Neck of femur fracture (NOF#) is the commonest reason for admission to an orthopaedic ward with 70-75,000 cases each year in the UK. 1. The femoral head is often sent to pathology if there is clinical suspicion of a malignant cause. There is limited evidence in the literature to support the efficacy of this. 2. The purpose of this project was to study the incidence of femoral head pathology analysis in NOF # patients with a background of malignancy and evaluate the impact this investigation has on guiding future management. Retrospective analysis of all neck of femur fractures admitted to the Queen Elizabeth University Hospital between 01/01/2021 and 31/12/2021. The electronic notes were accessed and for patients with past medical history of malignancy, it was confirmed whether femoral head or bone reamings were sent to pathology, resultant findings and the impact on subsequent management. In 2021, 784 patients were admitted to the QEUH with a NOF#. Of these, 770 (98.2%) underwent operative management, 138 (17.3%) of whom had a past medical history (PMH) of malignancy. Intra-operative pathology was sent from 19 (13.7%) of these 138 patients. No malignant cells were found in 13 (69%) samples, and in 6 (31%), the known active malignancy was confirmed. In all cases where samples were sent for pathology, none caused any change in management. In this retrospective study, pathological investigations in NOF# patients with a PMH of malignancy had no impact on further management. The authors would not advocate for sending pathology results in this cohort group

Neck of femur fractures are a common trauma presentation and patients with a history of malignancy are sent for long leg femur views (LLF), to exclude a distal lesion which would alter the management plan (Intra-medullary nail/Long stem Hemiarthroplasty). The aim of this is to identify incidence of malignancy on LLF views, the length of time in between each xray (XR) and to identify demographics. Data was retrospectively collected from 01/01/2021 to 31/01/2021 from a single centre. All patients admitted to the Queen Elizabeth University Hospital had their electronic records (Bluespier, PACS, Clinical Portal) accessed. These confirmed if patients had a past medical history of malignancy, if they had LLF view and the time differences between diagnostic pelvis XR and LLF XR. A total of 784 patients were identified in the specified time period. Of these, 138 were identified with a malignancy and there were 85 LLF views completed. LLF views diagnosed 1 patient with known prostate cancer that had a new distal femoral metastasis (Incidence = 1.28 cases per 1000). This patient underwent further imaging (MRI Femur) and received a long stem hip hemiarthroplasty. The average length of wait between the images was 9 hours 27 minutes. LLF views can alter management of patients with malignancy and are therefore useful to perform. There can be a long delay between each image. Therefore we recommend imaging tumour with common bony metastasis (Renal, Thyroid, Breast, Prostrate, Lung) and other remaining tumours with known secondary metastasis. Imaging primary low risk (eg basal cell carcinoma) can lead to long delays in a frail patient cohort and consideration should be given to rationalise appropriate use of resources

Abstract. Objectives. To analyse the costs and benefits of sending femoral head specimens for histopathological analysis and whether our practice had changed since the original study five years ago. Methods. The cohort definition was patients who had both undergone hip hemiarthroplasties (HHAs) and had femoral head specimens sent for histopathological analysis at our tertiary care institution from 2013 to 2016. Retrospective review of clinical and electronic notes was performed on these patients for history of malignancy, histopathological diagnosis of femoral head, indication for histopathological examination and radiological studies. The total number of HHAs performed at the centre and the costs involved in analysing each femoral head specimen were identified. Results. A total of 805 HHAs were performed at the centre. We identified 56 femoral head specimens from 56 patients (6.96%) that were sent for histopathological analysis after HHA. 29 patients (51.79%) had a known history of malignancy. Three femoral head specimens (10.34%) were histologically positive for malignancy. Two patients had hip radiographs demonstrating metastasis to the femoral head. The third patient had a PET-CT scan two weeks prior to fracture that did not show signs of metastatic disease. 27 patients (48.21%) had no history of malignancy. None of their femoral head specimens were found to be histologically positive for malignancy. 10 patients had no appropriate indication for histopathological analysis. The cost savings for 27 patients with no previous history of malignancy ranged from €2,295 to €9,504. The cost savings for 10 patients with no appropriate indication ranged from €850 to €3,520. Conclusions. Histopathological analysis of femoral head specimens after HHA has little benefit for patients without a history of malignancy. The practice at our institution had not changed significantly in the past five years. 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

