Aims:Accurate differentiation between periprosthetic joint infection (PJI) and aseptic osteolysis (AO) remains a major clinical challenge in revision joint arthroplasty. We examined bone biopsies for alterations in bone matrix integrity and the osteocyte lacunocanalicular network in order to differentiate between these pathologies. Methods:Bone biopsies were taken from areas of osteolysis in patients who underwent revision arthroplasty for either PJI, diagnosed according to 2018 International Consensus Meeting criteria, or AO. Controls were obtained during primary arthroplasty. Biopsies were assessed histologically by Masson's trichrome, silver and RGB trichrome staining, and for osteocyte lacunocanalicular characteristics. The diagnostic potential of histological measures was investigated using univariate area under the receiver operating characteristic curve (AUROC) analysis. Results:RGB trichrome staining was most effective for revealing both bone matrix and lacunocanalicular features. Osteocyte lacunar area and circularity were increased in PJI relative to both AO and Control bone. PJI bone exhibited reduced canalicular density compared to AO (p < 0.001), and shorter, narrower canaliculi than AO and Controls. Predictive univariate modelling revealed that all histological parameters measured except lacunar area were potentially diagnostic for PJI. Degraded bone matrix differentially predicted PJI (AUROC = 0.875, 88% sensitivity, 80% specificity) and AO (AUROC = 0.733, 63% sensitivity, 76% specificity), as did lacunar circularity (PJI: AUROC = 0.903, 75% sensitivity, 96% specificity; AO: AUROC = 0.797, 69% sensitivity, 84% specificity). Canalicular area fraction, length, and width all predicted PJI (AUROCs = 0.915, 0.982, 0.883), with canalicular length yielding 94% sensitivity and 100% specificity. Canalicular density differentially predicted PJI (AUROC = 0.770, 69% sensitivity, 80% specificity) and AO (AUROC = 0.757, 88% sensitivity, 60% specificity). Conclusion:Histological assessment of bone matrix degradation and the osteocyte lacunocanalicular network reveals new potential diagnostic markers for PJI to support clinical decision-making and provides novel measures for distinguishing PJI from AO, which may be particularly useful in cases of low-grade and chronic infections. Based on these findings, larger confirmatory studies are warranted.

Objective: A role for TWEAK expression and that of its receptor Fn14 by osteoblasts and synovial tissue has been proposed in the pathogenesis of osteoarthritis (OA). Here, we examined whether the cartilage in OA was also a source and target of TWEAK. Design: Articular cartilage samples from 24 patients undergoing hip or knee replacement surgery for OA were investigated for TWEAK and Fn14 expression by both immunostaining and real-time RT-PCR. Human primary chondrocytes isolated from OA cartilage were treated with combinations of recombinant TWEAK and TNF&alpha and examined for the regulation of TWEAK, FN14, RANKL, ADAM17 and SOST mRNA levels. Soluble RANKL levels were measured by ELISA. Results: TWEAK and Fn14 mRNA levels were elevated in grade 2 OA cartilage compared to grade 0 (p < 0.05). Immunostaining indicated that the majority (21/24) of OA cartilage samples expressed low levels of TWEAK protein and high levels of Fn14, however, expression did not appear to vary with respect to OA grade. Primary chondrocytes treated with TNF&alpha exhibited upregulated TWEAK and Fn14 mRNA expression. SOST mRNA expression was inhibited by both TWEAK and TNF&alpha treatment. TWEAK transiently induced RANKL mRNA expression while both TWEAK and TNF&alpha induced sRANKL release by chondrocytes. TWEAK and TNF&alpha also induced the mRNA expression of the RANKL sheddase, ADAM17. Conclusions: TWEAK and Fn14 mRNA expression is elevated in OA articular cartilage. The TWEAK and TNF&alpha induction of ADAM17 expression and sRANKL released by chondrocytes in vitro, as well as suppression of sclerostin/SOST, points to a potential catabolic role for these mediators in OA. ### Competing Interest Statement The authors have declared no competing interest. National Health and Medical Research Council, https://ror.org/011kf5r70, 1004871

