Introduction. There is widespread variation in the management of rare orthopaedic disease, in a large part owing to uncertainty. No individual surgeon or hospital is typically equipped to amass sufficient numbers of cases to draw robust conclusions from the information available to them. The programme of research will establish the British Orthopaedic Surgery Surveillance (BOSS) Study; a nationwide reporting structure for rare disease in orthopaedic surgery. Methods. The BOSS Study is a series of nationwide observational cohort studies of pre-specified orthopaedic disease. All relevant hospitals treating the disease are invited to contribute anonymised case details. Data will be collected digitally through REDCap, with an additional bespoke software solution used to regularly confirm case ascertainment, prompt follow-up reminders and identify potential missing cases from external sources of information (i.e. national administrative data). With their consent, patients will be invited to enrich the data collected by supplementing anonymised case data with patient reported outcomes. The study will primarily seek to calculate the incidence of the rare diseases under investigation, with 95% confidence intervals. Descriptive statistics will be used to describe the case mix, treatment variations and outcomes. Inferential statistical analysis may be used to analyze associations between presentation factors and outcomes. Types of analyses will be contingent on the disease under investigation. Discussion. This study builds upon other national rare disease supporting structures, particularly those in obstetrics and paediatric surgery. It is particularly focused on addressing the evidence base for quality and safety of surgery, and the design is influenced by the specifications of the IDEAL collaboration for the development of surgical research

The ability to edit DNA at the nucleotide level using clustered regularly interspaced short palindromic repeats (CRISPR) systems is a relatively new investigative tool that is revolutionizing the analysis of many aspects of human health and disease, including orthopaedic disease. CRISPR, adapted for mammalian cell genome editing from a bacterial defence system, has been shown to be a flexible, programmable, scalable, and easy-to-use gene editing tool. Recent improvements increase the functionality of CRISPR through the engineering of specific elements of CRISPR systems, the discovery of new, naturally occurring CRISPR molecules, and modifications that take CRISPR beyond gene editing to the regulation of gene transcription and the manipulation of RNA. Here, the basics of CRISPR genome editing will be reviewed, including a description of how it has transformed some aspects of molecular musculoskeletal research, and will conclude by speculating what the future holds for the use of CRISPR-related treatments and therapies in clinical orthopaedic practice. Cite this article: Bone Joint Res 2020;9(7):351–359

Mesenchymal stem cells-derived extracellular vesicles (MSC-EVs) have great promise in the field of orthopaedic nanomedicine due to their regenerative, as well as immunomodulatory and anti-inflammatory properties. Researchers are interested in harnessing these biologically sourced nanovesicles as powerful therapeutic tools with intrinsic bioactivity to help treat various orthopaedic diseases and defects. Recently, a new class of EV mimetics has emerged known as nanoghosts (NGs). These vesicles are derived from the plasma membrane of ghost cells, thus inheriting the surface functionalities and characteristics of the parent cell while at the same time allowing for a more standardized and reproducible production and significantly greater yield when compared to EVs. This study aims to investigate and compare the osteoinductive potential of MSC-EVs and MSC-NGs in vitro as novel tools in the field of bone tissue engineering and nanomedicine. To carry out this investigation, MSC-EVs were isolated from serum-free MSC conditioned media through differential ultracentrifugation. The remaining cells were treated with hypotonic buffer to produce MSC-ghosts that were then homogenized and serially extruded through 400 and 200 nm polycarbonate membranes to form the MSC-NGs. The concentration, size distribution, zeta potential, and protein content of the isolated nanoparticles were assessed. Afterwards, MSCs were treated with either MSC-EVs or MSC-NGs under osteogenic conditions, and their differentiation was assessed through secreted ALP assay, qPCR, and Alizarin Red mineralization staining. Isolation of MSC-EVs and MSC-NGs was successful, with relatively similar mean diameter size and colloidal stability. No effect on MSC viability and metabolic activity was observed with either treatment. Both MSC-EV and MSC-NG groups had enhanced osteogenic outcomes compared to the control; however, a trend was observed that suggests MSC-NGs as better osteoinductive mediators compared to MSC-EVs. Acknowledgements: The authors would like to acknowledge Canada Research Chair – Tier 1 in Regenerative Medicine and Nanomedicine, CHRP, and McGill's Faculty of Dental Medicine and Oral Health Sciences for their financial support

