The aim of this study was to investigate the occurrence of tissue hypoxia and apoptosis at different stages of tendinopathy and tears of the rotator cuff. We studied tissue from 24 patients with eight graded stages of either impingement (mild, moderate and severe) or tears of the rotator cuff (partial, small, medium, large and massive) and three controls. Biopsies were analysed using three immunohistochemical techniques, namely antibodies against HIF-1α (a transcription factor produced in a hypoxic environment), BNip3 (a HIF-1α regulated pro-apoptotic protein) and TUNEL (detecting DNA fragmentation in apoptosis). The HIF-1α expression was greatest in mild impingement and in partial, small, medium and large tears. BNip3 expression increased significantly in partial, small, medium and large tears but was reduced in massive tears. Apoptosis was increased in small, medium, large and massive tears but not in partial tears. These findings reveal evidence of hypoxic damage throughout the spectrum of pathology of the rotator cuff which may contribute to loss of cells by apoptosis. This provides a novel insight into the causes of degeneration of the rotator cuff and highlights possible options for treatment

Post-traumatic arthritis is a frequent consequence of articular fracture. The mechanisms leading to its development after such injuries have not been clearly delineated. A potential contributing factor is decreased viability of the articular chondrocytes. The object of this study was to characterise the regional variation in the viability of chondrocytes following joint trauma. A total of 29 osteochondral fragments from traumatic injuries to joints that could not be used in articular reconstruction were analysed for cell viability using the fluorescence live/dead assay and for apoptosis employing the TUNEL assay, and compared with cadaver control fragments. Chondrocyte death and apoptosis were significantly greater along the edge of the fracture and in the superficial zone of the osteochondral fragments. The middle and deep zones demonstrated significantly higher viability of the chondrocytes. These findings indicate the presence of both necrotic and apoptotic chondrocytes after joint injury and may provide further insight into the role of chondrocyte death in post-traumatic arthritis

The role of inflammatory cells and their products in tendinopathy is not completely understood. Pro-inflammatory cytokines are upregulated after oxidative and other forms of stress. Based on observations that increased cytokine expression has been demonstrated in cyclically-loaded tendon cells we hypothesised that because of their role in oxidative stress and apoptosis, pro-inflammatory cytokines may be present in rodent and human models of tendinopathy. A rat supraspinatus tendinopathy model produced by running overuse was investigated at the genetic level by custom micro-arrays. Additionally, samples of torn supraspinatus tendon and matched intact subscapularis tendon were collected from patients undergoing arthroscopic shoulder surgery for rotator-cuff tears and control samples of subscapularis tendon from ten patients with normal rotator cuffs undergoing arthroscopic stabilisation of the shoulder were also obtained. These were all evaluated using semiquantitative reverse transcription polymerase chain-reaction and immunohistochemistry. We identified significant upregulation of pro-inflammatory cytokines and apoptotic genes in the rodent model (p = 0.005). We further confirmed significantly increased levels of cytokine and apoptotic genes in human supraspinatus and subscapularis tendon harvested from patients with rotator cuff tears (p = 0.0008). These findings suggest that pro-inflammatory cytokines may play a role in tendinopathy and may provide a target for preventing tendinopathies

Chondrocytes at the lower zone of the growth plate must be eliminated to facilitate longitudinal growth; this is generally assumed to involve apoptosis. We attempted to provide definitive electron-microscopic evidence of apoptosis in chondrocytes of physes and chondroepiphyses in the rabbit. We were, however, unable to find a single chondrocyte with the ultrastructure of ‘classical’ apoptosis in vivo, although such a cell was found in vitro. Instead, condensed chondrocytes had a convoluted nucleus with patchy chromatin condensations while the cytoplasm was dark with excessive amounts of endoplasmic reticulum. These cells were termed ‘dark chondrocytes’. A detailed study of their ultrastructure combined with localisation methods in situ suggested a different mechanism of programmed cell death. In addition, another type of death was identified among the immature chondrocytes of the chondroepiphysis. These cells had the same nucleus as dark chondrocytes, but the lumen of the endoplasmic reticulum had expanded to fill the entire non-nuclear space, and all cytoplasm and organelles had been reduced to dark, worm-like inclusions. Since these cells appeared to be ‘in limbo’, they were termed ‘paralysed’ cells. It is proposed that ‘dark chondrocytes’ and ‘paralysed cells’ are examples of physiological cell death which does not involve apoptosis. It is possible that the confinement of chondrocytes within their lacunae, which would prevent phagocytosis of apoptotic bodies, necessitates different mechanisms of elimination

Systemic factors are believed to be pivotal for the development of heterotopic ossification in severely-injured patients. In this study, cell cultures of putative target cells (human fibroblastic cells, osteoblastic cells (MG-63), and bone-marrow stromal cells (hBM)) were incubated with serum from ten consecutive polytraumatised patients taken from post-traumatic day 1 to day 21 and with serum from 12 healthy control subjects. The serum from the polytraumatised patients significantly stimulated the proliferation of fibroblasts, MG-63 and of hBM cells. The activity of alkaline phosphatase in MG-63 and hBM cells was significantly decreased when exposed to the serum of the severely-injured patient. After three weeks in 3D cell cultures, matrix production and osteogenic gene expression of hBM cells were equal in the patient and control groups. However, the serum from the polytraumatised patients significantly decreased apoptosis of hBM cells compared with the control serum (4.3% vs 19.1%, p = 0.031). Increased proliferation of osteoblastic cells and reduced apoptosis of osteoprogenitors may be responsible for increased osteogenesis in severely-injured patients

