Painful OA is linked to CNS changes in pain processing. Temporal summation of pain (TSP) is a measure of one such CNS change, central sensitization. TSP is defined using a series (≥0.33Hz) of painful stimuli and is a predictor of postoperative pain, experienced by 20% of patients after total knee replacement (TKR) surgery. This study has developed a protocol to use functional MRI to assess CNS changes in OA pain processing. This pilot includes 3 participants with chronic knee OA pain awaiting TKR (62 ± 4.4) and 5 healthy volunteers (50 ± 13.6). 3-Tesla BOLD fMRI brain scans were recorded during short series of one second painful stimuli, applied using an automated inflatable cuff to the calf muscle of the leg with the affected knee or left side in healthy volunteers. The pain intensity at onset and during the 10 painful stimuli were recorded using a numerical rating scale. The pattern of brain activation was averaged across noxious stimuli, and the differential activation compared the 1st vs. 10th (last) stimulus. Bone marrow lesions (BMLs), synovitis and effusion size were scored from 3-Tesla knee MRI's using MOAKS scoring. TSP was raised in OA patients compared to control group (p=0.023). TSP brain activity in the chronic OA patients displayed higher signal within the subgenual anterior cingulate (sgACC) compared to healthy volunteers. Knee MRI identified OA patient's exhibited higher BML scores (p=0.038) and more knee effusion (p=0.018), but the lack of synovitis did not differ from control group (p=0.107). Enhanced TSP in chronic knee OA pain may be linked with augmented responses in emotional circuitry. BMLs and effusion size appear to contribute more with pain than synovitis. These results may help understand sensitization to improve outcomes for patients with knee OA undergoing TKR surgery

Though dentin matrix protein 1 (Dmp1) is known to play critical role in mediating bone mineralization, it has also been validated to be expressed in brain and helps maintain blood brain barrier (BBB). Our study aims to clarify the expression pattern of Dmp1 in mouse brain and explore whether intercellular mitochondrial transfer occurs between Dmp1 positive astrocytes (DPAs) and endothelial cells, and thus acting as a mechanism in maintaining BBB during aging. Single cell RNA sequencing (scRNAseq) of 1 month, 6 month, and 20 month old mice brain (n=1, respectively) was employed to identify Dmp1 positive cell types. Dmp1. Cre. -mGmT and Dmp1. Cre. -COX8a fluorescent mice were generated to visualize DPAs and investigate their mitochondrial activities. A 3D noncontact coculture system and mitochondrial transplantation were applied to study the role of mitochondrial transfer between astrocytes and bEnd.3 endothelial cells. Dmp1. Cre. -Mfn2. f/f. mice were generated by depleting the ER-mitochondria tethering protein Mfn2 in DPAs. Dmp1 was mainly expressed in astrocytes at different ages. GO analysis revealed that cell projection and adhesion of DPAs were upregulated. Confocal imaging on Dmp1. Cre. -mGmT mice indicated that DPAs are a cluster of astrocytes that closely adhere to blood vessels (n=3). Bioinformatics analysis revealed that mitochondrial activity of DPAs were compromised during aging. Enriched scRNAseq of fluorescent cells from Dmp1. Cre. -COX8a mice (n=2) and immunofluorescent imaging (n=3) validated the acquisition of extrinsic mitochondria in endothelial cells. 3D coculture of astrocytes and bEnd.3 and direct mitochondrial transplantation revealed the rescue effect of mitochondrial transfer on damaged bEnd.3. BBB was impaired after depleting Mfn2 in DPAs, expressing a similar phenotype with aging brain. Astrocytes that express Dmp1 play a significant role in maintaining BBB via transferring mitochondria to vascular endothelial cells. Compromised mitochondrial transfer between DPAs and endothelial cells might be the potential mechanism of impaired BBB during aging

