Aims. The aim of this study was to evaluate the diagnostic accuracy of the synovial alpha-defensin
Aims. This study aimed to investigate the role and mechanism of meniscal cell lysate (MCL) in fibroblast-like synoviocytes (FLSs) and osteoarthritis (OA). Methods. Meniscus and synovial tissue were collected from 14 patients with and without OA. MCL and FLS proteins were extracted and analyzed by liquid chromatography‒mass spectrometry (LC‒MS). The roles of MCL and adenine nucleotide translocase 3 (ANT3) in FLSs were examined by
Aims. It has been established that mechanical stimulation benefits tendon-bone (T-B) healing, and macrophage phenotype can be regulated by mechanical cues; moreover, the interaction between macrophages and mesenchymal stem cells (MSCs) plays a fundamental role in tissue repair. This study aimed to investigate the role of macrophage-mediated MSC chondrogenesis in load-induced T-B healing in depth. Methods. C57BL/6 mice rotator cuff (RC) repair model was established to explore the effects of mechanical stimulation on macrophage polarization, transforming growth factor (TGF)-β1 generation, and MSC chondrogenesis within T-B enthesis by immunofluorescence and
Aims. This study investigated the effects of β-caryophyllene (BCP) on protecting bone from vitamin D deficiency in mice fed on a diet either lacking (D-) or containing (D+) vitamin D. Methods. A total of 40 female mice were assigned to four treatment groups (n = 10/group): D+ diet with propylene glycol control, D+ diet with BCP, D-deficient diet with control, and D-deficient diet with BCP. The D+ diet is a commercial basal diet, while the D-deficient diet contains 0.47% calcium, 0.3% phosphorus, and no vitamin D. All the mice were housed in conditions without ultraviolet light. Bone properties were evaluated by X-ray micro-CT. Serum levels of klotho were measured by
Aims. Osteoarthritis (OA) is a common degenerative joint disease characterized by chronic inflammatory articular cartilage degradation. Long noncoding RNAs (lncRNAs) have been previously indicated to play an important role in inflammation-related diseases. Herein, the current study set out to explore the involvement of lncRNA H19 in OA. Methods. Firstly, OA mouse models and interleukin (IL)-1β-induced mouse chondrocytes were established. Expression patterns of IL-38 were determined in the synovial fluid and cartilage tissues from OA patients. Furthermore, the targeting relationship between lncRNA H19, tumour protein p53 (TP53), and IL-38 was determined by means of dual-luciferase reporter gene, chromatin immunoprecipitation, and RNA immunoprecipitation assays. Subsequent to gain- and loss-of-function assays, the levels of cartilage damage and proinflammatory factors were further detected using safranin O-fast green staining and
Aims. cAMP response element binding protein (CREB1) is involved in the progression of osteoarthritis (OA). However, available findings about the role of CREB1 in OA are inconsistent. 666-15 is a potent and selective CREB1 inhibitor, but its role in OA is unclear. This study aimed to investigate the precise role of CREB1 in OA, and whether 666-15 exerts an anti-OA effect. Methods. CREB1 activity and expression of a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS4) in cells and tissues were measured by immunoblotting and immunohistochemical (IHC) staining. The effect of 666-15 on chondrocyte viability and apoptosis was examined by cell counting kit-8 (CCK-8) assay, JC-10, and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling (TUNEL) staining. The effect of 666-15 on the microstructure of subchondral bone, and the synthesis and catabolism of cartilage, in anterior cruciate ligament transection mice were detected by micro-CT, safranin O and fast green (S/F), immunohistochemical staining, and
Aims. This study aimed to determine the expression and clinical significance of a cartilage protein, cartilage oligomeric matrix protein (COMP), in knee osteoarthritis (OA) patients. Methods. A total of 270 knee OA patients and 93 healthy controls were recruited. COMP messenger RNA (mRNA) and protein levels in serum, synovial fluid, synovial tissue, and fibroblast-like synoviocytes (FLSs) of knee OA patients were determined using
Aims. Poly (ADP-ribose) polymerase (PARP) inhibitor has been reported to attenuate inflammatory response in rat models of inflammation. This study was designed to investigate the effect of PARP signalling in osteoarthritis (OA) cartilage inflammatory response in an OA rat model. Methods. The OA model was established by anterior cruciate ligament transection with medial meniscectomy in Wistar rats. The poly (ADP-ribose) polymerase 1 (PARP-1) shRNA (short hairpin (sh)-PARP-1) and negative control shRNA (sh-NC) were delivered using a lentiviral vector and were intra-articularly injected into rats after surgery. The weight-bearing distribution of the hind limbs and the knee joint width were measured every two weeks. The expression levels of PARP-1, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) in cartilage were determined using real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) and Western blot. The serum concentrations of inflammatory cytokines were detected using
Aims. The aim of this study was to determine the diagnostic accuracy of α defensin (AD) lateral flow assay (LFA) and
Aims. Osteoarthritis (OA) is characterized by persistent destruction of articular cartilage. It has been found that microRNAs (miRNAs) are closely related to the occurrence and development of OA. The purpose of the present study was to investigate the mechanism of miR-486 in the development and progression of OA. Methods. The expression levels of miR-486 in cartilage were determined by quantitative real-time polymerase chain reaction (qRT-PCR). The expression of collagen, type II, alpha 1 (COL2A1), aggrecan (ACAN), matrix metalloproteinase (MMP)-13, and a disintegrin and metalloproteinase with thrombospondin motifs-4 (ADAMTS4) in SW1353 cells at both messenger RNA (mRNA) and protein levels was determined by qRT-PCR, western blot, and
