The Impact of Personal Protective Equipment on Speech Discrimination and Verbal Communication in the Operating Room and the Role of Audio Communication Devices.

INTRODUCTION: Recent work has highlighted communication difficulties when wearing personal protective equipment (PPE) in the clinical setting, but currently, there are little objective data on its effects. We assessed the impact of PPE on verbal communication in a simulated operating room and evaluated use of an audio communication device. METHODOLOGY: Frontline health professionals across specialties including surgery, anesthetics, and nursing undertook speech discrimination testing with and without standardized levels of PPE in a simulated operating room environment. Background noise (30- and 70-a-weighted decibel multitalker babble) at 2 distances (2 and 4 m) were selected representative of operating room environments. Bamford-Kowal-Bench (BKB) scoring (192 sentences per participant) was performed. A Digital Multichannel Transceiver System (DMTS) was evaluated. We assessed the effects of PPE use, distance, and use of the DMTS with pairwise comparisons, using a Bonferroni correction, and assessed participant experience via Likert scales. RESULTS: Thirty-one healthcare professionals were tested. Without PPE in 70-a-weighted decibel "babble," median BKB sentence scores were 90% and 76% at 2 and 4 m (adjusted P < 0.0005). The median BKB sentence scores dropped to 8% and 4% at 2 and 4 m in PPE (adjusted P < 0.0005). Improved speech discrimination was achieved with DMTS use to 70% and 76% at 2 and 4 m. Personal protective equipment led to a statistically significant reduction in BKB scores across all conditions compared with baseline. Overall participant confidence in PPE clinical communication was low. CONCLUSIONS: Addition of PPE dramatically impairs speech discrimination and communication in high levels of background noise characteristic of clinical environments, which can be significantly improved using DMTS. Measures should be taken by teams through both through reduction of background noise and consideration of assistive technologies maximizing patient safety. This may be further rehearsed in a simulation environment.

Septic arthritis in an in vivo murine model induced by Staphylococcus aureus : a comparison between actions of the haemolysin toxin and the effects of the host immune response.

AIMS: Staphylococcus aureus is a major cause of septic arthritis, and in vitro studies suggest α haemolysin (Hla) is responsible for chondrocyte death. We used an in vivo murine joint model to compare inoculation with wild type S. aureus 8325-4 with a Hla-deficient strain DU1090 on chondrocyte viability, tissue histology, and joint biomechanics. The aim was to compare the actions of S. aureus Hla alone with those of the animal's immune response to infection. METHODS: Adult male C57Bl/6 mice (n = 75) were randomized into three groups to receive 1.0 to 1.4 × 107 colony-forming units (CFUs)/ml of 8325-4, DU1090, or saline into the right stifle joint. Chondrocyte death was assessed by confocal microscopy. Histological changes to inoculated joints were graded for inflammatory responses along with gait, weight changes, and limb swelling. RESULTS: Chondrocyte death was greater with 8325-4 (96.2% (SD 5.5%); p < 0.001) than DU1090 (28.9% (SD 16.0%); p = 0.009) and both were higher than controls (3.8% (SD 1.2%)). Histology revealed cartilage/bone damage with 8325-4 or DU1090 compared to controls (p = 0.010). Both infected groups lost weight (p = 0.006 for both) and experienced limb swelling (p = 0.043 and p = 0.018, respectively). Joints inoculated with bacteria showed significant alterations in gait cycle with a decreased stance phase, increased swing phase, and a corresponding decrease in swing speed. CONCLUSION: Murine joints inoculated with Hla-producing 8325-4 experienced significantly more chondrocyte death than those with DU1090, which lack the toxin. This was despite similar immune responses, indicating that Hla was the major cause of chondrocyte death. Hla-deficient DU1090 also elevated chondrocyte death compared to controls, suggesting a smaller additional deleterious role of the immune system on cartilage.Cite this article: Bone Joint Res 2022;11(9):669-678.

The role of disrupted iron homeostasis in the development and progression of arthropathy.

Arthropathy or joint disease leads to significant pain and disability irrespective of etiology. Clinical and experimental evidence point to the presence of considerable links between arthropathy and iron overload. Previous work has suggested that iron accumulation in the joints is often associated with increased oxidative stress, disrupted matrix metabolism, and cartilage degeneration. However, key issues regarding the role of iron overload in the pathogenesis of arthropathy remain ambiguous. For example, significant gaps in our knowledge of the primary cellular targets of iron overload-induced damage and the exact molecular mechanism through which disrupted iron homeostasis leads to joint damage still exist. The exact signaling pathway that links iron metabolism and cellular damage in arthropathy also remains largely unmapped. In this review, we focus on the relationship between iron overload and arthropathy with special emphasis on the adversarial relationship between iron that accumulates in the joints over time and cartilage homeostasis. A better understanding of the mechanisms and pathways underlying iron-induced cartilage degeneration may help in defining new prognostic markers and therapeutic targets in arthropathy.

Iron Overload Induces Oxidative Stress, Cell Cycle Arrest and Apoptosis in Chondrocytes.