Based on Ilizarov's law of tension-stress principle, distraction histogenesis technique has been widely applied in orthopaedic surgery for decades. Derived from this technique, cranial bone transport technique was mainly used for treating cranial deformities and calvarial defects. Recent studies reported that there are dense short vascular connections between skull marrow and meninges for immune cells trafficking, highlighting complex and tight association between skull and brain. Alzheimer's disease (AD) is a progressive neurodegenerative disease and the most common cause of dementia without effective therapy. Meningeal lymphatics have been recognized as an important mediator in neurological diseases. The augmentation of meningeal lymphatic drainage might be a promising therapeutic target for AD. Our proof-of-concept study has indicated that cranial bone transport can promote ischemic stroke recovery via modulating meningeal lymphatic drainage function, providing a rationale for treating AD using cranial bone maneuver (CBM). This study aims to investigate the effects of CBM on AD and to further explore the potential mechanisms. Transgenic 5xFAD mice model was used in this study. After osteotomy, a bone flap was used to perform CBM without damaging the dura. Open filed test, novel object recognition test and Barn's maze test were used to evaluate neurological functions of 5xFAD mice after CBM treatment. Congo red and immunofluorescence staining were used to evaluate amyloid depositions and Aβ plaques in different brain regions. Lymphangiogenesis and the level of VEGF-C were examined after CBM treatment. OVA-A647 was intra-cisterna-magna injected to evaluate meningeal lymphatic drainage function after CBM treatment. CBM significantly improved memory functions and reduced amyloid depositions and Aβ plaques in the hippocampus of 5xFAD mice. A significant increase of meningeal lymphatic vessels in superior sagittal sinus and transverse sinus, and the upregulation of VEGF-C in meninges were observed in 5xFAD mice treated with CBM. Moreover, CBM remarkably enhanced meningeal lymphatic drainage function in 5xFAD mice (n=5-16 mice/group for all studies). CBM may promote meningeal lymphangiogenesis and lymphatic drainage function through VEGF-C-VEGFR3 pathway, and further reduce amyloid depositions and Aβ plaques and alleviate memory deficits in AD

Serial section electron microscopy (SSEM) was initially developed to map the neural connections in the brain. SSEM eventually led to the term ‘Connectomics’ to be coined to describe process of following a cell or structure through a volume of tissue. This permits the true three-dimensionality to be appreciated and relationships between cells and structures. The purpose of this study was to utilize this methodology to interrogate S. aureus infected bone. Bone samples were harvested from mice tibia infected with S. aureus and were fixed, decalcified, and osmicated. The samples were paraffin embedded and 5-micron sections were cut to identify regions of bacterial invasion into the osteocyte-lacuna-canalicular-network (OLCN). This area was cut from the paraffin block, deparaffinized, post-fixed and reprocessed into epoxy resin. Serial sections were cut at 60nm and collected onto Kapton tape utilizing the Automated Tape-collecting Ultramicrotome (ATUMtome) system. Samples were mounted onto 4” silicon wafers and post-stained with 2% uranyl acetate followed by 0.3% lead citrate and carbon coated. A ZEISS GeminiSEM 450 scanning electron microscope fitted with an electron backscatter diffusion detector was used to image the sections. The image stack was aligned and segmented using the open-source software, VASTlite. 264 serial sections were imaged, representing approximately 40 × 45 × 15-micron (x, y, z) volume of tissue. 70% of the canaliculi demonstrated infiltration by S. aureus. This study demonstrates that SSEM can be applied to the skeletal system and provide a new solution to investigate the OLCN system. It is feasible that this methodology could be implemented to investigate why some canaliculi are resistant to colonization and potentially opens up a new direction for the prevention of chronic osteomyelitis. In order to make this a realistic target, automated segmentation methodologies utilizing machine learning must be developed and applied to the bone tissue datasets

