ABSTRACT
OBJECTIVE: Intra-articular screws are used for internal fixation of osteochondral fragments after fracture or osteochondritis dissecans. This causes cartilage injury potentially leading to chondrocyte death. We have visualised/quantified the hole and zone of cell death (ZCD) in cartilage after drilling/insertion of various articular screws. METHOD: Using an ex vivo bovine model with transmitted light and confocal laser scanning microscopy (CLSM), the holes and ZCD following drilling/insertion of articular screws (cortical screw, headless variable pitch metallic screw, headless variable pitch bioabsorbable screw) were evaluated. In situ chondrocyte death was determined by live/dead cell viability assay. An imaging/quantification protocol was developed to compare hole diameter and ZCD from drilling/insertion of screws into cartilage. The effect of saline irrigation during drilling on the ZCD was also quantified. RESULTS: Screw insertion created holes in cartilage that were significantly (P ≤ 0.001) less than the diameters of the equipment used. With a 1.5 mm drill, a ZCD of 580.2 ± 124 µm was produced which increased to 637.0 ± 44 µm following insertion of a 2 mm cortical screw although this was not significant (P > 0.05). The ZCD from insertion of the variable pitch headless screws (diam. 3.5 mm) was lower for the metallic compared to the bioabsorbable design (800.9 ± 159 vs 1,236.4 ± 212 µm, respectively; P < 0.01). The ZCD from drilling was reduced â¼50% (P < 0.001) by saline irrigation. CONCLUSIONS: Cartilage injury during intra-articular screw fixation caused a ZCD around the hole irrespective of screw design. Saline irrigation significantly reduced the ZCD from drilling into cartilage.
Subject(s)
Bone Screws/adverse effects , Cartilage, Articular/injuries , Chondrocytes/pathology , Fracture Fixation, Internal/adverse effects , Animals , Cartilage, Articular/pathology , Cattle , Cell Death , Disease Models, Animal , Equipment Design , Fracture Fixation, Internal/instrumentation , Microscopy, Confocal/methods , Sodium Chloride , Therapeutic IrrigationABSTRACT
OBJECTIVE: To assess in situ chondrocyte viability following exposure to a laboratory strain and clinical isolates of Staphylococcus aureus. METHODS: Bovine cartilage explants were cultured in the presence of S. aureus 8325-4 (laboratory strain), clinical S. aureus isolates or non-infected culture medium of pH values 7.4, 6.4 and 5.4. All clinical isolates were isolated from the joint aspirates of patients presenting with S. aureus-induced septic arthritis (SA). At designated time points, in situ chondrocyte viability was assessed within defined regions-of-interest in the axial and coronal plane following live- and dead-cell image acquisition using the fluorescent probes 5-chloromethylfluorescein diacetate (CMFDA) and propidium iodide (PI), respectively, and confocal laser-scanning microscopy (CLSM). Cartilage water content, following S. aureus 8325-4 exposure, was obtained by measuring cartilage wet and dry weights. RESULTS: S. aureus 8325-4 and clinical S. aureus isolates rapidly reduced in situ chondrocyte viability (>45% chondrocyte death at 40 h). The increased acidity, observed during bacterial culture, had a minimal effect on chondrocyte viability. Chondrocyte death commenced within the superficial zone (SZ) and rapidly progressed to the deep zone (DZ). Simultaneous exposure of SZ and DZ chondrocytes to S. aureus 8325-4 toxins found SZ chondrocytes to be more susceptible to the toxins than DZ chondrocytes. Cartilage water content was not significantly altered compared to non-infected controls. CONCLUSIONS: Toxins released by S. aureus have a rapid and fatal action on in situ chondrocytes in this experimental model of SA. These data advocate the prompt and thorough removal of bacteria and their toxins during the treatment of SA.