We attempted to repair full-thickness defects in the articular cartilage of the trochlear groove of the femur in 30 rabbit knee joints using allogenic cultured chondrocytes embedded in a collagen gel. The repaired tissues were examined at 2, 4, 8, 12 and 24 weeks after operation using histological and histochemical methods. The articular defect filling index measurement was derived from safranin-O stained sections. Apoptotic cellular fractions were derived from analysis of apoptosis in situ using TUNEL staining, and was confirmed using caspase-3 staining along with quantification of the total cellularity. The mean articular defect filling index decreased with time. After 24 weeks it was 0.7 (. sd. 0.10), which was significantly lower than the measurements obtained earlier (p < 0.01). The highest mean percentage of apoptotic cells were observed at 12 weeks, although the total cellularity decreased with time. Because apoptotic cell death may play a role in delamination after chondrocyte transplantation, anti-apoptotic gene therapy may protect transplanted chondrocytes from apoptosis

We stably transfected early passage chondrocytes with an anti-apoptotic Bcl-2 gene in vitro using a retrovirus vector. Samples of articular cartilage were obtained from 11 patients with a mean age of 69 years (61 to 75) who were undergoing total knee replacement for osteoarthritis. The Bcl-2-gene-transfected chondrocytes were compared with non-transfected and lac-Z-gene-transfected chondrocytes, both of which were used as controls. All three groups of cultured chondrocytes were incubated with nitric oxide (NO) for ten days. Using the Trypan Blue exclusion assay, an enzyme-linked immunosorbent assay and flow cytometric analysis, we found that the number of apoptotic chondrocytes was significantly higher in the non-transfected and lac-Z-transfected groups than in the Bcl-2-transfected group (p < 0.05). The Bcl-2-transfected chondrocytes were protected from NO-induced impairment of proteoglycan synthesis.

We conclude that NO-induced chondrocyte death involves a mechanism which appears to be subject to regulation by an anti-apoptotic Bcl-2 gene. Therefore, Bcl-2 gene therapy may prove to be of therapeutic value in protecting human articular chondrocytes.

Osteonecrosis of the femoral head usually affects young individuals and is responsible for up to 12% of total hip arthroplasties. The underlying pathophysiology of the death of the bone cells remains uncertain. We have investigated nitric oxide mediated apoptosis as a potential mechanism and found that steroid- and alcohol-induced osteonecrosis is accompanied by widespread apoptosis of osteoblasts and osteocytes. Certain drugs or their metabolites may have a direct cytotoxic effect on cancellous bone of the femoral head leading to apoptosis rather than purely necrosis

Sex hormones play important roles in the regulation of the proliferation, maturation and death of chondrocytes in the epiphyseal growth plate. We have investigated the effects of male castration on the cell kinetics of chondrocytes as defined by the numbers of proliferating and dying cells. The growth plates of normal rabbits and animals castrated at eight weeks of age were obtained at 10, 15, 20 and 25 weeks of age. Our study suggested that castration led to an increase in apoptosis and a decrease in the proliferation of chondrocytes in the growth plate. In addition, the number of chondrocytes in the castrated rabbits was less than that of normal animals of the same age

Chondrocytes of the growth plate are generally assumed to undergo apoptosis, but the mechanisms which induce this cell death are not known. The Fas receptor is a mediator of the apoptotic signal in some systems. We studied its expression in situ in growth plates of rabbits aged from five to 20 weeks. In addition, we investigated the immunolocalisation in the growth plates of the bone proteins, osteonectin and osteocalcin, and the changes in their expression with age. The Fas-positive chondrocytes were found mostly in the hypertrophic zone, as were the osteonectin-positive and osteocalcin-positive cells. The percentage of Fas-positive cells increased with age whereas little change was found in the number of osteonectin-positive and osteocalcin-positive chondrocytes. Many of the Fas-positive chondrocytes were also TUNEL-positive. This strongly suggests that apoptosis in the growth plate is mediated through the Fas system. Double immunostaining for osteocalcin and Fas showed that not all hypertrophic chondrocytes were of the same cell type. Some chondrocytes stained for osteocalcin only, others for Fas only, while some were positive for both

Aims

Arthroscopic microfracture is a conventional form of treatment for patients with osteochondritis of the talus, involving an area of < 1.5 cm². However, some patients have persistent pain and limitation of movement in the early postoperative period. No studies have investigated the combined treatment of microfracture and shortwave treatment in these patients. The aim of this prospective single-centre, randomized, double-blind, placebo-controlled trial was to compare the outcome in patients treated with arthroscopic microfracture combined with radial extracorporeal shockwave therapy (rESWT) and arthroscopic microfracture alone, in patients with ostechondritis of the talus.

