Aims. This study explored the shared genetic traits and molecular interactions between postmenopausal osteoporosis (POMP) and sarcopenia, both of which substantially degrade elderly health and quality of life. We hypothesized that these motor system diseases overlap in pathophysiology and regulatory mechanisms. Methods. We analyzed microarray data from the Gene Expression Omnibus (GEO) database using weighted gene co-expression network analysis (WGCNA), machine learning, and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis to identify common genetic factors between POMP and sarcopenia. Further validation was done via differential gene expression in a new cohort. Single-cell analysis identified high expression cell subsets, with mononuclear macrophages in osteoporosis and muscle stem cells in sarcopenia, among others. A competitive endogenous
Aim. While metagenomic (microbial DNA) sequencing technologies can detect the presence of microbes in a clinical sample, it is unknown whether this signal represents dead or live organisms. Metatranscriptomics (sequencing of RNA) offers the potential to detect transcriptionally “active” organisms within a microbial community, and map expressed genes to functional pathways of interest (e.g. antibiotic resistance). We used this approach to evaluate the utility of metatrancriptomics to diagnose PJI and predict antibiotic resistance. Method. In this prospective study, samples were collected from 20 patients undergoing revision TJA (10 aseptic and 10 infected) and 10 primary TJA. Synovial fluid and peripheral blood samples were obtained at the time of surgery, as well as negative field controls (skin swabs, air swabs, sterile water). All samples were shipped to the laboratory for metatranscriptomic analysis. Following microbial
Hip and knee arthroplasty (HKA) are two of the most successful orthopaedic procedures. However, one major complication necessitating revision surgery is osteolysis causing aseptic loosening of the prosthesis. JAK-STAT has been demonstrated to influence bone metabolism and can be regulated by microRNA (miRNA). Adult patients with osteolysis or aseptic loosening undergoing revision HKA were recruited. Age and gender matched patients undergoing primary hip or knee arthroplasty were our controls. Samples of bone, tissue and blood were collected and
Osteoarthritis (OA) is the most common form of arthritis and one of the ten most disabling diseases in developed countries. Total joint replacement (TJR) is considered by far as the most effective treatment for end-stage OA patients. The majority of patients achieve symptomatic improvement following TJR. However, about 22% of the TJR patients either do not improve or deteriorate after surgery. Several potential non-genetic predictors for the TJR outcome have been investigated. However, the results were either inconclusive or had very limited predictive power. The aim of this study was to identify genetic variants for the poor outcome of TJR in primary OA patients by a genome-wide association study (GWAS). Study participants were total knee or hip replacement patients due to primary OA who were recruited to the Newfoundland Osteoarthritis Study (NFOAS) before 2017. The Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) was used to assess pain and functional impairment pre- and 3.99±1.38 years post-surgery. Two non-responder classification criteria were used in our study. One was defined by an absolute WOMAC change score. Participants with a change score less than 7/20 points for pain were considered as pain non-responders; and those with less than 22/68 points for function were classified as function non-responders. The second one was the Outcome Measures in Arthritis Clinical Trials and the Osteoarthritis Research Society International (OMERACT-OARSI) criteria. Blood DNA samples were genotyped using the Illumina GWAS microarrays genotyping platform. The quality control (QC) filtering was performed on GWAS data before the association of the genetic variants with non-responders to TJR was tested using the GenABEL package in R with adjustment for the relatedness of the study population and using the commonly accepted GWAS significance threshold p < 5*10. −8. to control multiple testing. In total, 316 knee and 122 hip OA patients (mean age 65.45±7.62 years, and 58% females) passed the QC check. These study participants included 368 responders and 56 non-responders to pain, and 364 responders and 68 non-responders to function based on the absolute WOMAC point score change classification. While 377 responders and 56 non-responders to pain, and 366 responders and 71 non-responders to function were identified by the OMERACT-OARSI classification criteria. Interestingly, the same results were obtained by both classification methods, and we found that the G allele of rs4797006 was significantly associated with pain non-responders with odds ratio (OR) of 5.12 (p<7.27×10. -10. ). This SNP is in intron one of the melanocortin receptor 5 (MC5R) gene on chr18. This gene plays central roles in immune response, pain sensitivity, and negative regulation of inflammatory response to antigenic stimulus. The A allele of rs200752023 was associated with function non-responders with OR of 4.41 (p<3.29×10. -8. ). The SNP is located in intron three of the
