Is RANKL a potential molecular target in osteoarthritis?

OBJECTIVE: Osteoarthritis (OA) is a disease of joints, in which the bone under the articular cartilage undergoes increased remodelling activity. The question is whether a better understanding of the causes and mechanisms of bone remodelling can predict disease-modifying treatments. DESIGN: This review summarises the current understanding of the aetiology of OA, with an emphasis on events in the subchondral bone (SCB), and the cells and cytokines involved, to seek an answer to this question. RESULTS: SCB remodelling across OA changes the microstructure of the SCB, which alters the load-bearing properties of the joint and seems to have an important role in the initiation and progression of OA. Bone remodelling is tightly controlled by numerous cytokines, of which Receptor Activator of NFκB ligand (RANKL) and osteoprotegerin are central factors in almost all known bone conditions. In terms of finding therapeutic options for OA, an important question is whether controlling the rate of SCB remodelling would be beneficial. The role of RANKL in the pathogenesis and progression of OA and the effect of its neutralisation remain to be clarified. CONCLUSIONS: This review further makes the case for SCB remodelling as important in OA and for additional study of RANKL in OA, both its pathophysiological role and its potential as an OA disease target.

Microstructural and cellular characterisation of the subchondral trabecular bone in human knee and hip osteoarthritis using synchrotron tomography.

OBJECTIVE: It is unclear if different factors influence osteoarthritis (OA) progression and degenerative changes characterising OA disease in hip and knee. We investigated the difference between hip OA and knee OA at the subchondral bone (SCB) tissue and cellular level, relative to the degree of cartilage degeneration. DESIGN: Bone samples were collected from 11 patients (aged 70.4 ± 10.7years) undergoing knee arthroplasty and 8 patients (aged 62.3 ± 13.4years) undergoing hip arthroplasty surgery. Trabecular bone microstructure, osteocyte-lacunar network, and bone matrix vascularity were evaluated using synchrotron micro-CT imaging. Additionally, osteocyte density, viability, and connectivity were determined histologically. RESULTS: The associations between severe cartilage degeneration and increase of bone volume fraction (%) [- 8.7, 95% CI (-14.1, -3.4)], trabecular number (#/mm) [- 1.5, 95% CI (-0.8, -2.3)], osteocyte lacunar density (#/mm3) [4714.9; 95% CI (2079.1, 7350.6)] and decrease of trabecular separation (mm) [- 0.07, 95% CI (0.02, 0.1)] were found in both knee and hip OA. When compared to knee OA, hip OA was characterised by larger (µm3) but less spheric osteocyte lacunae [47.3; 95% CI (11.2, 83.4), - 0.04; 95% CI (-0.06, -0.02), respectively], lower vascular canal density (#/mm3) [- 22.8; 95% CI (-35.4, -10.3)], lower osteocyte cell density (#/mm2) [- 84.2; 95% CI (-102.5, -67.4)], and less senescent (#/mm2) but more apoptotic osteocytes (%) [- 2.4; 95% CI (-3.6, -1.2), 24.9; 95% CI (17.7, 32.1)], respectively. CONCLUSION: SCB from hip OA and knee OA exhibits different characteristics at the tissue and cellular levels, suggesting different mechanisms of OA progression in different joints.

A Mild Case of Autosomal Recessive Osteopetrosis Masquerading as the Dominant Form Involving Homozygous Deep Intronic Variations in the CLCN7 Gene.

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.

Hip osteoarthritis: A novel network analysis of subchondral trabecular bone structures.