Total joint replacement (TJR) was one of the most revolutionary breakthroughs in joint surgery. The majority studies had shown that most implants could last about 25 years, anyway, there is still variation in the longevity of implants. In US, for all the hip revisions from 2012 to 2017 in the United States, 12.0% of the patients were diagnosed as aseptic loosening. Variable studies have showed that any factor that could cause a systemic or partial bone loss, might be the risk of periprosthetic osteolysis and aseptic loosening. Breast cancer is the most frequent malignancy in women, more than 2.1 million women were newly diagnosed with breast cancer, 626,679 women with breast cancer died in 2018. It's been reported that the mean incidence of THA was 0.29% for medicare population with breast cancer in USA, of which the incidence was 3.46% in Norwegian. However, the effects of breast cancer chemotherapy and hormonotherapy, such as aromatase inhibitors (AI), significantly increased the risk of osteoporosis, and had been proved to become a great threat to hip implants survival. In this case, a 46-year-old female undertook chemotherapy and hormonotherapy of breast cancer 3 years after her primary THA, was diagnosed with aseptic loosening of the hip prosthesis. Her treatment was summarized and analyzed. Breast cancer chemotherapy and hormonotherapy might be a threat to the stability of THA prosthesis. More attention should be paid when a THA paitent occurred with breast cancer. More studies about the effect of breast cancer treatments on skeleton are required

Breast cancer is the most frequent malignancy in women with an estimation of 2.1 million new diagnoses in 2018. Even though primary tumours are usually efficiently removed by surgery, 20–40% of patients will develop metastases in distant organs. Bone is one of the most frequent site of metastases from advanced breast cancer, accounting from 55 to 58% of all metastases. Currently, none of the therapeutic strategies used to manage breast cancer bone metastasis are really curative. Tailoring a suitable model to study and evaluate the disease pathophysiology and novel advanced therapies is one of the major challenges that will predict more effectively and efficiently the clinical response. Preclinical traditional models have been largely used as they can provide standardization and simplicity, moreover, further advancements have been made with 3D cultures, by spheroids and artificial matrices, patient derived xenografts and microfluidics. Despite these models recapitulate numerous aspects of tumour complexity, they do not completely mimic the clinical native microenvironment. Thus, to fulfil this need, in our study we developed a new, advanced and alternative model of human breast cancer bone metastasis as potential biologic assay for cancer research. The study involved breast cancer bone metastasis samples obtained from three female patients undergoing wide spinal decompression and stabilization through a posterior approach. Samples were cultured in a TubeSpin Bioreactor on a rolling apparatus under hypoxic conditions at time 0 and for up to 40 days and evaluated for viability by the Alamar Blue test, gene expression profile, histology and immunohistochemistry. Results showed the maintenance and preservation, at time 0 and after 40 days of culture, of the tissue viability, biological activity, as well as molecular markers, i.e. several key genes involved in the complex interactions between the tumour cells and bone able to drive cancer progression, cancer aggressiveness and metastasis to bone. A good tis sue morphological and microarchitectural preservation with the presence of lacunar osteolysis, fragmented trabeculae locally surrounded by osteoclast cells and malignant cells and an intense infiltration by tumour cells in bone marrow compartment in all examined samples. Histomorphometrical data on the levels of bone resorption and bone apposition parameters remained constant between T0 and T40 for all analysed patients. Additionally, immunohistochemistry showed homogeneous expression and location of CDH1, CDH2, KRT8, KRT18, Ki67, CASP3, ESR1, CD8 and CD68 between T0 and T40, thus further confirming the invasive behaviour of breast cancer cells and indicating the maintaining of the metastatic microenvironment. The novel tissue culture, set-up in this study, has significant advantages in comparison to the pre-existent 3D models: the tumour environment is the same of the clinical scenario, including all cell types as well as the native extracellular matrix; it can be quickly set-up employing only small samples of breast cancer bone metastasis tissue in a simple, ethically correct and cost-effective manner; it bypasses and/or decreases the necessity to use more complex preclinical model, thus reducing the ethical burden following the guiding principles aimed at replacing/reducing/refining (3R) animal use and their suffering for scientific purposes; it can allow the study of the interactions within the breast cancer bone metastasis tissue over a relatively long period of up to 40 days, preserving the tumour morphology and architecture and allowing also the evaluation of different biological factors, parameters and activities. Therefore, the study provides for the first time the feasibility and rationale for the use of a human-derived advanced alternative model for cancer research and testing of drugs and innovative strategies, taking into account patient individual characteristics and specific tumour subtypes so predicting patient specific responses

