We have examined the process of fusion of the intertransverse processes and bone graft in the rabbit by in situ hybridisation and evaluated the spatial and temporal expression of genes encoding pro-α1 (I) collagen (COL1A1), pro-α1 (II) collagen (COL2A1) and pro-α1 (X) collagen (COL10A1). Beginning at two weeks after operation, osteogenesis and chondrogenesis occurred around the transverse process and the grafted bone at the central portion of the area of the fusion mass. Osteoblasts and osteocytes at the newly-formed woven bone expressed COL1A1. At the cartilage, most chondrocytes expressed COL2A1 and some hypertrophic chondrocytes COL10A1. In some regions, co-expression of COL1A1 and COL2A1 was observed. At four weeks, such expressions for COL1A1, COL2A1 and COL10A1 became prominent at the area of the fusion mass. From four to six weeks, bone remodelling progressed from the area of the transverse processes towards the central zone. Osteoblasts lining the trabeculae expressed a strong signal for COL1A1. At the central portion of the area of the fusion mass, endochondral ossification progressed and chondrocytes expressed COL2A1 and COL10A1. Our findings show that the fusion process begins with the synthesis of collagens around the transverse processes and around the grafted bone independently. Various spatial and temporal osteogenic and chondrogenic responses, including intramembranous, endochondral and transchondroid bone formation, progress after bone grafting at the intertransverse processes. Bone formation through cartilage may play an important role in posterolateral spinal fusion

Periosteum is important for bone homoeostasis through the release of bone morphogenetic proteins (BMPs) and their effect on osteoprogenitor cells. Smoking has an adverse effect on fracture healing and bone regeneration. The aim of this study was to evaluate the effect of smoking on the expression of the BMPs of human periosteum. Real-time polymerase chain reaction was performed for BMP-2,-4,-6,-7 gene expression in periosteal samples obtained from 45 fractured bones (19 smokers, 26 non-smokers) and 60 non-fractured bones (21 smokers, 39 non-smokers). A hierarchical model of BMP gene expression (BMP-2 > BMP-6 > BMP-4 > BMP-7) was demonstrated in all samples. When smokers and non-smokers were compared, a remarkable reduction in the gene expression of BMP-2, -4 and -6 was noticed in smokers. The comparison of fracture and non-fracture groups demonstrated a higher gene expression of BMP-2, -4 and -7 in the non-fracture samples. Within the subgroups (fracture and non-fracture), BMP gene expression in smokers was either lower but without statistical significance in the majority of BMPs, or similar to that in non-smokers with regard to BMP-4 in fracture and BMP-7 in non-fracture samples. In smokers, BMP gene expression of human periosteum was reduced, demonstrating the effect of smoking at the molecular level by reduction of mRNA transcription of periosteal BMPs. Among the BMPs studied, BMP-2 gene expression was significantly higher, highlighting its role in bone homoeostasis

In order to clarify the role of cytokines in the remodelling of the grafted tendon for ligament reconstruction we compared the responses to interleukin (IL)-1β, platelet-derived growth factor (PDGF)-BB and transforming growth factor (TGF)-β1 of extrinsic fibroblasts infiltrating the frozen-thawed patellar tendon in rats with that of the normal tendon fibroblasts, in regard to the gene expression of matrix metalloproteinase (MMP)-13, using Northern blot analysis. We also examined, immunohistologically, the local expression of IL-1β, PDGF-BB, and TGF-β1 in fibroblasts infiltrating the frozen-thawed patellar tendon. Northern blot analysis showed that fibroblasts derived from the patellar tendon six weeks after the freeze-thaw procedure in situ showed less response to IL-1β than normal tendon fibroblasts with respect to MMP-13 mRNA gene expression. The immunohistological findings revealed that IL-1β was over-expressed in extrinsic fibroblasts which infiltrated the patellar tendon two and six weeks after the freeze-thaw procedure in situ, but neither PDGF-BB nor TGF-β1 was over-expressed in these extrinsic fibroblasts. Our findings indicated that IL-1β had a close relationship to matrix remodelling of the grafted tendon for ligament reconstruction, in addition to the commencement of inflammation during the tissue-healing process

