Propranolol Promotes Bone Formation and Limits Resorption Through Novel Mechanisms During Anabolic Parathyroid Hormone Treatment in Female C57BL/6J Mice.

Although the nonselective ß-blocker, propranolol, improves bone density with parathyroid hormone (PTH) treatment in mice, the mechanism of this effect is unclear. To address this, we used a combination of in vitro and in vivo approaches to address how propranolol influences bone remodeling in the context of PTH treatment. In female C57BL/6J mice, intermittent PTH and propranolol administration had complementary effects in the trabecular bone of the distal femur and fifth lumbar vertebra (L5 ), with combination treatment achieving microarchitectural parameters beyond that of PTH alone. Combined treatment improved the serum bone formation marker, procollagen type 1 N propeptide (P1NP), but did not impact other histomorphometric parameters relating to osteoblast function at the L5 . In vitro, propranolol amplified the acute, PTH-induced, intracellular calcium signal in osteoblast-like cells. The most striking finding, however, was suppression of PTH-induced bone resorption. Despite this, PTH-induced receptor activator of nuclear factor κ-B ligand (RANKL) mRNA and protein levels were unaltered by propranolol, which led us to hypothesize that propranolol could act directly on osteoclasts. Using in situ methods, we found Adrb2 expression in osteoclasts in vivo, suggesting ß-blockers may directly impact osteoclasts. Consistent with this, we found propranolol directly suppresses osteoclast differentiation in vitro. Taken together, this work suggests a strong anti-osteoclastic effect of nonselective ß-blockers in vivo, indicating that combining propranolol with PTH could be beneficial to patients with extremely low bone density. © 2022 American Society for Bone and Mineral Research (ASBMR).

Once daily calcium (1000 mg) and vitamin D (1000 IU) supplementation during military training prevents increases in biochemical markers of bone resorption but does not affect tibial microarchitecture in Army recruits.

Basic combat training (BCT) is a period of novel physical training including load carriage resulting in higher risk of stress fracture compared to any other time during military service. Prior trials reported a 20% reduction in stress fracture incidence with Ca and vitamin D (Ca + D) supplementation (2000 mg Ca, 800 IU vitamin D), and greater increases in tibia vBMD during BCT compared to placebo. The primary objective of this randomized, double-blind, placebo-controlled trial was to determine the efficacy of a lower dose of Ca (1000 mg/d Ca, 1000 IU vit D) on PTH, bone biomarkers and tibial microarchitecture during BCT. One hundred volunteers (50 males, 50 females; mean age 21.8 ± 3.5 y) were block randomized by race and sex to receive a daily Ca + D fortified food bar or placebo. Anthropometrics, dietary intake, fasted blood draws and high resolution pQCT scans of the distal and mid-shaft tibia were obtained at the start of BCT and 8 wks later at the conclusion of training. As compliance was 98% in both treatment groups, an intent-to-treat analysis was used. At the distal tibia, total vBMD, Tb.vBMD, Tb.N, Th.Th and Tb.BV/TV increased (+1.07 to 2.12% for all, p < 0.05) and Tb.Sp decreased (0.96 to 1.09%, p < 0.05) in both treatment groups. At the mid-shaft, Ct.Pm increased (+0.18 to 0.21%, p = 0.01) and Ct.vBMD decreased (-0.48 to -0.77%, p < 0.001) in both groups. Ca + D prevented increases in CTX and TRAP, which were observed in the placebo group (group-by-time, p < 0.05). Mean circulating 25OHD, BAP, P1NP and iCa increased and PTH decreased in both treatment groups (p < 0.05). These results, in agreement with other studies, suggest that bone microarchitectural changes indicative of bone formation occur during BCT. While Ca + D supplementation at lower doses than those tested in previous studies prevented increases in biochemical markers of bone resorption in this study, there were no significant changes in bone tissue after 8 wks of Army BCT.

Effects of Combination Denosumab and High-Dose Teriparatide Administration on Bone Microarchitecture and Estimated Strength: The DATA-HD HR-pQCT Study.