Purpose. To evaluate the clinical results of arthroscopic repair and open Ahlgren Larsson method in patients with chronic lateral ankle instability. Methods. We retrospectively evaluated 87 patients who were operated in our clinic between 2010 and 2018 with the diagnosis of chronic lateral ankle instability. 16 patients with osteochondral lesion, 5 patients with rheumatoid arthritis, 4 patients with ankle fractures of the same side, 2 patients with a history of active or previous malignancy were excluded. Preoperative and postoperative clinical evaluations were performed with AOFAS ankle-hindfoot score, FAOS and VAS scores. Results. Sixty patients with chronic lateral ankle instability were evaluated. 28 patients, treated with Ahlgren-Larsson method and 32 patients, treated with arthroscopic repair. 36 of the patients were female and 24 were male; the mean age of the arthroscopy group was 44 ± 9; the mean age of the open surgery group was 46 ± 11. There was no significant difference between the groups in terms of demographic features (age, sex, VKI). Postoperative clinical improvement was observed in both groups. There was no statistically significant difference between the groups in terms of functionality. However, there was a statistically significant difference in pain and satisfaction of VAS in favor of arthroscopy group. Conclusions. Ahlgren-Larsson method and arthroscopic repair technique are safe and effective for chronic lateral ankle instability. Arthroscopic technique may be preferred for pain and patient satisfaction as it is less invasive and less morbid

Sarcopenia is a progressive and generalized skeletal muscle disorder that involves loss of muscle mass and function. It is associated with increased adverse outcomes including falls, functional decline, frailty and mortality and affects 65% of people over the age of 65 more than half of people aged 80 and above. The factors that cause and worsen sarcopenia are categorised into two groups. The primary aetiological factor is ageing and the secondary factors include disease, physical inactivity, and poor nutrition. Sarcopenia is considered to be ‘primary' (or age-related) when no other specific cause is evident. However, a number of ‘secondary' factors may be present in addition to ageing. Sarcopenia can occur secondary to a systemic or inflammatory disease, including malignancy and organ failure. Physical inactivity is one of the major contributors to the development of sarcopenia, whether due to a sedentary lifestyle or to disease related immobility or disability. Furthermore, sarcopenia can develop as a result of inadequate protein consumption. Biomarkers are objective and quantifiable characteristics of physiological and pathophysiological processes. Biomarkers can be used to predict the development of sarcopenia in older susceptible adults and enable early interventions that can reduce the risk of physical disability, the co-morbidities associated with the loss of muscle mass and the poor health outcomes that result from sarcopenia. Non-invasive imaging technologies can be used as biomarkers to detect loss of skeletal muscle mass in sarcopenia include bone densitometry, computed tomography, ultrasound and magnetic resonance imaging. However, imaging requires sophisticated and expensive equipment that is not available in a resource poor setting. Therefore, markers of skeletal muscle strength and fitness and soluble biochemical markers in blood may be used as alternative biomarkers. Studies on sarcopenia have identified numerous soluble biochemical biomarkers. These biomarkers can be divided into two groups: “muscle-specific” and “non-muscle-specific” biomarkers. Since sarcopenia is associated with rapid skeletal muscle wasting, the skeletal muscle-specific isoform of troponin T may be considerate a useful biomarker of sarcopenia, since high troponin levels in blood are an expression of muscle wasting. Peptides derived from collagen type VI turnover may be potential biomarkers of sarcopenia. We have recently conducted a systematic review to summarize the data from recent mass-spectrometry based proteomic studies of the secretome of skeletal muscle cells in response to disease, exercise or metabolic stress in order to identify the proteins involved in muscle breakdown. Developing robust in vitro models for the study of sarcopenia using primary muscle cells is a high priority as is exploiting the in vitro models to understand catabolic and inflammatory processes and molecular mechanisms involved in sarcopenia. Co-cultures with adipose-derived and other cells may be used to screen for small molecules and biologicals capable of inhibiting the catabolic and inflammatory pathways involved in sarcopenia. This presentation reviews recent progress in this area and outlines opportunities for future research on sarcopenia

