Cortical Bone Volume Research Articles

D-mannose attenuates bone loss in mice via Treg cell proliferation and gut microbiota-dependent anti-inflammatory effects.

Background:D-mannose exhibits strong anti-inflammatory properties, but whether it has beneficial effects on preventing and treating osteoporosis remains unknown.Methods:Female, 12-month-old senile C57BL6/J mice (s-Man group) and 8-week-old ovariectomized C57BL6/J mice (OVX-Man group) were treated with D-mannose in drinking water for 2 months (six mice/group). Microcomputed tomography analysis and hematoxylin and eosin staining were performed to investigate the effect of D-mannose on attenuation of bone loss. Tartrate-resistant acid phosphatase staining of tissue sections, flow cytometry, enzyme-linked immunosorbent assay, quantitative real-time polymerase chain reaction, and gut microbiome biodiversity tests were used to explore the underlying mechanisms.Results:D-mannose-induced marked increases in cortical bone volume and trabecular bone microarchitecture in the s-Man and OVX-Man group compared with that in the s-CTRL (senile control) and OVX group, respectively. Moreover, D-mannose downregulated osteoclastogenesis-related cytokines in the bone marrow and expanded regulatory T cells in the spleen of mice. Furthermore, D-mannose reconstructed the gut microbiota and changed the metabolite composition.Conclusion:D-mannose attenuated bone loss induced by senility and estrogen deficiency in mice, and this effect may be mediated by D-mannose-induced proliferation of regulatory T cells and gut microbiota-dependent anti-inflammatory effects.

Pedicle Screws Challenged: Lumbar Cortical Density and Thickness Are Greater in the Posterior Elements Than in the Pedicles

Study Design:Controlled laboratory study.Objective:To measure the total bone mineral density (BMD), cortical volume, and cortical thickness in seven different anatomical regions of the lumbar spine.Methods:Using computed tomography (CT) images, 3 cadaveric spines were digitally isolated by applying filters for cortical and cancellous bone. Each spine model was separated into 5 lumbar vertebrae, followed by segmentation of each vertebra into 7 anatomical regions of interest using 3-dimensional software modeling. The average Hounsfield units (HU) was determined for each region and converted to BMD with calibration phantoms of known BMD. These BMD measurements were further analyzed by the total volume, cortical volume, and cancellous volume. The cortical thickness was also measured. A similar analysis was performed by vertebral segment. St Mary’s Medical Center’s Institutional Review Board approved this study. No external funding was received for this work.Results:The lamina and inferior articular process contained the highest total BMD, thickest cortical shell, and largest percent volumes of cortical bone. The vertebral body demonstrated the lowest BMD. The BMDs of the L4 and L5 segments were lower; however, there were no statistically significant differences in BMD between the L1-L5 vertebral segments.Conclusion:Extrapedicular regions of the lumbar vertebrae, including the lamina and inferior articular process, contain denser bone than the pedicles. Since screw pullout strength relies greatly on bone density, the lamina and inferior articular processes may offer stronger fixation of the lumbar spine.

Combination therapy in the Col1a2G610C mouse model of Osteogenesis Imperfecta reveals an additive effect of enhancing LRP5 signaling and inhibiting TGFβ signaling on trabecular bone but not on cortical bone.

Enhancing LRP5 signaling and inhibiting TGFβ signaling have each been reported to increase bone mass and improve bone strength in wild-type mice. Monotherapy targeting LRP5 signaling, or TGFβ signaling, also improved bone properties in mouse models of Osteogenesis Imperfecta (OI). We investigated whether additive or synergistic increases in bone properties would be attained if enhanced LRP5 signaling was combined with TGFβ inhibition. We crossed an Lrp5 high bone mass (HBM) allele (Lrp5A214V) into the Col1a2G610C/+ mouse model of OI. At 6-weeks-of-age we began treating mice with an antibody that inhibits TGFβ1, β2, and β3 (mAb 1D11), or with an isotype-matched control antibody (mAb 13C4). At 12-weeks-old, we observed that combining enhanced LRP5 signaling with inhibited TGFβ signaling produced an additive effect on femoral and vertebral trabecular bone volumes, but not on cortical bone volumes. Although enhanced LRP5 signaling increased femur strength in a 3-point bending assay in Col1a2G610C/+ mice, femur strength did not improve further with TGFβ inhibition. Neither enhanced LRP5 signaling nor TGFβ inhibition, alone or in combination, improved femur 3-point-bending post-yield displacement in Col1a2G610C/+ mice. These pre-clinical studies indicate combination therapies that target LRP5 and TGFβ signaling should increase trabecular bone mass in patients with OI more than targeting either signaling pathway alone. Whether additive increases in trabecular bone mass will occur in, and clinically benefit, patients with OI needs to be determined.