Hip osteoarthritis (HOA) is a degenerative joint disease that leads to the progressive destruction of subchondral bone and cartilage at the hip joint. Development of effective treatments for HOA remains an open problem, primarily due to the lack of knowledge of its pathogenesis and a typically late-stage diagnosis. We describe a novel network analysis methodology for microcomputed tomography (micro-CT) images of human trabecular bone. We explored differences between the trabecular bone microstructure of femoral heads with and without HOA. Large-scale automated extraction of the network formed by trabecular bone revealed significant network properties not previously reported for bone. Profound differences were discovered, particularly in the proximal third of the femoral head, where HOA networks demonstrated elevated numbers of edges, vertices, and graph components. When further differentiating healthy joint and HOA networks, the latter showed fewer small-world network properties, due to decreased clustering coefficient and increased characteristic path length. Furthermore, we found that HOA networks had reduced length of edges, indicating the formation of compressed trabecular structures. In order to assess our network approach, we developed a deep learning model for classifying HOA and control cases, and we fed it with two separate inputs: (i) micro-CT images of the trabecular bone, and (ii) the network extracted from them. The model with plain micro-CT images achieves 74.6% overall accuracy while the trained model with extracted networks attains 96.5% accuracy. We anticipate our findings to be a starting point for a novel description of bone microstructure in HOA, by considering the phenomenon from a graph theory viewpoint.

Greater heterogeneity of the bone mineralisation density distribution and low bone matrix mineralisation characterise tibial subchondral bone marrow lesions in knee osteoarthritis patients.

Tibial subchondral bone marrow lesions (BMLs) identified by MRI have been recognised as potential disease predictors in knee osteoarthritis (KOA), and may associate with abnormal bone matrix mineralisation and reduced bone quality. However, these tissue-level changes of BMLs have not been extensively investigated. Thus, the aim of this study was to quantify the degree of subchondral bone matrix mineralisation (both plate and trabeculae) in relation to histomorphometric parameters of bone remodelling and osteocyte lacunae (OL) characteristics in the tibial plateau (TP) of KOA patients with and without BMLs (OA-BML and OA No-BML, respectively) in comparison to nonOA cadaveric controls (CTL). Osteochondral (cartilage-bone) tissue was sampled from the BML signal region within the medial compartment for each OA-BML TP, and from a corresponding medial region for OA No-BML and CTL TPs. The tissue samples were embedded in resin, and sections stained with Von-Kossa Haematoxylin and Eosin (H&E) for quantitation of static indices of bone remodelling. Resin blocks were then further polished, and carbon-coated for quantitative backscattered electron imaging (qBEI) to determine the bone mineralisation density distribution (BMDD), as well as OL characteristics. It was found that OA-BML contained higher osteoid volume per tissue volume (OV/TV; %) and per bone volume (OV/BV; %) in both subchondral plate and trabecular bone compared to OA No-BML and CTL. The BMDD of OA-BML in both subchondral plate and trabecular bone was shifted toward a lower degree of mineralisation. Typically, an increase in both the heterogeneity of mineralisation density (Ca Width; wt%Ca) and the percentage of lower calcium (Ca Low; % B.Ar) in trabecular bone with OA-BML versus CTL was observed. Further, unmineralised OL density (#/mm2) in subchondral plate was distinctly higher in OA-BML samples compared to CTL. The KOA patients with and without BMLs had significantly decreased density of mineralised OL (#/mm2) in trabecular bone compared to CTL. Taken together, these findings indicate that tibial BMLs in advanced KOA patients are characterised by significantly hypo-mineralised subchondral bone compared with CTL. These differences associated with evidence of increased bone remodelling in OA-BML, and may influence the mechanical properties of the subchondral bone, with implications for the overlying cartilage.

Raman microspectroscopy demonstrates reduced mineralization of subchondral bone marrow lesions in knee osteoarthritis patients.