Intracapsular neck of femur fractures may be treated with fixation or arthroplasty, depending on fracture characteristics and patient factors. Two common methods of fixation are the sliding hip screw, with or without a de-rotation screw, and cannulated screws. Each has its merits, and to date there is controversy around which method is superior, with either method thought to risk avascular necrosis of the femoral head (AVN) rates in the region of 10–20%. Fixation with cannulated screws may be performed in various ways, with current paucity of evidence to show an optimum technique. There are a multitude of factors which are likely to affect patient outcomes: technique, screw configuration, fracture characteristics and patient factors. We present a retrospective case series analysis of 65 patients who underwent cannulated screw fixation of a hip fracture. Electronic operative records were searched from July 2014 until July 2019 for all patients with a neck of femur fracture fixed with cannulated screws: 68 were found. Three patients were excluded on the basis of them having a pathological fracture secondary to malignancy, cases were followed up for 2 years post-operatively. Electronic patient records and X-rays were reviewed for all included patients. All X-rays were examined by each team member twice, with a time interval of two weeks to improve inter-observer reliability. 65 patients were included with 2:1 female to male ratio and average age of 72 years. 36 patients sustained displaced fractures and 29 undisplaced. Ten patients sustained a high-energy injury, none of which developed AVN. Average time to surgery was 40 hours and 57 patients mobilised on day one post-operatively. All cases used either 7 or 7.3mm partially threaded screws in the following configurations: 2 in triangle apex superior, 39 triangle apex inferior, 22 rhomboid and 2 other, with 9 cases using washers. All reductions were performed closed. Five (8%) of our patients were lost to follow-up as they moved out of area, 48 (74%) had no surgical complications, seven (11%) had mild complications, three (5%) moderate and two (3%) developed AVN. Both of these sustained displaced fractures with low mechanism of injury, were female, ASA 2 and both ex-smokers. One received three screws in apex inferior configuration and one rhomboid, neither fixed with washers. Our AVN rate following intracapsular hip fracture fixation with cannulated screws is much lower than widely accepted. This study is under-powered to comment on factors which may contribute to the development of AVN. However, we can confidently say that our practice has led to low rates of AVN. This may be due to our method of fixation; we use three screws in an apex inferior triangle or four screws in a rhomboid, our consultant-led operations, closed reduction of all fractures, or our operative technique. We pass a short thread cannulated screw across the least comminuted aspect of the fracture first in order to achieve compression, followed by two or three more screws (depending on individual anatomy) to form a stable construct. Our series shows that fixation of intracapsular hip fractures with cannulated screws as we have outlined remains an excellent option. Patients retain their native hip, have a low rate of AVN, and avoid the risks of open reduction

Metal instrumentation (rods and screws) is used to stabilise the spine after trauma, malignancy or deformity. Approx 3% become infected often necessitating removal of metal. At surgery tissue samples and metal are removed for culture, but many clinical laboratories are not equipped to process metal or use simple culture methods. The causative bacteria exist as biofilms on the metal and they are often anaerobic and slow-growing, so conventional culture methods often fail to detect them. Also, they are common contaminants leading to diagnostic uncertainty. We have established a laboratory protocol to overcome these problems. Removed metalwork was sonicated and the sonicate centrifuged and the supernatant discarded. Quantitative aerobic and anaerobic culture of the resuspended pellet for 14 days and microscopy were carried out. Metalwork from 11 suspected infected cases was culture-positive (median 2857, 60–5000cfu/mL). Microscopy revealed an infection due to Candida albicans that would not have been detected otherwise. Bacteria were isolated from 8 of 10 non-infected cases (median 15, 0–35 cfu/mL). Conventionally processed samples failed to grow in 4 infected cases. (cfu/mL infected vs noninfected cases p=0.0093). Micro-organisms on spinal metalwork grow as biofilms and they require sonication to dislodge them. The causative bacteria are slow-growing and P acnes is anaerobic and requires prolonged incubation. S epidermidis and P acnes are common contaminants and quantitative culture helps to distinguish pathogens from contaminants, removing the diagnostic uncertainty that conventional methods give. Microscopy of the sonicate can reveal micro-organisms that fail to grow on culture. We recommend that sonication of metalwork, prolonged anaerobic incubation and quantitative culture be adopted to improve diagnostic clarity for spinal instrumentation infections

Introduction. We present our experience of the coned hemi-pelvis (‘ice-cream’ cone) implant, using an extended posterior approach to the hip joint, in the management of pelvic bone loss and pelvic discontinuity. Methods. Retrospective study conducted utilising a prospectively collected database. Patients who underwent an ice-cream cone reconstruction between August 2004 – September 2011 were identified. All had a posterior approach to the hip. Femur prepared in the standard fashion. A variety of femoral components used. Demographic data was recorded along with the indication for surgery and outcomes. Results. 16 patients identified. Mean age was 62.2 years. 5 (31.25%) male. 11 (69.75%) female. Indications included; multiple hip revision surgery 4(25%); post Gridlestones for severe hip dysplasia 1 (6.25%); peri-acetabular metastatic deposits 11 (68.75%) from breast, renal, endometrial, prostatic, myeloma primary malignancies. Mean follow-up was 32.06 months. Complications; 1 intra-operative death from tumour embolus; 1 dislocation; 1 superficial surgical site infection. 3 deaths from their primary malignancy. Mean time from prosthesis implantation to death was 14.5 months. All patients at last follow-up were mobilizing. No implant has needed to be revised. Discussion. Pelvic bone loss provides reconstructive challenges. The coned hemi-pelvis is simple to make, easy and versatile to use even when there is little pelvis remaining. It provides a method of negotiating hip reconstruction in patients with severe pelvic bone loss. Orthopaedic surgeons are familiar with the posterior approach to the hip. The ice-cream cone implant can therefore be placed with ease using this well-known approach to the hip

