Rapidly growing Brtl/+ mouse model of osteogenesis imperfecta improves bone mass and strength with sclerostin antibody treatment

Osteogenesis imperfecta (OI) is a heritable collagen-related bone dysplasia, characterized by brittle bones with increased fracture risk that presents most severely in children. Anti-resorptive bisphosphonates are frequently used to treat pediatric OI and controlled clinical trials have shown that bisphosphonate therapy improves vertebral outcomes but has little benefit on long bone fracture rate. New treatments which increase bone mass throughout the pediatric OI skeleton would be beneficial. Sclerostin antibody (Scl-Ab) is a potential candidate anabolic therapy for pediatric OI and functions by stimulating osteoblastic bone formation via the canonical Wnt signaling pathway. To explore the effect of Scl-Ab on the rapidly growing OI skeleton, we treated rapidly growing 3 week old Brtl/+ mice, harboring a typical heterozygous OI-causing Gly → Cys substitution on col1a1, for 5 weeks with Scl-Ab. Scl-Ab had anabolic effects in Brtl/+ and led to new cortical bone formation and increased cortical bone mass. This anabolic action resulted in improved mechanical strength to WT Veh levels without altering the underlying brittle nature of the material. While Scl-Ab was anabolic in trabecular bone of the distal femur in both genotypes, the effect was less strong in these rapidly growing Brtl/+ mice compared to WT. In conclusion, Scl-Ab was able to stimulate bone formation in a rapidly growing Brtl/+ murine model of OI, and represents a potential new therapy to improve bone mass and reduce fracture risk in pediatric OI.     

Effects of sclerostin antibody on bone healing

Promoting bone healing after a fracture has been a frequent subject of research. Recently, sclerostin antibody(Scl-Ab) has been introduced as a new anabolic agent for the treatment of osteoporosis. Scl-Ab activates the canonical Wnt(cWnt)-β-catenin pathway, leading to an increase in bone formation and decrease in bone resorption. Because of its rich osteogenic effects, preclinically, Scl-Ab has shown positive effects on bone healing in rodent models; researchers have reported an increase in bone mass, mechanical strength, histological bone formation, total mineralized callus volume, bone mineral density, neovascularization, proliferating cell nuclear antigen score, and bone morphogenic protein expression at the fracture site after Scl-Ab administration. In addition, in a rat critical-size femoral-defect model, the Scl-Ab-treated group demonstrated a higher bone healing rate. On the other hand, two clinical reports have researched Scl-Ab in bone healing and failed to show positive effects in the femur and tibia. This review discusses why Scl-Ab appears to be effective in animal models of fracture healing and not in clinical cases.     

Wnt signaling during fracture repair

Bone is one of the few tissues in the body with the capacity to regenerate and repair itself. Fractures usually are completely repaired in a relatively short time, but in a small percentage of cases, healing never occurs and nonunion is the result. Fracture repair and bone regeneration require the localized reactivation of signaling cascades that are crucial for skeletal development. The Wnt/β-catenin signaling pathway is one such developmental pathway whose role in bone formation and regeneration recently has been appreciated. During the past decade, much has been learned about how Wnt pathways regulate bone mass. Small molecules and biologics aimed at this pathway are now being tested as potential new anabolic agents. This article reviews recent data demonstrating that Wnt pathways are active during fracture repair and that increasing the activities of Wnt pathway components accelerates bone regeneration.     