In postmenopausal women at high risk of fracture, we previously reported that combined denosumab and high-dose (HD; 40 µg) teriparatide increased spine and hip bone mineral density (BMD) more than combination with standard-dose teriparatide (SD; 20 µg). To assess the effects of these combinations on bone microarchitecture and estimated bone strength, we performed high-resolution peripheral quantitative computed tomography (HR-pQCT) at the distal radius and distal tibia in these women, who were randomized to receive either teriparatide 20 µg (n = 39) or 40 µg (n = 37) during months 0 to 9 overlapped with denosumab 60 mg s.c. given at months 3 and 9, for a 15-month study duration. The 69 women who completed at least one study visit after baseline are included in this analysis. Over 15 months, increases in total BMD were higher in the HD-group than the SD-group at the distal tibia (5.3% versus 3.4%, p = 0.01) with a similar trend at the distal radius (2.6% versus 1.0%, p = 0.06). At 15 months, cortical porosity remained similar to baseline, with absolute differences of -0.1% and -0.7% at the distal tibia and -0.4% and -0.1% at the distal radius in the HD-group and SD-group, respectively; p = NS for all comparisons. Tibial cortical tissue mineral density increased similarly in both treatment groups (1.3% [p < 0.0001 versus baseline] and 1.5% [p < 0.0001 versus baseline] in the HD-group and SD-group, respectively; p = 0.75 for overall group difference). Improvements in trabecular microarchitecture at the distal tibia and estimated strength by micro-finite element analysis at both sites were numerically greater in the HD-group compared with SD-group but not significantly so. Together, these findings suggest that short-term treatment combining denosumab with either high- or standard-dose teriparatide improves HR-pQCT measures of bone density, microstructure, and estimated strength, with greater gains in total bone density observed in the HD-group, which may be of benefit in postmenopausal women with severe osteoporosis. © 2020 American Society for Bone and Mineral Research (ASBMR).

Effects of Estrogen Replacement on Bone Geometry and Microarchitecture in Adolescent and Young Adult Oligoamenorrheic Athletes: A Randomized Trial.

Oligoamenorrheic athletes (OAs) have lower bone mineral density (BMD) and greater impairment of bone microarchitecture, and therefore higher fracture rates compared to eumenorrheic athletes. Although improvements in areal BMD (aBMD; measured by dual-energy X-ray absorptiometry) in OAs have been demonstrated with transdermal estrogen treatment, effects of such treatment on bone microarchitecture are unknown. Here we explore effects of transdermal versus oral estrogen versus no estrogen on bone microarchitecture in OA. Seventy-five OAs (ages 14 to 25 years) were randomized to (i) a 100-µg 17ß-estradiol transdermal patch (PATCH) administered continuously with 200 mg cyclic oral micronized progesterone; (ii) a combined 30 µg ethinyl estradiol and 0.15 mg desogestrel pill (PILL); or (iii) no estrogen/progesterone (NONE) and were followed for 12 months. Calcium (≥1200 mg) and vitamin D (800 IU) supplements were provided to all. Bone microarchitecture was assessed using high-resolution peripheral quantitative CT at the distal tibia and radius at baseline and 1 year. At baseline, randomization groups did not differ by age, body mass index, percent body fat, duration of amenorrhea, vitamin D levels, BMD, or bone microarchitecture measurements. After 1 year of treatment, at the distal tibia there were significantly greater increases in total and trabecular volumetric BMD (vBMD), cortical area and thickness, and trabecular number in the PATCH versus PILL groups. Trabecular area decreased significantly in the PATCH group versus the PILL and NONE groups. Less robust differences between groups were seen at the distal radius, where percent change in cortical area and thickness was significantly greater in the PATCH versus PILL and NONE groups, and changes in cortical vBMD were significantly greater in the PATCH versus PILL groups. In conclusion, in young OAs, bone structural parameters show greater improvement after 1 year of treatment with transdermal 17ß-estradiol versus ethinyl estradiol-containing pills, particularly at the tibia. © 2019 American Society for Bone and Mineral Research.