The combined effects of teriparatide and anti-RANKL monoclonal antibody on bone defect regeneration in ovariectomized mice.

The purpose of this study was to investigate the combined effects of teriparatide (TPTD) and anti-murine receptor activator of nuclear factor-κB ligand monoclonal antibody (anti-RANKL Ab) on both cancellous and cortical bone healing in ovariectomized mice. Thirteen-week-old mice were divided into the sham-operated group (n=11) or the ovariectomized group (n=44). At 1 month post-operation, all mice underwent bone defect surgery on the left femoral metaphysis (cancellous bone healing model) and right femoral mid-diaphysis (cortical bone healing model). After creating the bone defects, all ovariectomized mice were assigned to one of four groups to receive 1) saline (5 times a week; CNT group), 2) TPTD (40μg/kg 5 times a week; TPTD group), 3) anti-RANKL Ab (5mg/kg once; Ab group), or 4) a combination of TPTD and anti-RANKL Ab (COMB group). The following analyses were performed: Time-course microstructural analysis of healing in both cancellous and cortical bone in the bone defect, measuring the volumetric bone mineral density and the cortical bone thickness of the tibia as a representative of whole body bone with the use of micro-computed tomography, and histological analysis. Regeneration of cancellous bone volume in the COMB group was the highest among the four groups, and combined treatment accelerated the formation of medullary callus during the early phase of bone regeneration. On the other hand, there were no significant differences in the regeneration of cortical bone volume during the early phase of bone regeneration among the four groups. Furthermore, lamellar bone was not well identified in the all four groups. Volumetric bone mineral density in the tibia in the COMB group was significantly higher compared with that in the CNT and TPTD groups and tended to be higher compared with that in the Ab group. The mean values of cortical bone thickness in the TPTD and COMB groups were significantly higher than that in the CNT group. In a mouse model of postmenopausal osteoporosis, combination therapy of TPTD and anti-RANKL Ab accelerates regeneration of cancellous bone more effectively than either agent alone during the early phase of bone regeneration.

Transcriptomic profiling of the myeloma bone-lining niche reveals BMP signalling inhibition to improve bone disease

Multiple myeloma is an incurable, bone marrow-dwelling malignancy that disrupts bone homeostasis causing skeletal damage and pain. Mechanisms underlying myeloma-induced bone destruction are poorly understood and current therapies do not restore lost bone mass. Using transcriptomic profiling of isolated bone lining cell subtypes from a murine myeloma model, we find that bone morphogenetic protein (BMP) signalling is upregulated in stromal progenitor cells. BMP signalling has not previously been reported to be dysregulated in myeloma bone disease. Inhibition of BMP signalling in vivo using either a small molecule BMP receptor antagonist or a solubilized BMPR1a-FC receptor ligand trap prevents trabecular and cortical bone volume loss caused by myeloma, without increasing tumour burden. BMP inhibition directly reduces osteoclastogenesis, increases osteoblasts and bone formation, and suppresses bone marrow sclerostin levels. In summary we describe a novel role for the BMP pathway in myeloma-induced bone disease that can be therapeutically targeted.

Monosodium Glutamate Supplementation Improves Bone Status in Mice Under Moderate Protein Restriction.

ABSTRACTAdequate protein intake during development is critical to ensure optimal bone gain and to attain a higher peak bone mass later. Using a mild protein restriction model in Balb/C mice consuming 6% of their total energy intake as soy protein (LP‐SOY)—for which we observed a significantly lower femoral cortical thickness, bone volume, trabecular number, and thickness reduction—we evaluated the effects of monosodium glutamate (MSG) supplementation at different concentrations (0.5, 1, 5, 10, and 20 g/kg of diet) on bone characteristics in LP‐SOY‐fed mice. After 6 and 12 weeks, LP‐SOY‐fed mice had lower BMD and reduced body weight related to lower lean mass, which was associated with a reduced IGF‐1 level. The negative effect of the LP‐SOY diet on BMD correlated with impaired bone formation. MSG supplementation, at 5, 10, and 20 g/kg of diet, and PTH injection, used as a positive control, were able to improve BMD and to increase osteoblast activity markers (P1NP and osteocalcin), as well as glutamine plasma concentration. An analysis of bone microarchitecture found that cortical bone was less sensitive to protein restriction than trabecular bone, and that MSG ingestion was able to preserve bone quality through an increase of collagen synthesis, although it did not allow normal bone growth. Our study reinforces the view that glutamate can act as a functional amino acid for bone physiology and support clinical investigation of glutamate supplementation in adults characterized by poor bone status, notably as a result of insufficient protein intake. © 2019 The Authors. JBMR Plus published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research.