Introduction and Objective. Klinefelter Syndrome (KS, karyotype 47,XXY) is the most frequent chromosomal aneuploidy in males, as well as the most common cause of infertility in men. Patients suffer from a lack of testosterone, i.e. hypergonadotropic hypogonadism provoking infertility, but KS men also show an increased predisposition to osteoporosis and a higher risk of bone fracture. In a mouse model for human KS, bone analysis of adult mice revealed a decrease in bone mass that could not be rescued by testosterone replacement, suggesting a gene dosage effect originating from the supernumerary X-chromosome on bone metabolism. Usually, X chromosome inactivation (XCI) compensates for the dosage imbalance of X-chromosomal genes between sexes. Some studies suggested that expression of genes that escape silencing of the supernumerary X-chromosome (e.g. androgen receptor) has an impact on sex differences, but may also cause pathological changes in males. As a promising new such candidate for a musculoskeletal escape gene, we identified the integral membrane protein (ITM) 2a, which is encoded on the X-chromosome and related to enchondral ossification. The aim of the project was to characterize systemic bone loss in the course of aging in our KS mouse model, and whether the supernumerary X-chromosome causes differences in expression of genes related to bone development. Materials and Methods. Bone structure of 24 month (=aged) old male wild type (WT) and 41, XXY mice (B6Ei.Lt-Y) were analysed by μCT. Afterwards bones were paraffin embedded and cut. In addition, tissue of brain, liver, kidney, lung and heart were also isolated and embedded for IHC staining. Using an anti-ITM2a antibody, expression and cellular localization of ITM2a was evaluated. IHC was also performed on musculoskeletal tissue of WT embryos (E18.5) and neonatal mice to determine possible age-related differences. Results. In 24 month old mice, the analysis of the lumbar vertebrae revealed a significantly lower BV/TV, trabecular bone volume and trabecular number in the XXY- group compared to WT. Trabecular thickness appeared lower but did not reach significance, with the cortical thickness being significantly higher in the XXY- group. High expression of ITM2a was detected in bone slices of both karyotypes in the chondrocytes inside the growth plate, as well as in megakaryocytes and leucocytes as well as endothelial cells of blood vessels inside the bone marrow. Osteocytes, along with erythrocytes and erythropoetic stem cells were negative for ITM2a. Other organs that showed ITM2a positive staining were kidney (blood vessels), heart (muscle) and brain (different structures). Liver and lung tissue were negative for ITM2a. No obvious difference in the intensity of the ITM2a-expression was observed between the WT and the XXY-karyotype. Analyses of embryotic bone tissue (WT) showed high expression of ITM2a in proliferating, hypertrophic and resting chondrocytes in the growth plates of tibia and femur. In comparison, the neonatal animals (WT) did not show any protein-expression in chondrocytes. Furthermore, within the metaphysis of both, embryotic and neonatal bones, endothelial cells and osteoblasts were ITM2a-positive. Further analyses of bones and tissues from young mice (4–6 month) are ongoing. Conclusions. Bone analyses revealed a significant reduction in trabecular bone mass along with fewer and thinner trabeculae in XXY mice compared to the WT, especially in the spine. ITM2a expression was visible in different cell types inside the bone, and in addition, different expression patterns at different stages of development (embryonic/neonatal) were observed. However, we have not found a significant difference in the quantity of ITM2a between tissues of XXY-karyotypes and WT. Further analyses of X-chromosomal encoded and therefore dysregulated modulators in XXY-karyotype mice and patients may reveal new sex chromosomal effector proteins in bone metabolism

Introduction. Low back pain (LBP) is a worldwide leading cause of disability. This preclinical study evaluated the safety of a combined advanced therapy medicinal product developed during the European iPSpine project (#825925) consisting of mesendoderm progenitor cells (MEPC), derived from human induced pluripotent stem cells, in combination with a synthetic poly(N-isopropylacrylamide) hydrogel (NPgel) in an ovine intervertebral disc degeneration (IDD) model. Method. IDD was induced through nucleotomy in 4 adult sheep, 5 lumbar discs each (n=20). After 5 weeks, 3 alternating discs were treated with NPgel (n=6) or NPgel+MEPC (n=6). Before sacrifice, animals were subjected to: MRI of lumbar spines (disc height and Pfirmann grading); blood sampling (hematological, biochemical, metabolic and lymphocyte/monocytes immunological). After 3 months the sheep were sacrificed. The spines were processed for: macroscopic morphology (Thompson grading), microscopic morphology (Histological grading), and glycosaminoglycan content (GAG, DMMB Assay). Furthermore, at sacrifice biodistribution of human MEPC was assessed by Alu-sequences quantification (qPCR) from three tissue samples of heart, liver, spleen, brain, lungs, and kidneys, and PBMCs collected to assess activation of systemic immune cells. To each evaluation, appropriate statistical analysis was applied. Result. Flow cytometry showed no induction of systemic activation of T cells or monocytes. Alu quantification did not give detection of any cells in any organ. Disc height index was slightly increased in discs treated with NPgel+MEPC. Pfirmann's and Thompson's classification showed that treatment with NPgel or NPgel+MEPC gave no adverse reactions. Histological grading showed similar degeneration in vertebrae treated with NPgel+MEPC or with NPgel alone. The amount of GAG was significantly increased in the nucleus pulposus following treatment with NPgel+MEPC compared to NPgel alone, in which a decrease was observed compared to untreated discs in both nucleus pulposus and annulus fibrosus. Conclusion. This study showed the safety of both NPgel+MEPC and NPgel treatments