Aims. This study aimed to examine the effects of tumour necrosis factor-alpha (TNF-α) on osteoblasts in metal wear-induced bone loss. Methods. TNF-α immunoexpression was examined in periprosthetic tissues of patients with failed metal-on-metal hip arthroplasties and also in myeloid MM6 cells after treatment with cobalt ions. Viability and function of human osteoblast-like SaOs-2 cells treated with recombinant TNF-α were studied by immunofluorescence, terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling (TUNEL) assay, western blotting, and
Aims. The role of N,N-dimethylformamide (DMF) in diabetes-induced osteoporosis (DM-OS) progression remains unclear. Here, we aimed to explore the effect of DMF on DM-OS development. Methods. Diabetic models of mice, RAW 264.7 cells, and bone marrow macrophages (BMMs) were established by streptozotocin stimulation, high glucose treatment, and receptor activator of nuclear factor-κB ligand (RANKL) treatment, respectively. The effects of DMF on DM-OS development in these models were examined by micro-CT analysis, haematoxylin and eosin (H&E) staining, osteoclast differentiation of RAW 264.7 cells and BMMs, H&E and tartrate-resistant acid phosphatase (TRAP) staining,
Aims. Osteoarthritis (OA) is a disabling joint disorder and mechanical loading is an important pathogenesis. This study aims to investigate the benefits of less mechanical loading created by intermittent tail suspension for knee OA. Methods. A post-traumatic OA model was established in 20 rats (12 weeks old, male). Ten rats were treated with less mechanical loading through intermittent tail suspension, while another ten rats were treated with normal mechanical loading. Cartilage damage was determined by gross appearance, Safranin O/Fast Green staining, and immunohistochemistry examinations. Subchondral bone changes were analyzed by micro-CT and tartrate-resistant acid phosphatase (TRAP) staining, and serum inflammatory cytokines were evaluated by
Aims. The aim of this study was to further evaluate the accuracy of ten promising synovial biomarkers (bactericidal/permeability-increasing protein (BPI), lactoferrin (LTF), neutrophil gelatinase-associated lipocalin (NGAL), neutrophil elastase 2 (ELA-2), α-defensin, cathelicidin LL-37 (LL-37), human β-defensin (HBD-2), human β-defensin 3 (HBD-3), D-dimer, and procalcitonin (PCT)) for the diagnosis of periprosthetic joint infection (PJI), and to investigate whether inflammatory joint disease (IJD) activity affects their concentration in synovial fluid. Methods. We included 50 synovial fluid samples from patients with (n = 25) and without (n = 25) confirmed PJI from an institutional tissue bank collected between May 2015 and December 2016. We also included 22 synovial fluid samples aspirated from patients with active IJD presenting to Department of Rheumatology, the first Medical Centre, Chinese PLA General Hospital. Concentrations of the ten candidate biomarkers were measured in the synovial fluid samples using standard
Aims. This study aimed to evaluate calprotectin in synovial fluid for diagnosing chronic prosthetic joint infection (PJI) . Methods. A total of 63 patients who were suspected of PJI were enrolled. The synovial fluid calprotectin was tested by an
Objectives. Prosthetic joint infection (PJI) diagnosis is a major challenge in orthopaedics, and no reliable parameters have been established for accurate, preoperative predictions in the differential diagnosis of aseptic loosening or PJI. This study surveyed factors in synovial fluid (SF) for improving PJI diagnosis. Methods. We enrolled 48 patients (including 39 PJI and nine aseptic loosening cases) who required knee/hip revision surgery between January 2016 and December 2017. The PJI diagnosis was established according to the Musculoskeletal Infection Society (MSIS) criteria. SF was used to survey factors by protein array and
Objectives. The aims of this study were to determine whether the administration of anti-inflammatory and antifibrotic agents affect the proliferation, viability, and expression of markers involved in the fibrotic development of the fibroblasts obtained from arthrofibrotic tissue in vitro, and to evaluate the effect of the agents on arthrofibrosis prevention in vivo. Methods. Dexamethasone, diclofenac, and decorin, in different concentrations, were employed to treat fibroblasts from arthrofibrotic tissue (AFib). Cell proliferation was measured by DNA quantitation, and viability was analyzed by Live/Dead staining. The levels of procollagen type I N-terminal propeptide (PINP) and procollagen type III N-terminal propeptide (PIIINP) were evaluated with
Objectives. The osteoprotegerin (OPG) and receptor activator of nuclear factor kappa-B ligand (RANKL) balance is of the utmost importance in fracture healing. The aim of this study was therefore to investigate the impact of nonosteogenic factors on OPG and RANKL levels. Methods. Serum obtained from 51 patients with long bone fractures was collected over 48 weeks. The OPG and serum sRANKL (soluble RANKL) concentrations were measured using
We stably transfected early passage chondrocytes with an anti-apoptotic Bcl-2 gene in vitro using a retrovirus vector. Samples of articular cartilage were obtained from 11 patients with a mean age of 69 years (61 to 75) who were undergoing total knee replacement for osteoarthritis. The Bcl-2-gene-transfected chondrocytes were compared with non-transfected and lac-Z-gene-transfected chondrocytes, both of which were used as controls. All three groups of cultured chondrocytes were incubated with nitric oxide (NO) for ten days. Using the Trypan Blue exclusion