Increased anabolic bone response in Dkk1 KO mice following tibial compressive loading.

A viable Dkk1 knockout (KO) mouse strain in which embryonic lethality is rescued by developmental Wnt3 heterozygosity (Dkk1-/-:Wnt3+/-) exhibits increased bone formation and a high bone mass phenotype. We hypothesized that in vivo mechanical loading would further augment the bone formation response in Dkk1 KO mice, comparable to results from Sost KO mice. A cyclic loading protocol was applied to Dkk1 KO mice, wild type mice (WT; Dkk1+/+:Wnt3+/+), and Wnt3 heterozygote (Wnt3+/-; Dkk1+/+:Wnt3+/-) controls. The left tibiae of 10-week-old female mice were dynamically loaded in vivo with 7N maximum compressive force 5 days/week for 2 weeks. Dkk1 KO bones were significantly stiffer, and so an additional group of Dkk1 KO received 12N maximum compressive force to achieve an equivalent +1200με strain at the mid-diaphysis. MicroCT and bone histomorphometry analyses were subsequently performed. All groups responded to tibial loading with increased mid-diaphyseal bone volume. The largest effect size was in the Dkk1 KO -12N group. Thus, Dkk1 KO animals had enhanced sensitivity to mechanical loading. Increases in cortical bone volume reflected increased periosteal bone formation. Bone volume and formation were not altered between WT and Wnt3+/- controls. These data support the concept that agonists of Wnt/β-catenin signaling can act synergistically with load-bearing exercise. Notably, Sost expression decreased with loading in Dkk1 KO and WT mice, independent of genotype. These data suggest that a compensatory downregulation of Sost in Dkk1 KO mice is not likely the primary mechanism for the augmented response to mechanical load.

Jump Exercise and Food Restriction on Bone Parameters in Young Female Rats.

We examined the effect of jump exercise on bone parameters in young female rats under food restriction. Seven-week-old female rats were divided into four groups: a sedentary and ad libitum feeding group (n = 10), a jump exercise and ad libitum feeding group (n = 9), a sedentary and 30% food restriction group (n = 9), and a jump exercise and 30% food restriction group (n = 10). The jump groups jumped 20 times/day, 5 times/week. The experiment lasted for 13weeks. There were no interactions of jump exercise and food restriction on bone. Jump exercise under food restriction conditions induced higher bone strength, bone mineral content, bone area, bone mineral density (BMD), and cortical bone volume in young female rats, similar to rats under ad libitum feeding conditions. Bone strength parameters were not significantly different between ad libitum intake and food restriction with jump exercise training; however, BMD, bone size, and bone mass in the food restriction groups did not reach the levels of those in the ad libitum conditions group with jump exercise training. Neither jump exercise nor food restriction had a significant effect on serum estradiol or IGF-1. Our study reveals jump exercise attenuates loss of biomechanical properties and some bone sites with food restriction in young female rats.

Effects of pentoxifylline and tocopherol on a rat-irradiated jaw model using micro-CT cortical bone analysis.

A combination of pentoxifylline (PTX) and tocopherol (TP) is believed to reduce chronic fibrosis and induce bone healing in osteoradionecrosis (ORN) of the mandible, but evidence of its therapeutic effectiveness for cortical bone is lacking. This study was designed to determine the effect of combined PTX and TP (PTX + TP) on mandibular cortical bone remodeling in a rat model of ORN, using micro-CT and histological analysis. Forty-eight 8-week-old male Sprague-Dawley rats were randomly divided into irradiated (n = 40) and non-irradiated (n = 8) groups. Animals in the irradiated group were divided into four sub-groups, including PTX, TP, PTX + TP, and normal saline. Three weeks after irradiation, mandibular posterior tooth extraction was performed, and animals were sacrificed 7weeks after irradiation. The mandibles were analyzed using micro-CT and histological evaluation. The alveolar bone height, cortical bone thickness, cortical bone volume, and total cortical bone surface of the PTX + TP group were significantly greater than those of other irradiated groups (p < 0.05). In 3D reconstructed images, the residual volumes of cortical and cancellous bone were inadequate in the irradiated groups. We found that a combination of PTX and TP improved quality and quantity of cortical bone in irradiated rat mandibles, thus providing supporting evidence of its utility as a treatment and prophylactic agent in ORN. We observed inadequate volumes of cortical and cancellous bone in ORN mandibles, suggesting that cortical bone could play an important role in further ORN studies.