Several experimental models have been used to produce intravascular fat embolism. We have developed a simple technique to induce fat embolism using corn oil emulsified with distilled water to form fatty micelles. Fat embolism was produced by intravenous administration of these fatty micelles in anaesthetised rats, causing alveolar oedema, haemorrhage and increased lung weight. Histopathological examination revealed fatty droplets and fibrin thrombi in the lung, kidney and brain. The arteriolar lumen was filled with fatty deposits. Following fat embolism, hypoxia and hypercapnia occurred. The plasma phospholipase A. 2. , nitrate/nitrite, methylguidanidine and proinflammatory cytokines were significantly increased. Mass spectrometry showed that the main ingredient of corn oil was oleic acid. This simple technique may be applied as a new animal model for the investigation of the mechanisms involved in the fat embolism syndrome

Osteoarthritis is the most prevalent joint disease, causing severe pain, deformity and a loss of mobility. Low back pain (LBP), frequently associated with degeneration of the intervertebral disc (IVD), is the No.1 cause of Years Lived with Disability. Age is a major risk factor for both conditions. However, the reasons why susceptibility to these conditions increases with age are poorly understood. The circadian (24 hourly) clocks in the brain and periphery direct key aspects of physiology through rhythmic control of tissue-specific sets of downstream genes. Work from our group focuses on the roles of circadian clocks in the articular cartilage and IVD. We show that the daily rhythm in these tissues becomes dampened and out-of-phase during ageing. Further, our data identify circadian clock disruption in cartilage and IVD as a new target of inflammation. Moreover, we show that mice with targeted knockout of an essential clock gene (BMAL1) in chondrocytes and disc cells have profound, yet tissue-specific degeneration in the articular cartilage and IVD. These findings implicate the local skeletal clock as a key regulatory mechanism for tissue homeostasis. This new avenue of research holds potential to better understand, and eventually treat these debilitating conditions

When joints sustain injury, the release of inflammation cytokines can cleavage matrix proteins and result in cartilage degradation and the subsequent osteoarthritis. RNA therapeutics emerging recently is a very promising approach to efficiently and specifically inhibit disease gene expression. However, the major challenge is how to deliver therapeutic RNA into joint and cartilage. Janus base nanotubes are self-assembled from synthetic Janus bases inspired from DNA base pairs. Based on the charge interaction, we are able to “sandwich” small RNAs among Janus base nanotubes to form tiny, nano-rod shaped delivery vehicles. Such vehicles can be engineered into different sizes and shapes. We have found that short and slim morphologies can greatly increase their penetration to extracellular matrix and delivery into “difficult-to-reach” tissues, such as cartilage and brain. Moreover, by delivering therapeutic siRNA, we have demonstrated its high-efficacy in inhibiting expression of an inflammatory regulator, Interleukin-1 receptor (IL-1R) in articular cartilage. Moreover, the inhibition effect is long-lasting so that joint inflammation and cartilage degradation caused by meniscus injury are greatly inhibited in a mouse model. Therefore, the Janus base nanotubes present a great potential in engineering into nano-structures for RNA delivery. Such approach may become an effective therapeutic against joint inflammation and arthritis