Abstract Background Vertebral mobility (V-mobility) has been used to diagnose fresh osteoporotic vertebral fractures (OVFs) and determine bone union by setting cutoff values for these purposes. V-mobility is the difference in vertebral height on dynamic radiographs taken in the sitting and lateral decubitus or supine positions. The dimensions for V-mobility were presented as anterior vertebral height (Ha; mm), wedge ratio (WR; %), and wedge angle (WA; °) in previous reports. This study was performed to obtain WR and WA values equivalent to V-mobility of 1.0 mm in Ha. Methods Lateral radiographs of 284 OVFs (grade 1–3 deformed vertebrae) from T11 to L2 were obtained from 77 patients with OVF. V-mobility presented as Ha, posterior vertebral height, and WA was obtained by the difference in these dimensions on dynamic radiographs. The WR and WA values equivalent to 1.0 mm in Ha were obtained by dividing the V-mobility values for WR and WA by that for Ha. Results The mean WR values corresponding to 1.0 mm in Ha for grade 1, 2, and 3 vertebrae were 3.2% ± 1.4%, 3.2% ± 0.9%, and 3.4% ± 1.0%, respectively, and the corresponding value for grade 1–3 vertebrae was 3.3% ± 1.0%. The mean WA values corresponding to 1.0 mm in Ha for grade 1, 2, and 3 vertebrae were 1.5° ± 0.8°, 1.5° ± 0.6°, and 1.5° ± 0.8°, respectively, and the corresponding value for grade 1–3 vertebrae was 1.5° ± 0.7°. Conclusions The WR and WA values equivalent to V-mobility of 1.0 mm in Ha were 3.3% and 1.5°, respectively, in grade 1–3 vertebrae. These findings may be useful to secure a reliable value of V-mobility of OVFs using simultaneous measurements in three dimensions (Ha, WR, and WA) in clinical practice and to establish cutoff values for V-mobility to determine bone union.

Osteopetrosis is a heterogeneous group of rare hereditary diseases characterized by increased bone mass of poor quality. Autosomal-dominant osteopetrosis type II (ADOII) is most often caused by mutation of the CLCN7 gene leading to impaired bone resorption. Autosomal recessive osteopetrosis (ARO) is a more severe form and is frequently accompanied by additional morbidities. We report an adult male presenting with classical clinical and radiological features of ADOII. Genetic analyses showed no amino-acid-converting mutation in CLCN7 but an apparent haploinsufficiency and suppression of CLCN7 mRNA levels in peripheral blood mononuclear cells. Next generation sequencing revealed low-frequency intronic homozygous variations in CLCN7, suggesting recessive inheritance. In silico analysis of an intronic duplication c.595-120_595-86dup revealed additional binding sites for Serine- and Arginine-rich Splicing Factors (SRSF), which is predicted to impair CLCN7 expression. Quantitative backscattered electron imaging and histomorphometric analyses revealed bone tissue and material abnormalities. Giant osteoclasts were present and additionally to lamellar bone, and abundant woven bone and mineralized cartilage were observed, together with increased frequency and thickness of cement lines. Bone mineralization density distribution (BMDD) analysis revealed markedly increased average mineral content of the dense bone (CaMean T-score + 10.1) and frequency of bone with highest mineral content (CaHigh T-score + 19.6), suggesting continued mineral accumulation and lack of bone remodelling. Osteocyte lacunae sections (OLS) characteristics were unremarkable except for an unusually circular shape. Together, our findings suggest that the reduced expression of CLCN7 mRNA in osteoclasts, and possibly also osteocytes, causes poorly remodelled bone with abnormal bone matrix with high mineral content. This together with the lack of adequate bone repair mechanisms makes the material brittle and prone to fracture. While the skeletal phenotype and medical history were suggestive of ADOII, genetic analysis revealed that this is a possible mild case of ARO due to deep intronic mutation.