The reliable production of _in vitro_ chondrocytes that faithfully recapitulate _in vivo_ development would be of great benefit for orthopaedic disease modelling and regenerative therapy(1,2). Current efforts are limited by off-target differentiation, resulting in a heterogeneous product, and by the lack of comparison to human tissue, which precludes detailed evaluation of _in vitro_ cells(3,4). We performed single-cell RNA-sequencing of long bones dissected from first-trimester fetal limbs to form a detailed ‘atlas’ of endochondral ossification. Through 100-gene in-situ sequencing, we placed each sequenced cell type into its anatomical context to spatially resolve the process of endochondral ossification. We then used this atlas to perform deconvolution on a series of previously published bulk transcriptomes generated from _in vitro_ chondrogenesis protocols to evaluate their ability to accurately produce chondrocytes. We then applied single-nuclear RNA-sequencing to cells from the best performing protocol collected at multiple time points to allow direct comparison between the differentiation of _in vitro_ and _in vivo_ cells. We captured 275,000 single fetal cells, profiling the development of chondrocytes from multipotent mesenchymal progenitors to hypertrophic cells at full transcriptomic breadth. Using this atlas as the ground truth for evaluating _in vitro_ cells, we found substantial variability in cell states produced by each protocol, with many showing little similarity to _in vivo_ cells, and all exhibiting off-target differentiation. Trajectory alignment between _in vivo_ and _in vitro_ single-cell data revealed key differences in gene expression dynamics between _in vitro_ and _in vivo cells,_ with several osteoblastic transcription factors erroneously unregulated _in vitro,_ including _FOXO1._. Using this information, we inhibited _FOXO1_ in culture to successfully increase chondrocyte yield _in vitro._. This study presents a new framework for evaluating tissue engineering protocols, using single-cell data to drive improvement and bring the prospect of true engineered cartilage closer to reality

Aims. Metabolic profiling is a top-down method of analysis looking at metabolites, which are the intermediate or end products of various cellular pathways. Our primary objective was to perform a systematic review of the published literature to identify metabolites in human synovial fluid (HSF), which have been categorized by metabolic profiling techniques. A secondary objective was to identify any metabolites that may represent potential biomarkers of orthopaedic disease processes. Methods. A systematic review was conducted in accordance with Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines using the MEDLINE, Embase, PubMed, and Cochrane databases. Studies included were case series, case control series, and cohort studies looking specifically at HSF. Results. The primary analysis, which pooled the results from 17 published studies and four meeting abstracts, identified over 200 metabolites. Seven of these studies (six published studies, one meeting abstract) had asymptomatic control groups and collectively suggested 26 putative biomarkers in osteoarthritis, inflammatory arthropathies, and trauma. These can broadly be categorized into amino acids plus related metabolites, fatty acids, ketones, and sugars. Conclusion. The role of metabolic profiling in orthopaedics is fast evolving with many metabolites already identified in a variety of pathologies. However, these results need to be interpreted with caution due to the presence of multiple confounding factors in many of the studies. Future research should include largescale epidemiological metabolic profiling studies incorporating various confounding factors with appropriate statistical analysis to account for multiple testing of the data. Cite this article:Bone Joint Res. 2020;9(3):108–119