Dkk1 haploinsufficiency requires expression of Bmp2 for bone anabolic activity

Bone fractures remain a serious health burden and prevention and enhanced healing of fractures have been obtained by augmenting either BMP or Wnt signaling. However, whether BMP and Wnt signaling are both required or are self-sufficient for anabolic and fracture healing activities has never been fully elucidated. Mice haploinsufficient for Dkk1 (Dkk1^+/−) exhibit a high bone mass phenotype due to an up-regulation of canonical Wnt signaling while mice lacking Bmp2 expression in the limbs (Bmp2^c/c;Prx1::cre) succumb to spontaneous fracture and are unable to initiate fracture healing; combined, these mice offer an opportunity to examine the requirement for activated BMP signaling on the anabolic and fracture healing activity of Wnts. When Dkk1^+/− mice were crossed with Bmp2^c/c;Prx1::cre mice, the offspring bearing both genetic alterations were unable to increase bone mass and heal fractures, indicating that increased canonical Wnt signaling is unable to exploit its activity in absence of Bmp2. Thus, our data suggest that BMP signaling is required for Wnt-mediated anabolic activity and that therapies aimed at preventing fractures and fostering fracture repair may need to target both pathways for maximal efficacy.     

Inhibition of beta‐catenin signaling by Pb leads to incomplete fracture healing

There is strong evidence in the clinical literature to suggest that elevated lead (Pb) exposure impairs fracture healing. Since Pb has been demonstrated to inhibit bone formation, and Wnt signaling is an important anabolic pathway in chondrocyte maturation and endochondral ossification, we investigated the impact of Wnt therapy on Pb‐exposed mice undergoing bone repair in a mouse tibial fracture model. We established that tibial fracture calluses from Pb‐treated mice were smaller and contained less mineralized tissue than vehicle controls. This resulted in the persistence of immature cartilage in the callus and decreased β‐catenin levels. Reduction of β‐catenin protein was concurrent with systemic elevation of LRP5/6 antagonists DKK1 and sclerostin in Pb‐exposed mice throughout fracture healing. β‐catenin stimulation by the GSK3 inhibitor BIO reversed these molecular changes and restored the amount of mineralized callus. Overall, Pb is identified as a potent inhibitor of endochondral ossification in vivo with correlated effects on bone healing with noted deficits in β‐catenin signaling, suggesting the Wnt/β‐catenin as a pivotal pathway in the influence of Pb on fracture repair. © 2014 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:1397–1405, 2014.     

Enhanced chondrogenesis and Wnt signaling in PTH-treated fractures.

Studies have shown that systemic PTH treatment enhanced the rate of bone repair in rodent models. However, the mechanisms through which PTH affects bone repair have not been elucidated. In these studies we show that PTH primarily enhanced the earliest stages of endochondral bone repair by increasing chondrocyte recruitment and rate of differentiation. In coordination with these cellular events, we observed an increased level of canonical Wnt-signaling in PTH-treated bones at multiple time-points across the time-course of fracture repair, supporting the conclusion that PTH responses are at least in part mediated through Wnt signaling. INTRODUCTION: Since FDA approval of PTH [PTH(1-34); Forteo] as a treatment for osteoporosis, there has been interest in its use in other musculoskeletal conditions. Fracture repair is one area in which PTH may have a significant clinical impact. Multiple animal studies have shown that systemic PTH treatment of healing fractures increased both callus volume and return of mechanical competence in models of fracture healing. Whereas the potential for PTH has been established, the mechanism(s) by which PTH produces these effects remain elusive. MATERIALS AND METHODS: Closed femoral fractures were generated in 8-wk-old male C57Bl/6 mice followed by daily systemic injections of either saline (control) or 30 microg/kg PTH(1-34) for 14 days after fracture. Bones were harvested at days 2, 3, 5, 7, 10, 14, 21, and 28 after fracture and analyzed at the tissue level by radiography and histomorphometry and at the molecular and biochemical levels level by RNase protection assay (RPA), real-time PCR, and Western blot analysis. RESULTS: Quantitative muCT analysis showed that PTH treatment induced a larger callus cross-sectional area, length, and total volume compared with controls. Molecular analysis of the expression of extracellular matrix genes associated with chondrogenesis and osteogenesis showed that PTH treated fractures displayed a 3-fold greater increase in chondrogenesis relative to osteogenesis over the course of the repair process. In addition, chondrocyte hypertrophy occurred earlier in the PTH-treated callus tissues. Analysis of the expression of potential mediators of PTH actions showed that PTH treatment significantly induced the expression of Wnts 4, 5a, 5b, and 10b and increased levels of unphosphorylated, nuclear localized beta-catenin protein, a central feature of canonical Wnt signaling. CONCLUSIONS: These results showed that the PTH-mediated enhancement of fracture repair is primarily associated with an amplification of chondrocyte recruitment and maturation in the early fracture callus. Associated with these cellular effects, we observed an increase in canonical Wnt signaling supporting the conclusion that PTH effects on bone repair are mediated at least in part through the activation of Wnt-signaling pathways.     