Congenital lipodystrophy induces severe osteosclerosis.

Berardinelli-Seip congenital generalized lipodystrophy is associated with increased bone mass suggesting that fat tissue regulates the skeleton. Because there is little mechanistic information regarding this issue, we generated "fat-free" (FF) mice completely lacking visible visceral, subcutaneous and brown fat. Due to robust osteoblastic activity, trabecular and cortical bone volume is markedly enhanced in these animals. FF mice, like Berardinelli-Seip patients, are diabetic but normalization of glucose tolerance and significant reduction in circulating insulin fails to alter their skeletal phenotype. Importantly, the skeletal phenotype of FF mice is completely rescued by transplantation of adipocyte precursors or white or brown fat depots, indicating that adipocyte derived products regulate bone mass. Confirming such is the case, transplantation of fat derived from adiponectin and leptin double knockout mice, unlike that obtained from their WT counterparts, fails to normalize FF bone. These observations suggest a paucity of leptin and adiponectin may contribute to the increased bone mass of Berardinelli-Seip patients.

High dose vitamin D supplementation does not rescue bone loss following Roux-en-Y gastric bypass in female rats.

Postoperative bone loss and increased fracture risk associated with Roux-en-Y gastric bypass (RYGB) have been attributed to vitamin D/calcium malabsorption and resultant secondary hyperparathyroidism (HPT). Adequate vitamin D supplementation (VDS), particularly in an older female population, reduces incidence of secondary HPT but the effect on bone loss and fracture risk remains unclear. To investigate whether VDS corrects the RYGB bone phenotype, 41 obese adult female rats were randomized to RYGB with 1000 IU (R1000) or 5000 IU (R5000) vitamin D/kg food or a sham surgical procedure with either paired (PF) or ad libitum (AL) feeding. Bone turnover markers, urinary calcium/creatinine ratio (CCR), and serum calciotropic and gut hormones were assessed throughout a 14-week postoperative period. Femurs were analyzed by micro-computed tomography (μCT), three-point bending test, and histomorphometry. 1000 IU animals had low 25‑hydroxyvitamin D (25(OH)D), high serum parathyroid hormone (PTH), and very low urine CCR levels. 5000 IU corrected the 25(OH)D and secondary HPT but did not increase urine CCR or serum levels of 1,25‑dihydroxyvitamin D (1,25(OH)D) significantly between RYGB groups. Compared to sham animals at 14 weeks, RYGB animals had significantly higher serum osteocalcin (OCN) and C-terminal telopeptide (CTX) levels. The gut hormone peptide tyrosine tyrosine hormone (PYY) was higher in the RYGB groups, and leptin was lower. μCT and biomechanical testing revealed RYGB females had decreased cortical and trabecular bone volume and weaker, stiffer bone than controls. Histomorphometry showed decreased bone volume and increased osteoid volume with increased mineral apposition rate in RYGB compared to controls. No differences in bone phenotype were identified between 1000 IU and 5000 IU groups, and osteoclast numbers were comparable across all four groups. Thus, in our model, 5000 IU VDS corrected vitamin D deficiency and secondary HPT but did not rescue RYGB mineralization rate nor the osteomalacia phenotype. Longer studies in this model are required to evaluate durability of these detrimental effects. Our findings not only underscore the importance of lifelong repletion of both calcium and vitamin D but also suggest that additional factors affect skeletal health in this population.