The poor prognosis of patients with soft-tissue sarcoma as not changed in the past several decades, highlighting the necessity for new therapeutic approaches. T-cell based immunotherapies are a promising alternative to traditional cancer treatments due to their ability to target only malignant cells, leaving benign cells unharmed. The development of successful immunotherapy requires the identification and characterization of targetable immunogenic tumor antigens. Cancer-testis antigens (CTA) are a group of highly immunogenic tumor-associated proteins that have emerged as potential targets for CD8+ T-cell recognition. In addition to identifying a targetable antigen, it is crucial to understand the tumor immune microenvironment. The level of immune infiltration and mechanisms of immune suppression within the tumor play important roles in the outcome of immunotherapy. The goal of this study is to identify targetable immunogenic antigens for T-cell based immunotherapy and to characterize the tumor immune microenvironment in human dedifferentiated liposarcoma (DDLS) by Nanostring and IHC. To assess the complexity of the human DDLS tumor immune microenvironment and to identify target antigens we used the nCounter NanoString platform to generate a gene expression profile for hundreds of genes from
Aims. To describe the incidence of adverse clinical outcomes related to COVID-19 infection following corticosteroid injections (CSI) during the COVID-19 pandemic. To describe the incidence of positive SARS-CoV-2 reverse transcriptase polymerase chain reaction (RT-PCR) testing, positive SARS-COV2 IgG antibody testing or positive imaging findings following CSI at our institution during the COVID-19 pandemic. Methods. A retrospective observational study was undertaken of consecutive patients who had CSI in our local hospitals between 1 February and 30June 2020. Electronic patient medical records (EPR) and radiology information system (RIS) database were reviewed. SARS-CoV-2 RT-PCR testing, SARS-COV2 IgG antibody testing, radiological investigations, patient management, and clinical outcomes were recorded. Lung findings were categorized according to the British Society of Thoracic Imaging (BSTI) guidelines. Reference was made to the incidence of lab-confirmed COVID-19 cases in our region. Results. Overall, 1,656 lab-confirmed COVID-19 cases were identified in our upper tier local authority (UTLA), a rate of 306.6 per 100,000, as of 30June 2020. A total of 504 CSI injections were performed on 443 patients between 1 February and 30June 2020. A total of 11 RT-PCR tests were performed on nine patients (2% of those who had CSI), all of which were negative for SARS-CoV-2
Osteoarthritis (OA) is a debilitating disease and the most common joint disorder worldwide. Although the development of OA is considered multifactorial, the mechanisms underlying its initiation and progression remain unclear. A prominent feature in OA is cartilage degradation typified by the progressive loss of extracellular matrix components - aggrecan and type II collagen (Col II). Cartilage homeostasis is maintained by the anabolic and catabolic activities of chondrocytes. Prolonged exposure to stressors such as mechanical loading and inflammatory cytokines can alter the phonotype of chondrocytes favoring cartilage catabolism, and occurs through decreased matrix protein synthesis and upregulation of catabolic enzymes such as aggrecanases (ADAMTS-) 4 and 5 and matrix metalloproteinases (MMPs). More recently, the endoplasmic reticulum (ER) stress response has been implicated in OA. The ER-stress response protects the cell from misfolded proteins however, excessive activation of this system can lead to chondrocyte apoptosis. Acute exposure of chondrocytes to IL-1β has been demonstrated to upregulate ER-stress markers (GADD153 and GRP78), however, it is unclear whether the ER-stress response plays a role on chronic IL-1β exposure. The purpose of this study was to determine whether modulating the ER stress response with tauroursodeoxycholic acid (TUDCA) in human OA chondrocytes during prolonged IL-1β exposure can alter its catabolic effects. Articular cartilage was isolated from donors undergoing total hip or knee replacement. Chondrocytes were recovered from the cartilage of each femoral head or knee by sequential digestion with Pronase followed by Collagenase, and expanded in DMEM-low glucose supplemented with 10% FBS. Chondrocytes were expanded in flasks for one passage before being prepared for micropellet culture. Chondrocyte pellets were cultured in regular growth medium (Control), medium supplemented with IL-1β [10 ng/mL], TUDCA [100 uM] or IL-1β + TUDCA for 12 days. Medium was replaced every three days. Cartilage explants were prepared from the donors undergoing knee replacement, and included cartilage with the cortical bone approximately 1 cm2 in dimension. Explants were cultured in the above mentioned media, however, the incubation period was extended to 21 days.