Objectives. Osteophytes are products of active endochondral and intramembranous ossification, and therefore could theoretically provide significant efficacy as bone grafts. In this study, we compared the bone mineralisation effectiveness of osteophytes and cancellous bone, including their effects on secretion of growth factors and anabolic effects on osteoblasts. Methods. Osteophytes and cancellous bone obtained from human patients were transplanted onto the calvaria of severe combined immunodeficient mice, with Calcein administered intra-peritoneally for fluorescent labelling of bone mineralisation. Conditioned media were prepared using osteophytes and cancellous bone, and growth factor concentration and effects of each graft on proliferation, differentiation and migration of osteoblastic cells were assessed using
The most common reason for revision surgery of total hip replacements is aseptic loosening of implants secondary to osteolysis, which is caused by immune-mediated reactions to implant debris. These debris can cause pseudotumour formation. As revision surgery is associated with higher mortality and infection, it is important to understand the pro-inflammatory process to improve implant survival. Toll-like receptor 4 (TLR4) has been shown to mediate immune responses to cobalt ions. Statin use in epidemiological studies has been associated with reduced risk of revision surgery. In-vitro studies have demonstrated the potential for statins to reduce orthopaedic debris-induced immune responses and there is evidence that statins can modulate TLR4 activity. This study investigates simvastatin's effect on orthopaedic biomaterial-mediated changes in protein expression of key inflammatory markers and soluble-ICAM-1 (sICAM-1), an angiogenic factor implicated in pseudotumour formation. Human macrophage THP-1 cells were pre-incubated with 50µM simvastatin for 2-hours or a vehicle control (VC), before being exposed to 0.75mM cobalt chloride, 50μm3 per cell zirconium oxide or LPS as a positive control, in addition to a further 24-hour co-incubation with 50µM simvastatin or VC. Interleukin −8 (IL-8), sICAM-1, chemokine ligand 2 (CCL2), CCL3 and CCL4 protein secretion was measured by
Objectives. Regenerative medicine is an emerging field aimed at the repair and regeneration of various tissues. To this end, cytokines (CKs), growth factors (GFs), and stem/progenitor cells have been applied in this field. However, obtaining and preparing these candidates requires invasive, costly, and time-consuming procedures. We hypothesised that skeletal muscle could be a favorable candidate tissue for the concept of a point-of-care approach. The purpose of this study was to characterize and confirm the biological potential of skeletal muscle supernatant for use in regenerative medicine. Methods. Semitendinosus muscle was used after harvesting tendon from patients who underwent anterior cruciate ligament reconstructions. A total of 500 milligrams of stripped muscle was minced and mixed with 1 mL of saline. The collected supernatant was analysed by
A growing number of recent investigations on the human genome, gut microbiome, and proteomics suggests that the loss of mucosal barrier function, particularly in the gastrointestinal tract, may substantially affect antigen trafficking, ultimately influencing the close bidirectional interaction between the gut microbiome and the immune system. This cross-talk is highly influential in shaping the host immune system and ultimately clinical infections. The hypothesis of the current study was that a change in microbiome and/or breach in GI epithelial barrier could be partially responsible for development of periprosthetic joint infections (PJI). Multiple biomarkers of gut barrier disruption were tested in parallel in plasma samples collected as part of a prospective cohort study of patients undergoing revision arthroplasty for aseptic failures or PJI (As defined by the 2018 ICM criteria). All blood samples were collected before any antibiotic was administered. Samples were tested for Zonulin, soluble CD14 (sCD14), and lipopolysaccharide (LPS) using commercially available
Aim. Synovial calprotectin point-of-care test (POC) has shown promising clinical value in diagnosing periprosthetic joint infections (PJIs). However, limited data are available in unclear cases. Moreover, cut-off values for calprotectin lateral flow assay (LFA) and
Aim. A growing number of recent investigations on the human genome, gut microbiome, and proteomics suggests that the loss of mucosal barrier function, particularly in the gastrointestinal tract, may substantially affect antigen trafficking, ultimately influencing the close bidirectional interaction between the gut microbiome and the immune system. This cross-talk is highly influential in shaping the host immune system function and ultimately shifting genetic predisposition to clinical outcome. Therefore, we hypothesized that a similar interaction could affect the occurrence of acute and chronic periprosthetic joint infections (PJI). Method. Multiple biomarkers of gut barrier disruption were tested in parallel in plasma samples collected as part of a prospective cohort study of patients undergoing revision arthroplasty for aseptic or PJI (As defined by the 2018 ICM criteria). All blood samples were collected before any antibiotic was administered. Samples were tested for Zonulin, soluble CD14 (sCD14), and lipopolysaccharide (LPS) using commercially available