Cortical bone vessel identification and quantification on contrast-enhanced MR images

Cortical bone porosity is a major determinant of bone strength. Despite the biomechanical importance of cortical bone porosity, the biological drivers of cortical porosity are unknown. The content of cortical pore space can indicate pore expansion mechanisms; both of the primary components of pore space, vessels and adipocytes, have been implicated in pore expansion. Dynamic contrast-enhanced MRI (DCE-MRI) is widely used in vessel detection in cardiovascular studies, but has not been applied to visualize vessels within cortical bone. In this study, we have developed a multimodal DCE-MRI and high resolution peripheral QCT (HR-pQCT) acquisition and image processing pipeline to detect vessel-filled cortical bone pores. For this in vivo human study, 19 volunteers (10 males and 9 females; mean age =63±5) were recruited. Both distal and ultra-distal regions of the non-dominant tibia were imaged by HR-pQCT (82 µm nominal resolution) for bone structure segmentation and by 3T DCE-MRI (Gadavist; 9 min scan time; temporal resolution =30 sec; voxel size 230×230×500 µm3) for vessel visualization. The DCE-MRI was registered to the HR-pQCT volume and the voxels within the MRI cortical bone region were extracted. Features of the DCE data were calculated and voxels were categorized by a 2-stage hierarchical kmeans clustering algorithm to determine which voxels represent vessels. Vessel volume fraction (volume ratio of vessels to cortical bone), vessel density (average vessel count per cortical bone volume), and average vessel volume (mean volume of vessels) were calculated to quantify the status of vessel-filled pores in cortical bone. To examine spatial resolution and perform validation, a virtual phantom with 5 channel sizes and an applied pseudo enhancement curve was processed through the proposed image processing pipeline. Overlap volume ratio and Dice coefficient was calculated to measure the similarity between the detected vessel map and ground truth. In the human study, mean vessel volume fraction was 2.2%±1.0%, mean vessel density was 0.68±0.27 vessel/mm3, and mean average vessel volume was 0.032±0.012 mm3/vessel. Signal intensity for detected vessel voxels increased during the scan, while signal for non-vessel voxels within pores did not enhance. In the validation phantom, channels with diameter 250 µm or greater were detected successfully, with volume ratio equal to 1 and Dice coefficient above 0.6. Both statistics decreased dramatically for channel sizes less than 250 µm. We have a developed a multi-modal image acquisition and processing pipeline that successfully detects vessels within cortical bone pores. The performance of this technique degrades for vessel diameters below the in-plane spatial resolution of the DCE-MRI acquisition. This approach can be applied to investigate the biological systems associated with cortical pore expansion.

SAT-530 Establishment of a Glucocorticoid (GC) Regimen to Induce Both Growth Retardation and Osteoporosis in C57BL/10 Muscular Dystrophy Mouse Models

Glucocorticoids (GCs) are associated with many adverse effects including osteoporosis and growth retardation. These are particular problems in Duchenne muscular dystrophy (DMD) where GCs form the mainstay of treatment. The mechanisms that underlie the undesirable effects of GCs on skeletal development are unclear and there is no proven treatment. We have shown previously (Wood et al. J Endocrinol 2018) that there is no established animal model to investigate GC effects on skeletal development. In this study, we examined this in greater detail and furthermore to mimic the clinical situation in DMD, we also aimed to find a suitable GC regimen to induce both growth retardation and osteoporosis in muscular dystrophy (mdx and mdx:cmah) mice and their C57BL/10 controls (WT). 6-8 male mice in each group were given a GC regimen for 28 days from 4 weeks of age. 5 regimens were trialled; 1) dexamethasone (dex) 2.5mg/kg and 2) dex 5mg/kg via SC injection, 3) soluble prednisolone (pred) 5mg via SC injection, 4) pred 10mg/kg and 5) 20mg/kg in cherry suspension via oral gavage. Regimens 2) and 5) were repeated in mdx and mdx:cmah mice. Control (C) mice received the appropriate vehicle. Bone mass and architecture were assessed by micro-CT and anthropometric measurements were used to assess growth. ELISA was used to analyse serum bone turnover markers and breaking strength was determined by 3-point bending. Regimens 1, 3 and 4 had little effect on any parameter measured. Results shown are for WT mice however the same trends were also seen in mdx and mdx:cmah. In the Pred 20 mice, body weight gain (GC: 12.9% v C: 59.2%, p<0.001), change in crown rump length/day (GC: 0.02cm v C:0.03cm, p=0.05) and growth plate width (GC:122.1 µm v C: 167µm, p=0.07) were all lower compared to control values. Cortical bone fraction (GC; 57.2% v C:63.5%, p<0.001), cortical bone volume (GC:0.48mm3 v C: 0.57 mm3, p<0.01), mean polar moment of inertia (GC:0.12mm4 v C: 0.14mm4, p<0.01) and serum bone turnover markers P1NP (GC: 159.9 pg/ml v C: 367.6 pg/ml, p=0.02) and CTX (GC: 75.7 pg/ml v 180.4 pg/ml, p=0.03) were also lower in the Pred 20 mice. Trabecular bone parameters, bone density and breaking strength were unchanged. Interestingly, trabecular number (GC: 5.67mm-1 v C: 5.51mm-1, p<0.001), was higher and cortical bone fraction (GC:59.4% v C: 65.2%, p<0.001) was lower in dex 5 mice. C57BL/10 mice appear fairly resistant to GC challenge; despite high doses no biomechanical or trabecular architecture changes were noted. Prednisolone 20mg/kg given by oral gavage appears to be the most effective regimen to induce growth retardation and osteoporosis in C57BL10 WT mice and DMD models over a 28-day period.