Intervertebral discs (IVDs) degeneration is one of the major causes of back pain. Upon degeneration, the IVDs tissue become inflamed, and this inflammatory microenvironment may cause discogenic pain. Cellular senescence is a state of stable cell cycle arrest in response to a variety of cellular stresses including oxidative stress and adverse load. The accumulation of senescent IVDs cells in the tissue suggest a crucial role in the initiation and development of painful IVD degeneration. Senescent cells secrete an array of cytokines, chemokines, growth factors, and proteases known as the senescence-associated secretory phenotype (SASP). The SASP promote matrix catabolism and inflammation in IVDs thereby accelerating the process of degeneration. In this study, we quantified the level of senescence in degenerate and non-degenerate IVDs and we evaluated the potential of two natural compounds to remove senescent cells and promote overall matrix production of the remaining cells. Human IVDs were obtained from organ donors. Pellet or monolayer cultures were prepared from freshly isolated cells and cultured in the presence or absence of two natural compounds: Curcumin and its metabolite vanillin. Monolayer cultures were analyzed after four days and pellets after 21 days for the effect of senolysis. A cytotoxicity study was performed using Alamar blue assay. Following treatment,
Aim. Diagnosing Orthopaedic infection is limited by the sensitivity of culture methods. Next generation sequencing (NGS) offers an alternative approach for detection of microorganisms from clinical specimens. However, the low ratio of pathogen DNA to human DNA often inhibits detection of microorganisms from specimens. Depletion of human DNA may enhance the detection of microbial DNA. 1. Our aim was to compare four DNA extraction methods for the recovery of microbial DNA from orthopaedic samples for NGS. Method. Simulated samples; pooled culture negative sample matrix was spiked with known concentrations of microorganisms, each panel consisting of 7 samples. Broth culture was performed on simulated samples for comparison with NGS. *. . DNA Extraction; total nucleic acid extraction was performed on an automated extraction platform. **. using the viral NA assay. Modifications included: (1) mechanical lysis (glass beads), (2) lysis of human cells (saponin 0.025%), turbo DNase treatment and (3) mechanical lysis and addition of MspJI enzyme post-extraction for methylated DNA digestion. Detection of human and microbial DNA; human endogenous (HE) gene rtPCR. ***. was utilised following manufacturer's recommendations. Microbial DNA was detected using SYBR green 16s ribosomal
Staphylococcus aureus gene locus sdr is encoding proteins responsible for adhesion to bone tissue and therefore for the invasiveness of this bacteria. The aim of the study was to determine possible mechanism of activation of this gene and its role in bone infections. S. aureus strain 838/05 was isolated from the bone adjacent to infected joint prostheses. The strain contains all sdrC,D, and E genes. It was grown in liquid TSB medium (Bio Merieux) with gentle aeration or on Columbia agar plates containing 5% sheep blood (Bio Merieux). The presence of putative promoters and transcriptional organization of the sdr region was detected using BPROM and FGENESB algorithms (. www.softberry.com. ) based on region 611262bp-623152bp (GeneBank number CP000730.1) of the Staphylococcus aureus subsp. aureus USA300_TCH1516 complete genome sequence. In order to determine interaction of S. aureus with human blood, 50 ml of bacteria grown in TSB medium to ML growth phase were collected and washed twice with sterile PBS to remove the medium and re-suspended in 50ml of sterile PBS. 10 ml of cells were mixed with ∼100ml of mixed fresh human blood collected from healthy volunteers. Immediately after mixing blood with bacteria, 30 ml of the sample (Time 0) were mixed with two volumes of