Osteopenia and impaired fracture healing in aged EP4 receptor knockout mice

The EP4 receptor, one of the subtypes of the prostaglandin E2 (PGE2) receptor, plays a critical role in the anabolic effects of PGE2 on bone. However, its role in the maintenance of bone mass in aged animals and its role in fracture healing is not well known. Our studies addressed these issues by characterizing the skeletal phenotype of aged, EP4 receptor knockout (KO) mice, and by comparing fracture healing in aged KO mice versus wild type (WT) mice. There was no significant difference in body weight and femoral length between KO and WT mice at 15 to 16 months of age. Lower bone mass was seen radiographically in both axial and long bones of KO mice relative to WT mice. Micro-CT images of the distal femurs showed thinner cortices, fewer trabeculae, and a deteriorated trabecular network in KO mice. Total bone content, trabecular content, and cortical content, as assessed by pQCT in the distal femur, were lower in KO mice than WT controls. Histomorphometric measurements showed that trabecular bone volume and bone formation rate were significantly decreased whereas osteoclast number on trabecular surface and eroded surface on endocortical surface were significantly increased in KO mice. These data indicated that deleting the EP4 receptor resulted in an imbalance in bone resorption over formation, leading to a negative bone balance. The lower bone formation rate in EP4 KO mice was primarily due to decreased mineralizing surface, suggesting that the defect in overall bone formation was mainly due to the defect in osteoblastogenesis. Fracture healing was examined in KO and WT mice subjected to a transverse femoral fracture. Callus formation was significantly delayed as evidenced both radiographically and histologically in the fractured femurs of KO mice compared with those of WT mice. KO mice had significant decreases in total callus area, cartilaginous callus area, and bony callus area 2 weeks after fracture. By 4 weeks, complete bony bridging was seen in WT mice but not in KO mice. These data demonstrate that the absence of the EP4 receptor decreases bone mass and impairs fracture healing in aged male mice. Our findings indicate that the EP4 receptor is a positive regulator in the maintenance of bone mass and fracture healing.     

A novel, non-prostanoid EP2 receptor-selective prostaglandin E2 agonist stimulates local bone formation and enhances fracture healing.

CP-533,536, a newly discovered, non-prostanoid EP2 receptor-selective PGE2 agonist, stimulates local bone formation and enhances fracture healing in rat models. INTRODUCTION: There is a significant medical need for agents that can stimulate local bone formation and enhance fracture healing. We tested the effects of CP-533,536, a newly discovered, non-prostanoid EP2 receptor-selective prostaglandin E2 (PGE2) agonist, in stimulating local bone formation and enhancing fracture healing in rat models. MATERIALS AND METHODS: In the first model, a single injection of CP-533,536 at doses of 0.3, 1, or 3 mg/kg to the proximal tibial metaphysis of 6-week-old male rats was given on day 1, and the local bone anabolic effect was determined on day 7. We then tested the effects of this compound in inducing bone formation on rat periosteum of the femur. A single dose of 0.3 mg of CP-533,536 incorporated in a poly-(D,L-lactide-co-glycolide) (PLGH) matrix was injected onto the periosteum of the femur in 3-week-old male rats, and local bone formation was determined on day 14. Finally, the ability of CP-533,536 in PLGH matrix in enhancing fracture healing was tested using the rat femoral fracture model. CP-533,536 in PLGH matrix at doses of 0.05, 0.5, or 5 mg was delivered to the local fracture site on the same day of fracture, and its efficacy was evaluated on day 21. RESULTS AND CONCLUSIONS: A single injection of CP-533,536 at doses of 0.3, 1, or 3 mg/kg to the proximal tibial metaphysis dose-dependently stimulated local lamellar bone formation on trabecular, endocortical, and periosteal surfaces, and thus increased bone mineral content and bone strength at the injected site. Similarly, a single injection of 0.3 mg of CP-533,536 incorporated in PLGH matrix onto the periosteum of the femur induced significantly local bone formation. In the rat femoral fracture model, CP-533,536 in PLGH matrix at doses of 0.05, 0.5, and 5 mg dose-dependently increased callus size, density, and strength compared with PLGH matrix alone. These results show that CP-533,536 stimulates new bone formation on trabecular, endocortical, and periosteal surfaces and enhances fracture healing. These data reveal that EP2 receptor-selective agonists provide therapeutic potential for local bone augmentation, bone repair, and bone healing in humans.     