Gene therapy with insulin-like growth factor-1 (IGF-1) increases matrix production and enhances chondrocyte proliferation and survival in vitro. The purpose of this study was to determine whether arthroscopically-grafted chondrocytes genetically modified by an adenovirus vector encoding equine IGF-1 (AdIGF-1) would have a beneficial effect on cartilage healing in an equine femoropatellar joint model. A total of 16 horses underwent arthroscopic repair of a single 15 mm cartilage defect in each femoropatellar joint. One joint received 2 × 10. 7. AdIGF-1 modified chondrocytes and the contralateral joint received 2 × 10. 7. naive (unmodified) chondrocytes. Repairs were analysed at four weeks, nine weeks and eight months after surgery. Morphological and histological appearance, IGF-1 and collagen type II gene expression (polymerase chain reaction, in situ hybridisation and immunohistochemistry), collagen type II content (cyanogen bromide and sodium dodecyl sulphate-polyacrylamide gel electrophoresis), proteoglycan content (dimethylmethylene blue assay), and gene expression for collagen type I, matrix metalloproteinase (MMP)-1, MMP-3, MMP-13, aggrecanase-1, tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) and TIMP-3 were evaluated. Genetic modification of chondrocytes significantly increased IGF-1 mRNA and ligand production in repair tissue for up to nine weeks following transplantation. The gross and histological appearance of IGF-1 modified repair tissue was improved over control defects. Gross filling of defects was significantly improved at four weeks, and a more hyaline-like tissue covered the lesions at eight months. Histological outcome at four and nine weeks post-transplantation revealed greater tissue filling of defects transplanted with genetically modified chondrocytes, whereas repair tissue in control defects was thin and irregular and more fibrous. Collagen type II expression in IGF-1 gene-transduced defects was increased 100-fold at four weeks and correlated with increased collagen type II immunoreaction up to eight months. Genetic modification of chondrocytes with AdIGF-1 prior to transplantation improved early (four to nine weeks), and to a lesser degree long-term, cartilage healing in the equine model. The equine model of cartilage healing closely resembles human clinical cartilage repair. The results of this study suggest that cartilage healing can be enhanced through genetic modification of chondrocytes prior to transplantation

MicroRNAs (miRNAs ) are small non-coding RNAs that regulate gene expression. We hypothesised that the functions of certain miRNAs and changes to their patterns of expression may be crucial in the pathogenesis of nonunion. Healing fractures and atrophic nonunions produced by periosteal cauterisation were created in the femora of 94 rats, with 1:1 group allocation. At post-fracture days three, seven, ten, 14, 21 and 28, miRNAs were extracted from the newly generated tissue at the fracture site. Microarray and real-time polymerase chain reaction (PCR) analyses of day 14 samples revealed that five miRNAs, miR-31a-3p, miR-31a-5p, miR-146a-5p, miR-146b-5p and miR-223-3p, were highly upregulated in nonunion. Real-time PCR analysis further revealed that, in nonunion, the expression levels of all five of these miRNAs peaked on day 14 and declined thereafter. . Our results suggest that miR-31a-3p, miR-31a-5p, miR-146a-5p, miR-146b-5p and miR-223-3p may play an important role in the development of nonunion. These findings add to the understanding of the molecular mechanism for nonunion formation and may lead to the development of novel therapeutic strategies for its treatment. Cite this article: Bone Joint J 2015; 97-B:1144–51