A prospective field study of U.S. Army trainees to identify the physiological bases and key factors influencing musculoskeletal injuries: a study protocol.

BACKGROUND: Musculoskeletal injuries (MSKIs) are common in military trainees and present a considerable threat to occupational fitness, deployability, and overall military readiness. Despite the negative effects of MSKIs on military readiness, comprehensive evaluations of the key known and possible risk factors for MSKIs are lacking. The U.S. Army Research Institute of Environmental Medicine (ARIEM) is initiating a large-scale research effort, the ARIEM Reduction in Musculoskeletal Injury (ARMI) Study, to better understand the interrelationships among a wide range of potential MSKI risk factors in U.S. Army trainees in order to identify those risk factors that most contribute to MSKI and may be best targeted for effective mitigation strategies. METHODS: This prospective study aims to enroll approximately 4000 (2000 male and 2000 female) U.S. Army trainees undergoing Basic Combat Training (BCT). Comprehensive in-person assessments will be completed at both the beginning and end of BCT. Participants will be asked to complete surveys of personal background information, medical history, physical activity, sleep behaviors, and personality traits. Physical measurements will be performed to assess anthropometrics, tibial microarchitecture and whole body bone mineral density, muscle cross-sectional area, body composition, and muscle function. Blood sampling will be also be conducted to assess musculoskeletal, genetic, and nutritional biomarkers of risk. In addition, participants will complete weekly surveys during BCT that examine MSKI events, lost training time, and discrete risk factors for injury. Participants' medical records will be tracked for the 2 years following graduation from training to identify MSKI events and related information. Research hypotheses focus on the development of a multivariate prediction model for MSKI. DISCUSSION: Results from this study are expected to inform current understanding of known and potential risk factors for MSKIs that can be incorporated into solutions that optimize Soldier health and enhance military readiness.

In vitro injection of osteoporotic cadaveric femurs with a triphasic calcium-based implant confers immediate biomechanical integrity.

Current pharmaceutical therapies can reduce hip fractures by up to 50%, but compliance to treatment is low and therapies take up to 18 months to reduce risk. Thus, alternative or complementary approaches to reduce the risk of hip fracture are needed. The AGN1 local osteo-enhancement procedure (LOEP) is one such alternative approach, as it is designed to locally replace bone lost due to osteoporosis and provide immediate biomechanical benefit. This in vitro study evaluated the initial biomechanical impact of this treatment on human cadaveric femurs. We obtained 45 pairs of cadaveric femurs from women aged 77.8 ± 8.8 years. One femur of each pair was treated, while the contralateral femur served as an untreated control. Treatment included debridement, irrigation/suction, and injection of a triphasic calcium-based implant (AGN1). Mechanical testing of the femora was performed in a sideways fall configuration 24 h after treatment. Of the 45 pairs, 4 had normal, 16 osteopenic, and 25 osteoporotic BMD T-scores. Altogether, treatment increased failure load on average by 20.5% (p < 0.0001). In the subset of osteoporotic femurs, treatment increased failure load by 26% and work to failure by 45% (p < 0.01 for both). Treatment did not significantly affect stiffness in any group. These findings provide evidence that local delivery of the triphasic calcium-based implant in the proximal femur is technically feasible and provides immediate biomechanical benefit. Our results provide strong rationale for additional studies investigating the utility of this approach for reducing the risk of hip fracture. © 2019 The Authors. Journal of Orthopaedic Research® Published by Wiley Periodicals, Inc. on behalf of Orthopaedic Research Society.

Oestrogen replacement improves bone mineral density in oligo-amenorrhoeic athletes: a randomised clinical trial.