Introduction and Objective. Total joint replacement is indicated for osteoarthritis where conservative treatment has failed, and in the UK the number of patients requiring hip and knee replacements is set to increase with an ageing population. Survival of total hip replacements is around 85% at 20 years with the most common reason for revision being aseptic loosening of the implant secondary to osteolysis, which is caused by immune-mediated reactions to implant debris. These debris can also cause pseudotumour formation. As revision surgery is associated with higher morbidity, mortality, infection rates, venous thromboembolism, resource demand and poorer subsequent function it is important to understand the mechanisms underlying the pro-inflammatory process to improve implant survival. Toll-like receptor 4 (TLR4), an innate immune receptor, has been demonstrated to mediate deleterious immune responses by the Tyson-Capper research group, including inflammatory cytokine interleukin-8 (IL-8) secretion. Statin use in epidemiological studies has been associated with reduced overall risk of revision surgery after hip replacement. In-vitro studies have demonstrated the potential for statins to reduce orthopaedic debris-induced immune responses which can lead to osteolysis and pseudotumour formation. As literature from cardiological investigations demonstrate that statins can reduce the expression and responsiveness of TLR4, this could be an exciting mechanism to exploit to reduce the host immune response to orthopaedic wear debris, thereby improving implant survival by reducing immune mediated osteolysis. This ongoing study investigates simvastatin's effect on cobalt ion-mediated changes in gene and protein expression of interleukin-8 and soluble-ICAM-1 (sICAM-1) which is an angiogenic factor implicated in pseudotumour formation. Materials and Methods. TLR4-expressing human monocyte/macrophage THP-1 cells were pre-incubated with 50μM simvastatin for 2-hours or a vehicle control, before being exposed to exposed to 0.75mM cobalt chloride, in addition to a further 24-hour co-incubation with 50μM simvastatin or vehicle control. IL-8 protein and sICAM-1 secretion was measured by
Introduction and Objective. Intervertebral disc (IVD) degeneration is one of the major contributors to low back pain, the leading cause of disability worldwide. This multifactorial pathological process involves the degradation of the extracellular matrix, inflammation, and cell loss due to apoptosis and senescence. While the deterioration of the extracellular matrix and cell loss lead to structural collapse of the IVD, increased levels of inflammation result in innervation and the development of pain. Amongst the known regulators of inflammation, toll-like receptors (TLRs) and more specifically TLR-2 have been shown to be specifically relevant in IVD degeneration. As strong post-transcriptional regulators, microRNAs (miRNAs) and their dysregulation has been connected to multiple pathologies, including degenerative diseases such as osteoarthritis and IVD degeneration. However, the role of miRNAs in TLR signalling in the IVD is still poorly understood and was hence investigated in this study. Materials and Methods. Human Nucleus pulposus (hNP) and Annulus fibrosus (hAF) cells (n=5) were treated with the TLR-2/6 specific agonist PAM2CSK4 (100 ng/mL for 6 hours) in order to activate the TLR2 signalling pathway. After the activation both miRNA and mRNA were isolated, followed by next-generation sequencing and qPCR analysis of proinflammatory cytokines respectively. Furthermore, cell supernatants were used to analyze the secretion of proinflammatory cytokines with
Although 80% of fractures typically heal without any problems, there is a small proportion (<20%) that suffer complications such as delayed healing and potential progression to non-union. In patients with healing complications, the coordinated regulation between pro- and anti-inflammatory cytokines, such as interleukin-1β (IL-1β) and interleukin-1 receptor antagonist (IL-1Ra) respectively, is often dysregulated. The aim of this study is to develop a therapeutic strategy based on the local delivery of genes to reparative mesenchymal stromal cells (MSCs) migrating into the local fracture microenvironment, thereby promoting a more favourable healing environment to enhance fracture repair. Our approach involves the local delivery of nanoparticles complexing the non-viral vector polyethyleneimine (PEI) with therapeutic plasmid DNA (pDNA) encoding for IL-1Ra. pDNA encoding green fluorescent protein and Gaussia luciferase were used as reporter genes to determine the transfection efficiency of both rat and human MSCs using flow cytometry and to assess the transgene expression profile using a luciferase expression assay. The effect of transfection with PEI on the viability of MSCs was assessed using the metabolic assay Cell Titer Blue and dsDNA quantification. Levels of IL-1Ra produced by cells following transfection with nanoparticles encoding IL-1Ra was assessed using
Increased revision rates and early failure of Metal-on-Metal (MoM) hip replacements are often due to adverse reaction to metal debris (ARMD). Cobalt is a major component of MoM joints and can initiate an immune response via activation of the innate immune receptor Toll-like receptor 4 (TLR4). This leads to increased secretion of inflammatory cytokines/chemokines e.g. CCL3 and CCL4. The aim of this study was to evaluate whether TLR4-specific neutralising antibodies can prevent cobalt-mediated activation of TLR4. MonoMac 6 (MM6) cells, a human macrophage cell line, were treated with two different TLR4-specific monoclonal antibodies followed by 0.75mM of cobalt chloride (CoCl2). Lipopolysaccharide (LPS), a known TLR4 agonist was used as a positive control.