Systemic administration of pharmacological agents and bone repair: what can we expect

Pharmacologic agents that modulate bone formation and bone remodelling are in broad use and development for the treatment of osteoporosis and other disorders of bone fragility. There is significant interest into the effect these agents may have on bone repair and fracture healing and whether these agents may be beneficial or detrimental to bone repair. Bisphosphonates delay callus remodelling, but increased callus volume seen during endochondral bone repair with bisphosphonate use allows for equivalent biomechanical properties for the fractured bone. Teripartide stimulates bone formation and in bone repair appears to have the potential to accelerate fracture callus formation and remodelling, potentially accelerating fracture healing. Animal models of fracture healing have demonstrated accelerated healing with larger callus volume, more rapid remodelling to mature bone, and improved biomechanical properties of the fractured bone. Clinical data with teriparatide has shown mixed results for its ability to stimulate fracture healing. Wnt signalling is one of the major pathways through which cartilage and bone formation is regulated during development. This same pathway has been identified as one of the ways that teriparatide stimulates bone formation. Antibodies to downstream proteins in this pathway, Dkk-1 and sclerostin, show significant promise of accelerating even normal fracture healing in preclinical animal models.     

BMP‐7–induced ectopic bone formation and fracture healing is impaired by systemic NSAID application in C57BL/6‐mice

Nonsteroidal antiinflammatory drugs (NSAIDs) are known to potentially impair the fracture healing process. The aim of the present study was to determine if the impairment of bone healing by systemic NSAID application is, at least in part, due to an interaction of NSAIDs with the bone anabolic BMP‐7 pathway. Therefore, we first analyzed fracture healing in control and diclofenac‐treated mice, where we not only found a significant impairment of fracture healing due to diclofenac treatment as assessed by biomechanical testing and µCT imaging, but also found high coexpression of bone morphogenetic protein‐7 (BMP‐7) and cyclooxygenase‐2 (COX‐2) within the fracture callus of both groups. To experimentally address the possible interaction between BMP‐7 and COX‐2, we then induced ectopic bone formation in control (n = 10) and diclofenac‐treated mice (n = 10) by application of BMP‐7 (recombinant human OP‐1, rhOP‐1) into the hamstring muscles. After 20 days of treatment, each ectopic bone nodule was analyzed by contact‐radiography, µCT, histology, and histomorphometry. Diclofenac application decreased the trabecular number and bone mass in the ectopic bone nodules significantly due to reduced osteoblast number and activity. These data demonstrate that the bone anabolic effect of BMP‐7 and fracture healing is impaired by diclofenac application, and suggest that the potential negative impact of NSAIDs on fracture healing is, at least in part, due to interference with BMP‐7 signaling. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:785–791, 2010     

Distinct frequency dependent effects of whole‐body vibration on non‐fractured bone and fracture healing in mice