The growth plates of the femoral head of Japanese white rabbits aged 5, 10, 15 and 20 weeks were stained for apoptotic and proliferating chondrocytes using the TUNEL and PCNA antibody staining techniques. Both TUNEL- and PCNA-positive chondrocytes were detected in all of the specimens. The positive ratios of both stainings were calculated for the whole plate and for the resting, proliferating and hypertrophic zones. The highest ratios in both stainings occurred in the hypertrophic zone in all age groups. With growth, the TUNEL-positive ratio increased whereas the proliferating ratio decreased. We suggest that the increase in chondrocytic death by apoptosis and the decrease in cell proliferation potential led to closure of the growth plate

Using in situ hybridisation and the terminal deoxynucleotidyl transferase-mediated biotin-dUTP nick end-labelling (TUNEL) reaction in rats with osteonecrosis of the femoral head we have studied the effect of ischaemia on the gene expression of the stress proteins oxygen-regulated protein 150 (ORP150) and haemoxygenase 1 (HO1) and the death mechanism of the cells involved in osteonecrosis. Both ORP150 and HO1 have been reported to have important roles in the successful adaptation to oxygen deprivation. ORP150 and HO1 mRNA expression was induced by ischaemia in osteoblasts and osteocytes. In proliferative chondrocytes, these signals were detected constitutively. During the development of ischaemic osteonecrosis, the mechanism of cell death was apoptosis as indicated by DNA fragmentation and the presence of apoptotic bodies in osteocytes, chondrocytes and bone-marrow cells. After the initial ischaemic event, expression of ORP150 and HO1 mRNA, the TUNEL-positive reaction and empty lacunae were found sequentially. These findings were exclusive and may be considered to be markers for each stage in the development of osteonecrosis

Aims

A limited number of investigations with conflicting results have described perivascular lymphocytic infiltration (PVLI) in the setting of total knee arthroplasty (TKA). The purpose of this study was to determine if PVLI found in TKAs at the time of aseptic revision surgery was associated with worse clinical outcomes and survivorship.

Aims

Currently, periprosthetic fractures are excluded from the American Society for Bone and Mineral Research (ASBMR) definition of atypical femoral fracture (AFFs). This study aims to report on a series of periprosthetic femoral fractures (PFFs) that otherwise meet the criteria for AFFs. Secondary aims were to identify predictors of periprosthetic atypical femoral fractures (PAFFs) and quantify the complications of treatment.

Stem cells are defined by their potential for self-renewal and the ability to differentiate into numerous cell types, including cartilage and bone cells. Although basic laboratory studies demonstrate that cell therapies have strong potential for improvement in tissue healing and regeneration, there is little evidence in the scientific literature for many of the available cell formulations that are currently offered to patients. Numerous commercial entities and ‘regenerative medicine centres’ have aggressively marketed unproven cell therapies for a wide range of medical conditions, leading to sometimes indiscriminate use of these treatments, which has added to the confusion and unpredictable outcomes. The significant variability and heterogeneity in cell formulations between different individuals makes it difficult to draw conclusions about efficacy. The ‘minimally manipulated’ preparations derived from bone marrow and adipose tissue that are currently used differ substantially from cells that are processed and prepared under defined laboratory protocols. The term ‘stem cells’ should be reserved for laboratory-purified, culture-expanded cells. The number of cells in uncultured preparations that meet these defined criteria is estimated to be approximately one in 10 000 to 20 000 (0.005% to 0.01%) in native bone marrow and 1 in 2000 in adipose tissue. It is clear that more refined definitions of stem cells are required, as the lumping together of widely diverse progenitor cell types under the umbrella term ‘mesenchymal stem cells’ has created confusion among scientists, clinicians, regulators, and our patients. Validated methods need to be developed to measure and characterize the ‘critical quality attributes’ and biological activity of a specific cell formulation. It is certain that ‘one size does not fit all’ – different cell formulations, dosing schedules, and culturing parameters will likely be required based on the tissue being treated and the desired biological target. As an alternative to the use of exogenous cells, in the future we may be able to stimulate the intrinsic vascular stem cell niche that is known to exist in many tissues. The tremendous potential of cell therapy will only be realized with further basic, translational, and clinical research.

Cite this article: Bone Joint J 2019;101-B:361–364.

Aims

The aim of this study was to investigate the effects of preoperative bisphosphonate treatment on the intra- and postoperative outcomes of arthroplasty of the shoulder. The hypothesis was that previous bisphosphonate treatment would adversely affect both intra- and postoperative outcomes.

Ketamine has been used in combination with a variety of other agents for intra-articular analgesia, with promising results. However, although it has been shown to be toxic to various types of cell, there is no available information on the effects of ketamine on chondrocytes.

We conducted a prospective randomised controlled study to evaluate the effects of ketamine on cultured chondrocytes isolated from rat articular cartilage. The cultured cells were treated with 0.125 mM, 0.250 mM, 0.5 mM, 1 mM and 2 mM of ketamine respectively for 6 h, 24 hours and 48 hours, and compared with controls. Changes of apoptosis were evaluated using fluorescence microscopy with a 490 nm excitation wavelength. Apoptosis and eventual necrosis were seen at each concentration. The percentage viability of the cells was inversely proportional to both the duration and dose of treatment (p = 0.002 and p = 0.009). Doses of 0.5 mM, 1 mM and 2mM were absolutely toxic.

We concluded that in the absence of solid data to support the efficacy of intra-articular ketamine for the control of pain, and the toxic effects of ketamine on cultured chondrocytes shown by this study, intra-articular ketamine, either alone or in combination with other agents, should not be used to control pain.

Cite this article: Bone Joint J 2014; 96-B:989–94.

Recently, the use of metal-on-metal articulations in total hip arthroplasty (THA) has led to an increase in adverse events owing to local soft-tissue reactions from metal ions and wear debris. While the majority of these implants perform well, it has been increasingly recognised that a small proportion of patients may develop complications secondary to systemic cobalt toxicity when these implants fail. However, distinguishing true toxicity from benign elevations in cobalt ion levels can be challenging.

The purpose of this two part series is to review the use of cobalt alloys in THA and to highlight the following related topics of interest: mechanisms of cobalt ion release and their measurement, definitions of pathological cobalt ion levels, and the pathophysiology, risk factors and treatment of cobalt toxicity. Historically, these metal-on-metal arthroplasties are composed of a chromium-cobalt articulation.