INTRODUCTION: Bone marrow lesions (BMLs) are frequently identified by MRI in the subchondral bone in knee osteoarthritis (KOA). BMLs are known to be closely associated with joint pain, loss of the cartilage and structural changes in the subchondral trabecular bone (SCTB). Despite this, understanding of the nature of BMLs at the trabecular tissue level is incomplete. Thus, we used Raman microspectroscopy to examine the biochemical properties of SCTB from KOA patients with presence or absence of BMLs (OA-BML, OA No-BML; respectively), in comparison with age-matched cadaveric non-symptomatic controls (Non-OA CTL). METHODS: Tibial plateau (TP) specimens were collected from 19 KOA arthroplasty patients (6-Male, 13-Female; aged 56-74 years). BMLs were identified ex-vivo by MRI, using PDFS- and T1-weighted sequences. The KOA specimens were then categorized into an OA-BML group (n = 12; containing a BML within the medial condyle only) and an OA No-BML group (n = 7; with no BMLs identified in the TP). The control (CTL) group consisted of Non-OA cadaveric TP samples with no BMLs and no macroscopic or microscopic evidence of OA-related changes (n = 8; 5-Male, 3-Female; aged 44-80 years). Confocal Raman microspectroscopy, with high spatial resolution, was used to quantify the biochemical properties of SCTB tissue of both the medial and the lateral condyle in each group. RESULTS: The ratios of peak intensity and integrated area of bone matrix mineral (Phosphate (v1), Phosphate (v2) and Phosphate (v4)), to surrogates of the organic phase of bone matrix (Amide I, Proline and Amide III), were calculated. Within the medial compartment, the mineral:organic matrix ratios were significantly lower for OA-BML, compared to Non-OA CTL. These ratios were also significantly lower for the OA-BML medial compartment, compared to the OA-BML lateral compartment. There were no group or compartmental differences for Carbonate:Phosphate (v1, v2 and v4), Amide III (α-helix):Amide III (random-coil), Hydroxyproline:Proline, or Crystallinity. CONCLUSION: As measured by Raman microspectroscopy, SCTB tissue in BML zones in KOA is significantly less mineralized than the corresponding zones in individuals without OA. These data are consistent with those obtained using other methods (e.g. Fourier transform infrared spectroscopy; FTIR) and with the increased rate of bone remodeling observed in BML zones. Reduced mineralization may change the biomechanical properties of the trabecular bone in BMLs and the mechanical interaction between subchondral bone and its overlying cartilage, with potential implications for the development and progression of OA.

Evidence for Gender-Specific Bone Loss Mechanisms in Periprosthetic Osteolysis.

Osteolysis adjacent to total hip replacement (THR) prostheses is a major cause of their eventual failure. Periprosthetic osteolysis is associated with the production of bioactive particles, produced by the wear of articulating prosthesis surfaces. Wear particles invade the periprosthetic tissue, inducing inflammation and bone resorption. Previous studies have shown that osteocytes, the most numerous cell type in mineralised bone, can respond to wear particles of multiple orthopaedic material types. Osteocytes play important roles in bone resorption, regulating bone resorption by osteoclasts and directly through osteocytic osteolysis, also known as perilacunar remodelling. In this study, we perform a histological analysis of bone biopsies obtained from cohorts of male and female patients undergoing either primary THR surgery or revision THR surgery for aseptic loosening. The osteocyte lacunae area (Ot.Lac.Ar) and percentage lacunar area/bone area (%Ot.Lac.Ar/B.Ar) were significantly larger overall in revision THR bone than bone from similar sites in primary THR. Analysis by patient gender showed that increased Ot.Lac.Ar, indicative of increased perilacunar remodelling, was restricted to female revision samples. No significant differences in osteoclast parameters were detectable between the cohorts. These findings suggest previously unrecognised gender-specific mechanisms of bone loss in orthopaedic wear particle-induced osteolysis in humans.

Osteocytes respond to particles of clinically-relevant conventional and cross-linked polyethylene and metal alloys by up-regulation of resorptive and inflammatory pathways.