Summary Statement. Survivin is a member of the inhibitor of apoptosis family, which may contribute to the progression of human MFH via inhibiting the mitochondrial apoptosis, and may be considered as a potent therapeutic target for the treatment of human MFH. Introduction. Survivin is a member of the inhibitor of apoptosis (IAP) family, which usually expresses in the embryonic lung and fetal organs in the developmental stages, but is undetectable in normal adult tissues other than thymus, placenta, CD34. +. stem cells, and basal colonic epitherial cells. However, several studies reported that survivin is highly expressed in various human malignancies, including sarcomas, and increased expression of survivin is an unfavorable prognostic marker correlating with decreased overall survival in cancer patients. We have previously reported that survivin was strongly expressed in human malignant fibrous histiocyoma (MFH), however, the roles of survivin in human MFH have not been studied. The aim of this study was to evaluate the effect of survivin inhibition on apoptotic activity in human MFH cells. Methods. Nara-H, a human MFH cell line which expresses the high levels of survivin, was used in this study. Cells were cultured in DMEM supplemented with 10% FBS and 1% penicillin/streptomycin at 37°C in a humidified atmosphere containing 5% CO. 2. To evaluate the effect of survivin inhibition on MFH cell apoptosis, cells were transfected with either a survivin specific siRNA (survivin-siRNA) or a non-specific control siRNA (control-siRNA) by lipofection method. After siRNA transfection, the efficiency of siRNA knockdown of survivin was assessed by quantitative real time PCR. Expressions of apoptosis-related proteins, such as caspase-3, caspase-9 and PARP, were assessed by immunoblot analysis, and the apoptotic activity was evaluated by flow cytometric analysis. Results. Transfection of survivin-siRNA strongly suppressed the expression of survivin compared with control-siRNA. Immunoblot analyses revealed that expressions of cleaved forms of caspase-3, caspase-9 and PARP were increased in survivin-siRNA transfected cells, while the expressions were barely detected in control cells. In flow cytometric analysis, the number of apoptotic cells was significantly increased in survivin-siRNA transfected cells compared with that in control cells. Discussion/Conclusion. Previous studies revealed that survivin regulates the mitochondrial apoptotic pathway, and that overexpression of survivin is associated with tumor growth, progression, and resistance to conventional targeted anticancer agents in various human malignancies. In the current study, we demonstrated that siRNA knockdown of survivin induced the cleavage of caspase-3, caspase-9 and PARP, and increased the apoptotic activity in human MFH cells. The findings in this study strongly suggest that survivin may contribute to the progression of human MFH via inhibiting the mitochondrial apoptosis in human MFH, and may be considered as a potent therapeutic target for the treatment of human MFH

We performed positron emission tomography (PET) with . 18. fluorine-fluoro-2-deoxy-D-glucose (FDG) on 55 patients with tumours involving the musculoskeletal system in order to evaluate its role in operative planning. The standardised uptake value (SUV) of FDG was calculated and, to distinguish malignancies from benign lesions, the cases were divided into high (≥ 1.9) and low (< 1.9) SUV groups. The sensitivity of PET for correctly diagnosing malignancy was 100% with a specificity of 76.9% and an overall accuracy of 83.0%. The mean SUV for metastatic lesions was twice that for primary sarcomas (p < 0.0015). Our results suggest that the SUV may be useful in differentiating malignant tumours from benign lesions. However, some of the latter, such as schwannomas, had high SUVs so that biopsy or wide resection was selected as the first operation. Thus, some other quantitative analysis may be required for preoperative planning in cases of high-SUV neurogenic benign tumours. The reverse transcription-polymerase chain reaction revealed that the RNA message of a key enzyme in glucose metabolism, phosphohexose isomerase (PHI)/autocrine motility factor, was augmented in only high FDG-uptake lesions, suggesting that a high expression of the PHI message may be associated with accumulation of FDG in musculoskeletal tumours