Abstract. Cranial cruciate ligament (CrCL) disease/rupture is a highly prevalent orthopaedic disease in dogs and common cause of pain, lameness, and secondary joint osteoarthritis (OA). Previous experiments investigating the role of glutamate receptors (GluR) in arthritic degeneration and pain revealed that OA biomarkers assessing early bone turnover and inflammation, including osteoprotegerin (OPG) and the receptor activator of nuclear factor kappa-B ligand (RANKL) are more likely to be influenced by glutamate signalling. Moreover, interleukin-6 (IL-6) has a complex and potentially bi directional (beneficial and detrimental) effect, and it is a critical mediator of arthritic pain, OA progression and joint destruction. Objectives. 1) to recruit dogs undergoing CrCL disease/rupture surgery and obtain discarded synovial fluid (SF) and serum/plasma (ethics approval, RCVS:2017/14/Alves); 2) to quantify the biomarkers listed above in the SF and serum/plasma by enzyme linked immunosorbent assay (ELISA); 3) to assess radiographic OA at the time of surgery and correlate it with the biomarkers and clinical findings. Methods. Abnova, Abcam and AMSBIO ELISA kits were tested using a validation protocol relating the standard curve to a dilution series of SF and serum/plasma (1× to 1/50×), with and without SF hyaluronidase treatment to evaluate linearity, specificity and optimal dilutions. Validated ELISA kits were used to measure [IL-6], glutamate [glu], [RANKL] and [OPG] in SF and serum/plasma. For each dog, CrCL disease pre-operative lameness scores were graded as: (1) mild, (2) moderate (easily visible), (3) marked (encumbered), (4) non-weightbearing lameness. Blinded OA scoring was performed on radiographs [15–60, normal-severe OA]. Results. canine population (n=14) was of various breeds, aged between 2–10 years and weighing 17.1–45.5Kg; 42.86% male; 57.14% female; 83.33% males and 62.5% females were neutered. Lameness scores varied from 1 and 4 (average 2.07±1.12) and radiographic OA scores from 18 and 36 (average 27.86±5.11). Individual correlations in concentrations with respect to age, weight, lameness score (1–4) and OA scores (15–60) were tested. SF [glu] and lameness score were inversely correlated with higher levels of lameness corresponding to lower SF [glu] (P=0.0141). SF [RANKL] inversely correlated with weight (P=0.0045) and lameness score (P=0.0135), and serum [RANKL] inversely correlated with weight (P=0.0437). There was also a negative correlation between SF and serum [OPG] and weight (P=0.0165 and P=0.0208, respectively). No other significant correlations were detected. Overall, [glu] and [IL-6] are increased in SF compared to serum/plasma, by 12.84 and 1.28, respectively, whereas all the remaining biomarkers are higher (2–3 times) in the serum/plasma compared to SF. Principal component analysis (PCA) and Pearson correlation coefficient matrix [IL-6/glu/RANKL/OPG] (n=7) showed SF [IL-6] correlates with SF [glu] (rs=0.64) and strong positive correlations between SF/serum [RANKL] and SF/serum [OPG] (rs 0.68–0.96). Conclusions. Dogs with CrCL disease show an association between the bone remodelling markers RANKL and OPG, and the inflammatory cytokine IL-6, and to a lesser extent SF [glu]. Therapeutics targeting bone remodelling, IL-6 or GluR/[glu] may be of interest for the management of OA in dogs. Declaration of Interest. (b) declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported:I declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research project

Abstract. Cranial cruciate ligament (CrCL) disease/rupture causes pain and osteoarthritis (OA) in dogs. α-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA)-2 and kainate (KA)-1 glutamate receptors (GluR) and the excitatory amino acid transporter-1 (EAAT-1) and EAAT-3 are expressed in joint tissues from OA patients and rodent arthritis models and represent potential therapeutic targets. Objectives. To evaluate glutamate signalling in canine diseased and normal CrCL and meniscus by immunohistochemistry (IHC). Methods. Surgical waste (CrCL, n=5 and medial meniscus, n=3) were obtained from canines with CrCL disease (RCVS ethics approval:2017/14/Alves) and normal analogous tissues (n=2). IHC optimization was performed for rabbit polyclonal (AMPA-2:ab52176, KA-1:ab67402, EAAT-1:ab416) and monoclonal (EAAT-3:ab124802) antibodies from Abcam. IHC was optimised over antibody dilutions from 1:100 to 1:5000 alongside equivalent IgG isotype controls (ab37415 and ab172730) and negative controls (TBS/Tween buffer without primary antibodies). IHC staining was compared in diseased and normal tissues and disclosed with 3,3’-Diaminobenzidine (DAB). Results. Specific immunostaining was observed for all primary antibodies, at concentrations between 2.0×10. −4. mg/mL to 1.0×10. −2. mg/mL, depending on the tissue and primary antibody. All GluR and transporters were expressed in the cellular membrane, in the normal and diseased CrCL and meniscus. Healthy CrCL showed a well-organized microstructure, with normal positively labelled ligamentocytes, whereas diseased CrCL microstructure was disrupted, with many positively stained fibroblastic cells in the epiligamentous region and evident neovascularization, indicative of ongoing repair. The normal and diseased meniscal tissues showed similar chondrocytes-like cells labelling and microstructure. Negative controls demonstrated no labelling. Conclusions. GluR and transporters expression is altered in canine diseased CrCLs, implicating glutamate signalling in this pathology. Since AMPA/KA GluR antagonists alleviate joint degeneration in post-traumatic OA in rodent models, they may be useful for the treatment of CrCL disease in dogs, as well as translated to other veterinary and human orthopaedic diseases