Periprosthetic osteolysis is a major cause of implant failure in total hip replacements. Aseptic loosening caused by osteolytic lesions is associated with the production of bioactive wear particles from the articulations of implants. Wear particles infiltrate the surrounding tissue of implants, promoting inflammation as well as bone resorption. Osteocytes have been shown to both regulate physiological osteoclastogenesis and directly remodel their perilacunar bone matrix by the process of osteocytic osteolysis. We hypothesise that osteocytes respond to wear debris of orthopaedic implant materials by adopting a pro-catabolic phenotype and thus contribute to periprosthetic osteolysis through the known pathways of bone loss. Osteocyte responses to particles derived from clinically relevant materials, ultra-high molecular weight polyethylene (UHMWPE), highly cross-linked polyethylene (XLPE) and metal alloys, Ti6Al4V and CoCrMo, were examined in vitro in human primary osteocyte-like cultures. Osteocyte-like cells exposed to both polyethylene and metal wear particle types showed upregulated expression of catabolic markers associated with osteocytic osteolysis, MMP13, carbonic anhydrase 2 (CA2) and cathepsin K (CTSK). In addition, pro-osteoclastogenesis markers RANKL and M-CSF were induced, as well as the expression of pro-inflammatory cytokines, IL-6 and TNFα, albeit with different kinetics. These findings suggest a previously unrecognised action of wear particles of multiple orthopaedic materials on osteocytes, and suggest a multifaceted role for osteocytes in periprosthetic osteolysis. STATEMENT OF SIGNIFICANCE: This study addresses periprosthetic osteolysis, a major clinical problem leading to aseptic loosening of orthopaedic implants. It is well accepted that wear particles of polyethylene and of other implant materials stimulate the activity of bone resorbing osteoclasts. Our recent work provided evidence that commercial particles of ultra-high molecular weight polyethylene (UHMWPE) stimulated osteocytes to adopt a bone catabolic state. In this study we demonstrate for the first time that particles derived from materials in clinical use, conventional UHMWPE, highly cross-linked polyethylene (XLPE), and Ti6Al4V and CoCrMo metal alloys, all stimulate human osteocyte activities of osteocyte-regulated osteoclastogenesis, osteocytic osteolysis, proinflammatory responses, osteocyte apoptosis, albeit to varying extents. This study provides further mechanistic insight into orthopaedic wear particle mediated bone disease in terms of the osteocyte, the most abundant and key controlling cell type in bone.

Novel Insights into Staphylococcus aureus Deep Bone Infections: the Involvement of Osteocytes.

Periprosthetic joint infection (PJI) is a potentially devastating complication of orthopedic joint replacement surgery. PJI with associated osteomyelitis is particularly problematic and difficult to cure. Whether viable osteocytes, the predominant cell type in mineralized bone tissue, have a role in these infections is not clear, although their involvement might contribute to the difficulty in detecting and clearing PJI. Here, using Staphylococcus aureus, the most common pathogen in PJI, we demonstrate intracellular infection of human-osteocyte-like cells in vitro and S. aureus adaptation by forming quasi-dormant small-colony variants (SCVs). Consistent patterns of host gene expression were observed between in vitro-infected osteocyte-like cultures, an ex vivo human bone infection model, and bone samples obtained from PJI patients. Finally, we confirm S. aureus infection of osteocytes in clinical cases of PJI. Our findings are consistent with osteocyte infection being a feature of human PJI and suggest that this cell type may provide a reservoir for silent or persistent infection. We suggest that elucidating the molecular/cellular mechanism(s) of osteocyte-bacterium interactions will contribute to better understanding of PJI and osteomyelitis, improved pathogen detection, and treatment.IMPORTANCE Periprosthetic joint infections (PJIs) are increasing and are recognized as one of the most common modes of failure of joint replacements. Osteomyelitis arising from PJI is challenging to treat and difficult to cure and increases patient mortality 5-fold. Staphylococcus aureus is the most common pathogen causing PJI. PJI can have subtle symptoms and lie dormant or go undiagnosed for many years, suggesting persistent bacterial infection. Osteocytes, the major bone cell type, reside in bony caves and tunnels, the lacuno-canalicular system. We report here that S. aureus can infect and reside in human osteocytes without causing cell death both experimentally and in bone samples from patients with PJI. We demonstrate that osteocytes respond to infection by the differential regulation of a large number of genes. S. aureus adapts during intracellular infection of osteocytes by adopting the quasi-dormant small-colony variant (SCV) lifestyle, which might contribute to persistent or silent infection. Our findings shed new light on the etiology of PJI and osteomyelitis in general.

Bone matrix microdamage and vascular changes characterize bone marrow lesions in the subchondral bone of knee osteoarthritis.