Compression of morbidity in a progeroid mouse model through the attenuation of myostatin/activin signalling.

BackgroundOne of the principles underpinning our understanding of ageing is that DNA damage induces a stress response that shifts cellular resources from growth towards maintenance. A contrasting and seemingly irreconcilable view is that prompting growth of, for example, skeletal muscle confers systemic benefit.MethodsTo investigate the robustness of these axioms, we induced muscle growth in a murine progeroid model through the use of activin receptor IIB ligand trap that dampens myostatin/activin signalling. Progeric mice were then investigated for neurological and muscle function as well as cellular profiling of the muscle, kidney, liver, and bone.ResultsWe show that muscle of Ercc1 Δ/− progeroid mice undergoes severe wasting (decreases in hind limb muscle mass of 40–60% compared with normal mass), which is largely protected by attenuating myostatin/activin signalling using soluble activin receptor type IIB (sActRIIB) (increase of 30–62% compared with untreated progeric). sActRIIB‐treated progeroid mice maintained muscle activity (distance travel per hour: 5.6 m in untreated mice vs. 13.7 m in treated) and increased specific force (19.3 mN/mg in untreated vs. 24.0 mN/mg in treated). sActRIIb treatment of progeroid mice also improved satellite cell function especially their ability to proliferate on their native substrate (2.5 cells per fibre in untreated progeroids vs. 5.4 in sActRIIB‐treated progeroids after 72 h in culture). Besides direct protective effects on muscle, we show systemic improvements to other organs including the structure and function of the kidneys; there was a major decrease in the protein content in urine (albumin/creatinine of 4.9 sActRIIB treated vs. 15.7 in untreated), which is likely to be a result in the normalization of podocyte foot processes, which constitute the filtration apparatus (glomerular basement membrane thickness reduced from 224 to 177 nm following sActRIIB treatment). Treatment of the progeric mice with the activin ligand trap protected against the development of liver abnormalities including polyploidy (18.3% untreated vs. 8.1% treated) and osteoporosis (trabecular bone volume; 0.30 mm3 in treated progeroid mice vs. 0.14 mm3 in untreated mice, cortical bone volume; 0.30 mm3 in treated progeroid mice vs. 0.22 mm3 in untreated mice). The onset of neurological abnormalities was delayed (by ~5 weeks) and their severity reduced, overall sustaining health without affecting lifespan.ConclusionsThis study questions the notion that tissue growth and maintaining tissue function during ageing are incompatible mechanisms. It highlights the need for future investigations to assess the potential of therapies based on myostatin/activin blockade to compress morbidity and promote healthy ageing.

Combination Therapy With Zoledronic Acid and FES-Row Training Mitigates Bone Loss in Paralyzed Legs: Results of a Randomized Comparative Clinical Trial.

Spinal cord injury (SCI) results in rapid, severe osteoporosis and an increased risk of lower extremity fractures. Despite the medical complications associated with these fractures, there is no standard of care to prevent osteoporotic fractures following SCI. Functional electrical stimulation‐ (FES‐) assisted rowing is a promising intervention to improve bone health in SCI because of its ability to generate a muscular contraction in conjunction with mechanical loading of the lower extremity long bones. Combination therapy consisting of FES‐rowing plus zoledronic acid (ZA) may be a superior treatment via inhibition of bone resorption and stimulation of new bone formation. We studied participants enrolled in a randomized clinical trial comparing FES‐rowing alone with FES‐rowing plus ZA to improve bone health in SCI. Volumetric CT scans at the distal femur and proximal tibial metaphyses were performed. Bone geometric properties (cortical thickness index [CTI], cortical compressive strength index [CSI], buckling ratio [BR], bending strength index) and mineral (cortical bone volume [CBV], cortical bone mineral density, cortical bone mineral content) indices were determined. In models adjusting for baseline values, we found that the CBV (p = 0.05 to 0.006), the CTI (p = 0.009), and the BR (p = 0.001) at both the distal femoral and proximal tibial metaphyses were greater in the ZA plus rowing group compared with the rowing‐only group. Similarly, there was a significant positive association between the total rowing work completed and the BR at the proximal tibia (p = 0.05). A subgroup analysis of the rowing‐only arm showed that gains in the CSI at the tibial metaphysis varied in a dose‐dependent fashion based on the total amount of exercise performed (p = 0.009). These findings demonstrate that the osteogenic response to FES‐rowing is dose‐dependent. Combination therapy with ZA and FES‐row training has therapeutic potential to improve bone quality, and perhaps reduce fracture risk at the most common fracture site following SCI. © 2019 The Authors. JBMR Plus Published by Wiley Periodicals, Inc. on behalf of the American Society for Bone and Mineral Research.

