients with skeletal disorders. However, the effects of long‐term bisphosphonate therapy and cessation of therapy during growth are unclear. Thus, studies were undertaken to determine the effects of alendronate discontinuation after treatment of C57Bl/6 mice during the period of rapid skeletal growth. Compared with vehicle‐treated mice, 16 weeks of alendronate treatment starting at age 18 days resulted in a 3.7‐fold increase in trabecular bone in the setting of suppressed bone formation. Alendronate therapy for 8 weeks followed by 8 weeks of vehicle treatment resulted in a more pronounced increase in trabecular bone compared with mice treated with alendronate for 16 weeks (1.7‐fold) and to vehicle‐treated controls (6.5‐fold). Mice that received alendronate for 8 weeks followed by 8 weeks of vehicle exhibited increased osteoblast surface (2.5‐fold), mineralizing surface (5.7‐fold), and bone formation rate (5.1‐fold) compared with mice treated continuously with alendronate. However, these parameters were not restored to the levels observed in the vehicle‐treated mice. Thus, partial resumption of bone formation upon cessation of bisphosphonate therapy leads to a greater increase in trabecular bone than that found when bisphosphonates are administered continuously to growing mice. These data suggest that intermittent administration of bisphosphonates may optimize their beneficial effects on the growing skeleton. © 2017 American Society for Bone and Mineral Research.     

Understanding Bone Strength Is Not Enough

Increases in fracture risk beyond what are expected from bone mineral density (BMD) are often attributed to poor “bone quality,” such as impaired bone tissue strength. Recent studies, however, have highlighted the importance of tissue material properties other than strength, such as fracture toughness. Here we review the concepts behind failure properties other than strength and the physical mechanisms through which they cause mechanical failure: strength describes failure from a single overload; fracture toughness describes failure from a modest load combined with a preexisting flaw or damage; and fatigue strength describes failure from thousands to millions of cycles of small loads. In bone, these distinct failure mechanisms appear to be more common in some clinical fractures than others. For example, wrist fractures are usually the result of a single overload, the failure mechanism dominated by bone strength, whereas spinal fractures are rarely the result of a single overload, implicating multiple loading cycles and increased importance of fatigue strength. The combination of tissue material properties and failure mechanisms that lead to fracture represent distinct mechanistic pathways, analogous to molecular pathways used to describe cell signaling. Understanding these distinct mechanistic pathways is necessary because some characteristics of bone tissue can increase fracture risk by impairing fracture toughness or fatigue strength without impairing bone tissue strength. Additionally, mechanistic pathways to failure associated with fracture toughness and fatigue involve multiple loading events over time, raising the possibility that a developing fracture could be detected and interrupted before overt failure of a bone. Over the past two decades there have been substantial advancements in fracture prevention by understanding bone strength and fractures caused by a single load, but if we are to improve fracture risk prevention beyond what is possible now, we must consider material properties other than strength. © 2017 American Society for Bone and Mineral Research.     

Proximal Femur Volumetric Bone Mineral Density and Mortality: 13 Years of Follow‐Up of the AGES‐Reykjavik Study

Bone mineral density (BMD) has been linked to mortality, but little is known about the independent contribution of each endosteal bone compartment and also the rate of bone loss to risk of mortality. We examined the relationships between (1) baseline trabecular and cortical volumetric BMD (vBMD) at the proximal femur, and (2) the rate of trabecular and cortical bone loss and all‐cause mortality in older adults from the AGES‐Reykjavik study. The analysis of trabecular and cortical vBMD and mortality was based on the baseline cohort of 4654 participants (aged ≥66 years) with a median follow‐up of 9.4 years; the association between rate of bone loss and mortality was based on 2653 participants with bone loss data (median follow‐up of 5.6 years). Analyses employed multivariable Cox‐proportional models to estimate hazard ratios (HRs) with time‐varying fracture status; trabecular and cortical variables were included together in all models. Adjusted for important confounders, Cox models showed that participants in the lowest quartile of trabecular vBMD had an increased risk of mortality compared to participants in other quartiles (HR = 1.12; 95% confidence interval (CI), 1.01 to 1.25); baseline cortical vBMD was not related to mortality (HR = 1.08; 95% CI, 0.97 to 1.20). After adjustment for time‐dependent fracture status, results were attenuated and not statistically significant. A faster loss (quartile 1 versus quartiles 2–4) in both trabecular and cortical bone was associated with higher mortality risk (HR = 1.37 and 1.33, respectively); these associations were independent of major potential confounders including time‐dependent incident fractures (HR = 1.32 and 1.34, respectively). Overall, data suggest that faster bone losses over time in both the trabecular and cortical bone compartments are associated with mortality risk and that measurements of change in bone health may be more informative than single‐point measurements in explaining mortality differences in older adults. © 2017 American Society for Bone and Mineral Research.     