Dura mater is a thick membrane that is the outermost of the three layers of the meninges that surround the brain and spinal cord. Appropriate dural healing is crucial to prevent cerebrospinal fluid leaks but the entire process has been barely understood so far. Understanding of dural healing and tissue neoformation over the dural grafts, which are usually used for duraplasty, is still partial. Therefore, implantation of decellular dura mater (DM) to recipient from different donor and vitalization with recipient”s mesenchymal stem cells for the treatment of tissue on transplantation process is significant approach. This approach prevents immunological reactions and provides long-term stabilization. According to this study, it is believed that this approach will provide DM healing and become crucial in DM transplantation. The aim of this study was to develop a new construct by tissue engineering of the human DM based on a decellular allograft. Thus human DM collected from forensic medicine and decellularized using the detergent sodium dodecyl sulfate (SDS) in the multiple process of physical, enzimatic and chemical steps. Decellularization were exposing the tissue to freeze-thaw cycles, incubation in hypotonic tris-HCl buffer, 0.1% (w/v) SDS in hypotonic buffer and hypertonic buffer followed by disinfection using 0.1% (v/v) peracetic acid and final washing in phosphate-buffered saline. As a result of all these processes, cellular components of DM were removed by preserving the extracellular matrix without any significant loss in mechanical properties. Based on the histological analysis of the decellularized DM revealed the absence of visible whole cells. Collagen and glycosaminoglycan (GAG) contents of decellular DM evaluated histological staining by Masson Trichrome and Alcian blue respectively. Also biochemical tests were carried out by spectrophotometry (Quickzym Biosciences, The Netherlands) and total GAG content were analyzed by 1.9 dimethylmethylene blue assay. The histoarchitecture was unchanged, and there were no significant changes of total collagen and GAG content. Biomechanical properties were determined by tensile tests, which has confirmed the retention of biomechanical properties following decellularization. The mean tensile strengths were 7,424±4,20 MPa for control group, 5,254±2,068 MPa for decellularization group. There was no statistically significant difference between tensile strength (p=0,277) and tissue thickness (p=0, 520) for both group. In conclusion, this study has developed biomechanically functional decellularized DM scaffold for use in DM repair. In addition, this study is a part of the progressing study and additional studies investigating the biocompatibility performance of the decellularized DM scaffold and there is need for in vivo studies. Keywords. Dura mater, Decellularization, Allografts, Scaffolds, Tissue Engineering

Background. Traditionally, a Surgical Tourniquet (ST) is used during Total Knee Replacement Surgery (TKRS) to prevent blood flow to the leg and improve the surgical field of view. The use of a ST is known to increase the risk of venous thromboembolism. Echogenic material, suggestive of emboli has been observed in the brain following ST deflation in TKRS despite the absence of a patent foramen ovale, likely through pulmonary shunts. The aim of this study was to assess whether cerebral emboli result from tourniquet use in TKRS and the sequelae of any emboli. Methods. 11 subjects from a single centre undergoing routine TKRS with a ST gave informed consent. Each participant had diffusion weighted MR brain imaging prior to, and within 48 hours after TKRS and completed pre and post-operative mini-mental state examinations (MMSE). Results. Pre and post-operative MR imaging were assessed by a senior radiologist and the results were reported according to Age and Cognitive Performance Research Centre (ACPRC) rating scale. There were no changes from pre and post-operative MR scans. 4 participants (36%) had no change in MMSE scores. These participants had a score of 1 or 0 in both MR scans. 7 participants (63%) had a score of 2 or 3. These participants had mean 2.8 point (9.5%) drop on MMSE (IQR = 1) within 48 hours. Conclusions. In this small, exploratory study we found no evidence of discrete cerebral emboli occurring with the use of ST in TKRS. Interestingly, participants noted to have minimal or no pre-operative ischaemic change appeared to maintain more cognitive function post-operatively than those with higher levels of pre-existing ischaemia. Subtle changes in ischaemic load in patients with pre-existing ischaemia may lead to impaired cognitive function, however further evidence is required to confirm this theory. Level of Evidence. III. Approval. Protocol approved by NRES Committee Yorkshire & The Humber, Leeds West and The Research, Development & Innovation Department University Hospitals Coventry & Warwickshire