Low‐magnitude high‐frequency vibration (LMHFV) provokes anabolic effects in non‐fractured bone; however, in fracture healing, inconsistent results were reported and optimum vibration conditions remain unidentified. Here, we investigated frequency dependent effects of LMHFV on fracture healing. Twelve‐week‐old, female C57BL/6 mice received a femur osteotomy stabilized using an external fixator. The mice received whole‐body vibrations (20 min/day) with 0.3g peak‐to‐peak acceleration and a frequency of either 35 or 45 Hz. After 10 and 21 days, the osteotomized femurs and intact bones (contra‐lateral femurs, lumbar spine) were evaluated using bending‐testing, µ‐computed tomography, and histomorphometry. In non‐fractured trabecular bone, vibration with 35 Hz significantly increased the relative amount of bone (+28%) and the trabecular number (+29%), whereas cortical bone was not influenced. LMHFV with 45 Hz failed to provoke anabolic effects in trabecular or cortical bone. Fracture healing was not significantly influenced by whole‐body vibration with 35 Hz, whereas 45 Hz significantly reduced bone formation (?64%) and flexural rigidity (?34%) of the callus. Although the exact mechanisms remain open, our results suggest that small vibration setting changes could considerably influence LMHFV effects on bone formation in remodeling and repair, and even disrupt fracture healing, implicating caution when treating patients with impaired fracture healing. © 2014 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:1006–1013, 2014.

     

Adult Brtl/+ mouse model of osteogenesis imperfecta demonstrates anabolic response to sclerostin antibody treatment with increased bone mass and strength

Summary

Treatments to reduce fracture rates in adults with osteogenesis imperfecta are limited. Sclerostin antibody, developed for treating osteoporosis, has not been explored in adults with OI. This study demonstrates that treatment of adult OI mice respond favorably to sclerostin antibody therapy despite retention of the OI-causing defect.

Introduction

Osteogenesis imperfecta (OI) is a heritable collagen-related bone dysplasia, characterized by brittle bones with increased fracture risk. Although OI fracture risk is greatest before puberty, adults with OI remain at risk of fracture. Antiresorptive bisphosphonates are commonly used to treat adult OI, but have shown mixed efficacy. New treatments which consistently improve bone mass throughout the skeleton may improve patient outcomes. Neutralizing antibodies to sclerostin (Scl-Ab) are a novel anabolic therapy that have shown efficacy in preclinical studies by stimulating bone formation via the canonical wnt signaling pathway. The purpose of this study was to evaluate Scl-Ab in an adult 6 month old Brtl/+ model of OI that harbors a typical heterozygous OI-causing Gly?>?Cys substitution on Col1a1.

Methods

Six-month-old WT and Brtl/+ mice were treated with Scl-Ab (25 mg/kg, 2×/week) or Veh for 5 weeks. OCN and TRACP5b serum assays, dynamic histomorphometry, microCT and mechanical testing were performed.

Results

Adult Brtl/+ mice demonstrated a strong anabolic response to Scl-Ab with increased serum osteocalcin and bone formation rate. This anabolic response led to improved trabecular and cortical bone mass in the femur. Mechanical testing revealed Scl-Ab increased Brtl/+ femoral stiffness and strength.

Conclusion

Scl-Ab was successfully anabolic in an adult Brtl/+ model of OI.     

Improved healing of experimental long bone fractures in rabbits by delayed internal fixation.

Experiments in rabbits were conducted to test the clinical propositions that a) the risk of nonunion in certain long bone fractures can be sharply reduced by open reduction and internal fixation, and b) that there is an optimum critical period following trauma in which this procedure should be carried out to facilitate healing. Bilateral standard open fractures of rabbit radii were internally fixed by intramedullary K-wires immediately or at 5, 10, and 17 days after trauma. These bones were subjected to stress and histometric analyses at 4-56 days after the initial fractures. The observations suggested that the pace of fracture healing could be optimized by time-delay surgery. The most exuberant and strongest calluses developed when the bones were fixed 10 days after fracture, and the rate of healing was improved. Delay times of 5 or 17 days did not provide better healing than that achieved by an immediate operation. Callus quality, in terms of the proportion of fiber bone, lamellar bone, and cartilage, was similar in all groups. The tensile strengths of the bones during callus formation were highly correlated with the ratio of callus/cortical bone areas (p less than 0.01).     