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

Aims. Chronic conditions of the wrist may be difficult to manage because pain and psychiatric conditions are correlated with abnormal function of the hand. Additionally, intra-articular inflammatory cytokines may cause pain. We aimed to validate the measurement of inflammatory cytokines in these conditions and identify features associated with symptoms. Patients and Methods. The study included 38 patients (18 men, 20 women, mean age 43 years) with a chronic condition of the wrist who underwent arthroscopy. Before surgery, the Self-Rating Depression Scale (SDS), Hand20 questionnaire and a visual analogue scale (VAS) for pain were used. Cytokine and chemokine levels in the synovial fluid of the wrist were measured using enzyme-linked immunosorbent assays and correlations between the levels with pain were analysed. Gene expression profiles of the synovial membranes were assessed using quantitative polymerase chain reaction. Results. Older patients had high pre-operative Hand20 scores. One-year post-operative Hand20 and VAS scores and pre-operative VAS scores correlated with SDS scores. Post-operative VAS scores negatively correlated with the expression of nerve growth factor and SDS scores positively correlated with the expression of tumour necrosis factor-alpha and negatively correlated with the expression of tumour necrosis factor-converting enzyme. Conclusion. There was a positive correlation between depression and chronic conditions of the wrist. Levels of some cytokines correlate with pain and depression. Additionally, cytokines may be important in the assessment and treatment of chronic conditions of the wrist and depression. Cite this article: Bone Joint J 2016;98-B:961–8

Prediction tools are instruments which are commonly used to estimate the prognosis in oncology and facilitate clinical decision-making in a more personalized manner. Their popularity is shown by the increasing numbers of prediction tools, which have been described in the medical literature. Many of these tools have been shown to be useful in the field of soft-tissue sarcoma of the extremities (eSTS). In this annotation, we aim to provide an overview of the available prediction tools for eSTS, provide an approach for clinicians to evaluate the performance and usefulness of the available tools for their own patients, and discuss their possible applications in the management of patients with an eSTS.

Cite this article: Bone Joint J 2022;104-B(9):1011–1016.

Adolescent idiopathic scoliosis (AIS), defined by an age at presentation of 11 to 18 years, has a prevalence of 0.47% and accounts for approximately 90% of all cases of idiopathic scoliosis. Despite decades of research, the exact aetiology of AIS remains unknown. It is becoming evident that it is the result of a complex interplay of genetic, internal, and environmental factors. It has been hypothesized that genetic variants act as the initial trigger that allow epigenetic factors to propagate AIS, which could also explain the wide phenotypic variation in the presentation of the disorder. A better understanding of the underlying aetiological mechanisms could help to establish the diagnosis earlier and allow a more accurate prediction of deformity progression. This, in turn, would prompt imaging and therapeutic intervention at the appropriate time, thereby achieving the best clinical outcome for this group of patients.

Cite this article: Bone Joint J 2022;104-B(8):915–921.

Periprosthetic bone loss after total joint arthroplasty is a major clinical problem resulting in aseptic loosening of the implant. Among many cell types, osteoblasts play a crucial role in the development of peri-implant osteolysis. In this study, we tested the effects of calcitriol (1α,25-dihydroxy-vitamin-D. 3. ) and the bisphosphonate pamidronate on titanium-particle- and TNF-α-induced release of interleukin-6 and suppression of osteoblast-specific gene expressions in bone-marrow-derived stromal cells with an osteoblastic phenotype. We monitored the expression of procollagen α1[1], osteocalcin, osteonectin and alkaline phosphatase mRNAs by Northern blots and real-time reverse transcription and polymerase chain reaction analyses. The release of various cytokines was also analysed by ELISA. We found that calcitriol or pamidronate could only partially recover the altered functions of osteoblasts when added alone. Only a combination of these compounds restored all the tested functions of osteoblasts. The local delivery of these drugs may have therapeutic potential to prevent or to treat periprosthetic osteolysis and aseptic loosening of implants

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

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

It has been suggested that matrix metalloproteinase-3 (MMP-3, stromelysin-1) has an important role in the degeneration of intervertebral discs (IVDs). A human MMP-3 promoter 5A/6A polymorphism was reported to be involved in the regulation of MMP-3 gene expression. We suggest that IVD degeneration is associated with 5A/6A polymorphism. We studied 54 young and 49 elderly Japanese subjects. Degeneration of the lumbar discs was graded using MRI in the younger group and by radiography in the elderly. 5A/6A polymorphism was determined by polymerase-chain reaction-based assays. We found that the 5A5A and 5A6A genotype in the elderly was associated with a significantly larger number of degenerative IVDs than the 6A6A (p < 0.05), but there was no significant difference in the young. In the elderly, the IVD degenerative scores were also distributed more highly in the 5A5A and 5A6A genotypes (p = 0.0029). Our findings indicate that the 5A allele is a possible risk factor for the acceleration of degenerative changes in the lumbar disc in the elderly