Background Conservative treatments for osteoporotic vertebral fractures (OVFs) have not been standardized, and criteria for determining bone union have not been established. To determine bone union, we have adopted a cutoff value of 1.0 mm for vertebral mobility (V-mobility), defined as the difference in anterior vertebral height (Ha) between lateral radiographs taken in weight-bearing and non-weight-bearing positions. The present study aimed to investigate the usefulness of V-mobility for determining bone union and predicting bone union at 6 months after OVF onset. Methods The study included 54 acute OVFs from T11 to L3 in 53 patients (12 males, 41 females; mean age 82 years; age range 55–97 years) who were hospitalized at ≤ 3 weeks after OVF onset. Vertebral deformity (V-deformity) and V-mobility were evaluated in accordance with Ha on lateral radiographs taken in the sitting position (SIT), lateral decubitus position (DEC), and supine position (SUP). OVFs showing V-mobility of ≤ 1.0 mm between SIT and DEC radiographs and no intravertebral cleft on DEC radiograph were defined as semi-union, while those showing V-mobility of ≤ 1.0 mm between SIT and SUP radiographs and no intravertebral cleft on SUP radiograph were defined as bone union. We calculated the bone union rates including semi-unions associated with V-mobility cutoff values of 1.0 mm, 1.5 mm, and 2.0 mm and estimated cutoff values for V-mobility at 5 weeks after OVF onset to predict bone union at 6 months after OVF onset. Results The cumulative number of bone unions including semi-unions was more influenced by the different V-mobility cutoff values in Ha for determining bone union in the earlier period compared with the later period in the time course of OVF. Receiver-operating characteristic curve analyses revealed that V-mobility cutoff value of 2.1 mm in Ha between SIT and DEC radiographs at 5 weeks after OVF had moderate accuracy for predicting bone union including semi-union at 6 months after OVF. The mean V-deformity value on SIT radiographs did not progress significantly. Conclusion V-mobility in the early stage after OVF can predict bone union at 6 months after OVF and is a useful quantitative indicator for determining bone union.

ABSTRACTWe investigated if time between injury and surgery affects cancellous bone properties in patients suffering tibial plateau fractures (TPF), in terms of structural integrity and gene expression controlling bone loss. A cohort of 29 TPF, operated 1–17 days post‐injury, had biopsies from the fracture and an equivalent contralateral limb site, at surgery. Samples were assessed using micro‐computed tomography and real‐time RT‐PCR analysis for the expression of genes known to be involved in bone remodeling and fracture healing. Significant decreases in the injured vs control side were observed for bone volume fraction (BV/TV, −13.5 ± 6.0%, p = 0.011), trabecular number (Tb.N, −10.5 ± 5.9%, p = 0.041) and trabecular thickness (Tb.Th, −4.6 ± 2.5%, p = 0.033). Changes in these parameters were more evident in patients operated 5–17 days post‐injury, compared to those operated in the first 4 days post‐injury. A significant negative association was found between Tb.Th (r = −0.54, p < 0.01) and BV/TV (r = −0.39, p < 0.05) in relation to time post‐injury in the injured limb. Both BV/TV and Tb.Th were negatively associated with expression of key molecular markers of bone resorption, CTSK, ACP5, and the ratio of RANKL:OPG mRNA. These structure/gene expression relationships did not exist in the contralateral tibial plateau of these patients. This study demonstrated that there is a significant early time‐dependent bone loss in the proximal tibia after TPF. This bone loss was significantly associated with altered expression of genes typically involved in the process of osteoclastic bone resorption but possibly also bone resorption by osteocytes. The mechanism of early bone loss in such fractures should be a subject of further investigation. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:2865–2875, 2018.

There is an ongoing demand for new approaches for treating localized bone pathologies. Here we propose a new strategy for treatment of such conditions, via local delivery of hormones/drugs to the trauma site using drug releasing nano-engineered implants. The proposed implants were prepared in the form of small Ti wires/needles with a nano-engineered oxide layer composed of array of titania nanotubes (TNTs). TNTs implants were inserted into a 3D collagen gel matrix containing human osteoblast-like, and the results confirmed cell migration onto the implants and their attachment and spread. To investigate therapeutic efficacy, TNTs/Ti wires loaded with parathyroid hormone (PTH), an approved anabolic therapeutic for the treatment of severe bone fractures, were inserted into 3D gels containing osteoblast-like cells. Gene expression studies revealed a suppression of SOST (sclerostin) and an increase in RANKL (receptor activator of nuclear factor kappa-B ligand) mRNA expression, confirming the release of PTH from TNTs at concentrations sufficient to alter cell function. The performance of the TNTs wire implants using an example of a drug needed at relatively higher concentrations, the anti-inflammatory drug indomethacin, is also demonstrated. Finally, the mechanical stability of the prepared implants was tested by their insertion into bovine trabecular bone cores ex vivo followed by retrieval, which confirmed the robustness of the TNT structures. This study provides proof of principle for the suitability of the TNT/Ti wire implants for localized bone therapy, which can be customized to cater for specific therapeutic requirements.

