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Background and purpose — Displaced fractures of the talar neck are associated with a high risk of structural collapse. In this observational analysis we hypothesized that pharmacological inhibition of osteoclast function might reduce the risk of structural collapse through a reduction in bone resorption during revascularization of the injured bone. Patients and methods — Between 2002 and 2014 we treated 19 patients with displaced fractures of the talar neck with open reduction and internal fixation. Of these, 16 patients were available for final follow-up between January and November 2017 (median 12 years, IQR 7–13). Among these, 6 patients with Hawkins type 3 fractures and 2 patients with Hawkins type 2b fractures received postoperative antiresorptive treatment (7 alendronate, 1 denosumab) for 6 to 12 months. The remaining 8 patients received no antiresorptive treatment. The self-reported foot and ankle score (SEFAS) was available in all patients and 15 patients had undergone computed tomography (CT) at final follow-up, which allowed evaluation of structural collapse of the talar dome and signs of post-traumatic osteoarthritis. Results — The risk for partial collapse of the talar dome was equal in the 2 groups (3 in each group) and post-traumatic arthritis was observed in all patients. The SEFAS in patients with antiresorptive treatment was lower, at 21 points (95% CI 15–26), compared with those without treatment, 29 points (CI 22–35). Interpretation — Following a displaced fracture of the talar neck, we found no effect of antiresorptive therapy on the rate of talar collapse, post-traumatic osteoarthritis, and patient-reported outcomes.

Background and purpose — Teriparatide accelerates fracture healing in animals and probably in man. Abaloparatide is a new drug with similar although not identical effects on the teriparatide receptor. Given at 4 times the teriparatide dose in a human osteoporosis trial, abaloparatide increased bone density more than teriparatide, and both reduced fracture risk. We investigated in mice whether abaloparatide stimulates fracture healing, and if it does so with the suggested dose effect relation (4:1). Patients and methods — In a validated mouse model for metaphyseal healing (burr hole with screw pull-out), 96 mice were randomly allocated to 11 groups: control (saline), teriparatide or abaloparatide, where teriparatide and abaloparatide were given at 5 different doses each. In a femoral shaft osteotomy model, 24 mice were randomly allocated to 3 groups: control (saline), teriparatide (15 µg/kg) or abaloparatide (60 µg/kg). Each treatment was given daily via subcutaneous injections. Results were evaluated by mechanical testing and microCT. Results — In the metaphyseal model, a dose-dependent increase in screw pull-out force could be seen. In a linear regression analysis (r = 0.78) each increase in ln(dose) by 1 (regardless of drug type) was associated with an increase in pull-out force by 1.50 N (SE 0.18) (p < 0.001). Changing drug from teriparatide to abaloparatide increased the force by 1.41 N (SE 0.60; p = 0.02). In the diaphyseal model, the callus density was 23% (SD 10), 38% (SD 10), and 47% (SD 2) for control, for teriparatide and abaloparatide respectively. Both drugs were significantly different from controls (p = 0.001 and p = 0.008), but not from each other. Interpretation — Both drugs improve fracture healing, but in these mouse models, the potency per µg of abaloparatide seems only 2.5 times that of teriparatide, rather than the 4:1 relation chosen in the clinical abaloparatide–teriparatide comparison trial.

We have previously shown that changes in the microbiome influence how the healing tendon responds to different treatments. The aim of this study was to investigate if changes in the microbiome influence the response to mechanical loading during tendon healing. 90 Sprague-Dawley rats were used. Specific Opportunist and Pathogen Free (SOPF) rats were co-housed with Specific Pathogen Free (SPF) rats, carrying Staphylococcus aureus and other opportunistic microbes. After 6 weeks of co-housing, the SOPF rats were contaminated which was confirmed by Staphylococcus aureus growth. Clean SOPF rats were used as controls. The rats were randomized to full loading or partial unloading by Botox injections in their calf muscles followed by complete Achilles tendon transection. Eight days later, the healing tendons were tested mechanically. The results were analysed by a 2-way ANOVA with interaction between loading and contamination on peak force as the primary outcome and there was an interaction for both peak force (p = 0.049) and stiffness (p = 0.033). Furthermore, partial unloading had a profound effect on most outcome variables. In conclusion, the response to mechanical loading during tendon healing is influenced by changes in the microbiome. Studies aiming for clinical relevance should therefore consider the microbiome of laboratory animals.