OBJECTIVE: Normal-weight oligo-amenorrhoeic athletes (OAA) are at risk for low bone mineral density (BMD). Data are lacking regarding the impact of oestrogen administration on bone outcomes in OAA. Our objective was to determine the effects of transdermal versus oral oestrogen administration on bone in OAA engaged in weight-bearing activity. METHODS: 121 patients with OAA aged 14-25 years were randomised to receive: (1) a 17ß-estradiol transdermal patch continuously with cyclic oral micronised progesterone (PATCH), (2) a combined ethinyl estradiol and desogestrel pill (PILL) or (3) no oestrogen/progesterone (NONE). All participants received calcium and vitamin D supplementation. Areal BMD was assessed at the lumbar spine, femoral neck, total hip and total body less head using dual-energy X-ray absorptiometry at baseline, 6 and 12 months. Intention-to-treat (ITT) and completers analyses were performed. RESULTS: Randomised groups did not differ for age, body mass index or BMD Z-scores at baseline. For ITT analysis, spine and femoral neck BMD Z-scores significantly increased in the PATCH versus PILL (p=0.011 and p=0.021, respectively) and NONE (p=0.021 and p=0.033, respectively) groups, and hip BMD Z-scores significantly increased in the PATCH versus PILL group (p=0.018). Similar findings were noted in completers analysis. CONCLUSION: Transdermal estradiol over 12 months improves BMD in young OAA, particularly compared with an ethinyl estradiol-containing contraceptive pill/oral contraceptives. TRIAL REGISTRATION NUMBER: NCT00946192; Pre-results.

1,25-Dihydroxyvitamin D Alone Improves Skeletal Growth, Microarchitecture, and Strength in a Murine Model of XLH, Despite Enhanced FGF23 Expression.

X-linked hypophosphatemia (XLH) is characterized by impaired renal tubular reabsorption of phosphate owing to increased circulating FGF23 levels, resulting in rickets in growing children and impaired bone mineralization. Increased FGF23 decreases renal brush border membrane sodium-dependent phosphate transporter IIa (Npt2a) causing renal phosphate wasting, impairs 1-α hydroxylation of 25-hydroxyvitamin D, and induces the vitamin D 24-hydroxylase, leading to inappropriately low circulating levels of 1,25-dihydroxyvitamin D (1,25D). The goal of therapy is prevention of rickets and improvement of growth in children by phosphate and 1,25D supplementation. However, this therapy is often complicated by hypercalcemia and nephrocalcinosis and does not always prevent hyperparathyroidism. To determine if 1,25D or blocking FGF23 action can improve the skeletal phenotype without phosphate supplementation, mice with XLH (Hyp) were treated with daily 1,25D repletion, FGF23 antibodies (FGF23Ab), or biweekly high-dose 1,25D from d2 to d75 without supplemental phosphate. All treatments maintained normocalcemia, increased serum phosphate, and normalized parathyroid hormone levels. They also prevented the loss of Npt2a, α-Klotho, and pERK1/2 immunoreactivity observed in the kidneys of untreated Hyp mice. Daily treatment with 1,25D decreased urine phosphate losses despite a marked increase in bone FGF23 mRNA and in circulating FGF23 levels. Daily 1,25D was more effective than other treatments in normalizing the growth plate and metaphyseal organization. In addition to being the only therapy that normalized lumbar vertebral height and body weight, daily 1,25D therapy normalized bone geometry and was more effective than FGF23Ab in improving trabecular bone structure. Daily 1,25D and FGF23Ab improved cortical microarchitecture and whole-bone biomechanical properties more so than biweekly 1,25D. Thus, monotherapy with 1,25D improves growth, skeletal microarchitecture, and bone strength in the absence of phosphate supplementation despite enhancing FGF23 expression, demonstrating that 1,25D has direct beneficial effects on the skeleton in XLH, independent of its role in phosphate homeostasis. © 2016 American Society for Bone and Mineral Research.

Nocturnal oxytocin secretion is lower in amenorrheic athletes than nonathletes and associated with bone microarchitecture and finite element analysis parameters.