The aim of this study was to analyze the prevalence of culture-negative periprosthetic joint infections (PJIs) when adequate methods of culture are used, and to evaluate the outcome in patients who were treated with antibiotics for a culture-negative PJI compared with those in whom antibiotics were withheld. A multicentre observational study was undertaken: 1,553 acute and 1,556 chronic PJIs, diagnosed between 2013 and 2018, were retrospectively analyzed. Culture-negative PJIs were diagnosed according to the Muskuloskeletal Infection Society (MSIS), International Consensus Meeting (ICM), and European Bone and Joint Society (EBJIS) definitions. The primary outcome was recurrent infection, and the secondary outcome was removal of the prosthetic components for any indication, both during a follow-up period of two years.Aims
Methods
Aim. The aim of this study was to gain insight into the in vivo expression of virulence and metabolic genes of Staphylococcus aureus in a prosthetic joint infection in a human subject. Method. Deep
The new COVID-19 variant was reported by the authorities of the UK to the World Health Organization (WHO) on 14 December 2020. We aim to describe the clinical characteristics and nosocomial infection rates in major trauma and orthopaedic patients comparing the first and second wave of COVID-19 infection. A retrospective analysis of a prospectively collected trauma database was reviewed at a level 1 major trauma centre from 1 December 2020 to 18 February 2021 looking at demographics, clinical characteristics, and nosocomial infections and compared to our previously published first wave data (26 January 2020 to 14 April 2020).Aims
Methods
Introduction. Macrophages phagocytes implant wear debris and produce various cytokines to evoke inflammation and periprosthetic osteolysis of aseptic loosening. It had been reported that expression of Toll-like receptor (TLR) 2 and other TLRs increased in periprosthetic tissues of aseptic loosening. Pathogen-associated molecular patterns (PAMPs) and damaged-associated molecular patterns (DAMPs) have been known as ligands of TLRs and considered to be involved in the osteolytic reactions via TLRs. Another type of immune sensors, nucleotide-binding and oligomerization domain (NOD)-like receptors (NLR) with a pyrin domain 3 (NLRP3) can also recognize PAMPs and DAMPs as their lignds, which has been presumed to participate in the local host response of macrophage cascade via phagocytosis of implant wear particles. However, the contribution of NLRP3 in periprosthetic tissues of aseptic loosening and the correlation between TLR2 and NLRP3 are still unclear. Materials and methods. TLR1, TLR2, TLR6, NLRP3, TNF-α and IL-1β of macrophages in aseptic loose periprosthetic tissues were immnohistorically evaluated and compared to osteoarthritic synovium. RAW264.7 cells, macrophagic cell line, were stimulated by titanium particles (Ti) and lipoteichoic acid (LTA)-coated Ti. The celluar reaction associated with TLR2 and NLRP3 and the correlation of them were analyzed at mRNA expression levels with small-interfering
A randomised controlled trial was conducted using a rabbit model of a complex contaminated extremity war wound. Compared to saline soaked gauze dressings Inadine (iodine) and Acticoat (nanocrystalline silver) had significantly lower levels of Staphylococcus aureus after 7 days while Activon Tulle (Manuka honey) had significantly higher levels. Molecular level analysis of the wound was conducted. Plasma cytokines of interest were assayed using ELISA and levels of expression of relevant tissue genes measured using PCR following
Introduction. The ability of activated platelets to induce cellular proliferation is well recognised. In a previous diffusion model, platelets combined with Tri-calcium phosphate (TCP) led to an osteoprogenitor mitogenic response followed by cellular differentiation. This study was designed to look at osteoprogenitor responses when cultured directly onto TCP granules combined with activated platelets. Method. Human osteoprogenitors were loaded onto TCP with activated platelets at a low seeding density and high seeding densities. Cellular proliferation was assessed using the pico-green DNA content analysis. Differentiation towards osteoblastic phenotype was assessed using an alkaline phosphatase assay.