Mechanical loading affects tendon healing and recovery. However, our understanding about how physical loading affects recovery of viscoelastic functions, collagen production and tissue organisation is limited. The objective of this study was to investigate how different magnitudes of loading affects biomechanical and collagen properties of healing Achilles tendons over time. Achilles tendon from female Sprague Dawley rats were cut transversely and divided into two groups; normal loading (control) and reduced loading by Botox (unloading). The rats were sacrificed at 1, 2- and 4-weeks post-injury and mechanical testing (creep test and load to failure), small angle x-ray scattering (SAXS) and histological analysis were performed. The effect of unloading was primarily seen at the early time points, with inferior mechanical and collagen properties (SAXS), and reduced histological maturation of the tissue in unloaded compared to loaded tendons. However, by 4 weeks no differences remained. SAXS and histology revealed heterogeneous tissue maturation with more mature tissue at the peripheral region compared to the center of the callus. Thus, mechanical loading advances Achilles tendon biomechanical and collagen properties earlier compared to unloaded tendons, and the spatial variation in tissue maturation and collagen organization across the callus suggests important regional (mechano-) biological activities that require more investigation.

Background and purpose — Fracture healing in the shaft is usually described as a sequence of events, starting with inflammation, which triggers mesenchymal tissue formation in successive steps. Most clinical fractures engage cancellous bone. We here describe fracture healing in cancellous bone, focusing on the timing of inflammatory and mesenchymal cell type appearance at the site of injury Material and methods — Rats received a proximal tibial drill hole. A subgroup received clodronate-containing liposomes before or after surgery. The tibiae were analyzed with micro-CT and immunohistochemistry 1 to 7 days after injury. Results — Granulocytes (myeloperoxidase) appeared in moderate numbers within the hole at day 1 and then gradually disappeared. Macrophage expression (CD68) was seen on day 1, increased until day 3, and then decreased. Mesenchymal cells (vimentin) had already accumulated in the periphery of the hole on day 1. Mesenchymal cells dominated in the entire lesion on day 3, now producing extracellular matrix. A modest number of preosteoblasts (RUNX2) were seen on day 1 and peaked on day 4. Osteoid was seen on day 4 in the traumatized region, with a distinct border to the uninjured surrounding marrow. Clodronate liposomes given before the injury reduced the volume of bone formation at day 7, but no reduction in macrophage numbers could be detected. Interpretation — The typical sequence of events in shaft fractures was not seen. Mesenchymal cells appeared simultaneously with granulocyte and macrophage arrival. Clodronate liposomes, known to reduce macrophage numbers, seemed to be associated with the delineation of the volume of tissue to be replaced by bone. Most fracture healing studies in animal models concern cortical bone in shafts. However, most fractures in patients occur in cancellous bone in the metaphysis, such as the distal radius or in the vertebrae. A growing body of evidence suggests that there are important differences between the healing processes in cortical and cancellous bone.

Background and purpose — [18F]Fluoride ([18F]NaF) PET scan is frequently used for estimation of bone healing rate and extent in cases of bone allografting and fracture healing. Some authors claim that [18F]NaF uptake is a measure of osteoblastic activity, calcium metabolism, or bone turnover. Based on the known affinity of fluoride to hydroxyapatite, we challenged this view. Methods — 10 male rats received crushed, frozen allogeneic cortical bone fragments in a pouch in the abdominal wall on the right side, and hydroxyapatite granules on left side. [18F]NaF was injected intravenously after 7 days. 60 minutes later, the rats were killed and [18F]NaF uptake was visualized in a PET/CT scanner. Specimens were retrieved for micro CT and histology. Results — MicroCT and histology showed no signs of new bone at the implant sites. Still, the implants showed a very high [18F]NaF uptake, on a par with the most actively growing and remodeling sites around the knee joint. Interpretation — [18F]NaF binds with high affinity to dead bone and calcium phosphate materials. Hence, an [18F]NaF PET/CT scan does not allow for sound conclusions about new bone ingrowth into bone allograft, healing activity in long bone shaft fractures with necrotic fragments, or remodeling around calcium phosphate coated prostheses