OBJECTIVE: Preclinical data indicate that oxytocin, a hormone produced in the hypothalamus and secreted into the peripheral circulation, is anabolic to bone. Oxytocin knockout mice have severe osteoporosis, and administration of oxytocin improves bone microarchitecture in these mice. Data suggest that exercise may modify oxytocin secretion, but this has not been studied in athletes in relation to bone. We therefore investigated oxytocin secretion and its association with bone microarchitecture and strength in young female athletes. DESIGN: Cross-sectional study of 45 females, 14-21 years (15 amenorrheic athletes (AA), 15 eumenorrheic athletes (EA), and 15 nonathletes (NA)), of comparable bone age and BMI. METHODS: We used high-resolution peripheral quantitative CT to assess bone microarchitecture and finite element analysis to estimate bone strength at the weight-bearing distal tibia and non-weight-bearing ultradistal radius. Serum samples were obtained every 60  min, 2300-0700  h, and pooled for an integrated measure of nocturnal oxytocin secretion. Midnight and 0700  h samples were used to assess diurnal variation of oxytocin. RESULTS: Nocturnal oxytocin levels were lower in AA and EA than in NA. After controlling for estradiol, the difference in nocturnal oxytocin between AA and NA remained significant. Midnight and 0700  h oxytocin levels did not differ between groups. At the tibia and radius, AA had impaired microarchitecture compared with NA. In AA, nocturnal oxytocin correlated strongly with trabecular and cortical microarchitecture, particularly at the non-weight-bearing radius. In regression models that include known predictors of microarchitecture in AA, oxytocin accounted for a substantial portion of the variability in microarchitectural and strength parameters. CONCLUSIONS: Nocturnal oxytocin secretion is low in AA compared with NA and associated with site-dependent microarchitectural parameters. Oxytocin may contribute to hypoestrogenemic bone loss in AA.

Insights from the conduct of a device trial in older persons: low magnitude mechanical stimulation for musculoskeletal health.

BACKGROUND: Osteoporosis is a common complication of aging. Alternatives to pharmacologic treatment are needed for older adults. Nonpharmacologic treatment with low magnitude, high frequency mechanical stimulation has been shown to prevent bone loss in animal and human studies. METHODS: The VIBES (Vibration to Improve Bone Density in Elderly Subjects) study is a randomized, double-blind, sham-controlled trial of the efficacy of low magnitude, high frequency mechanical stimulation in 200 men and women aged 60 years and older with bone mineral density T-scores by dual X-ray absorptiometry between -1 and -2.5 at entry. Participants are healthy, cognitively intact residents of independent living communities in the Boston area who receive free calcium and Vitamin D supplements. They are randomly assigned to active or sham treatment and stand on their assigned platform once daily for 10 min. All platforms have adherence data collection software downloadable to a laptop computer. Adverse events are closely monitored. 174 participants were randomized and will be followed for 2 years. Almost all active subjects have attained 1 year of follow-up. Bone mineral density is measured by both dual X-ray absorptiometry and quantitative computed tomography at baseline and annually. The main analysis will compare mean changes from baseline in volumetric bone density by quantitative computed tomography in active and sham groups. Adherence and treatment effect magnitude will also be evaluated. Secondary analyses will compare changes in two biochemical markers of bone turnover as well as longitudinal comparisons of muscle and balance endpoints. RESULTS: The VIBES trial has completed its first year of data collection and encountered multiple challenges leading to valuable lessons learned about the areas of recruitment from independent living communities, deployment of multiuser mechanical devices using radio frequency identification cards and electronic adherence monitoring, organization of transportation for imaging at a central site, and the expansion of study aims to include additional musculoskeletal outcomes. CONCLUSIONS: These lessons will guide future investigations in studies of individuals of advanced age.

High-dose zoledronic acid impacts bone remodeling with effects on osteoblastic lineage and bone mechanical properties.