The release of cobalt is due to the mechanical and oxidative stresses placed on the prosthetic joint. It exerts its pathological effects through direct cellular toxicity.

This manuscript will highlight the pathophysiology of cobalt toxicity in patients with metal-on-metal hip arthroplasties.

Take home message: Patients with new or evolving hip symptoms with a prior history of THA warrant orthopaedic surgical evaluation. Increased awareness of the range of systemic symptoms associated with cobalt toxicity, coupled with prompt orthopaedic intervention, may forestall the development of further complications.

Cite this article: Bone Joint J 2016;98-B:6–13.

Peri-prosthetic osteolysis and subsequent aseptic loosening is the most common reason for revising total hip replacements. Wear particles originating from the prosthetic components interact with multiple cell types in the peri-prosthetic region resulting in an inflammatory process that ultimately leads to peri-prosthetic bone loss. These cells include macrophages, osteoclasts, osteoblasts and fibroblasts. The majority of research in peri-prosthetic osteolysis has concentrated on the role played by osteoclasts and macrophages. The purpose of this review is to assess the role of the osteoblast in peri-prosthetic osteolysis.

In peri-prosthetic osteolysis, wear particles may affect osteoblasts and contribute to the osteolytic process by two mechanisms. First, particles and metallic ions have been shown to inhibit the osteoblast in terms of its ability to secrete mineralised bone matrix, by reducing calcium deposition, alkaline phosphatase activity and its ability to proliferate. Secondly, particles and metallic ions have been shown to stimulate osteoblasts to produce pro inflammatory mediators in vitro. In vivo, these mediators have the potential to attract pro-inflammatory cells to the peri-prosthetic area and stimulate osteoclasts to absorb bone. Further research is needed to fully define the role of the osteoblast in peri-prosthetic osteolysis and to explore its potential role as a therapeutic target in this condition.

Cite this article: Bone Joint J 2013;95-B:1021–5.

It has been proposed that intervertebral disc degeneration might be caused by low-grade infection. The purpose of the present study was to assess the incidence of herpes viruses in intervertebral disc specimens from patients with lumbar disc herniation. A polymerase chain reaction based assay was applied to screen for the DNA of eight different herpes viruses in 16 patients and two controls. DNA of at least one herpes virus was detected in 13 specimens (81.25%). Herpes Simplex Virus type-1 (HSV-1) was the most frequently detected virus (56.25%), followed by Cytomegalovirus (CMV) (37.5%). In two patients, co-infection by both HSV-1 and CMV was detected. All samples, including the control specimens, were negative for Herpes Simplex Virus type-2, Varicella Zoster Virus, Epstein Barr Virus, Human Herpes Viruses 6, 7 and 8. The absence of an acute infection was confirmed both at the serological and mRNA level.

To our knowledge this is the first unequivocal evidence of the presence of herpes virus DNA in intervertebral disc specimens of patients with lumbar disc herniation suggesting the potential role of herpes viruses as a contributing factor to the pathogenesis of degenerative disc disease.

Methods

In this study of patients who underwent internal fixation without fusion for a burst thoracolumbar or lumbar fracture, we compared the serial changes in the injured disc height (DH), and the fractured vertebral body height (VBH) and kyphotic angle between patients in whom the implants were removed and those in whom they were not. Radiological parameters such as injured DH, fractured VBH and kyphotic angle were measured. Functional outcomes were evaluated using the Greenough low back outcome scale and a VAS scale for pain.

We hypothesised that cells obtained via a Reamer–Irrigator–Aspirator (RIA) system retain substantial osteogenic potential and are at least equivalent to graft harvested from the iliac crest. Graft was harvested using the RIA in 25 patients (mean age 37.6 years (18 to 68)) and from the iliac crest in 21 patients (mean age 44.6 years (24 to 78)), after which ≥ 1 g of bony particulate graft material was processed from each. Initial cell viability was assessed using Trypan blue exclusion, and initial fluorescence-activated cell sorting (FACS) analysis for cell lineage was performed. After culturing the cells, repeat FACS analysis for cell lineage was performed and enzyme-linked immunosorbent assay (ELISA) for osteocalcin, and Alizarin red staining to determine osteogenic potential. Cells obtained via RIA or from the iliac crest were viable and matured into mesenchymal stem cells, as shown by staining for the specific mesenchymal antigens CD90 and CD105. For samples from both RIA and the iliac crest there was a statistically significant increase in bone production (both p < 0.001), as demonstrated by osteocalcin production after induction.

Medullary autograft cells harvested using RIA are viable and osteogenic. Cell viability and osteogenic potential were similar between bone grafts obtained from both the RIA system and the iliac crest.

Cite this article: Bone Joint J 2013;95-B:1269–74.

The belief that an intervertebral disc must degenerate before it can herniate has clinical and medicolegal significance, but lacks scientific validity. We hypothesised that tissue changes in herniated discs differ from those in discs that degenerate without herniation. Tissues were obtained at surgery from 21 herniated discs and 11 non-herniated discs of similar degeneration as assessed by the Pfirrmann grade. Thin sections were graded histologically, and certain features were quantified using immunofluorescence combined with confocal microscopy and image analysis. Herniated and degenerated tissues were compared separately for each tissue type: nucleus, inner annulus and outer annulus.