Subject(s)
Arthritis, Infectious/microbiology , Bacterial Toxins/pharmacology , Cartilage, Articular/pathology , Chondrocytes/drug effects , Staphylococcus aureus/pathogenicity , Animals , Arthritis, Infectious/pathology , Body Water/metabolism , Cartilage, Articular/chemistry , Cattle , Cell Death/drug effects , Chondrocytes/pathology , Culture Media/chemistry , Disease Models, Animal , Humans , Hydrogen-Ion Concentration , Microscopy, Confocal , Staphylococcus aureus/classification , Staphylococcus aureus/isolation & purification , Tissue Culture Techniques , VirulenceABSTRACT
OBJECTIVES: Tranexamic acid (TXA) is an anti-fibrinolytic medication commonly used to reduce perioperative bleeding. Increasingly, topical administration as an intra-articular injection or perioperative wash is being administered during surgery. Adult soft tissues have a poor regenerative capacity and therefore damage to these tissues can be harmful to the patient. This study investigated the effects of TXA on human periarticular tissues and primary cell cultures using clinically relevant concentrations. METHODS: Tendon, synovium, and cartilage obtained from routine orthopaedic surgeries were used for ex vivo and in vitro studies using various concentrations of TXA. The in vitro effect of TXA on primary cultured tenocytes, fibroblast-like synoviocytes, and chondrocytes was investigated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cell viability assays, fluorescent microscopy, and multi-protein apoptotic arrays for cell death. RESULTS: There was a significant (p < 0.01) increase in cell death within all tissue explants treated with 100 mg/ml TXA. MTT assays revealed a significant (p < 0.05) decrease in cell viability in all tissues following treatment with 50 mg/ml or 100 mg/ml of TXA within four hours. There was a significant (p < 0.05) increase in cell apoptosis after one hour of exposure to TXA (100 mg/ml) in all tissues. CONCLUSION: The current study demonstrates that TXA caused significant periarticular tissue toxicity ex vivo and in vitro at commonly used clinical concentrations.Cite this article: M. McLean, K. McCall, I. D. M. Smith, M. Blyth, S. M. Kitson, L. A. N. Crowe, W. J. Leach, B. P. Rooney, S. J. Spencer, M. Mullen, J. L. Campton, I. B. McInnes, M. Akbar, N. L. Millar. Tranexamic acid toxicity in human periarticular tissues. Bone Joint Res 2019;8:11-18. DOI: 10.1302/2046-3758.81.BJR-2018-0181.R1.
ABSTRACT
OBJECTIVES: Staphylococcus aureus (S. aureus) is the most commonly implicated organism in septic arthritis, a condition that may be highly destructive to articular cartilage. Previous studies investigating laboratory and clinical strains of S. aureus have demonstrated that potent toxins induced significant chondrocyte death, although the precise toxin or toxins that were involved was unknown. In this study, we used isogenic S. aureus mutants to assess the influence of alpha (Hla)-, beta (Hlb)-, and gamma (Hlg)-haemolysins, toxins considered important for the destruction of host tissue, on in situ bovine chondrocyte viability. METHODS: Bovine cartilage explants were cultured with isogenic S. aureus mutants and/or their culture supernatants. Chondrocyte viability was then assessed within defined regions of interest in the axial and coronal plane following live- and dead-cell imaging using the fluorescent probes 5-chloromethylfluorescein diacetate and propidium iodide, respectively, and confocal laser-scanning microscopy. RESULTS: Hla-producing mutants caused substantial chondrocyte death compared with the toxin-deficient control (Hla-Hlb-Hlg-), whilst mutants producing Hlb and Hlg in the absence of Hla induced minimal chondrocyte death. Coronal studies established that Hla-induced chondrocyte death started in the superficial zone of cartilage and spread to deeper layers, whereas Hlb and Hlg toxins were without significant effect. CONCLUSION: This study identified Hla as a highly potent S. aureus toxin that caused rapid chondrocyte death in bovine cartilage, with other toxins or metabolic products produced by the bacteria playing a minor role. The identification of Hla in mediating chondrocyte death may assist in the development of therapeutic strategies aimed at reducing the extent of cartilage damage during and after an episode of septic arthritis.Cite this article: I. D. M. Smith, K. M. Milto, C. J. Doherty, S. G. B. Amyes, A. H. R. W. Simpson, A. C. Hall. A potential key role for alpha-haemolysin of Staphylococcus aureus in mediating chondrocyte death in septic arthritis. Bone Joint Res 2018;7:457-467. DOI: 10.1302/2046-3758.77.BJR-2017-0165.R1.
ABSTRACT
In Scotland, the number of primary total knee replacements performed annually has been increasing steadily. The price of the implant is fixed but the length of hospital stay is variable. We prospectively investigated all patients who underwent primary unilateral total knee replacement in the Scottish region of Fife, between December 1994 and February 2007 and assessed their recorded pre-operative details. The data were analysed using univariate and multiple linear regression statistical analysis. Data on the length of stay were available from a total of 2106 unilateral total knee replacements. The median length of hospital stay was eight days. The significant pre-operative risk factors for an increased length of stay were the year of admission, details of the consultant looking after the patient, the stair score, the walking-aid score and age. Awareness of the pre-operative factors which increase the length of hospital stay may provide the opportunity to influence them favourably and to reduce the time in hospital and the associated costs of unilateral total knee replacement.