Background. Multiple Myeloma is a hematological malignancy of terminally differentiated plasma cells associated with increased osteoclast activity and decreased osteoblast functions. Systemic antiproliferative treatment includes proteasome inhibitors such as bortezomib, a clinical potent antimyeloma agent. Local delivery of biological active molecules via biomaterial composite implants to the site of the lesion has been shown to be beneficial for bone and implant-associated infections. In anticancer treatment local delivery of anticancer agents to the neoplasia via biomaterial carriers has never been reported before. The purpose of the current is to present the concepts and the first in vivo results for proteasome inhibitor composite biomaterials for local delivery of bortezomib to proliferative multiple myeloma bone lesions including concentration measurements at different anatomical regions in a rat model. Methods. 80 female Sprague-Dawley rats were randomised into five different treatment groups (n=16/group): 1) Empty (2) Xerogel-granulat: XG (3) Xerogel-granulat+100mgbortezomib [b]: XG100b (4) Xerogel-granulat+500mgb:XG500b (5) Xerogel-granulat+2500mgb:XG2500b. A 2.5 mm drill hole was then created in the metaphysis of the left femur. The defect was then either filled with the previously mentioned substitutes or left empty to serve as a control. After 4 weeks femora were harvested followed by histological, histomorphometrical and immunohistochemical (BMP2; bone-morphogenic protein 2, OPG; osteoprotegerin, RANKL; Receptor activator of nuclear factor kappa-B ligand, ASMA; alpha smooth muscle actin, ED1;CD68 antibody). TOF-SIMS was used to assess the distribution of released strontium ions. Statistical analysis was done using SPSS software. Data was not found normally distributed and hence Mann-Whitney U with bonferroni correction was used. To avoid type I errors due to unequal variances and group sizes Games-Howell test was also performed

The glycolytic-based metabolism of cancers promotes an acidic microenvironment that is responsible for increased aggressiveness. However, the effects of acidosis on tumour metabolism have been almost unexplored, and the metabolic adaptation of cancer cells to acidosis has never been compared with the metabolic response of normal cells. In this study, to pinpoint for the first time the different metabolic profiles between osteosarcoma (OS) cells and normal human fibroblasts (Fb) under short-term acidosis, we used capillary electrophoresis with time-of-flight mass spectrometry (CE-TOFMS). We also screened alterations of the epigenetic profiles – DNA methylation and histone acetylation – of OS cells and compared it with those of normal Fb. Using CE-TOFMS, we observed a significant metabolic difference associated with glycolysis repression (dihydroxyacetone phosphate), increase of amino acid catabolism (phosphocreatine and glutamate) and urea cycle enhancement (arginino succinic acid) in OS cells compared with normal Fb. Noteworthy, metabolites associated with chromatin modification, like UDP-glucose and N. 8. -acetylspermidine, decreased more in OS cells than in normal Fb. Further, combined bisulfite restriction analysis (COBRA) and acetyl-H3 immunoblotting indicated an epigenetic stability in OS cells than in normal Fb, and OS cells were more sensitive to an HDAC inhibitor under acidosis than under neutral condition. Our data suggest that acidosis promotes a metabolic reprogramming that can contribute to the epigenetic maintenance under acidosis only in OS cells, and then the acidic microenvironment should be considered for future therapeutic approaches. The application of epigenetic modulators will be able to become an effectively therapeutic option to selectively target malignancies under the acidic microenvironment

Background. Post-traumatic immunosuppression (PTI) after surgery increases vulnerability to nosocomial infections, sepsis, and death. Knee arthroplasty offers a sterile clinical model to characterise PTI and explore its underlying mechanisms. Methods. This prospective non-randomised cohort study of primary total knee arthroplasty was approved by the Local Ethics Committee. Exclusion criteria included revision-arthroplasty, pre-existing infections, blood-transfusions, malignancy, and auto-immune disease. 48 recruited patients fell into two groups, the first received unwashed anti-coagulated autologous salvaged blood transfusions after surgery (ASBT cohort, n=25). The second received no salvaged blood transfusions (NSBT cohort, n=18). Venous blood was sampled pre-operatively and within 3–7 days post-operatively. Salvaged blood was sampled at one and six hours post-operatively. Biomarkers of immune status included: interleukins (IL) or cytokines (x15), chemokines (x3), Damage-Associated-Molecular-Patterns (DAMPS) (x5), anti-microbial proteins (x3), CD24, and Sialic-acid-binding-Immunoglobulin-type-Lectin-10 (Siglec-10). Results were expressed as fold-change over pre-operative values. Only significant changes are described. Results. Certain biomarkers associated with sterile trauma were common to all 43 patients, including supra-normal: IL-6, IL-1-Receptor-Antagonist, IL-8, Heat-Shock-Protein-70 (HSP70), Calprotectin, CD24 and Siglec-10. But, whereas in NSBT patients post-operative pro-inflammatory biomarkers were sub-normal consistent with PTI, they were supra-normal in ASBT patients implying its reversal. These PTI-biomarkers included: IL-1β, IL-2, IL-17A, Interferon-gamma (IFN-γ), Tumour-Necrosis-Factor-alpha (TNF-α), and Annexin-A2. Reversal of PTI by salvaged blood was further endorsed in ASBT by sub-normal levels of the anti-inflammatory biomarkers: IL-4, IL-5, IL-10, and IL-13. Salvaged blood analyses revealed sustained supra-normal levels of DAMPs, CD24 and Siglec-10; and increasingly elevated levels of cytokines and chemokines during the six hour collection period. Interestingly, plasma CD24, Siglec-10, HSP70 and Calprotectin levels were significantly correlated, implying physical association within the circulation. Conclusions. Several anti-inflammatory processes triggered by traumatised tissue induce systemic PTI, thereby increasing vulnerability to infections. Reversal of PTI by re-infusion of anti-coagulated salvaged blood suggests a novel source of immuno-stimulants