Periprosthetic osteolysis (PO) leading to aseptic loosening, is the most common cause of failure of total hip replacement (THR) in the mid- to long-term. Polyethylene (PE) particulates from the wear of prosthesis liners are bioactive and are implicated in the initiation and or progression of osteolysis. Evidence exists that cells of the osteoblast/osteocyte lineage are affected by PE particles and contribute to the catabolic response by promoting osteoclastic bone resorption. In this study, we hypothesised that osteocytes contribute directly to PO by removing bone from their perilacunar matrix. Osteocyte responses to ultra-high molecular weight PE (UHMWPE) particles were examined in vitro in human primary osteocyte-like cultures, in vivo in the mouse calvarial osteolysis model, and in the acetabulum of patients undergoing revision total hip replacement (THR) surgery for PO. Osteocytes exposed to UHMWPE particles showed upregulated expression of catabolic markers, MMP-13, carbonic anhydrase 2 (CA2), cathepsin (CTSK) and tartrate resistant acid phosphatase (TRAP), with no effect on cell viability, as assessed by Caspase 3 activity. Consistent with this catabolic activity causing perilacunar bone loss, histological analysis of calvarial sections from mice exposed to UHMWPE revealed a significant (p < 0.001) increase in osteocyte lacunar area (Lac.Ar) compared to sham-operated animals. Furthermore, acetabular biopsies from patients with PO also showed significantly (p < 0.001) increased osteocyte lacunar size in trabecular bone adjacent to PE particles, compared with osteocyte lacunar size in bone from primary THR patients. Together, these findings suggest a previously unrecognised action of UHMWPE wear particles on osteocytes, which directly results in a loss of osteocyte perilacunar bone. This action may exacerbate the indirect pro-osteoclastic action of UHMWPE-affected osteocytes, previously shown to contribute to aseptic loosening of orthopaedic implants.Statement of SignificanceThis study addresses the clinical problem of periprosthetic osteolysis, bone loss in response to polyethylene wear particles derived from materials used in orthopaedic implants. Periprosthetic osteolysis has been thought to be due largely to wear particles stimulating the activity of bone resorbing osteoclasts. However, in this study we demonstrate for the first time that polyethylene particles stimulate another type of bone loss, mediated by the direct activity of bone mineral embedded osteocytes, termed osteocytic osteolysis or osteocyte perilacunar remodelling. This study provides new mechanistic insight into wear particle mediated bone loss and represents a new paradigm for the way in which bone cells, namely osteocytes, the key controlling cell type in bone, react to biomaterials. (C) 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

The success of implantation of materials into bone is governed by effective osseointegration, requiring biocompatibility of the material and the attachment and differentiation of osteoblastic cells. To enhance cellular function in response to the implant surface, micro- and nano-scale topography have been suggested as essential. In this study, we present bone implants based on 3D-printed titanium alloy (Ti6Al4V), with a unique dual topography composed of micron-sized spherical particles and vertically aligned titania nanotubes. The implants were prepared by combination of 3D-printing and anodization processes, which are scalable, simple and cost-effective. The osseointegration properties of fabricated implants, examined using human osteoblasts, showed enhanced adhesion of osteoblasts compared with titanium materials commonly used as orthopaedic implants. Gene expression studies at early (day 7) and late (day 21) stages of culture were consistent with the Ti substrates inducing an osteoblast phenotype conducive to effective osseointegration. These implants with the unique combination of micro- and nano-scale topography are proposed as the new generation of multi-functional bone implants, suitable for addressing many orthopaedic challenges, including implant rejection, poor osseointegration, inflammation, drug delivery and bone healing. Copyright © 2016 John Wiley & Sons, Ltd.

Sclerostin, the SOST gene product, is a negative regulator of bone formation and a positive regulator of bone resorption. In this study, treatment of human primary osteoblasts, including cells differentiated to an osteocyte-like stage, with 1 alpha,25-dihydroxyvitaminD(3) (1,25D) resulted in the dose-dependent increased expression of SOST mRNA. A similar effect was observed in human trabecular bone samples cultured ex vivo, and in osteocyte-like cultures of differentiated SAOS2 cells. Treatment of SAOS2 cells with 1,25D resulted in the production and secretion of sclerostin protein. In silica analysis of the human SOST gene revealed a single putative DR3-type vitamin D response element (VDRE) at position 6216 bp upstream of the transcription start site (TSS). This sequence was confirmed to have strong VDRE activity by luciferase reporter assays and electrophoretic mobility shift analysis (EMSA). Sequence substitution in the VDR/RXR half-sites abolished VDRE reporter activity and binding of nuclear proteins. A 63 kb fragment of the human proximal SOST promoter demonstrated responsiveness to 1,25D. The addition of the evolutionary conserved region 5 (ECR5), a known bone specific enhancer region, ahead of the 63 kb fragment increased basal promoter activity but did not increase 1,25D responsiveness. Site-specific mutagenesis abolished the responsiveness of the 6.3 kb promoter to 1,25D. We conclude that 1,25D is a direct regulator of human SOST gene and sclerostin protein expression, extending the pathways of control of sclerostin expression. At least some of this responsiveness is mediated by the identified classical VDRE however the nature of the transcriptional regulation by 1,25D warrants further investigation. (C) 2015 Elsevier Ireland Ltd. All rights reserved.