Effect of Direct Electrical Current on Bones Infected with Staphylococcus epidermidis.

ABSTRACTWe are developing electrical approaches to treat biofilm‐associated orthopedic foreign‐body infection. Although we have previously shown that such approaches have antibiofilm activity, the effects on bone have not been assessed. Herein, low‐amperage 200 μA fixed direct current (DC) was compared with no current, in a rat femoral foreign‐body infection model. In the infected group, a platinum implant seeded with S. epidermidis biofilm (105 CFU/cm2), plus 50 μL of a 109 CFU suspension of bacteria, were placed in the femoral medullary cavity of 71 rats. One week later, rats were assigned to one of four groups: infected with no current or DC, or uninfected with no current or DC. After 2 weeks, bones were removed and subjected to histopathology, micro‐computed tomography (μCT), and strength testing. Histopathology showed no inflammation or bony changes/remodeling in the uninfected no current group, and some osteoid formation in the DC group; bones from the infected no current group had evidence of inflammation without bony changes/remodeling; along with inflammation, there was moderate osteoid present in the DC group. μCT showed more cortical bone volume and density, trabecular thickness, and cancellous bone volume in the DC group compared with the no current group, for both uninfected and infected bones (p < 0.05). There was no difference in torsional strength or stiffness between the no current versus DC groups, for both infected and uninfected bones (p > 0.05). © 2018 The Authors. JBMR Plus Published by Wiley Periodicals, Inc. on behalf of the American Society for Bone and Mineral Research.

Vitamin A decreases the anabolic bone response to mechanical loading by suppressing bone formation.

Increased vitamin A consumption is associated with decreased cortical bone mass and increased fracture risk in humans. Rodent studies have demonstrated that hypervitaminosis A increases cortical bone resorption, whereas the importance of the effects on bone formation is less well defined. We used an experimental model of increased bone formation by loading of the tibiae to investigate the effect of vitamin A on bone formation. Control [retinol activity equivalents (RAE) 4.5 µg/g chow] or vitamin A (RAE 60 µg/g chow) diets were given to female C57BL/6N mice for 4 wk, after which the tibiae were subjected to axial loading on alternate days for 2 wk, while the diets were continued. Vitamin A inhibited the loading-induced increase in trabecular and cortical bone volume. This was attributed to inhibition of loading-induced increase in osteoblast number and activity, and expression of osteoblastic genes Sp7, Alpl, and Col1a1 in cortical bone. Vitamin A, loading, and combination thereof also resulted in site-specific effects on bone composition measured by Raman spectroscopy. In summary, a clinically relevant dose of vitamin A suppresses the loading-induced gain of bone mass by decreasing bone formation. These observations may have implications for regulation of bone mass caused by physical activity and the risk of osteoporosis in humans.—Lionikaite, V., Henning, P., Drevinge, C., Shah, F. A., Palmquist, A., Wikström, P., Windahl, S. H., Lerner, U. H. Vitamin A decreases the anabolic bone response to mechanical loading by suppressing bone formation.