INTRODUCTION: Bone marrow lesions (BMLs) in the subchondral bone in osteoarthritis (OA) are suggested to be multifactorial, although the pathogenic mechanisms are unknown. Bone metabolism and cardiovascular risk factors associate with BML in epidemiologic studies. However, there are no studies at the tissue level investigating the relationship between these processes and BML. The aim of this study was to investigate the relationship between BMLs in the tibial plateau (TP) of knee OA and bone matrix microdamage, osteocyte density and vascular changes. METHODS: TP were obtained from 73 patients at total knee replacement surgery and BMLs were identified ex vivo in TP tissue using MRI. Comparator 'No BML' tissue was from matched anatomical sites to the BMLs. Quantitative assessment was made of subchondral bone microdamage, bone resorption indices, osteocyte cellularity, and vascular features. RESULTS: Several key parameters were different between BML and No BML tissue. These included increased microcrack burden (p = .01, p = .0001), which associated positively with bone resorption and negatively with cartilage volume, and greater osteocyte numerical density (p = .02, p = .01), in the subchondral bone plate and subchondral trabeculae, respectively. The marrow tissue within BML zones contained increased arteriolar density (p = .04, p = .0006), and altered vascular characteristics, in particular increased wall thickness (p = .007) and wall:lumen ratio (wall thickness over internal lumen area) (p = .001), compared with No BML bone. CONCLUSIONS: Increased bone matrix microdamage and altered vasculature in the subchondral bone of BMLs is consistent with overloading and vascular contributions to the formation of these lesions. Given the important role of BMLs in knee OA, these contributing factors offer potential targets for the treatment and prevention of knee OA.

Bone marrow lesions in osteoarthritis: What lies beneath.

Osteoarthritis (OA) is the most common joint disease in the United States, affecting more than 30 million people, and is characterized by cartilage degeneration in articulating joints. OA can be viewed as a group of overlapping disorders, which result in functional joint failure. However, the precise cellular and molecular events within which lead to these clinically observable changes are neither well understood nor easily measurable. It is now clear that multiple factors, in multiple joint tissues, contribute to degeneration. Changes in subchondral bone are recognized as a hallmark of OA, but are normally associated with late-stage disease when degeneration is well established. However, early changes such as Bone Marrow Lesions (BMLs) in OA are a relatively recent discovery. BMLs are patterns from magnetic resonance images (MRI) that have been linked with pain and cartilage degeneration. Their potential utility in predicting progression, or as a target for therapy, is not yet fully understood. Here, we will review the current state-of-the-art in this field under three broad headings: (i) BMLs in symptomatic OA: malalignment, joint pain, and disease progression; (ii) biological considerations for bone-cartilage crosstalk in joint disease; and (iii) mechanical factors that may underlie BMLs and drive their communication with other joint tissues. Thus, this review will provide insights on this topic from a clinical, biological, and mechanical perspective. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:1818-1825, 2018.

Both ligand and VDR expression levels critically determine the effect of 1α,25-dihydroxyvitamin-D3 on osteoblast differentiation.

Previous studies have shown that 1α,25-dihydroxyvitamin D3 (1,25D) through vitamin D receptor (VDR) signalling has both catabolic and anabolic effects on osteoblast differentiation. However, the mechanism of these differential effects by 1,25D is not fully understood. In this study, mice with three different genetic backgrounds, representing a normal VDR level (wild-type, WT), VDR over-expression specifically in mature osteoblasts (ObVDR-B6) and global VDR knockout (VDRKO), were utilised to generate primary osteoblast-like cultures to further elucidate the effects of 1,25D on osteoblast differentiation. Our data confirm the importance of VDR in the late stage of osteogenic differentiation and also for the expression of factors critical for osteoblastic support of osteoclast formation. This study also demonstrates the differential effects of a pharmacological level of 1,25D (1nM) on the expression of osteogenic differentiation markers, including Ocn and Sost, depending on the relative level of VDR. Our findings suggest that 1,25D plays an inhibitory role in matrix mineralisation, possibly through the modulation of the tissue non-specific alkaline phosphatase to ectonucleotide pyrophosphatase/phosphodiesterase 1 axis, in a VDR level-dependent manner. We conclude that the relative VDR level and the 1,25D availability to cells, are important co-determinants for whether 1,25D plays a promoting or suppressive role in osteoblast-mediated osteogenic activity.