Alterations to the Gut Microbiome Impair Bone Strength and Tissue Material Properties

Alterations in the gut microbiome have been associated with changes in bone mass and microstructure, but the effects of the microbiome on bone biomechanical properties are not known. Here we examined bone strength under two conditions of altered microbiota: (1) an inbred mouse strain known to develop an altered gut microbiome due to deficits in the immune system (the Toll‐like receptor 5–deficient mouse [TLR5KO]); and (2) disruption of the gut microbiota (ΔMicrobiota) through chronic treatment with selected antibiotics (ampicillin and neomycin). The bone phenotypes of TLR5KO and WT (C57Bl/6) mice were examined after disruption of the microbiota from 4 weeks to 16 weeks of age as well as without treatment (n = 7 to 16/group, 39 animals total). Femur bending strength was less in ΔMicrobiota mice than in untreated animals and the reduction in strength was not fully explained by differences in bone cross‐sectional geometry, implicating impaired bone tissue material properties. Small differences in whole‐bone bending strength were observed between WT and TLR5KO mice after accounting for differences in bone morphology. No differences in trabecular bone volume fraction were associated with genotype or disruption of gut microbiota. Treatment altered the gut microbiota by depleting organisms from the phyla Bacteroidetes and enriching for Proteobacteria, as determined from sequencing of fecal 16S rRNA genes. Differences in splenic immune cell populations were also observed; B and T cell populations were depleted in TLR5KO mice and in ΔMicrobiota mice (p < 0.001), suggesting an association between alterations in bone tissue material properties and immune cell populations. We conclude that alterations in the gut microbiota for extended periods during growth may lead to impaired whole‐bone mechanical properties in ways that are not explained by bone geometry. © 2017 American Society for Bone and Mineral Research.     

Integrin‐Linked Kinase Regulates Bone Formation by Controlling Cytoskeletal Organization and Modulating BMP and Wnt Signaling in Osteoprogenitors

Cell‐matrix interactions constitute a fundamental aspect of skeletal cell biology and play essential roles in bone homeostasis. These interactions are primarily mediated by transmembrane integrin receptors, which mediate cell adhesion and transduce signals from the extracellular matrix to intracellular responses via various downstream effectors, including integrin‐linked kinase (ILK). ILK functions as adaptor protein at focal adhesion sites, linking integrins to the actin cytoskeleton, and has been reported to act as a kinase phosphorylating signaling molecules such as GSK‐3β and Akt. Thereby, ILK plays important roles in cellular attachment, motility, proliferation and survival. To assess the in vivo role of ILK signaling in osteoprogenitors and the osteoblast lineage cells descending thereof, we generated conditional knockout mice using the Osx‐Cre:GFP driver strain. Mice lacking functional ILK in osterix‐expressing cells and their derivatives showed no apparent developmental or growth phenotype, but by 5 weeks of age they displayed a significantly reduced trabecular bone mass, which persisted into adulthood in male mice. Histomorphometry and serum analysis indicated no alterations in osteoclast formation and activity, but provided evidence that osteoblast function was impaired, resulting in reduced bone mineralization and increased accumulation of unmineralized osteoid. In vitro analyses further substantiated that absence of ILK in osteogenic cells was associated with compromised collagen matrix production and mineralization. Mechanistically, we found evidence for both impaired cytoskeletal functioning and reduced signal transduction in osteoblasts lacking ILK. Indeed, loss of ILK in primary osteogenic cells impaired F‐actin organization, cellular adhesion, spreading, and migration, indicative of defective coupling of cell‐matrix interactions to the cytoskeleton. In addition, BMP/Smad and Wnt/β‐catenin signaling was reduced in the absence of ILK. Taken together, these data demonstrate the importance of integrin‐mediated cell‐matrix interactions and ILK signaling in osteoprogenitors in the control of osteoblast functioning during juvenile bone mass acquisition and adult bone remodeling and homeostasis. © 2017 American Society for Bone and Mineral Research.     

Sclerostin antibody improves skeletal parameters in a Brtl/+ mouse model of osteogenesis imperfecta

Osteogenesis imperfecta (OI) is a genetic bone dysplasia characterized by osteopenia and easy susceptibility to fracture. Symptoms are most prominent during childhood. Although antiresorptive bisphosphonates have been widely used to treat pediatric OI, controlled trials show improved vertebral parameters but equivocal effects on long‐bone fracture rates. New treatments for OI are needed to increase bone mass throughout the skeleton. Sclerostin antibody (Scl‐Ab) therapy is potently anabolic in the skeleton by stimulating osteoblasts via the canonical wnt signaling pathway, and may be beneficial for treating OI. In this study, Scl‐Ab therapy was investigated in mice heterozygous for a typical OI‐causing Gly→Cys substitution in col1a1. Two weeks of Scl‐Ab successfully stimulated osteoblast bone formation in a knock‐in model for moderately severe OI (Brtl/+) and in WT mice, leading to improved bone mass and reduced long‐bone fragility. Image‐guided nanoindentation revealed no alteration in local tissue mineralization dynamics with Scl‐Ab. These results contrast with previous findings of antiresorptive efficacy in OI both in mechanism and potency of effects on fragility. In conclusion, short‐term Scl‐Ab was successfully anabolic in osteoblasts harboring a typical OI‐causing collagen mutation and represents a potential new therapy to improve bone mass and reduce fractures in pediatric OI. © 2013 American Society for Bone and Mineral Research