The principal challenge for bone therapy is to deliver an effective dose of therapeutic agent (for example antibiotic or anti-cancer drug) to the affected site within bone, while sparing other organs. The solution to this dilemma is to deliver drug locally within the bone; hence various surface/therapeutic modifications of the conventional bone implants have been suggested to achieve this. Implants composed of biocompatible materials and loaded with active therapeutics thus provide one possible option for effective bone therapy. This chapter showcases the challenges that an electrochemically nano-engineered bone implant based on titania nanotubes must overcome to survive and deliver therapeutics in conditions such as infections and cancer of bone. The fabrication of titania nanotubes, the therapeutic loading and release, ex vivo and in vivo investigations; all are reviewed in terms of effectiveness for therapeutic action. Also discussed are the potential advances of titania nanotube technology and the future research directions to address additional clinical problems.

The association between increased serum 25-hydroxyvitamin D (25D) and reduced osteoclastic bone resorption is well known. Previously, we have demonstrated that mechanism by which this occurs, may include the conversion of 25D to 1,25-dihydroxyvitamin D (1,25D) by osteoclasts, catalysed by the CYP27B1 enzyme. Local 1,25D synthesis in osteoclasts was shown to regulate osteoclastogenesis and moderating resorptive activity. Thus, we hypothesised that osteoclasts differentiated from mice with global deletion of the Cyp27b1 gene (Cyp27b1 KO) would display enhanced resorptive capacity due to the lack of an ameliorating effect of 1,25D. Splenocytes isolated from Cyp27b1 KO mice or their wild-type (WT) littermates between 6 and 8 weeks of age were cultured under osteoclast-forming conditions for up to 14 days. Osteoclast formation was measured by staining for the osteoclast marker tartrate resistant acid phosphatase (TRAP). Bone resorption activity was measured by plating the cells on a bone-like substrate. In Cyp27b1 KO cultures, osteoclastogenesis was reduced, as indicated by fewer TRAP-positive multinucleated cells at all time points measured (p<0.05) when compared to wild-type (WT) levels. However, Cyp27b1 KO osteoclasts demonstrated greater resorption on a per cell basis than their WT counterparts (p<0.03). In addition, the ratio of expression of the pro-apoptotic gene Bax to the pro-survival gene Bcl-2 was decreased in Cyp27b1 KO cultures, implying that these smaller osteoclasts survive longer than WT osteoclasts. Our data indicate abnormal osteoclastogenesis due to the absence of CYP27B1 expression, consistent with the notion that endogenous metabolism of 25D optimises osteoclastogenesis and ameliorates the resulting activity of mature osteoclasts.

To treat skeletal conditions such as bone infections, osteoporotic fractures, and osteosarcoma, it would be ideal to introduce drugs directly to the affected site. Localized drug delivery from the bone implants is a promising alternative to systemic drug administration. In this study we investigated electrochemically nanoengineered Ti wire implants with titania nanotubes (TNTs), as minimally invasive drug-releasing implants for the delivery of drugs directly into the bone tissue. Since trabecular bone in vivo contains a highly interconnected bone marrow, we sought to determine the influence of marrow on drug release and diffusion. Electrochemical anodization of Ti wires (length 10 mm) was performed to create an oxide layer with TNTs on the surface, followed by loading with a fluorescent model drug, Rhodamine B (RhB). Cores of bovine trabecular bone were generated from the sternum of a young steer, and were processed to have an intact bone marrow, or the marrow was removed. RhB-loaded TNTs/Ti wires were inserted into the bone cores, which were then cultured ex vivo using the ZetOS bioreactor system to maintain bone viability. Release and diffusion of RhB inside the bone was monitored using fluorescence imaging and different patterns of drug transport in the presence or absence of marrow were observed. Scanning electron microscopy of the implants after retrieval from bone cores confirmed survival of the TNTs structures. Histological investigation showed the presence of bone cells adherent on the implants. This study shows a potential of Ti drug-releasing implants based on TNTs technology towards localized bone therapy. (c) 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 714-725, 2016.