Anticancer efficacy of the hypoxia-activated prodrug evofosfamide is enhanced in combination with proapoptotic receptor agonists against osteosarcoma.

Tumor hypoxia is a major cause of treatment failure for a variety of malignancies. However, hypoxia also leads to treatment opportunities as demonstrated by the development of compounds that target regions of hypoxia within tumors. Evofosfamide is a hypoxia-activated prodrug that is created by linking the hypoxia-seeking 2-nitroimidazole moiety to the cytotoxic bromo-isophosphoramide mustard (Br-IPM). When evofosfamide is delivered to hypoxic regions of tumors, the DNA cross-linking toxin, Br-IPM, is released leading to cell death. This study assessed the anticancer efficacy of evofosfamide in combination with the Proapoptotic Receptor Agonists (PARAs) dulanermin and drozitumab against human osteosarcoma in vitro and in an intratibial murine model of osteosarcoma. Under hypoxic conditions in vitro, evofosfamide cooperated with dulanermin and drozitumab, resulting in the potentiation of cytotoxicity to osteosarcoma cells. In contrast, under the same conditions, primary human osteoblasts were resistant to treatment. Animals transplanted with osteosarcoma cells directly into their tibiae developed mixed osteosclerotic/osteolytic bone lesions and consequently developed lung metastases 3 weeks post cancer cell transplantation. Tumor burden in the bone was reduced by evofosfamide treatment alone and in combination with drozitumab and prevented osteosarcoma-induced bone destruction while also reducing the growth of pulmonary metastases. These results suggest that evofosfamide may be an attractive therapeutic agent, with strong anticancer activity alone or in combination with either drozitumab or dulanermin against osteosarcoma.

Musculoskeletal Health in the Context of Spinal Cord Injury.

PURPOSE OF REVIEW: This review assembles recent understanding of the profound loss of muscle and bone in spinal cord injury (SCI). It is important to try to understand these changes, and the context in which they occur, because of their impact on the wellbeing of SC-injured individuals, and the urgent need for viable preventative therapies. RECENT FINDINGS: Recent research provides new understanding of the effects of age and systemic factors on the response of bone to loading, of relevance to attempts to provide load therapy for bone in SCI. The rapidly growing dataset describing the biochemical crosstalk between bone and muscle, and the cell and molecular biology of myokines signalling to bone and osteokines regulating muscle metabolism and mass, is reviewed. The ways in which this crosstalk may be altered in SCI is summarised. Therapeutic approaches to the catabolic changes in muscle and bone in SCI require a holistic understanding of their unique mechanical and biochemical context.

Postoperative weight bearing and patient reported outcomes at one year following tibial plateau fractures.

Tibial plateau fractures are complex and the current evidence for postoperative rehabilitation is weak, especially related to the recommended postoperative weight bearing. The primary aim of this study was to investigate if loading in the first 12 weeks of recovery is associated with patient reported outcome measures at 26 and 52 weeks postoperative. We hypothesized that there would be no association between loading and patient reported outcome measures. Seventeen patients, with a minimum of 52-week follow-up following fragment-specific open reduction and internal fixation for tibial plateau fracture, were selected for this retrospective analysis. Postoperatively, patients were advised to load their limb to a maximum of 20kg during the first 6 weeks. Loading data were collected during walking using force platforms. A ratio of limb loading (affected to unaffected) was calculated at 2, 6 and 12 weeks postoperative. Knee Injury and Osteoarthritis Scores were collected at 6, 12, 26 and 52 weeks postoperative. The association between loading ratios and patient reported outcomes were investigated. Compliance with weight bearing recommendations and changes in the patient reported outcome measures are described. Fracture reduction and migration were assessed on plain radiographs. No fractures demonstrated any measurable postoperative migration at 52 weeks. Significant improvements were seen in all patient reported outcome measures over the first 52 weeks, despite poor adherence to postoperative weight bearing restrictions. There were no associations between weight bearing ratio and patient reported outcomes at 52 weeks postoperative. Significant associations were identified between the loading ratio at 2 weeks and knee-related quality of life at six months (R2=0.392), and between the loading ratio at 6 weeks combined with injury severity and knee-related quality of life at 26 weeks (R2=0.441). In summary, weight bearing as tolerated does not negatively affect the results of tibial plateau fracture and may therefore be safe for postoperative management. These findings should be taken in context of the sample size, which was not sufficient for sub-group analysis to investigate the role of impaction grafting.