Herniated tissues showed significantly greater proteoglycan loss (outer annulus), neovascularisation (annulus), innervation (annulus), cellularity/inflammation (annulus) and expression of matrix-degrading enzymes (inner annulus) than degenerated discs. No significant differences were seen in the nucleus tissue from herniated and degenerated discs. Degenerative changes start in the nucleus, so it seems unlikely that advanced degeneration caused herniation in 21 of these 32 discs. On the contrary, specific changes in the annulus can be interpreted as the consequences of herniation, when disruption allows local swelling, proteoglycan loss, and the ingrowth of blood vessels, nerves and inflammatory cells.

In conclusion, it should not be assumed that degenerative changes always precede disc herniation.

Cite this article: Bone Joint J 2013;95-B:1127–33.

We have studied the effects of bupivacaine on human and bovine articular chondrocytes in vitro. Time-lapse confocal microscopy of human articular chondrocytes showed > 95% cellular death after exposure to 0.5% bupivacaine for 30 minutes. Human and bovine chondrocytes exposed to 0.25% bupivacaine had a time-dependent reduction in viability, with longer exposure times resulting in higher cytotoxicity. Cellular death continued even after removal of 0.25% bupivacaine. After exposure to 0.25% bupivacaine for 15 minutes, flow cytometry showed bovine chondrocyte viability to be 41% of saline control after seven days. After exposure to 0.125% bupivacaine for up to 60 minutes, the viability of both bovine and human chondrocytes was similar to that of control groups.

These data show that prolonged exposure 0.5% and 0.25% bupivacaine solutions are potentially chondrotoxic.

We present two cases of metastatic lung cancer which occurred at the site of a previously united tibial fracture. Both patients were treated with a locked intramedullary nail. The patients presented with metastases at the site of their initial fracture approximately 16 and 13 months after injury respectively.

We discuss this unusual presentation and review the relevant literature. We are unaware of any previous reports of a metastatic tumour occurring at the site of an orthopaedic implant used to stabilise a non-pathological fracture. These cases demonstrate the similar clinical presentation of infection and malignancy: a diagnosis which should always be considered in such patients.

Implant-associated infection is a major source of morbidity in orthopaedic surgery. There has been extensive research into the development of materials that prevent biofilm formation, and hence, reduce the risk of infection. Silver nanoparticle technology is receiving much interest in the field of orthopaedics for its antimicrobial properties, and the results of studies to date are encouraging. Antimicrobial effects have been seen when silver nanoparticles are used in trauma implants, tumour prostheses, bone cement, and also when combined with hydroxyapatite coatings. Although there are promising results with in vitro and in vivo studies, the number of clinical studies remains small. Future studies will be required to explore further the possible side effects associated with silver nanoparticles, to ensure their use in an effective and biocompatible manner. Here we present a review of the current literature relating to the production of nanosilver for medical use, and its orthopaedic applications.

Cite this article: Bone Joint J 2015; 97-B:582–9.

The incidence of acute and chronic conditions of the tendo Achillis appear to be increasing. Causation is multifactorial but the role of inherited genetic elements and the influence of environmental factors altering gene expression are increasingly being recognised. Certain individuals’ tendons carry specific variations of genetic sequence that may make them more susceptible to injury. Alterations in the structure or relative amounts of the components of tendon and fine control of activity within the extracellular matrix affect the response of the tendon to loading with failure in certain cases.

This review summarises present knowledge of the influence of genetic patterns on the pathology of the tendo Achillis, with a focus on the possible biological mechanisms by which genetic factors are involved in the aetiology of tendon pathology. Finally, we assess potential future developments with both the opportunities and risks that they may carry.

Cite this article: Bone Joint J 2013;95-B:305–13.

We have examined the deterioration of implant fixation after withdrawal of parathyroid hormone (PTH) in rats. First, the pull-out force for stainless-steel screws in the proximal tibia was measured at different times after withdrawal. The stimulatory effect of PTH on fixation was lost after 16 days. We then studied whether bisphosphonates could block this withdrawal effect. Mechanical and histomorphometric measurements were conducted for five weeks after implantation. Subcutaneous injections were given daily. Specimens treated with either PTH or saline during the first two weeks showed no difference in the mechanical or histological results (pull-out force 76 N vs 81 N; bone volume density 19% vs 20%). Treatment with PTH for two weeks followed by pamidronate almost doubled the pull-out force (152 N; p < 0.001) and the bone volume density (37%; ANOVA, p < 0.001). Pamidronate alone did not have this effect (89 N and 25%, respectively). Thus, the deterioration can be blocked by bisphosphonates. The clinical implications are discussed.

When transferring tissue regenerative strategies involving skeletal stem cells to human application, consideration needs to be given to factors that may affect the function of the cells that are transferred. Local anaesthetics are frequently used during surgical procedures, either administered directly into the operative site or infiltrated subcutaneously around the wound. The aim of this study was to investigate the effects of commonly used local anaesthetics on the morphology, function and survival of human adult skeletal stem cells.

Cells from three patients who were undergoing elective hip replacement were harvested and incubated for two hours with 1% lidocaine, 0.5% levobupivacaine or 0.5% bupivacaine hydrochloride solutions. Viability was quantified using WST-1 and DNA assays. Viability and morphology were further characterised using CellTracker Green/Ethidium Homodimer-1 immunocytochemistry and function was assessed by an alkaline phosphatase assay. An additional group was cultured for a further seven days to allow potential recovery of the cells after removal of the local anaesthetic.

A statistically significant and dose dependent reduction in cell viability and number was observed in the cell cultures exposed to all three local anaesthetics at concentrations of 25% and 50%, and this was maintained even following culture for a further seven days.