Background and purpose — Healing of shaft fractures is commonly described as regards external callus. We wanted to clarify the role of the bone marrow compartment in the healing of stable shaft fractures. Patients and methods — A longitudinal furrow was milled along the longitudinal axis of the femoral shaft in mice. The exposed bone marrow under the furrow was scooped out. The mice were then randomized to no further treatment, or to receiving 2 silicone plugs in the medullary canal distal and proximal to the defect. The plugs isolated the remaining marrow from contact with the defect. Results were studied with histology and flow cytometry. Results — Without silicone plugs, the marrow defect was filled with new bone marrow-like tissue by day 5, and new bone was seen already on day 10. The new bone was seen only at the level of the cortical injury, where it seemed to form simultaneously in the entire region of the removed cortex. The new bone seemed not to invade the marrow compartment, and there was a sharp edge between new bone and marrow. The regenerated marrow was similar to uninjured marrow, but contained considerably more cells. In the specimens with plugs, the marrow compartment was either filled with loose scar tissue, or empty, and there was only minimal bone formation, mainly located around the edges of the cortical injury. Interpretation — Marrow regeneration in the defect seemed to be a prerequisite for normal cortical healing. Shaft fracture treatment should perhaps pay more attention to the local bone marrow.

Wear debris particles released from prosthetic bearing surfaces and mechanical instability of implants are two main causes of periprosthetic osteolysis. While particle-induced loosening has been studied extensively, mechanisms through which mechanical factors lead to implant loosening have been less investigated. This study compares the transcriptional profiles associated with osteolysis in a rat model for aseptic loosening, induced by either mechanical instability or titanium particles. Rats were exposed to mechanical instability or titanium particles. After 15 min, 3, 48 or 120 h from start of the stimulation, gene expression changes in periprosthetic bone tissue was determined by microarray analysis. Microarray data were analyzed by PANTHER Gene List Analysis tool and Ingenuity Pathway Analysis (IPA). Both types of osteolytic stimulation led to gene regulation in comparison to unstimulated controls after 3, 48 or 120 h. However, when mechanical instability was compared to titanium particles, no gene showed a statistically significant difference (fold change ≥ ±1.5 and adjusted p-value ≤ 0.05) at any time point. There was a remarkable similarity in numbers and functional classification of regulated genes. Pathway analysis showed several inflammatory pathways activated by both stimuli, including Acute Phase Response signaling, IL-6 signaling and Oncostatin M signaling. Quantitative PCR confirmed the changes in expression of key genes involved in osteolysis observed by global transcriptomics. Inflammatory mediators including interleukin (IL)-6, IL-1β, chemokine (C-C motif) ligand (CCL)2, prostaglandin-endoperoxide synthase (Ptgs)2 and leukemia inhibitory factor (LIF) showed strong upregulation, as assessed by both microarray and qPCR. By investigating genome-wide expression changes we show that, despite the different nature of mechanical implant instability and titanium particles, osteolysis seems to be induced through similar biological and signaling pathways in this rat model for aseptic loosening. Pathways associated to the innate inflammatory response appear to be a major driver for osteolysis. Our findings implicate early restriction of inflammation to be critical to prevent or mitigate osteolysis and aseptic loosening of orthopedic implants.