Introduction. Albania is one of the poorest countries in Western European with a GDP per capita standing at 26 percent of the EU average in 2010. Whilst there is government-funded universal free provision of healthcare, it is accepted that delivery is patchy, not accessible to all and lacking expertise for more complex paediatric orthopaedic conditions. With the sponsorship of a UK-based charity, we have set up and completed 5 visits to Albania (3 assessment and 2 operative) to provide additional expertise for paediatric orthopaedic disorders running parallel to and utilising currently available local services. We present the results of this treatment and training programme to date. Patients and methods. Between 2008 and 2011, we assessed 204 children and adolescents with paediatric orthopaedic disorders in Tirana and Durres on 3 separate visits. Of these, 28 were listed for surgical procedures whilst the rest were treated non-operatively. Of the listed patients, 14 patients underwent surgical intervention (total of 18 procedures). Results. The most common diagnoses were developmental dysplasia of the hip, club feet, cerebral palsy and scoliosis. Most patients were treated non-operatively with advice and/or reassurance. Of those listed for surgery, the reasons for cancellation included problems with access to the treating hospital and failure to establish patient contact on the day of admission. Of the operated patients, the procedures performed, the perioperative challenges and significant complications (2/18) will be discussed. Conclusion. Although it is viable to establish parallel service delivery of paediatric orthopaedic surgical services in countries such as Albania, the perioperative and social challenges must be considered

Developmental Dysplasia of the Hip (DDH) is the most common orthopaedic disorder in newborns. Whilst the Pavlik harness is one of the most frequently used treatments for DDH, there is immense variability in treatment parameters reported in the literature and in clinical practice, leading to difficulties in standardising teaching and comparing outcomes. In the absence of definitive quantitative evidence for the optimal Pavlik harness management strategy in DDH, we addressed this problem by scientifically obtaining international expert-based consensus on the same. An initial list of items relevant to Pavlik harness treatment was derived by systematic review of the literature according to PRISMA criteria and reviewed by two expert clinicians in DDH management. Delphi methodology was used to guide serial rounds of surveying and feedback to content matter experts from the International Hip Dysplasia Institute (IHDI), a collaborative group of paediatric orthopaedic surgeons with expertise in the management of DDH. Rounds of surveying continued in the same manner until consensus was reached. Importance ratings were derived from each round of surveying by calculating median score responses on the 5-point Likert scale for each item. Items requiring clarification or those with a median score of below 4 (“agree”) were modified as needed prior to each subsequent round. Consensus was considered reached when 90% or more of the items had an interquartile range (IQR) of ≤ 1. This value indicates low sample deviation and is accepted as having achieved consensus. This was followed by a corroboration of face validity to derive the final set of management principles. The literature search and expert review identified an initial list of 66 items in 8 categories relevant to Pavlik harness management. Four rounds of structured surveying were required to reach consensus. Following a final round of face validity, a definitive list of 33 items in 8 categories met consensus by the experts. These items were tabulated and presented as “General Principles of Pavlik Harness Treatment for DDH” and “Pavlik Harness Treatment by Severity of Hip Dysplasia”. Furthermore, highly contentious items were identified as important future areas of study and will be discussed. We have developed a comprehensive set of principles derived by expert consensus to assist clinicians, and for use as a teaching resource, in the non-operative management of DDH using the Pavlik harness. We have gained consensus on both the general principles of Pavlik harness treatment as well as the detailed treatment of hip subtypes seen across the spectrum of pathology of DDH. Furthermore, this study has also served to generate a list of the most controversial areas in the non-operative management of DDH which should be considered high priority for future study to further refine and optimise the outcomes of children with developmental hip dysplasia