This study indicates that certain local anaesthetic agents in widespread clinical use are deleterious to skeletal progenitor cells when studied in vitro; this might have relevance in clinical applications.

Surgical reconstruction of articular surfaces by transplantation of osteochondral autografts has shown considerable promise in the treatment of focal articular lesions. During mosaicplasty, each cylindrical osteochondral graft is centred over the recipient hole and delivered by impacting the articular surface. Impact loading of articular cartilage has been associated with structural damage, loss of the viability of chondrocytes and subsequent degeneration of the articular cartilage. We have examined the relationship between single-impact loading and chondrocyte death for the specific confined-compression boundary conditions of mosaicplasty and the effect of repetitive impact loading which occurs during implantation of the graft on the resulting viability of the chondrocytes.

Fresh bovine and porcine femoral condyles were used in this experiment. The percentage of chondrocyte death was found to vary logarithmically with single-impact energy and was predicted more strongly by the mean force of the impact rather than by the number of impacts required during placement of the graft. The significance of these results in regard to the surgical technique and design features of instruments for osteochondral transplantation is discussed.

The pathophysiology of intervertebral disc degeneration has been extensively studied. Various factors have been suggested as influencing its aetiology, including mechanical factors, such as compressive loading, shear stress and vibration, as well as ageing, genetic, systemic and toxic factors, which can lead to degeneration of the disc through biochemical reactions. How are these factors linked? What is their individual importance? There is no clear evidence indicating whether ageing in the presence of repetitive injury or repetitive injury in the absence of ageing plays a greater role in the degenerative process. Mechanical factors can trigger biochemical reactions which, in turn, may promote the normal biological changes of ageing, which can also be accelerated by genetic factors. Degradation of the molecular structure of the disc during ageing renders it more susceptible to superimposed mechanical injuries.

This review supports the theory that degeneration of the disc has a complex multifactorial aetiology. Which factors initiate the events in the degenerative cascade is a question that remains unanswered, but most evidence points to an age-related process influenced primarily by mechanical and genetic factors.

We undertook a study of the anti-tumour effects of hyperthermia, delivered via magnetite cationic liposomes (MCLs), on local tumours and lung metastases in a mouse model of osteosarcoma. MCLs were injected into subcutaneous osteosarcomas (LM8) and subjected to an alternating magnetic field which induced a heating effect in MCLs. A control group of mice with tumours received MCLs but were not exposed to an AMF. A further group of mice with tumours were exposed to an AMF but had not been treated with MCLs. The distribution of MCLs and local and lung metastases was evaluated histologically. The weight and volume of local tumours and the number of lung metastases were determined. Expression of heat shock protein 70 was evaluated immunohistologically. Hyperthermia using MCLs effectively heated the targeted tumour to 45°C. The mean weight of the local tumour was significantly suppressed in the hyperthermia group (p = 0.013). The mice subjected to hyperthermia had significantly fewer lung metastases than the control mice (p = 0.005). Heat shock protein 70 was expressed in tumours treated with hyperthermia, but was not found in those tumours not exposed to hyperthermia.

The results demonstrate a significant effect of hyperthermia on local tumours and reduces their potential to metastasise to the lung.

Fracture repair occurs by two broad mechanisms: direct healing, and indirect healing with callus formation. The effects of bisphosphonates on fracture repair have been assessed only in models of indirect fracture healing.

A rodent model of rigid compression plate fixation of a standardised tibial osteotomy was used. Ten skeletally mature Sprague–Dawley rats received daily subcutaneous injections of 1 µg/kg ibandronate (IBAN) and ten control rats received saline (control). Three weeks later a tibial osteotomy was rigidly fixed with compression plating. Six weeks later the animals were killed. Fracture repair was assessed with mechanical testing, radiographs and histology.

The mean stress at failure in a four-point bending test was significantly lower in the IBAN group compared with controls (8.69 Nmm^-2 (sd 7.63) vs 24.65 Nmm^-2 (sd 6.15); p = 0.017). On contact radiographs of the extricated tibiae the mean bone density assessment at the osteotomy site was lower in the IBAN group than in controls (3.7 mmAl (sd 0.75) vs 4.6 mmAl (sd 0.57); p = 0.01). In addition, histological analysis revealed progression to fracture union in the controls but impaired fracture healing in the IBAN group, with predominantly cartilage-like and undifferentiated mesenchymal tissue (p = 0.007).

Bisphosphonate treatment in a therapeutic dose, as used for risk reduction in fragility fractures, had an inhibitory effect on direct fracture healing. We propose that bisphosphonate therapy not be commenced until after the fracture has united if the fracture has been rigidly fixed and is undergoing direct osteonal healing.

Cite this article: Bone Joint J 2013;95-B:1263–8.

Lateral epicondylitis, or ’tennis elbow’, is a common condition that usually affects patients between 35 and 55 years of age. It is generally self-limiting, but in some patients it may continue to cause persistent symptoms, which can be refractory to treatment. This review discusses the mechanism of disease, symptoms and signs, investigations, current management protocols and potential new treatments.

Cite this article: Bone Joint J 2013;95-B:1158–64.

Successful healing of a nine-year tibial nonunion resistant to six previous surgical procedures was achieved by tissue engineering. We used autologous bone marrow stromal cells (BMSCs) expanded to 5 × 10⁶ cells after three weeks’ tissue culture. Calcium sulphate (CaSO₄) in pellet form was combined with these cells at operation. The nonunion was clinically and radiologically healed two months after implantation.

This is the description of on healing of a long-standing tibial nonunion by tissue engineering. The successful combination of BMSCs and CaSO₄ has not to our knowledge been reported in a clinical setting.