Background and purpose — There is increasing evidence that several commonly performed surgical procedures provide little advantage over nonoperative treatment, suggesting that doctors may sometimes be inappropriately optimistic about surgical benefit when suggesting treatment for individual patients. We investigated whether attitudes to risk influenced the choice of operative treatment and nonoperative treatment. Methods — 946 Swedish orthopedic surgeons were invited to participate in an online survey. A radiograph of a 4-fragment proximal humeral fracture was presented together with 5 different patient characteristics, and the surgeons could choose between 3 different operative treatments and 1 nonoperative treatment. This was followed by an economic risk-preference test, and then by an instrument designed to measure 6 attitudes to surgery that are thought to be hazardous. We then investigated if choice of non-operative treatment was associated with risk aversion, and thereafter with the other variables, by regression analysis. Results — 388 surgeons responded. Nonoperative treatment for all cases was suggested by 64 of them. There was no significant association between risk aversion and tendency to avoid surgery. However, there was a statistically significant association between suggesting to operate at least 1 of the cases and a "macho" attitude to surgery or resignation regarding the chances of influencing the outcome of surgery. Choosing nonoperative treatment for all cases was associated with long experience as a surgeon. Interpretation — The discrepancy between available evidence for surgery and clinical practice does not appear to be related to risk preference, but relates to hazardous attitudes. It appears that choosing nonoperative treatment requires experience and a feeling that one can make a difference (i.e. a low score for resignation). There is a need for better awareness of available evidence for surgical indications.

Background and purpose — In 3 papers in Acta Orthopaedica 10 years ago, we described that platelet-rich plasma (PRP) improves tendon healing in a rat Achilles transection model. Later, we found that microtrauma has similar effects, probably acting via inflammation. This raised the suspicion that the effect ascribed to growth factors within PRP could instead be due to unspecific influences on inflammation. While testing this hypothesis, we noted that the effect seemed to be related to the microbiota. Material and methods — We tried to reproduce our old findings with local injection of PRP 6 h after tendon transection, followed by mechanical testing after 11 days. This failed. After fruitless variations in PRP production protocols, leukocyte concentration, and physical activity, we finally tried rats carrying potentially pathogenic bacteria. In all, 242 rats were used. Results — In 4 consecutive experiments on pathogen-free rats, no effect of PRP on healing was found. In contrast, apparently healthy rats carrying Staphylococcus aureus showed increased strength of the healing tendon after PRP treatment. These rats had higher levels of cytotoxic T-cells in their spleens. Interpretation — The failure to reproduce older experiments in clean rats was striking, and the difference in response between these and Staphylococcus-carrying rats suggests that the PRP effect is dependent on the immune status. PRP functions may be more complex than just the release of growth factors. Extrapolation from our previous findings with PRP to the situation in humans therefore becomes even more uncertain.

Background and purpose — Aseptic loosening is a main cause of late revision in total knee replacement (TKR). Teriparatide, a recombinant parathyroid hormone (PTH), stimulates osteoblasts and has been suggested to improve cancellous bone healing in humans. This might also be relevant for prosthesis fixation. We used radiostereometric analysis (RSA) to investigate whether teriparatide influences prosthesis fixation. Early migration as measured by RSA can predict future loosening. Patients and methods — In a randomized controlled trial with blind evaluation, 50 patients with osteoarthritis of the knee were allocated to a teriparatide treatment group (Forsteo, 20 μg daily for 2 months postoperatively) or to an untreated control group. RSA was performed postoperatively and at 6 months, 12 months, and 24 months. The primary effect variable was maximal total point motion (MTPM) from 12 to 24 months. Results — Median maximal total point motion from 12 to 24 months was similar in the 2 groups (teriparatide: 0.14 mm, 10% and 90% percentiles: 0.08 and 0.24; control: 0.13 mm, 10% and 90% percentiles: 0.09 and 0.21). [Authors: this is perhaps better than using "10th" and "90th", which looks ugly in print./language editor] The 95% confidence interval for the difference between group means was −0.03 to 0.04 mm, indicating that no difference occurred. Interpretation — We found no effect of teriparatide on migration in total knee replacement. Other trials using the same dosing have suggested a positive effect of teriparatide on human cancellous fracture healing. Thus, the lack of effect on migration may have been due to something other than the dose. In a similar study in this issue of Acta Orthopaedica, we found that migration could be reduced with denosumab (Ledin et al. ). The difference in response between the anabolic substance teriparatide and the antiresorptive denosumab suggests that resorption has a more important role during the postoperative course than any deficit in bone formation.