Purpose: Ligament laxity is a common feature of trisomy 21 and is incriminated in most of the orthopaedic disorders observed. Early diagnosis and management is essential. C1-C2 instability is a recognised manifestation in trisomy 21 and is associated, at least theoretically, with significant risk of cord complications. The purpose of this work was to provide a descriptive analysis of the C1-C2 joint in trisomy 21 and to analyse instability factors in order to determine the tolerable C1-C2 distance. Material and methods: Within the framework of a French national epidemiology survey of trisomy 21, we focused on the C1-C2 joint. A total of 472 children with trisomy 21 were identified; 458 who were examined were included in this study. Careful history taking and a detailed physical examination with neurological tests (search for even minimal signs of neurological disorders) was conducted. The Carter and Wilkinson method was used to assess joint laxity. The same specialist searched for other orthopaedic disorders. Patients were divided into two groups depending on the presence or absence of neurological signs. Two groups were also distinguished according to the presence or absence of generalised laxity (Carter and Wilkinson). Lateral x-rays centred on C1-C2 were performed by the same technician on the same machine with the patient in a neutral position, hyperflexion and hyperextension. The same technique inspired by the method described by Singer et al. and modified for simplification was used in all cases. The same observer interpreted the images using a single-blinded protocol to search for congenital malformations and signs of degeneration, measure the C1-C2 distance the minimal sagittal diameter and the C1-C2 angle (not reported in the literature and described for this study). These measures were then compared with data in the literature as available and correlated by age, gender, presence of neurological signs and joint laxity. Seven patients were excluded from the study due to insufficient cooperation for the x-rays and nine because of incomplete clinical or radiological data. The statistical analysis was performed on data from 442 patients. Quantitative variables were compared with the Pearson test and parameteric ANOVA was used to search for correlations of quantitative and qualitative variables. Significance was set at p< 0.05. Results: Mean patient age was 13.8 years. There were 184 girls and 258 boys. Minor neurological anomalies were found in 42% of the patients. There were no cases of major motor deficit. Generalised laxity as defined by Carter and Wilkinson was observed in 24% of patients. Other orthopaedic problems, basically of the foot, were found in 85%. The radiograms revealed a very wide range of measures were thus expressed as means. The C1-C2 distance was greater than 4 mm in 34 patients on the flexion films (limit established in the literature for instability in trisomy 21). The maximal C1-C2 distance in the neutral position was 8 mm, 9.6 mm in flexion. The lowest minimal sagittal distance was 8 mm in flexion and 10 mm in the neutral position (the lower limit reported in the literature before considering the cord to be threatened in 14 mm). The greatest variability was found for the C1-C2 angle. Ligament laxity and atlantoaxial distance were inversely proportional to patient age, but there was no significant correlation between atlantoaxial instability (C1-C2 distance > 4 mm) and gender or generalised hyperlaxity. There was no significant correlation between C1-C2 instability or laxity and neurological signs. Discussion and conclusion: Compared with earlier publications, our series offers the advantage of a large unselected population providing epidemiological data on trisomy 21. A standard radiography protocol was used. The large majority of the radiographic measures reported in the literature do not take into account the magnification effect nor position variability between patients. Our findings confirm certain data in the literature and also provide new information suggesting it could be useful to revisit certain pathogenic hypotheses about C1-C2 instability and its neurological consequences in trisomy 21. Two important observations were the absence of a correlation between general laxity and C1-C2 instability and the absence of correlation between C1-C2 instability and the presence of neurological signs