The mammalian growth plate is a complex structure which is essential for the elongation of long bones. However, an understanding of how the growth plate functions at the cellular level is lacking. This review, summarises the factors involved in growth-plate regulation, its failure and the consequence of injury. We also describe some of the cellular mechanisms which underpin the increase in volume of the growth-plate chondrocyte which is the major determinant of the rate and extent of bone lengthening. We show how living in situ chondrocytes can be imaged using 2-photon laser scanning microscopy to provide a quantitative analysis of their volume. This approach should give better understanding of the cellular control of bone growth in both healthy and failed growth plates.

This article reviews the current knowledge of the intervertebral disc (IVD) and its association with low back pain (LBP). The normal IVD is a largely avascular and aneural structure with a high water content, its nutrients mainly diffusing through the end plates. IVD degeneration occurs when its cells die or become dysfunctional, notably in an acidic environment. In the process of degeneration, the IVD becomes dehydrated and vascularised, and there is an ingrowth of nerves. Although not universally the case, the altered physiology of the IVD is believed to precede or be associated with many clinical symptoms or conditions including low back and/or lower limb pain, paraesthesia, spinal stenosis and disc herniation.

New treatment options have been developed in recent years. These include biological therapies and novel surgical techniques (such as total disc replacement), although many of these are still in their experimental phase. Central to developing further methods of treatment is the need for effective ways in which to assess patients and measure their outcomes. However, significant difficulties remain and it is therefore an appropriate time to be further investigating the scientific basis of and treatment of LBP.

Bone loss involving articular surface is a challenging problem faced by the orthopaedic surgeon. In the hand and wrist, there are articular defects that are amenable to autograft reconstruction when primary fixation is not possible. In this article, the surgical techniques and clinical outcomes of articular reconstructions in the hand and wrist using non-vascularised osteochondral autografts are reviewed.

We performed a retrospective review of all patients admitted to two large University Hospitals in the United Kingdom over a 24-month period from January 2008 to January 2010 to identify the incidence of atypical subtrochanteric and femoral shaft fractures and their relationship to bisphosphonate treatment. Of the 3515 patients with a fracture of the proximal femur, 156 fractures were in the subtrochanteric region. There were 251 femoral shaft fractures. The atypical fracture pattern was seen in 27 patients (7%) with 29 femoral shaft or subtrochanteric fractures. A total of 22 patients with 24 atypical fractures were receiving bisphosphonate treatment at the time of fracture. Prodromal pain was present in nine patients (11 fractures); 11 (50%) of the patients on bisphosphonates suffered 12 spontaneous fractures, and healing of these fractures was delayed in a number of patients. This large dual-centre review has established the incidence of atypical femoral fractures at 7% of the study population, 81% of whom had been on bisphosphonate treatment for a mean of 4.6 years (0.04 to 12.1).

This study does not advocate any change in the use of bisphosphonates to prevent fragility fractures but attempts to raise awareness of this possible problem so symptomatic patients will be appropriately investigated. However, more work is required to identify the true extent of this new and possibly increasing problem.

The most frequent cause of failure after total hip replacement in all reported arthroplasty registries is peri-prosthetic osteolysis. Osteolysis is an active biological process initiated in response to wear debris. The eventual response to this process is the activation of macrophages and loss of bone.

Activation of macrophages initiates a complex biological cascade resulting in the final common pathway of an increase in osteolytic activity. The biological initiators, mechanisms for and regulation of this process are beginning to be understood. This article explores current concepts in the causes of, and underlying biological mechanism resulting in peri-prosthetic osteolysis, reviewing the current basic science and clinical literature surrounding the topic.

Articular cartilage repair remains a challenge to surgeons and basic scientists. The field of tissue engineering allows the simultaneous use of material scaffolds, cells and signalling molecules to attempt to modulate the regenerative tissue. This review summarises the research that has been undertaken to date using this approach, with a particular emphasis on those techniques that have been introduced into clinical practice, via in vitro and preclinical studies.

A 70-year-old man with an uncemented metal-on-polyethylene total hip prosthesis underwent revision arthroplasty 33 months later because of pain, swelling and recurrent dislocation. There appeared to be corrosion and metal release from the prosthetic head, resulting in pseudotumour formation and severe local soft-tissue destruction. The corrosion occurred at the junction between the titanium-molybdenum-zirconium-iron taper and the cobalt-chrome-molybdenum head, but the mechanism was unproven.

Microfracture is frequently used as the first line of treatment for the repair of traumatic cartilage defects. We present the clinical and histological results 18 months to two-years after treatment in a 26-year-old male with a post-traumatic chondral defect of the medial femoral condyle managed by microfracture covered with chondrotissue, a cell-free cartilage implant made of a resorbable polyglycolic acid felt and hyaluronic acid.

There are 33 million people worldwide currently infected with human immunodeficiency virus (HIV). This complex disease affects many of the processes involved in wound and fracture healing, and there is little evidence available to guide the management of open fractures in these patients. Fears of acute and delayed infection often inhibit the use of fixation, which may be the most effective way of achieving union.

This study compared fixation of open fractures in HIV-positive and -negative patients in South Africa, a country with very high rates of both HIV and high-energy trauma. A total of 133 patients (33 HIV-positive) with 135 open fractures fulfilled the inclusion criteria. This cohort is three times larger than in any similar previously published study.