Aims. Chronic conditions of the wrist may be difficult to manage because
pain and psychiatric conditions are correlated with abnormal function
of the hand. Additionally, intra-articular inflammatory cytokines
may cause pain. We aimed to validate the measurement of inflammatory cytokines
in these conditions and identify features associated with symptoms. Patients and Methods. The study included 38 patients (18 men, 20 women, mean age 43
years) with a chronic condition of the wrist who underwent arthroscopy.
Before surgery, the Self-Rating Depression Scale (SDS), Hand20 questionnaire
and a visual analogue scale (VAS) for pain were used. Cytokine and
chemokine levels in the synovial fluid of the wrist were measured
using
Background. Adverse reactions to metal debris are implicated in the failure of metal-on-metal hip arthroplasty. The peri-implant tissues are often infiltrated by leukocytes which may cause observed immunological effects, including soft tissue necrosis and osteolysis. Cobalt ions from orthopaedic implants aberrantly activate the innate immune receptor human toll-like receptor-4 (TLR4), leading to inflammatory cytokine release including interleukin-8 (IL-8). IL-8 has been shown to increase expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). These factors are essential for leukocyte adhesion to endothelium, which is required for leukocyte migration into tissues. This study investigates cobalt's effect on gene and protein changes in IL-8, ICAM-1 and VCAM-1 to determine their potential role in immune cell infiltration of peri-implant tissues. Methods. TLR4-expressing human dermal microvascular endothelial cells (HMEC-1) were treated with a range of clinically relevant cobalt ion concentrations. IL-8 protein secretion was measured by
Objectives. Osteoarthritis (OA) is the most common form of arthritis, affecting approximately 15% of the human population. Recently, increased concentration of nitric oxide in serum and synovial fluid in patients with OA has been observed. However, the exact role of nitric oxide in the initiation of OA has not been elucidated. The aim of the present study was to investigate the role of nitric oxide in innate immune regulation during OA initiation in rats. Methods. Rat OA was induced by performing meniscectomy surgery while cartilage samples were collected 0, 7, and 14 days after surgery. Cartilage cytokine levels were determined by using
We undertook a prospective study to evaluate the prognostic significance of the serum levels of vascular endothelial growth factor (VEGF) in predicting the survival of patients with osteosarcoma. The levels were measured by an
Background. Increased revision rates and early failure of Metal-on-Metal (MoM) hip replacements are often due to adverse reaction to metal debris (ARMD). ARMD describes numerous symptoms in patients such as pain, osteolysis and soft tissue damage. Cobalt is a major component of MoM joints and can initiate an immune response via activation of the innate immune receptor Toll-like receptor 4 (TLR4). This leads to increased secretion of inflammatory cytokines e.g. interleukin-8 (IL-8). This study investigates whether TLR4-specific antagonists inhibit the inflammatory response to cobalt using IL-8 gene expression and protein secretion as a marker of TLR4 activation. Methods. MonoMac 6 (MM6) cells, a human macrophage cell line, were treated with TLR4-specific antagonists followed by 0.75mM of cobalt chloride. Lipopolysaccharide (LPS), a known TLR4 agonist was used as a positive control.