Background and purpose — Aseptic loosening is a main cause of late revision in total knee replacement (TKR). Migration of implants as measured by radiostereometric analysis (RSA) can predict future loosening. This migration is associated with bone resorption. Denosumab is a human monoclonal antibody that binds to receptors on osteoclast precursors and osteoclasts. This prevents osteoclast formation, resulting in less bone resorption in cortical and trabecular bone. We investigated whether denosumab can reduce migration of TKR, as measured with RSA. Patients and methods — In this 2-center, randomized, double-blind placebo-controlled trial, 50 patients with osteoarthritis of the knee were treated with an injection of either denosumab (60 mg) or placebo 1 day after knee replacement surgery and again after 6 months. RSA was performed postoperatively and after 6, 12, and 24 months. The primary effect variable was RSA maximal total point motion (MTPM) after 12 months. We also measured other RSA variables and the knee osteoarthritis outcome score (KOOS). Results — The primary effect variable, MTPM after 12 months, showed that migration in the denosumab group was statistically significantly less than in the controls. Denosumab MTPM 12 months was reduced by one-third (denosumab: median 0.24 mm, 10% and 90% percentiles: 0.15 and 0.41; placebo: median 0.36 mm, 10% and 90% percentiles: 0.20 and 0.62). The secondary MTPM variables (6 and 24 months) also showed a statistically significant reduction in migration. There was no significant difference in MTPM for the period 12–24 months. KOOS sub-variables were similiar between denosumab and placebo after 12 and 24 months. Interpretation — Denosumab reduces early migration in total knee replacement, as in previous trials using bisphosphonates. As migration is related to the risk of late loosening, denosumab may be beneficial for long-term results.

Background and purpose — Fracture healing involves different inflammatory cells, some of which are not part of the traditional bone field, such as B-cells and cytotoxic T-cells. We wanted to characterize bone healing by flow cytometry using 15 different inflammatory cell markers in a mouse model of metaphyseal injury, and incidentally discovered a previously unknown general skeletal reaction to trauma. Material and methods — A bent needle was inserted and twisted to traumatize the cancellous bone in the proximal tibia of C57/Bl6 female mice. This is known to induce vivid bone formation locally in the marrow compartment. Cells were harvested from the injured region, the uninjured contralateral tibia, and the humerus. The compositions of the immune cell populations were compared to those in untraumatized control animals. Results — Tibial metaphyseal injury led to substantial changes in the cell populations over time. Unexpectedly, similar changes were also seen in the contralateral tibia and in the humerus, despite the lack of local trauma. Most leukocyte subsets were affected by this generalized reaction. Interpretation — A relatively small degree of injury to the proximal tibia led to systemic changes in the immune cell populations in the marrow of unrelated bones, and probably in the entire skeleton. The few changes that were specific for the injury site appeared to relate to modulatory functions.

Background and purpose — Studies of fracture healing have mainly dealt with shaft fractures, both experimentally and clinically. In contrast, most patients have metaphyseal fractures. There is an increasing awareness that metaphyseal fractures heal partly through mechanisms specific to cancellous bone. Several new models for the study of cancellous bone healing have recently been presented. This review summarizes our current knowledge of cancellous fracture healing. Methods — We performed a review of the literature after doing a systematic literature search. Results — Cancellous bone appears to heal mainly via direct, membranous bone formation that occurs freely in the marrow, probably mostly arising from local stem cells. This mechanism appears to be specific for cancellous bone, and could be named inter-trabecular bone formation. This kind of bone formation is spatially restricted and does not extend more than a few mm outside the injured region. Usually no cartilage is seen, although external callus and cartilage formation can be induced in meta­physeal fractures by mechanical instability. Inter-trabecular bone formation seems to be less sensitive to anti-inflammatory treatment than shaft fractures. Interpretation — The unique characteristics of inter-trabecular bone formation in metaphyseal fractures can lead to differences from shaft healing regarding the effects of age, loading, or drug treatment. This casts doubt on generalizations about fracture healing based solely on shaft fracture models.