Osteosarcoma (OS) is an aggressive bone malignancy with a high relapse rate despite combined treatment with surgery and multiagent chemotherapy. As for other cancers, OS-associated microenvironment may contribute to tumor initiation, growth, and metastasis. We consider mesenchymal stromal cells (MSC) as a relevant cellular component of OS microenvironment, and have previously found that the interaction between MSC and tumor cells is bidirectional: tumor cells can modulate their peripheral environment that in turn becomes more favourable to tumor growth through metabolic reprogramming (1). Stem-like cells were derived from HOS osteosarcoma cell line by using the spherogenic system (2). CSC isolated from HOS (HOS-CSC) were co-coltured with MSC isolated from bone marrow. Cell lysates and supernatants were collected for the analysis of RNA expression and of secreted cytokines, by Q-RT-PCR and specific ELISA assays, respectively. Here, we determined the effects of MSC on OS stemness and migration, two major features associated with recurrence and chemoresistance. Recruitment of MSC to the tumor environment leads to enhanced proliferation of OS stem cells, which increase the expression levels of TGFβ1. The latter, in turn, could be responsible for the activation of NF-kB genes and IL-6 secretion by MSC. Pro-tumorigenic effects of MSC, via IL-6, including induction of HOS-CSC migration and sphere growth, can be counteracted by IL-6 neutralizing antibody. The presence of MSC is also responsible for increased expression of adhesion molecules involved in intra- or extra-vasation. Stromal cells in combination with OS spheres exploit a vicious cycle where the presence of CSC stimulates mesenchymal cytokine secretion, which in turn increases stemness, proliferation, migration, and metastatic potential of CSC. Furthermore, for the first time we identified a novel OS stem cell marker, the Met proto-oncogene, that is frequently overexpressed and is pathogenetically relevant in OS (2 and 3). Altogether, our data corroborates the concept that a comprehensive knowledge of the interplay between tumor and stroma that also includes the stem-like fraction of tumor cells is needed to develop novel and effective anti-cancer therapies

Extracorporeal irradiation and re-implantation of a bone segment is a technique employed in bone sarcoma surgery for limb salvage in the setting of reasonable bone stock. There is neither consensus nor rationale given for the dosage of irradiation used in previous studies, with values of up to 300Gy applied. We investigated the influence of extracorporeal irradiation on the elastic and viscoelastic properties of bone. Bone specimens were extracted from mature cattle and subdivided into thirteen groups; twelve groups exposed to increasing levels of irradiation and a control group. The specimens, once irradiated, underwent mechanical testing in saline at 37°C. Mechanical properties were calculated by experimental means which included Young's Modulus, Storage Modulus and Loss Modulus. These were all obtained for comparison of the irradiated specimens to the control group. There were non-significant negligible changes in all of the mechanical properties of bone that were assessed with increasing dosage of irradiation. Therefore, we conclude that the overall mechanical effect of high levels of extracorporeal irradiation (300Gy) is minute, and can be administered to reduce the risk of malignancy recurrence

This case report describes a patient with thoracic plasmacytoma, an uncommon haematological malignancy, who presented with neck pain. Plasmacytoma is a neoplastic proliferation of B cell lineage but is much less common than multiple myeloma. The histological examination of multiple myeloma and plasmacytoma is identical however in plasmacytoma there is a solitary lesion with negative skeletal survey, negative bone marrow aspirate and little or no myeloma protein detected in the blood. This makes it more challenging to diagnose and a high index of suspicion is required