Coagulase-negative staphylococci produce an exocellular glycolipid antigen which has potential as a serological marker of infection in bone. The value of this newly detected antigen was investigated by
Summary. RNAi targeting p110β reduces TNF-alpha production and osteolysis in response to wear particles. Introduction. Aseptic joint loosening is a key factor that reduces the life span of joint prosthesis. Prosthetic wear particles are thought to play a central role in the initiation and development of periprosthetic osteolysis, leading to aseptic loosening of prostheses. This study aims to explore the effect of p110β-targeted small interfering RNA (siRNA) and lentivirus on particle-induced inflammatory cytokine expression in murine macrophage. Methods. siRNA and lentivirus targeting p110β were transfected and infected prior to particle stimulation, respectively. Ceramic and titanium particles of different sizes were prepared to stimulate macrophages. Fluorescence microscopy showed that the siRNA transfection and lentivirus infection efficiency were 74.2 ± 4.2% and 92.3 ± 2.6%, respectively. Results. Real-time polymerase chain reaction (PCR) showed that the levels of tumor necrosis factor-alpha (TNF-alpha) mRNA in the particle stimulation plus RNA interference (RNAi) groups were significantly lower compared with the particle stimulation-only groups (P<0.05), respectively. Similarly,
Introduction: We have investigated the accuracy of a serological marker to distinguish between septic and aseptic loosening of Total Hip Replacements (THR). We present the preliminary results of our on-going prospective study. Methods: After obtaining Ethical Committee approval, 46 patients were collected in 3 groups; “control” primary THR, revision THR for aseptic loosening, and revision THR for infection. Serum IgG responses to an exocellular bacterial antigen (Lipid S) were determined by
Introduction and Aims: We have investigated the accuracy of a serological marker to distinguish between septic and aseptic loosening of Total Hip Replacements (THR). We present the preliminary results of our ongoing prospective study. Method: After obtaining Ethical Committee approval, 46 patients were collected in three groups: ‘control’ primary THR, revision THR for aseptic loosening and revision THR for infection. Serum IgG responses to an exocellular bacterial antigen (LipidS) were determined by
Introduction: We have investigated the accuracy of a serological marker to distinguish between septic and aseptic loosening of Total Hip Replacements (THR). We present the preliminary results of our on-going prospective study. Methods: After obtaining Ethical Committee approval, 46 patients were collected in 3 groups; “control” primary THR, revision THR for aseptic loosening, and revision THR for infection. Serum IgG responses to an exocellular bacterial antigen (Lipid S) were determined by
Summary. RNAi targeting TNF-alpha inhibits particle-induced inflammation and osteolysis. Introduction. Over 1000,000 joint prostheses are implanted every year in the world. Aseptic joint loosening is a key factor that reduces the longevity of joint prosthesis. Prosthetic wear particles are thought to play a central role in the initiation and development of periprosthetic osteolysis, leading to aseptic loosening of prostheses. This study aims to investigate the effect of RNA interference (RNAi) targeting tumor necrosis factor-alpha (TNF-α) gene on particle-induced inflammation and osteolysis in macrophages in vitro and in vivo. Methods. An in vitro-transcribed small interfering (siRNA) sequences targeting mouse TNF-alpha gene from four candidates was screened and identified and then a lentivirus vector expressing short hairpin RNA (shRNA) was constructed to allow an efficient expression of TNF-alpha-siRNA. Lentivirus-mediated shRNA was transduced into mouse macrophages cells line RAW 264.7. Ceramic particles and titanium particles were added to stimulate cells 24 h after lentivirus transduction. TNF-alpha expression at both mRNA and protein levels were detected quantitatively by real-time polymerase chain reaction (PCR) and
Objectives. To evaluate the contribution of leptin, an adipose-derived hormone, to the pathophysiology of osteoarthritis (OA), by determining leptin in both synovial fluid and cartilage specimens from human joints, and by investigating the effect on cartilage of intra-articular injection of leptin in rat. Methods. Leptin levels were measured by
Background. Intervertebral disc cells exist in a challenging physiological environment. Disc degeneration occurs early in life implying that disc cells may no longer be able to maintain a functional tissue. We hypothesise that disc cells have a stress response different from most other cells because of the disc environment. We have compared the stress response of freshly isolated and cultured bovine nucleus pulposus (NP) cells with bovine dermal fibroblasts, representative of cells from a vascularised tissue. Methods. Freshly isolated and passaged bovine NP cells and dermal fibroblasts were cultured for 3 days then subjected to either thermal stress at 45°C for 1h followed by recovery times of 6, 24 and 48h or nutrient stress involving culture without serum for 6, 24 and 48 h. At each time point, cell number and viability were assessed and heat shock protein 70 (Hsp70) measured in cell lysates by an