A Novel Oral Anti-osteoporosis Drug with Osteogenesis-promoting Effects via Osteoblast Differentiation

Osteoporosis increases the risk of bone fractures (e.g., the femur), reduces a person's activities of daily living (ADL) and increases the likelihood of being bedridden. Therapeutic drugs for osteoporosis include oral bisphosphonates and intravenous receptor activator of nuclear factor-κB ligand (RANKL) antibodies, both of which suppress osteoclast activity, as well as the subcutaneously administered recombinant human parathyroid hormone (PTH), which activates osteoblasts. However, there is currently no oral osteogenesis-promoting drug. In the present study, we found a low-molecular-weight compound, KY-273, with osteogenesis promoting effects. KY-273 induced osteoblast differentiation in ST2 cells and in rat bone marrow-derived mesenchymal stem cells at a dose of 0.1 μM. On the other hand, KY-273 did not clearly exert differentiation effects in osteoclasts, chondrocytes, adipocytes, or myoblasts. In ovariectomized rats, KY-273 clearly increased serum bone alkaline phosphatase (ALP) by at a dose of 3 mg/kg for 8 weeks, and increased both the cortical bone volume and medullary volume of the diaphyseal and epiphyseal regions of femoral bone, but did not affect trabecular bone. Although alendronate (used to decrease bone loss) increased trabecular bone, it did not have any significant effects on cortical bone. PTH increased epiphysis cortical and trabecular bone volume, and reduced medullary volume. KY-273 also displayed good oral absorption in rats. In conclusion, KY-273 is a promising candidate for use as an oral anti-osteoporosis drug with osteogenesis promoting effects.

Effects of pentoxifylline and alendronate on fracture healing in ovariectomy-induced osteoporosis in rats.

Osteoporosis is determined by decreased bone strength that increases the threat of fractures. The aim of this study was to evaluate the effects of pentoxifylline (PTX) and alendronate (ALN), on the stereological parameters, and gene expression in callus of fracture in an experimental rat model of ovariectomy-induced osteoporosis (OVX). The OVX was induced in 90 female rats. Fourteen weeks later, a complete fracture on the right femur was made. Rats were divided into five groups: 1) control: no treatment; 2) sham: received daily distilled water; 3) daily 3.00 mg kg-1 ALN subcutaneously (SC); 4) daily 200 mg kg-1 PTX (SC) and 5) daily PTX (SC) + ALN (same doses). The osteoclast count was significantly lower in all treatment groups, at 21 and 56 days post-surgery, compared to the control and sham groups. The PTX significantly increased total callus volume at 21 and 56 days post-surgery, compared to the other groups. The PTX+ALN treatment significantly increased both cortical bone volume on day 21, and osteocyte and osteoblast numbers on day 56, compared to the control and sham groups. It can be concluded that PTX and ALN have antiresorptive effects, in OVX rats. Also, PTX has increased the extracellular matrix on both 21 and 56 days after surgery, compared to the other groups. PTX+ALN elevated cortical bone volume on day 21, and osteocyte and osteoblast numbers compared to the control and sham groups on day 56.

Lactobacillus rhamnosus GG attenuates tenofovir disoproxil fumarate-induced bone loss in male mice via gut-microbiota-dependent anti-inflammation.

Background:Although antiretroviral agents trigger bone loss in human immunodeficiency virus patients, tenofovir disoproxil fumarate (TDF) induces more severe bone damage, such as osteoporosis. While, the mechanisms are unclear, probiotic supplements may be effective against osteoporosis.Methods:C57BL6/J mice were administered with Lactobacillus rhamnosus GG (LGG)+TDF, TDF, and zoledronic acid+TDF, respectively. Bone morphometry and biomechanics were evaluated using microcomputed tomography, bone slicing, and flexural tests. The lymphocyte, proinflammatory cytokines, and intestinal permeability levels were detected using enzyme-linked immunosorbent assays, quantitative real-time polymerase chain reaction, and flow cytometry. The gut microbiota composition and metabolomics were analyzed using 16S recombinant deoxyribonucleic acid pyrosequencing and ultra-performance liquid-chromatography–quadrupole time-of-flight mass spectrometry.Results:LGG administered orally induced marked increases in trabecular bone microarchitecture, cortical bone volume, and biomechanical properties in the LGG+TDF group compared with that in the TDF-only group. Moreover, LGG treatment increased intestinal barrier integrity, expanded regulatory T cells, decreased Th17 cells, and downregulated osteoclastogenesis-related cytokines in the bone marrow, spleen, and gut. Furthermore, LGG reconstructed the gut microbiota and changed the metabolite composition, especially lysophosphatidylcholine levels. However, the amount of N-acetyl-leukotriene E4 was the highest in the TDF-only group.Conclusion:LGG reconstructed the community structure of the gut microbiota, promoted the expression of lysophosphatidylcholines, and improved intestinal integrity to suppress the TDF-induced inflammatory response, which resulted in attenuation of TDF-induced bone loss in mice. LGG probiotics may be a safe and effective strategy to prevent and treat TDF-induced osteoporosis.