Multiple myeloma (MM) is an incurable hematological tumor stemming from malignant plasma cells. MM cells accumulate in the bone marrow (BM) and shape the BM niche by establishing complex interactions with normal BM cells, boosting osteoclasts (OCLs) differentiation and causing bone disease. This unbalance in bone resorption promotes tumor survival and the development of drug resistance. The communication between tumor cells and stromal cells may be mediated by: 1) direct cell-cell contact; 2) secretion of soluble factors, i.e. chemokines and growth factors; 3) release of extracellular vesicles/exosomes (EVs) which are able to deliver mRNAs, miRNAs, proteins and metabolites in different body district. Primary CD138+ MM cells were isolated from patients BM aspirates. MM cell lines were cultured alone in complete RPMI-1640 medium or co-cultured with murine (NIH3T3) or human (HS5) BMSC cell lines or murine Raw264.7 monocytes in DMEM medium supplemented with 10% V/V FBS. Silencing of Jagged1 and Jagged2 was obtained by transient expression of specific siRNAs or by lentiviral transduction using a Dox-inducible system (pTRIPZ). EVs were isolated using differential ultracentrifugation. EVs concentration and size were analyzed using Nano Track Analysis (NTA) system. The uptake of PKH26-labelled MM-derived EVs by HS5 or Raw264.7 was measured after 48 hours by confocal microscopy and flow cytometry. Osteoclast (OCL) differentiation of Raw264.7 cells was induced by 50ng/ml mRANKL, co-culturing with MM cells, CM or EVs. OCLs were stained by TRAP Kit and counted. Bone resorption was assessed by Osteo Assay Surface plates. Flow cytometric detection of apoptotic cells was performed after staining with Annexin V. Gene expression was analyzed by qRT-PCR, while protein levels were determined using flow cytometry ELISA or WB. Notch oncogenic signaling is dysregulated in several hematological and solid malignancies. Notch receptors and ligands are key players in the crosstalk between tumor cells and BM cells. We have demonstrated that: 1) the dysregulated Jagged ligands on MM cells trigger the activation of Notch receptors in the nearby stromal cells by cell-cell contact. This results in the release of anti-apoptotic and growth stimulating factors, i.e. IL6 and SDF1; 2) MM cells promote the development of bone lesions boosting osteoclast differentiation by secreting soluble factors (i.e. RANKL) and by the activation of Notch signaling mediated by direct contact with osteoclast precursors; 3) Finally, we present evidences that EVs play a crucial role in the dysregulated interactions of MM cells with the microenvironment and that Notch signaling regulates their release and participate in this cross-talk. These evidences supports the hypothesis that Jagged targeting on MM cells may interrupt the communication between tumor cells and the surrounding milieu, blocking the activation of the oncogenic Notch pathway and finally resulting in the a reduction of MM-associated bone disease and drug resistance

Summary Statement. In this study we suggested a possible role of prion proteins genes in osteosarcoma. Therefore, the inhibition of prion proteins expression must be tested because it could represent a new approach to the molecular treatment of osteosarcoma. Introduction. Although osteosarcoma is the most common bone malignancy, the molecular and cellular mechanisms influencing its pathogenesis have remained elusive. Prion proteins (PRNP and PRND), known mostly for its involvement in neurodegenerative spongiform encephalopathies, have been recently demonstrated to be involved in resistance to apoptosis, tumorigenesis, proliferation and metastasis. Patients & Methods. The main aim of research was to study whether prion proteins were over-expressed in human osteosarcoma, and if prion proteins could have a role also in osteosarcomas. We evaluated differential gene expression between 22 cases of osteosarcoma and 40 cases of normal bone specimens through cDNA microarray analysis spanning a substantial fraction of the human genome. Results. PRNP and PRND are significantly over-expressed in osteosarcoma. PRNP and PRND appear involved with some important genes related to tumorigenesis and apoptosis. PRNP is linked to PTK2, RBBP9 and TGFB1 while PRND is linked to TNFSF10, BCL2A1, NFKB2 and TP53RK. Discussion/Conclusion. Increased expression on Affymetrix arrays of prion proteins seems to be associated with the development of osteosarcoma. Prions seem to induce a negative regulation of apoptosis, thus promoting osteosarcoma development and progression. Osteosarcoma is a very aggressive tumor and even after modern chemotherapy and excision of tumors efforts are needed to improve clinical outcome. Since Prion proteins seem to be related to osteosarcoma development, their inhibition could represent a new approach to the molecular treatment of osteosarcoma