PURPOSE: The increasing incidence of osteonecrosis of the jaw and its possible association with high cumulative doses of bisphosphonate led us to study the effects of high doses of zoledronic acid (ZA) on bone remodeling. EXPERIMENTAL DESIGN: Five-week-old C57BL6 mice were treated with saline or ZA weekly for 3 weeks at increasing doses (0.05-1 mg/Kg). Effects of ZA on bone remodeling were studied using standard assays. RESULTS: We observed an increase in bone mineral density and content in treated animals at doses of 0.05 mg/Kg, which was not further enhanced at higher doses of ZA. Trabecular bone volume at the proximal tibia and the distal femur assessed by histomorphometry and microCT, respectively, increased significantly in ZA-treated groups. There was however no difference between 0.5 and 1 mg/kg, suggesting a ceiling effect for ZA. ZA led to decreased numbers of osteoclasts and osteoblasts per bone perimeter that paralleled a significant reduction of serum levels of TRAC5b and osteocalcin in vivo. Effects on osteoblasts were confirmed in in vitro assays. Mechanical testing of the femur showed increased brittleness in ZA-treated mice. CONCLUSIONS: High doses of ZA inhibit both osteoclast and osteoblasts function and bone remodeling in vivo interfering with bone mechanical properties. No dose response was noted beyond 0.5 mg/kg suggesting that lower doses of ZA may be adequate in inhibiting bone resorption. Our data may help inform future studies of ZA use with respect to alternate and lower doses in the treatment of patients with cancer bone disease.

Healing of segmental bone defects by direct percutaneous gene delivery: effect of vector dose.

Previous studies have demonstrated the ability of direct adenoviral BMP-2 (Ad.BMP-2) gene delivery to enhance bone repair. Nevertheless, in studies using a rat segmental defect model, it has not proved possible to achieve reliably full osseous union in all animals. To address this issue, we evaluated the effect of vector dose on healing. Critical-size defects were created in the right femora of 27 Sprague-Dawley rats. The defects received a single, intralesional, percutaneous injection of 2.7 x 10(7) (low dose), 2.7 x 10(8) (medium dose), or 2.7 x 10(9) (high dose) plaque-forming units of Ad.BMP-2. After 8 weeks, femora were evaluated by X-ray, dual-energy X-ray absorptiometry, microcomputed tomography (microCT), and histology. The high dose of vector bridged 100%, the medium dose 11%, and the low dose 25% of the defects, as evaluated by X-ray and microCT imaging. Bone mineral content and bone volume of the defects receiving the high dose of vector were significantly higher than those of both groups receiving lower doses. Histologically, defects treated with the high dose were filled by trabecular bone and small amounts of cartilage, whereas large areas of fibrous tissue and cartilage remained in the defects receiving lower doses. However, the newly formed bone lacked the structural organization of native bone, suggesting that further maturation is necessary.

Impact of seafood and fruit consumption on bone mineral density.

OBJECTIVES: Over the past decade, dietary choices and nutrition have proven to be major modulators of bone mineral density (BMD) in men and women. We investigated environmental determinants, specifically dietary habits, of BMD by using multiple regression models in a rural Chinese population. METHODS: BMDs were measured at the hip and total body in 5848 men and 6207 women, aged 25-64. Dietary and supplemental intakes were assessed by a simple, one-page questionnaire tailored to collect nutritional information from large rural populations. Another questionnaire was used to collect information on the subjects' age, disease history, smoking, alcohol consumption, physical activity as well as women's menstrual status and reproductive history. Multiple regression models were used to assess the relationships among dietary variables and BMD, after adjusting for age, BMI (body mass index), weight, occupation, smoking status, and alcohol consumption. RESULTS: Increasing seafood consumption was significantly associated with greater BMD in women (p<0.001), especially those consuming more than 250 g per week of seafood. One thousand and three hundred and twenty-four men and 1479 women consumed >250 g of fruit per week. Higher fruit intake was found to be significantly associated with higher BMD in both sexes (p<0.05). High vegetable consumption, however, did not positively impact BMD. CONCLUSIONS: This study with its large population size has identified preventive measures, as well as some risk factors, involved in bone loss and osteoporosis. Our results highlight the importance of several dietary variables as significant determinants of BMD. It also emphasizes the role of dietary intake in general and shows that specific foods, such as fruits and seafood, can positively impact BMD.