MSCs (mesenchymal stem cells) are bone marrow-derived cells capable of replication and differentiation in-vitro into several tissues including bone, cartilage, stroma, fat, muscle and tendon. MSCs can be isolated by relatively simple procedures and then expanded without losing the ability to differentiate into multiple lineages. As such, these cells have immense clinical potential in regenerative medicine and in orthopaedics for repair or replacement of damaged tissues. In this work we investigated the interaction between magnetic carbon nanotubes (CNTs) and MSCs and their ability to guide these cells injected intravenously in living mice by using an external magnetic field. CNTs did not affect cell viability and their ability to differentiate. Both the CNTs and the magnetic field did not alter cell growth rate, phenotype and cytoskeletal conformation. CNTs, when exposed to magnetic fields, are able to shepherd MSCs towards the magnetic source in vitro. Moreover, the application of a magnetic field alters the biodistribution of CNT-labelled MSCs after intravenous injection into rats. We demonstrated that CNTs hold the potential for use as nano-devices to improve therapeutic protocols for transplantation and homing of stem cells in vivo. This could pave the way for the development of new strategies for manipulation/guidance of MSCs in regenerative medicine and cell transplantation for the treatment of many orthopaedic diseases

Magnetic resonance images (MRI) were obtained of 10 healthy volunteers and 70 patients suffering from various orthopaedic disorders. Selected images of soft tissue, joint, bone and spinal abnormalities are presented and their interpretation is described. Although we have been using MRI for only a very short time, it is already possible to see its advantages: it provides good images of soft-tissues, detailed pictures of bone marrow, and excellent visualisation of the spine and spinal cord. The decision-making process in surgical procedures will in the future be influenced by this technique

A treatment regime using electrical stimulation in association with a variety of surgical procedures has improved the prognosis in congenital pseudarthrosis of the tibia--one of the most challenging of all orthopaedic disorders. The technique consists of correction of the tibial deformity, intramedullary fixation and cancellous bone grafting, augmented by electrical stimulation using an implanted bone-growth stimulator. Experience with 27 pseudarthroses in 25 patients is presented; of those, 20 have joined. The cases have been reviewed and the causes of failure analysed. These results offer encouragement to the orthopaedic surgeon treating this difficult condition

In vitro studies on isolated bone cells were undertaken to investigate the presence of transplantation (histocompatibility) antigens. Bone cells were cultured with allogeneic lymphocytes and exposed to cytotoxic sera containing antibodies against transplantation antigens, to determine their antigenic profile. Preliminary results suggest that bone cells may not express lymphocyte stimulating antigens in an active form, at least after the isolation procedure performed. On the other hand, bone cells were killed by cytotoxic antibodies in a specific way, providing evidence for the presence of serologically defined (SD) transplantation antigens on the cell surface. Additional studies with absorbed sera suggest "sharing" of histocompatibility antigens between bone cells and lymphocytes. The relevance of the surface antigens of bone cells to clinical fields such as bone allotransplantation, susceptibility to various orthopaedic diseases and skeletal sarcomata is discussed

Aims

The aim of this study was to produce clinical consensus recommendations about the non-surgical treatment of children with Perthes’ disease. The recommendations are intended to support clinical practice in a condition for which there is no robust evidence to guide optimal care.

Aims

Lumbar spinal stenosis (LSS) is a common skeletal system disease that has been partly attributed to genetic variation. However, the correlation between genetic variation and pathological changes in LSS is insufficient, and it is difficult to provide a reference for the early diagnosis and treatment of the disease.

Aims

We investigated the prevalence of late developmental dysplasia of the hip (DDH), abduction bracing treatment, and surgical procedures performed following the implementation of universal ultrasound screening versus selective ultrasound screening programmes.

Aims

It is well described that patients with bone and joint infections (BJIs) commonly experience significant functional impairment and disability. Published literature is lacking on the impact of BJIs on mental health. Therefore, the aim of this study was to assess health-related quality of life (HRQoL) and the impact on mental health in patients with BJIs.