The results suggest that HIV is not a contraindication to internal or external fixation of open fractures in this population, as HIV is not a significant risk factor for acute wound/implant infection. However, subgroup analysis of grade I open fractures in patients with advanced HIV and a low CD4 count (< 350) showed an increased risk of infection; we suggest that grade I open fractures in patients with advanced HIV should be treated by early debridement followed by fixation at an appropriate time.

Lyme disease is a vector-borne multisystem inflammatory disease caused by the spirochete Borrelia burgdorferi sensu lato. This disease is frequently seen in North America and to a lesser degree in Europe. However, its presence in England is uncommon and we present a case in which the patient developed a palsy of the common peroneal nerve

Limb salvage involving wide resection and reconstruction is now well established for managing musculoskeletal sarcomas. However, involvement of major nerves and vessels with a large volume of muscle and skin may result in a useless limb, contributing to depression and a low quality of life. We have been studying alternative treatments for musculoskeletal sarcoma since 1990, and have recently established a regime using photodynamic surgery with cells labelled with acridine orange, photodynamic therapy with cells treated similarly and radiodynamic treatment using the effect of X-rays on such cells.

These techniques have been used after marginal or intralesional resection of tumours since 1999 and have enabled maintenance of excellent limb function in patients with sarcomas.

Several bisphosphonates are now available for the treatment of osteoporosis. Porous hydroxyapatite/collagen (HA/Col) composite is an osteoconductive bone substitute which is resorbed by osteoclasts. The effects of the bisphosphonate alendronate on the formation of bone in porous HA/Col and its resorption by osteoclasts were evaluated using a rabbit model. Porous HA/Col cylinders measuring 6 mm in diameter and 8 mm in length, with a pore size of 100 μm to 500 μm and 95% porosity, were inserted into a defect produced in the lateral femoral condyles of 72 rabbits. The rabbits were divided into four groups based on the protocol of alendronate administration: the control group did not receive any alendronate, the pre group had alendronate treatment for three weeks prior to the implantation of the HA/Col, the post group had alendronate treatment following implantation until euthanasia, and the pre+post group had continuous alendronate treatment from three weeks prior to surgery until euthanasia. All rabbits were injected intravenously with either saline or alendronate (7.5 μg/kg) once a week. Each group had 18 rabbits, six in each group being killed at three, six and 12 weeks post-operatively. Alendronate administration suppressed the resorption of the implants. Additionally, the mineral densities of newly formed bone in the alendronate-treated groups were lower than those in the control group at 12 weeks post-operatively. Interestingly, the number of osteoclasts attached to the implant correlated with the extent of bone formation at three weeks.

In conclusion, the systemic administration of alendronate in our rabbit model at a dose-for-weight equivalent to the clinical dose used in the treatment of osteoporosis in Japan affected the mineral density and remodelling of bone tissue in implanted porous HA/Col composites.

We studied the effects of hyperbaric oxygen (HBO) and zoledronic acid (ZA) on posterior lumbar fusion using a validated animal model. A total of 40 New Zealand white rabbits underwent posterior lumbar fusion at L5–6 with autogenous iliac bone grafting. They were divided randomly into four groups as follows: group 1, control; group 2, HBO (2.4 atm for two hours daily); group 3, local ZA (20 μg of ZA mixed with bone graft); and group 4, combined HBO and local ZA. All the animals were killed six weeks after surgery and the fusion segments were subjected to radiological analysis, manual palpation, biomechanical testing and histological examination.

Five rabbits died within two weeks of operation. Thus, 35 rabbits (eight in group 1 and nine in groups 2, 3 and 4) completed the study. The rates of fusion in groups 3 and 4 (p = 0.015) were higher than in group 1 (p < 0.001) in terms of radiological analysis and in group 4 was higher than in group 1 with regard to manual palpation (p = 0.015). We found a statistically significant difference in the biomechanical analysis between groups 1 and 4 (p = 0.024). Histological examination also showed a statistically significant difference between groups 1 and 4 (p = 0.036).

Our results suggest that local ZA combined with HBO may improve the success rate in posterior lumbar spinal fusion.

The scarcity of mesenchymal stem cells (MSCs) in iliac crest bone marrow aspirate (ICBMA), and the expense and time in culturing cells, has led to the search for alternative harvest sites. The reamer-irrigation-aspirator (RIA) provides continuous irrigation and suction during reaming of long bones. The aspirated contents pass via a filter, trapping bony fragments, before moving into a ‘waste’ bag from which MSCs have been previously isolated. We examined the liquid and solid phases, performed a novel digestion of the solid phase, and made a comparative assessment in terms of number, phenotype and differentiation capacity with matched ICBMA.

The solid fraction from the filtrate was digested for 60 minutes at 37°C with collagenase. Enumeration was performed via the colony-forming unit fibroblast (CFU-F) assay. Passage (P2) cells were differentiated towards osteogenic, adipogenic and chondrogenic lineages, and their phenotypes assessed using flow cytometry (CD33, CD34, CD45, CD73, CD90, and CD105).

MSCs from the RIA phases were able to differentiate at least as well as those from ICBMA, and all fractions had phenotypes consistent with other established sources. The median number of colonies for the three groups was: ICBMA = 8.5 (2 to 86), RIA-liquid = 19.5 (4 to 90), RIA-solid = 109 (67 to 200) per 200 μl. The mean total yield of cells for the three groups was: ICBMA = 920 (0 to 4275), RIA-liquid = 114 983 (16 500 to 477 750), RIA-solid = 12 785 (7210 to 28 475).

The RIA filtrate contains large numbers of MSCs that could potentially be extracted without enzymatic digestion and used for bone repair without prior cell expansion.