Introduction. Changes in central nervous system (CNS) pathways controlling trunk and leg muscles in patients with low back pain(LBP) and lumbar radiculopathy have been observed and this study investigated whether surgery impacts upon these changes in the long term. Methods. 80 participants were recruited into the following groups: 25 surgery(S), 20 chronic LBP(CH), 14 spinal injection(SI), and 21 controls(C). Parameters of corticospinal control were examined before, at 6, 26 and 52 weeks following lumbar decompression surgery and equivalent intervals. Electromyographic(EMG) activity was recorded from tibialis anterior(TA), soleus(SOL), rectus abdominis(RA), external oblique(EO) and erector spinae(ES) muscles at the T12&L4 levels in response to transcranial magnetic stimulation of the motor cortex. Motor evoked potentials (MEP) and cortical silent periods(cSP) recruitment curves(RC) were analysed. Results. Trunk muscles in all patients had reduced raw EMG (P<0.001), increased motor thresholds (MTh;P<0.001) and MEP RC slopes. MTh in ESL4 correlated with back pain in all patients (r=0.201, P=0.016) and soleus MTh laterality with disability in surgery patients (r=0.49, P=0.018). S&SI patients displayed bilaterally increased soleus cSP (p<0.001), MEP latencies on the painful side (P<0.001), and cSP asymmetry (cSPA;P<0.001). cSPA resulted from abnormal soleus late responses on the painful side, indicating compromised agonist-antagonist control in patients with radiculopathy. In contrast to SI, surgery significantly reduced soleus cSPA and MEP latencies at 6 weeks (P≤0.034). Discussion. These results show long term changes in CNS control of trunk and leg muscles in radiculopathy and LBP, which are only partly reversed by surgery, and may provide future therapeutic targets to address the altered inhibitory processes within the brain. No conflicts of Interest. Sources of funding: The DISCS foundation. This abstract has not been previously published in whole or substantial part nor has it been presented previously at a national meeting

Summary Statement. Conventional culture techniques have poor sensitivity for detecting bacteria growing in biofilms, which can result in under-diagnosis of infections. Sonication of biofilm colonised orthopaedic biomaterials can render bacteria in biofilm more culturable, thereby improving diagnosis of orthopaedic implant infections. Introduction. Prosthetic joint infection (PJI) is a potentially devastating complication in arthroplasty. Biofilm formation is central to PJI offering protection to the contained bacteria against host defence system and antimicrobials. Orthopaedic biomaterials generally have a proclivity to biofilm colonisation. Conventional culture technique has a low sensitivity for detecting bacteria in biofilm. Sonication can disrupt bacteria biofilms aggregations and dislodge them from colonised surfaces, rendering them culturable and consequently improve the diagnosis of otherwise culture-negative PJI. We investigated the effect of ultrasonication on biofilms adherent to poylmethylmethacrylate PMMA cement. Method. Identical PMMA cement beads were aseptically prepared using 7mm bead templates. Each sample comprised of two beads and with multiple replicates made for each sample. Two proficient biofilm forming strains of Staphylococcus epidermidis (5179-R1 and 1457) were used for the experiments. Each set of cement sample was immersed in Brain Heart Infusion broth inoculated with a pre-culture of the chosen bacteria strains (final concentration approximately 4 × 10. 6. CFU/ml). All samples were then incubated for 24 hours at 37°C to allow for biofilm growth and colonisation of the cement surfaces, as well as for biofilm maturity. After incubation, each sample was washed twice with sterile phosphate buffer saline (PBS) to remove non-adherent and loosely adherent bacteria. The cement beads were transferred to a fresh sterile bottle at each stage of the experiment, while ensuring the maintenance of asepsis. After the final wash, 10ml of sterile PBS was added to the cement beads and each sample was sonicated for varying periods: 0min, 5min, 10min, 20min and 40min. Sonicate fluid were collected after each period of sonication, with which culture plates were inoculated for the purpose of viable bacteria counting. Results. The optimum sonication period was between 5min and10 min. The mean pre-sonication CFU/ml were 4.7 × 10. 5. and 8.3 × 10. 5. for bacteria strains 5179-R1 and 1457 respectively, while the mean CFU/ml after 10min of sonication were 1.4 × 10. 7. and 0.74 × 10. 7. for bacteria strains the respective bacteria strains. Discussion / Conclusion. Our study showed a significant increase (almost 100 fold) in bacteria culture yield following sonication. We were also able to demonstrate that the optimum duration for sonication (using comparable sonicators) was approximately 10min. Sonication was able to completely remove adherent bacteria from the surfaces of our cement samples allowing them to be cultured. Our result suggests that sonication of bone cement can be instrumental in improving the diagnosis of biofilm associated PJI

Objectives

In order to ensure safety of the cell-based therapy for bone regeneration, we examined in vivo biodistribution of locally or systemically transplanted osteoblast-like cells generated from bone marrow (BM) derived mononuclear cells.