Despite the COVID-19 pandemic, incidence of hip fracture has not changed. Evidence has shown increased mortality rates associated with COVID-19 infection. However, little is known about the outcomes of COVID-19 negative patients in a pandemic environment. In addition, the impact of vitamin D levels on mortality in COVID-19 hip fracture patients has yet to be determined. This multicentre observational study included 1,633 patients who sustained a hip fracture across nine hospital trusts in North West England. Data were collected for three months from March 2020 and for the same period in 2019. Patients were matched by Nottingham Hip Fracture Score (NHFS), hospital, and fracture type. We looked at the mortality outcomes of COVID-19 positive and COVID-19 negative patients sustaining a hip fracture. We also looked to see if vitamin D levels had an impact on mortality.Aims
Methods
The aim of this study is to report the 30 day COVID-19 related morbidity and mortality of patients assessed as SARS-CoV-2 negative who underwent emergency or urgent orthopaedic surgery in the NHS during the peak of the COVID-19 pandemic. A retrospective, single centre, observational cohort study of all patients undergoing surgery between 17 March 2020 and 3May 2020 was performed. Outcomes were stratified by British Orthopaedic Association COVID-19 Patient Risk Assessment Tool. Patients who were SARS-CoV-2 positive at the time of surgery were excluded.Introduction
Method
Background. Current treatments for the prevention of thromboembolism include heparin and low-molecular weight heparins (LMWHs). A number of studies have suggested that long term administration of these drugs may adversely affect osteoblasts and therefore, bone metabolism. Xarelto(tm) (Rivaroxaban) is a new anti-thrombotic drug for the prevention of venous thromboembolism in adult patients undergoing elective hip and knee replacement surgery. The aim of this in vitro study was to investigate the possible effects of rivaroxaban on osteoblast proliferation, function, matrix mineralisation and gene expression compared to enoxaparin, a commonly used LMWH. Methods. Primary human osteoblast cultures were treated with varying concentrations of rivaroxaban (0.013, 0.13, 1.3 and 13 μg/ml) or enoxaparin (0.1, 1.0 and 10 international units/ml). The effect of each drug on osteoblast function and matrix mineralisation was evaluated by measuring alkaline phosphatase activity and calcium deposition, respectively. The MTS assay was used to assess the effect of drug treatments on cell proliferation. Changes in osteocalcin, Runx2 and BMP-2 messenger
Introduction. Osteoporosis is a common skeletal disorder characterised by a reduced bone mass and a progressive micro-architectural deterioration in bone tissue leading to bone fragility and susceptibility to fracture. With a progressively aging population, osteoporosis is becoming an increasingly important public health issue. The Wnt/β-catenin pathway is a major signalling cascade in bone biology, playing a key role in regulating bone development and remodelling, with aberrations in signalling resulting in disturbances in bone mass. Objectives. To assess the effects of silencing the expression of the Wnt antagonist Dickkopf-1 (Dkk1) on the bone profile of primary human osteoblasts exposed in vitro to 10-8M dexamethasone. Methods. Primary human osteoblasts (HOBs) were cultured in vitro and exposed to 10-8M dexamethasone over a time course of 4hr, 12hr and 24hr. Dkk1 expression was silenced using small interfering