Fibroblast growth factor-23 (FGF23), produced by osteocytes, is the key physiological regulator of phosphate homeostasis. Sepsis patients often experience transient hypophosphataemia, suggesting the regulation of FGF23 levels by pro-inflammatory factors. Here, we used the osteocyte-like cell line IDG-SW3 to investigate the effect of pro-inflammatory stimuli on FGF23 production. In differentiated IDG-SW3 cultures, basal Fgf23 mRNA was dose-dependently up-regulated by pro-inflammatory cytokines TNF, IL-1β and TWEAK, and bacterial LPS. Similar effects were observed in human bone samples. TNF- and IL-1β-induced Fgf23 expression was NF-κB-dependent. Conversely, mRNA encoding negative regulators of FGF23, Phex, Dmp1 and Enpp1, were suppressed by TNF, IL-1β, TWEAK and LPS, independent of NF-κβ signalling. Galnt3, the protein product of which protects intact FGF23 protein from furin/furin-like proprotein convertase cleavage, increased in response to these treatments. C-terminal FGF23 and intact FGF23 protein levels also increased, the latter only in the presence of Furin inhibitors, suggesting that enzymatic cleavage exerts critical control of active FGF23 secretion by osteocytes. Our results demonstrate in principle that pro-inflammatory stimuli are capable of increasing osteocyte secretion of FGF23, which may contribute to hypophosphataemia during sepsis and possibly other inflammatory conditions.

Introduction Sclerostin has been implicated in mechanotransduction in bone and recent data show a lack of response to loading in the sclerostin transgenic mouse. Sclerostin, the protein product of the SOST gene, is an attractive therapeutic target for low bone mass conditions, including osteoporosis. It is expressed exclusively by mature osteocytes in bone and we have shown that sclerostin targets pre-osteocytes/osteocytes to regulate bone mineralization and osteoclast activity, as well as inducing catabolic gene expression in osteocytes themselves and promoting osteocyte-mediated bone loss (osteocytic osteolysis). The aim of this study was to examine the direct effects of sclerostin on anabolic responses to loading in bone ex vivo. Methods 10 × 5mm bovine sternum trabecular bone cores were perfused with osteogenic media at 37°C for up to 3 weeks in individual bone culture chambers. The cores were divided into 3 groups; a) mechanically loaded (300 cycles, 4000 μstrain, 1 Hz/day), b) identical loading regime with continuous perfusion of 50 ng/ml recombinant human sclerostin and c) unloaded controls. Loading was accomplished using a second-generation Zetos™ bone loading system. Daily measurements of bone stiffness (Young9s modulus), media pH and ionic calcium concentrations were made. Histomorphometric assessment, including fluorochrome labelling analysis, was made of resin-embedded, non-decalcified samples at the end of the experiment. Gene expression in the bovine bone was examined by real-time RT-PCR. Results Bovine bone cores showed a steady increase in Young9s modulus with daily application of mechanical loading. This increase in stiffness was blocked by the co-addition of sclerostin. Sclerostin also induced bone acidification and a net release of bone calcium, indicated by the decrease in media pH and the relative increase in ionic calcium concentrations in the presence of sclerostin. Sclerostin also completely abrogated loading-induced calcium/calcein uptake. Sclerostin induced an increase in the expression of the bone resorption genes, tartrate resistant acid phosphatase (TRAP), carbonic anhydrase and cathepsin K and induced the release of β-CTX. Histological examination revealed a significant increase in the size of the osteocyte lacunae in sclerostin-treated bone cores, suggesting a role for osteocytic osteolysis in this effect. Discussion/Conclusion The observation that sclerostin abrogated the loading-induced increase in bone stiffness constitutes direct evidence for a negative effect of sclerostin on the anabolic response to mechanical loading. Our findings may be explained in part by the observation that sclerostin negatively controls mineralization by late osteoblasts and pre-osteocytes (1). It is also possible that osteocytes themselves are capable of releasing bone mineral in response to sclerostin. This study demonstrates that sclerostin directly antagonises the anabolic effects of mechanical loading in the absence of external (circulating, neural, hormonal) influences. The mechanisms, by which sclerostin exerts these effects, warrant further study.