Objectives

Cellular movement and relocalisation are important for many physiologic properties. Local mesenchymal stem cells (MSCs) from injured tissues and circulating MSCs aid in fracture healing. Cytokines and chemokines such as Stromal cell-derived factor 1(SDF-1) and its receptor chemokine receptor type 4 (CXCR4) play important roles in maintaining mobilisation, trafficking and homing of stem cells from bone marrow to the site of injury. We investigated the differences in migration of MSCs from the femurs of young, adult and ovariectomised (OVX) rats and the effect of CXCR4 over-expression on their migration.

High-intensity narrow-spectrum (HINS) light is a novel violet-blue light inactivation technology which kills bacteria through a photodynamic process, and has been shown to have bactericidal activity against a wide range of species. Specimens from patients with infected hip and knee arthroplasties were collected over a one-year period (1 May 2009 to 30 April 2010). A range of these microbial isolates were tested for sensitivity to HINS-light. During testing, suspensions of the pathogens were exposed to increasing doses of HINS-light (of 123mW/cm² irradiance). Non-light exposed control samples were also used. The samples were then plated onto agar plates and incubated at 37°C for 24 hours before enumeration. Complete inactivation (greater than 4-log₁₀ reduction) was achieved for all of the isolates. The typical inactivation curve showed a slow initial reaction followed by a rapid period of inactivation. The doses of HINS-light required ranged between 118 and 2214 J/cm². Gram-positive bacteria were generally found to be more susceptible than Gram-negative.

As HINS-light uses visible wavelengths, it can be safely used in the presence of patients and staff. This unique feature could lead to its possible use in the prevention of infection during surgery and post-operative dressing changes.

Cite this article: Bone Joint J 2015;97-B:283–8.

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.

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.

Platelet-leucocyte gel (PLG), a new biotechnological blood product, has hitherto been used primarily to treat chronic ulcers and to promote soft-tissue and bone regeneration in a wide range of medical fields. In this study, the antimicrobial efficacy of PLG against Staphylococcus aureus (ATCC 25923) was investigated in a rabbit model of osteomyelitis. Autologous PLG was injected into the tibial canal after inoculation with Staph. aureus. The prophylactic efficacy of PLG was evaluated by microbiological, radiological and histological examination. Animal groups included a treatment group that received systemic cefazolin and a control group that received no treatment.

Treatment with PLG or cefazolin significantly reduced radiological and histological severity scores compared to the control group. This result was confirmed by a significant reduction in the infection rate and the number of viable bacteria. Although not comparable to cefazolin, PLG exhibited antimicrobial efficacy in vivo and therefore represents a novel strategy to prevent bone infection in humans.

A rat model of lumbar root constriction with an additional sympathectomy in some animals was used to assess whether the sympathetic nerves influenced radicular pain. Behavioural tests were undertaken before and after the operation.

On the 28th post-operative day, both dorsal root ganglia and the spinal roots of L4 and L5 were removed, frozen and sectioned on a cryostat (8 μm to 10 μm). Immunostaining was then performed with antibodies to tyrosine hydroxylase (TH) according to the Avidin Biotin Complex method. In order to quantify the presence of sympathetic nerve fibres, we counted TH-immunoreactive fibres in the dorsal root ganglia using a light microscope equipped with a micrometer graticule (10 x 10 squares, 500 mm x 500 mm). We counted the squares of the graticule which contained TH-immunoreactive fibres for each of five randomly-selected sections of the dorsal root ganglia.

The root constriction group showed mechanical allodynia and thermal hyperalgesia. In this group, TH-immunoreactive fibres were abundant in the ipsilateral dorsal root ganglia at L5 and L4 compared with the opposite side. In the sympathectomy group, mechanical hypersensitivity was attenuated significantly.

We consider that the sympathetic nervous system plays an important role in the generation of radicular pain.