Peroxidase enzymes inhibit osteoclast differentiation and bone resorption.

Myeloperoxidase (MPO) and eosinophil peroxidase (EPO) are heme-containing enzymes, well known for their antimicrobial activity, are released in abundance by innate immune infiltrates at sites of inflammation and injury. We have discovered new and previously unrecognised roles for heme peroxidases in extracellular matrix biosynthesis, angiogenesis, and bone mineralisation, all of which play an essential role in skeletal integrity. In this study we used in vitro models of osteoclastogenesis to investigate the effects of heme peroxidase enzymes on osteoclast differentiation and bone resorbing activity, pertinent to skeletal development and remodelling. Receptor activator of nuclear factor kappa B-ligand (RANKL) stimulates the formation of tartate-resistant acid phosphatase (TRAP) positive multinucleated cells and increases bone resorption when cultured with human peripheral blood mononuclear cells (PBMCs) or the RAW264.7 murine monocytic cell line. When RANKL was added in combination with either MPO or EPO, a dose-dependent inhibition of osteoclast differentiation and bone resorption was observed. Notably, peroxidases had no effect on the bone resorbing activity of mature osteoclasts, suggesting that the inhibitory effect of the peroxidases was limited to osteoclast precursor cells. Mechanistically, we observed that osteoclast precursor cells readily internalize peroxidases, and inhibited the phosphorylation of JNK, p38 MAPK and ERK1/2, important signalling molecules central to osteoclastogenesis. Our findings suggest that peroxidase enzymes, like MPO and EPO, may play a fundamental role in inhibiting RANKL-induced osteoclast differentiation at inflammatory sites of bone fracture and injury. Therefore, peroxidase enzymes could be considered as potential therapeutic agents to treat osteolytic bone disease and aberrant bone resorption.

Anodized 3D-printed titanium implants with dual micro- and nano-scale topography promote interaction with human osteoblasts and osteocyte-like cells.

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.

Bone-cartilage crosstalk: a conversation for understanding osteoarthritis.

Although cartilage degradation is the characteristic feature of osteoarthritis (OA), it is now recognized that the whole joint is involved in the progression of OA. In particular, the interaction (crosstalk) between cartilage and subchondral bone is thought to be a central feature of this process. The interface between articular cartilage and bone of articulating long bones is a unique zone, which comprises articular cartilage, below which is the calcified cartilage sitting on and intercalated into the subchondral bone plate. Below the subchondral plate is the trabecular bone at the end of the respective long bones. In OA, there are well-described progressive destructive changes in the articular cartilage, which parallel characteristic changes in the underlying bone. This review examines the evidence that biochemical and biomechanical signaling between these tissue compartments is important in OA disease progression and asks whether such signaling might provide possibilities for therapeutic intervention to halt or slow disease development.

Drug-releasing nano-engineered titanium implants: therapeutic efficacy in 3D cell culture model, controlled release and stability.

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.

Titania nanotubes for orchestrating osteogenesis at the bone-implant interface.

Titanium implants can fail due to inappropriate biomechanics at the bone-implant interface that leads to suboptimal osseointegration. Titania nanotubes (TNTs) fabricated on Ti implants by the electrochemical process have emerged as a promising modification strategy to facilitate osseointegration. TNTs enable augmentation of bone cell functions at the bone-implant interface and can be tailored to incorporate multiple functionalities including the loading of active biomolecules into the nanotubes to target anabolic processes in bone conditions such as osteoporotic fractures. Advanced functions can be introduced, including biopolymers, nanoparticles and electrical stimulation to release growth factors in a desired manner. This review describes the application of TNTs for enhancing osteogenesis at the bone-implant interface, as an alternative approach to systemic delivery of therapeutic agents.