Anti-DKK1 antibody promotes bone fracture healing through activation of β-catenin signaling

In this study we investigated if Wnt/β-catenin signaling in mesenchymal progenitor cells plays a role in bone fracture repair and if DKK1-Ab promotes fracture healing through activation of β-catenin signaling. Unilateral open transverse tibial fractures were created in CD1 mice and in β-catenin^Prx1ER conditional knockout (KO) and Cre-negative control mice (C57BL/6 background). Bone fracture callus tissues were collected and analyzed by radiography, micro-CT (μCT), histology, biomechanical testing and gene expression analysis. The results demonstrated that treatment with DKK1-Ab promoted bone callus formation and increased mechanical strength during the fracture healing process in CD1 mice. DKK1-Ab enhanced fracture repair by activation of endochondral ossification. The normal rate of bone repair was delayed when the β-catenin gene was conditionally deleted in mesenchymal progenitor cells during the early stages of fracture healing. DKK1-Ab appeared to act through β-catenin signaling to enhance bone repair since the beneficial effect of DKK1-Ab was abrogated in β-catenin^Prx1ER conditional KO mice. Further understanding of the signaling mechanism of DKK1-Ab in bone formation and bone regeneration may facilitate the clinical translation of this anabolic agent into therapeutic intervention.     

Heterotopic ossification prophylaxis with indomethacin increases the risk of long-bone nonunion

Indomethacin is commonly administered for the prophylaxis of heterotopic ossification (HO) after the surgical treatment of acetabular fractures. Non-steroidal anti-inflammatory drugs such as indomethacin, have been associated with delayed healing of fractures and mechanically weaker callus. Our aim was to determine if patients with an acetabular fracture, who received indomethacin for prophylaxis against HO, were at risk of delayed healing or nonunion of any associated fractures of long bones. We reviewed 282 patients who had had open reduction and internal fixation of an acetabular fracture. Patients at risk of HO were randomised to receive either radiation therapy (XRT) or indomethacin. Of these patients, 112 had sustained at least one concomitant fracture of a long bone; 36 needed no prophylaxis, 38 received focal radiation and 38 received indomethacin. Fifteen patients developed 16 nonunions. When comparing patients who received indomethacin with those who did not, a significant difference was noted in the rate of nonunion (26% v 7%; p = 0.004). Patients with concurrent fractures of the acetabulum and long bones who receive indomethacin have a significantly greater risk of nonunion of the fractures of the long bones when compared with those who receive XRT or no prophylaxis.     

Inhibin A enhances bone formation during distraction osteogenesis

Given the aging population and the increased incidence of fracture in the elderly population, the need exists for agents that can enhance bone healing, particularly in situations of delayed fracture healing and/or non‐union. Our previous studies demonstrated that overexpression of the gonadal peptide, human inhibin A (hInhA), in transgenic mice enhances bone formation and strength via increased osteoblast activity. We tested the hypothesis that hInhA can also exert anabolic effects in a murine model of distraction osteogenesis (DO), using both transgenic hInhA overexpression and administration of normal physiological levels of hInhA in adult male Swiss‐Webster mice. Tibial osteotomies and external ring fixation were performed, followed by a 3‐day latency period, 14‐day distraction, and sacrifice on day 18. Supraphysiological levels of hInhA in transgenic mice, but not normal physiological levels of hInhA, significantly increased endosteal bone formation and mineralized bone area in the distraction gap, as determined by radiographic and µCT analysis. Significantly, increased PCNA and osteocalcin expression in the primary matrix front suggested that hInhA increased osteoblast proliferation. This mechanism is consistent with the effects of other agents and pathologies that modulate bone formation during DO, and demonstrates the potential of hInhA to enhance bone repair and regeneration. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 30:288–295, 2012     