BACKGROUND:Mechanical loading stimulates Achilles tendon healing. However, various degrees of loading appear to have different effects on the mechanical properties of the healing tendon, and strong loading might create microdamage in the tissue. This suggests that different mechanisms might be activated depending on the magnitude of loading. The aim of this study was to investigate these mechanisms further. METHODS:Female rats had their right Achilles tendon cut transversely and divided into three groups: 1) unloading (calf muscle paralysis by Botox injections, combined with joint fixation by a steel-orthosis), 2) mild loading (Botox only), 3) strong loading (free cage activity). Gene expression was analyzed by PCR, 5 days post-injury, and mechanical testing 8 days post-injury. The occurrence of microdamage was analyzed 3, 5, or 14 days post-injury, by measuring leakage of injected fluorescence-labelled albumin in the healing tendon tissue. RESULTS:Peak force, peak stress, and elastic modulus of the healing tendons gradually improved with increased loading as well as the expression of extracellular matrix genes. In contrast, only strong loading increased transverse area and affected inflammation genes. Strong loading led to higher fluorescence (as a sign of microdamage) compared to mild loading at 3 and 5 days post-injury, but not at 14 days. DISCUSSION:Our results show that strong loading improves both the quality and quantity of the healing tendon, while mild loading only improves the quality. Strong loading also induces microdamage and alters the inflammatory response. This suggests that mild loading exert its effect via mechanotransduction mechanisms, while strong loading exert its effect both via mechanotransduction and the creation of microdamage. CONCLUSION:In conclusion, mild loading is enough to increase the quality of the healing tendon without inducing microdamage and alter the inflammation in the tissue. This supports the general conception that early mobilization of a ruptured tendon in patients is advantageous.

Computational models of Achilles tendons can help understanding how healthy tendons are affected by repetitive loading and how the different tissue constituents contribute to the tendon's biomechanical response. However, available models of Achilles tendon are limited in their description of the hierarchical multi-structural composition of the tissue. This study hypothesised that a poroviscoelastic fibre-reinforced model, previously successful in capturing cartilage biomechanical behaviour, can depict the biomechanical behaviour of the rat Achilles tendon found experimentally.We developed a new material model of the Achilles tendon, which considers the tendon's main constituents namely: water, proteoglycan matrix and collagen fibres. A hyperelastic formulation of the proteoglycan matrix enabled computations of large deformations of the tendon, and collagen fibres were modelled as viscoelastic. Specimen-specific finite element models were created of 9 rat Achilles tendons from an animal experiment and simulations were carried out following a repetitive tensile loading protocol. The material model parameters were calibrated against data from the rats by minimising the root mean squared error (RMS) between experimental force data and model output.All specimen models were successfully fitted to experimental data with high accuracy (RMS 0.42-1.02). Additional simulations predicted more compliant and soft tendon behaviour at reduced strain-rates compared to higher strain-rates that produce a stiff and brittle tendon response. Stress-relaxation simulations exhibited strain-dependent stress-relaxation behaviour where larger strains produced slower relaxation rates compared to smaller strain levels. Our simulations showed that the collagen fibres in the Achilles tendon are the main load-bearing component during tensile loading, where the orientation of the collagen fibres plays an important role for the tendon's viscoelastic response. In conclusion, this model can capture the repetitive loading and unloading behaviour of intact and healthy Achilles tendons, which is a critical first step towards understanding tendon homeostasis and function as this biomechanical response changes in diseased tendons.