Septic arthritis induced by Staphylococcus aureus causes a rapid destruction of joint cartilage and periarticular bone. The mechanisms behind this phenomenon are not fully understood. Toll-like receptors (TLRs) are essential in host defense against pathogens by virtue of their capacity to detect microbes and initiate the immune response. TLR2 is seen as the most important receptor for gram-positive bacteria. TLR2 signaling can lead to the activation of NF-kB through myeloid differentiation factor 88 (MyD88) dependent pathway. The purpose of this study was to examine the catabolic role of TLR2 mediated by the NF-kB pathway in human septic arthritic chondrocytes. Septic arthritic (SA) chondrocytes (n=7) and fibroblast-like synoviocytes (n=7) infected by gram-positive bacteria, mainly Staphylococcus aureus, as well as chondrocytes from healthy individuals (n=5) were used for this study. The expression of TLR2 in septic articular cartilage and normal cartilage was analyzed by real time reverse transcription polymerase chain reaction as well western blot analysis. Production of matrix metalloproteinase MMP- 13 and IL-1b was evaluated by
Alloys of titanium, aluminium, vanadium, iron and other metals are traditional materials used in bone tissue surgery. The anchorage of the metallic materials into the surrounding tissue depends of their mechanical and other physical and chemical properties. The integration of metallic material with the surrounding tissue can be markedly improved by appropriate physicochemical surface properties of the material, such as roughness, topography, wettability or presence of certain chemical functional groups. In the present study the first step the surface roughness of samples of pure Ti or Ti6Al4V alloy. In order to influence the adhesion, growth and presence of bone differentiation markers in human osteoblast-like MG 63 cells, we modified as machining or subsequent polishing by diamond paste was performed. In addition, we investigated the interaction of these cells with a newly developed material for construction of bone-anchoring parts of bone implants. These tested materials were treated either with electro-erosion or plasma-spraying with Ti. After the cells seeding (MG63, human osteoblast-like cells of the line MG 63, derived from osteosarcoma of a 13-year-old boy, on different surfaces, the basic parametrs of adhesion and the viability of bone cells were detected, the cell were analysed and cultered for 1–8 days, during 3 different time intervals(expl.1. 4. and 7 day). Cells number, were detected and analyzed in a ViCell XR analyzer. The concentration of molecules participating in cell adhesion, osteoblastic differentiation, was determined semi-quantitatively by the
INTRODUCTION. Ultra-High Molecular Weight Polyethylene (UHMWPE) wear debris is thought to be a main factor in the development of osteolysis (1). However, the method for the evaluation of the biological response to UHMWPE particles has not yet been standardized. In this study, four different types of UHMWPE particles were generated using a mechanized pulverizing method and the biological responses of macrophages to the particles were investigated using an inverted cell culturing process (2). MATERIALS & METHODS. Virgin samples were manufactured via Direct Compression Molding (DCM) technique from UHMWPE GUR1050 resin powder (Ticona, USA). For vitamin E (VE)-blended sample, the resin was mixed with VE at 0.3 wt% and the mixture was then molded using DCM. The crosslinked virgin samples were made by gamma ray irradiation to UHMWPE GUR1020 resin sheet (Meditech, USA) with doses of 95kGy ±10% and annealed. The VE-blended crosslinked samples were made by electron beam irradiation to VE-blended samples with doses of 300kGy and annealed. The material conditions were summarized in Figure 1. To pulverize the samples, the Multi-Beads Shocker (Yasui Kikai, Japan) was used. After pulverization, samples were dispersed in an ethanol solution and sequentially filtered through polycarbonate filters. Over 100 sections of the filter were selected randomly and images of the particles were analyzed using scanning electron microscope (SEM). To analyze the macrophage biological response, an inverted cell culturing process was used (2). The mouse macrophage-like cells were seeded at densities of 4×105cells per well in a 96-well culture plate and incubated for 1h. UHMWPE particles suspended in the culture medium were then added to each well in the appropriate amount. After that, fresh medium was added to fill the wells, and a sealing film was used to cover the culture plate. The culture plate was then inverted to cause the UHMWPE particles interact with the adhered macrophages. The inverted culture plate was incubated for 8h. The amount of TNF-α was measured by
Summary. Hyaluronan suppressed lipopolysaccharide-stimulated prostaglandin E. 2. production via intercellular adhesion molecule-1 through down-regulation of nuclear factor-κB. Administration of hyaluronan into rheumatoid joints may decrease prostaglandin E. 2. production by activated macrophages, which could result in improvement of arthritic pain. Introduction. Prostaglandin E. 2. (PGE. 2. ) is one of the key mediators of inflammation in rheumatoid arthritis (RA) joints. Intra-articular injection of high molecular weight hyaluronan (HA) into RA knee joints relieves arthritic pain. Although HA has been shown to inhibit PGE. 2. production in cytokine-stimulated synovial fibroblasts, it remains unclear how HA suppresses PGE. 2. production in catabolically activated cells. Furthermore, HA effect on macrophages has rarely been investigated in spite of their contribution to RA joint pathology. Objectives. This study was aimed to investigate the inhibitory mechanism of HA on lipopolysaccharide (LPS)-stimulated PGE. 2. in U937 human macrophage culture system. Methods. With or without pretreatment with one of HA, NS-398, and BAY11-7085, differentiated U937 macrophages were stimulated with LPS. In another set of experiments, the cells were incubated with anti-ICAM-1 antibody or non-specific IgG before pretreatment with HA. PGE. 2. concentrations of the cell-free supernatants were determined using an