Local Strategies to Prevent and Treat Osteoporosis

Despite advances in systemic osteoporosis therapeutic outcomes, management of fragility fractures and implant fixation in osteoporotic bone remain difficult clinical challenges. Low initial bone density and a prolonged healing response can lead to fracture nonunion and aseptic implant loosening. Local treatment strategies could be used to prevent fracture, accelerate healing, and increase implant fixation by locally stimulating anabolic pathways or inhibiting catabolic pathways. Local strategies under investigation include direct drug release from injectable materials or implant surface coatings. Common locally delivered drugs include bisphosphonates, parathyroid hormone, and bone morphogenetic proteins, yet additional compounds targeting novel pathways in bone biology are also being actively explored. Mechanical stimulation via low intensity pulsed ultrasound, alone or in combination with drug therapy, may also prove effective to promote local bone healing and implant fixation within osteoporotic bone.     

Rejuvenation of the inflammatory system stimulates fracture repair in aged mice

Age significantly reduces the regenerative capacity of the skeleton, but the underlying causes are unknown. Here, we tested whether the functional status of inflammatory cells contributes to delayed healing in aged animals. We created chimeric mice by bone marrow transplantation after lethal irradiation. In this model, chondrocytes and osteoblasts in the regenerate are derived exclusively from host cells while inflammatory cells are derived from the donor. Using this model, the inflammatory system of middle‐aged mice (12 month old) was replaced by transplanted bone marrow from juvenile mice (4 weeks old), or age‐matched controls. We found that the middle‐aged mice receiving juvenile bone marrow had larger calluses and more bone formation during early stages and faster callus remodeling at late stages of fracture healing, indicating that inflammatory cells derived from the juvenile bone marrow accelerated bone repair in the middle‐aged animals. In contrast, transplanting bone marrow from middle‐aged mice to juvenile mice did not alter the process of fracture healing in juvenile mice. Thus, the roles of inflammatory cells in fracture healing may be age‐related, suggesting the possibility of enhancing fracture healing in aged animals by manipulating the inflammatory system. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:1000–1006, 2010     

Mechanically Driven Counter-Regulation of Cortical Bone Formation in Response to Sclerostin-Neutralizing Antibodies

Sclerostin (Scl) antibodies (Scl-Ab) potently stimulate bone formation, but these effects are transient. Whether the rapid inhibition of Scl-Ab anabolic effects is due to a loss of bone cells’ capacity to form new bone or to a mechanostatic downregulation of Wnt signaling once bone strength exceeds stress remains unclear. We hypothesized that bone formation under Scl-Ab could be reactivated by increasing the dose of Scl-Ab and/or by adding mechanical stimuli, and investigated the molecular mechanisms involved in this response, in particular the role of periostin (Postn), a co-activator of the Wnt pathway in bone. For this purpose, C57Bl/6, Postn^−/− and Postn^+/+ mice were treated with vehicle or Scl-Ab (50 to 100 mg/kg/wk) for various durations and subsequently subjected to tibia axial compressive loading. In wild-type (WT) mice, Scl-Ab anabolic effects peaked between 2 and 4 weeks and declined thereafter, with no further increase in bone volume and strength between 7 and 10 weeks. Doubling the dose of Scl-Ab did not rescue the decline in bone formation. In contrast, mechanical stimulation was able to restore cortical bone formation concomitantly to Scl-Ab treatment at both doses. Several Wnt inhibitors, including Dkk1, Sost, and Twist1, were upregulated, whereas Postn was markedly downregulated by 2 to 4 weeks of Scl-Ab. Mechanical loading specifically upregulated Postn gene expression. In turn, Scl-Ab effects on cortical bone were more rapidly downregulated in Postn^−/− mice. These results indicate that bone formation is not exhausted by Scl-Ab but inhibited by a mechanically driven downregulation of Wnt signaling. Hence, increasing mechanical loads restores bone formation on cortical surfaces, in parallel with Postn upregulation. © 2020 American Society for Bone and Mineral Research (ASBMR).