We compared the changes in the ratio of type-I and type-II collagen in monolayer cultures of human articular chondrocytes (HAC). HAC were isolated from samples of cartilage from normal joints and cultivated in monolayer for up to 46 days. Expression of collagen type-I and type-II was determined by immunocytochemistry, Western blotting, and the nested reverse transcription polymerase chain reaction (RT-PCR), and quantified by real-time PCR. The transition from a spherical morphology to the flattened morphology of an anchorage-dependent culture was accompanied by a rapid change in the collagen phenotype with the replacement of collagen type II by collagen type I. This was confirmed by immunocytochemistry and Western blotting between days 21 and 28. Using techniques for the analysis of gene transcription (nested RT-PCR and real-time PCR), a complete switch of collagen gene expression was not observed. Expression of collagen type I increased 100-fold during the culture time. That of collagen type II was found during the entire period and decreased more than 100-fold. The main finding was that expression of the genes encoding collagen type I and II was highly time-dependent and the ratio of collagen type II to I (CII/CI), defined as an index of cell differentiation, was significantly higher (215- to 480-fold) at the beginning of the culture. At the end of the experimental culture time, ratios between 0.1 and 1 were reached

Aims

Options for the treatment of intra-articular ligament injuries are limited, and insufficient ligament reconstruction can cause painful joint instability, loss of function, and progressive development of degenerative arthritis. This study aimed to assess the capability of a biologically enhanced matrix material for ligament reconstruction to withstand tensile forces within the joint and enhance ligament regeneration needed to regain joint function.

Aims

The impact of tobacco use on readmission and medical and surgical complications has been documented in hip and knee arthroplasty. However, there remains little information about the effect of smoking on the outcome after total shoulder arthroplasty (TSA). We hypothesized that active smokers are at an increased risk of poor medical and surgial outcomes after TSA.

Aims

Stiffness is a common complication after total knee arthroplasty (TKA). Pathogenesis is not understood, treatment options are limited, and diagnosis is challenging. The aim of this study was to investigate if MRI can be used to visualize intra-articular scarring in patients with stiff, painful knee arthroplasties.

Aims

The purpose of this study was to use pharmacogenetics to determine the frequency of genetic variants in our total knee arthroplasty (TKA) patients that could affect postoperative pain medications. Pharmacogenetic testing evaluates patient DNA to determine if a drug is expected to have a normal clinical effect, heightened effect, or no effect at all on the patient. It also predicts whether patients are likely to experience side effects from medicine. We further sought to determine if changing the multimodal programme based on these results would improve pain control or reduce side effects.

Abnormal wear of cobalt-containing metal-on-metal joints is associated with inflammatory pseudotumours. Cobalt ions activate human toll-like receptor 4 (TLR4), which normally responds to bacterial lipopolysaccharide (LPS) in sepsis. Activation of TLR4 by LPS increases the expression of chemokines IL-8 and CXCL10, which recruit leukocytes and activated T-cells, respectively. This study was designed to determine whether cobalt induces a similar inflammatory response to LPS by promoting the expression of IL-8 and CXCL10. A human monocytic cell line, derived from acute monocytic leukaemia, was treated with cobalt ions and expression of IL-8 and CXCL10 measured at mRNA and protein levels. Cobalt-treated macrophages showed a 60-fold increase in IL-8 mRNA, and an eightfold increase in production of the mature chemokine (both p < 0.001); expression of the CXCL10 gene and protein was also significantly increased by cobalt (both p < 0.001). Experiments were also performed in the presence of CLI-095, a TLR4-specific antagonist which abrogated the cobalt-mediated increase in IL-8 and CXCL10 expression.

These findings suggest that cobalt ions induce inflammation similar to that observed during sepsis by the simultaneous activation of two TLR4-mediated signalling pathways. These pathways result in increased production of IL-8 and CXCL10, and may be implicated in pseudotumour formation following metal-on-metal replacement.

Cite this article: Bone Joint J 2014; 96-B:1172–7.