Introduction: Despite a resurgence in cobalt-chromium metal-on-metal arthroplasty and hip resurfacing, the potential toxicity of cobalt ions in the periprosthetic area remains a cause for concern. Cytotoxic effects have been demonstrated in macrophages with cobalt ions inducing apoptosis and TNF-α secretion. A similar cytotoxic effect has been demonstrated in osteoblast-like cells. However, these studies assessed the acute cellular response to cobalt ions over 48 hours. To date, the effect on osteoblasts of chronic exposure to cobalt ions is unknown. Aim: In this study we investigated the effect on osteoblasts of chronic exposure to cobalt ions. Specifically we investigated the chemokine response and effect on osteoblast function. We also investigated for a change in osteoblast phenotype to a less differentiated mesenchymal cell type. Methods. Primary human osteoblasts were cultured and treated with cobalt (10ppm) over 21 days. Secreted chemokines (IL-8, MCP-1, TNF-α) were
Summary Statement. An autologous thrombin activated 3-fold PRP, mixed with a biphasic calcium phosphate at a 1mL:1cc ratio, is beneficial for early bone healing in older age sheep. Introduction. The management of bone defects continues to present challenges. Upon activation, platelets secrete an array of growth factors that contribute to bone regeneration. Therefore, combining platelet rich plasma (PRP) with bone graft substitutes has the potential to reduce or replace the reliance on autograft. The simple, autologous nature of PRP has encouraged its use. However, this enthusiasm has failed to consistently translate to clinical expediency. Lack of standardisation and improper use may contribute to the conflicting outcomes reported within both pre-clinical and clinical investigations. This study investigates the potential of PRP for bone augmentation in an older age sheep model. Specifically, PRP dose is controlled to provide clearer indications for its clinical use. Methods. Eighty 11mm diameter defects of 20mm in depth were created in the cancellous bone within the epiphyseal region of the medial proximal tibia and distal femur of twenty five-year-old sheep. The defects were treated with three doses of an autologous thrombin activated PRP combined with a biphasic calcium phosphate (BCP). Activated platelet poor plasma (PPP) and the BCP alone provided reference groups, while the autograft and empty defects served as controls. All animals were sacrificed at four weeks post-operatively for radiographic assessment, micro-computed tomography quantification, histological assessment, histomorphometric quantification of new bone area and bone ingrowth, and weekly fluorochrome bone label quantification. TGF-β1 concentrations were quantified using
Introduction: Selective COX-2 (Cyclooxygenase-2) inhibitors have been found to impede fracture healing. The effect of selective COX-2 inhibitors on tendon healing in a bone tunnel, however, is unknown. Methods: The authors performed bilateral anterior cruciate ligament reconstructions in 32 rabbits and used peripheral quantitative computed tomography (pQCT) to compare tendon-to-bone healing between tunnel aperture and midtunnel regarding bone mineral density (BMD) and ingrowth of new bone. Each animal was assigned to one of four groups. Two groups received selective COX-2 inhibitors orally for 3 weeks (Cele-coxib; 10 mg/kg/d), the two other groups received no COX-2 inhibitors (controls). The animals were sacrificed 3 and 6 weeks after surgery. In biomechanical testing maximum load to failure and stiffness of the tendon grafts were calculated from the load displacement curve and failure modes were recorded. To assess indirectly the effect on local COX-2 activity the synovial content of Prostaglandin E2 (PGE2), the major metabolite of arachnidonic acid metabolism and catalyzed by COX-2, was measured by
Exosomes (exo) are involved in the progression of osteoarthritis (OA). This study aimed to investigate the function of dysfunctional chondrocyte-derived exo (DC-exo) on OA in rats and rat macrophages. Rat-derived chondrocytes were isolated, and DCs induced with interleukin (IL)-1β were used for exo isolation. Rats with OA (n = 36) or macrophages were treated with DC-exo or phosphate-buffered saline (PBS). Macrophage polarization and autophagy, and degradation and chondrocyte activity of cartilage tissues, were examined. RNA sequencing was used to detect genes differentially expressed in DC-exo, followed by RNA pull-down and ribonucleoprotein immunoprecipitation (RIP). Long non-coding RNA osteoarthritis non-coding transcript (OANCT) and phosphoinositide-3-kinase regulatory subunit 5 (PIK3R5) were depleted in DC-exo-treated macrophages and OA rats, in order to observe macrophage polarization and cartilage degradation. The PI3K/AKT/mammalian target of rapamycin (mTOR) pathway activity in cells and tissues was measured using western blot.Aims
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