Clinical and experimental evidence point to the presence of considerable links between arthropathy, osteoarthritis (OA) in particular, and iron overload possibly due to oxidative stress and tissue damage. However, the specific cellular targets of iron overload-related oxidative stress in OA remain ambiguous. We examined the effects of iron overload on chondrocyte health using the C-20/A4 chondrocyte cell line. Cells were treated with increasing concentrations of ferric ammonium citrate (FAC) to mimic iron overload in vitro. Treated cells were assessed for cell viability, cycling, apoptosis, collagen II synthesis, and oxidative stress along with cellular iron content and the expression of key iron regulatory genes. FAC treatment resulted in an increase in ferritin expression and a significant decrease in the expression of hepcidin, ferroportin, transferrin receptors 1 (TfR1) and TfR2. Increased labile iron content was also evident, especially in cells treated with high FAC at 24 h. High doses of FAC treatment also induced higher levels of reactive oxygen species, reduced collagen II production, disrupted cell cycle and higher cell death as compared with untreated controls. In conclusion, findings presented here demonstrate that iron overload disrupts cellular iron homeostasis, which compromises the functional integrity of chondrocytes and leads to oxidative stress and apoptosis.

Anatomical, functional and biomechanical review of the glenoid labrum.

The glenohumeral joint is the most mobile joint in the human skeleton, supported by both active and passive stabilisers. As one of the passive stabilisers, the glenoid labrum has increasingly been recognised to play an important role in stability of the glenohumeral joint, acting to maintain intraarticular pressure, centralise the humeral head and contribute to concavity-compression stability. Several studies have investigated the macro- and micro-anatomical features of the labrum as well as its biomechanical function. However, in order to better understand the role of the labrum and its mechanics, a comprehensive anatomical, functional and biomechanical review of these studies is needed. Therefore, this article reviews the current literature detailing anatomical descriptions of the glenoid labrum, with an emphasis on its function(s) and biomechanics, as well as its interaction with neighbouring structures. The intimate relationship between the labrum and the surrounding structures was found to be important in glenohumeral stability, which owes further investigation into the microanatomy of labrum to better understand this relationship.

Cell-associated type I collagen in nondegenerate and degenerate human articular cartilage.

Chondrocytes with abnormal morphology are present in nondegenerate human cartilage suggesting dedifferentiation to a fibroblastic phenotype and production of a mechanically-weakened matrix of unknown composition. We determined the relationship between in situ chondrocyte morphology, chondrocyte clusters, and levels of cell-associated collagen type I. Chondrocyte morphology in fresh femoral head cartilage from 19 patients with femoral neck fracture and collagen type I labelling was identified with Cell TrackerTM fluorescence and immunofluorescence, respectively, in axial/coronal orientations using confocal microscopy with images analysed by ImarisTM . In axial images of grade 0 cartilage, 87 ± 8% were normal chondrocytes with a small (10 ± 6%) abnormal population possessing ≥1 cytoplasmic process. More normal chondrocytes (78 ± 11%) were collagen type I negative than those labelling positively (p < 0.001). For abnormal chondrocytes, 81 ± 14% labelled negatively for collagen type I compared to those labelling positively (19 ± 3%; p = 0.007; N(n)=11(3)). Overall, approximately 9% of the cells in normal cartilage labelled for collagen type I. With degeneration, the percentage of normal chondrocytes decreased (p < 0.001) but increased for abnormal cells (p = 0.036) and clusters (p = 0.003). A larger percentage of normal, abnormal and clustered chondrocytes now demonstrated collagen type I labelling (p = 0.004; p = 0.009; p = 0.001 respectively). Coronal images exhibited increased (p = 0.001) collagen type I labelling in the superficial zone of mildly degenerate cartilage with none in the mid or deep zones. These results show that collagen type I was identified around normal and abnormal chondrocytes in nondegenerate cartilage, which increased with degeneration. This suggested the presence of mechanically weak fibro-cartilaginous repair tissue in otherwise macroscopically nondegenerate human cartilage which progressed with degeneration as occurs in osteoarthritis.

Optimization and Validation of a Human Ex Vivo Femoral Head Model for Preclinical Cartilage Research and Regenerative Therapies.

OBJECTIVE: Articular cartilage is incapable of effective repair following injury or during osteoarthritis. While there have been developments in cartilage repair technologies, there is a need to advance biologically relevant models for preclinical testing of biomaterial and regenerative therapies. This study describes conditions for the effective ex vivo culture of the whole human femoral head. DESIGN: Fresh, viable femoral heads were obtained from femoral neck fractures and cultured for up to 10 weeks in (a) Dulbecco's modified Eagle's medium (DMEM); (b) DMEM + mixing; (c) DMEM + 10% human serum (HS); (d) DMEM + 10% HS + mixing. The viability, morphology, volume, and density of fluorescently labelled in situ chondrocytes and cartilage surface roughness were assessed by confocal microscopy. Cartilage histology was studied for glycosaminoglycan content using Alcian blue and collagen content using picrosirius red. RESULTS: Chondrocyte viability remained at >95% in DMEM + 10% HS. In DMEM alone, viability remained high for ~4 weeks and then declined. For the other conditions, superficial zone chondrocyte viability fell to <35% at 10 weeks with deeper zones being relatively unaffected. In DMEM + 10% HS at 10 weeks, the number of chondrocytes possessing cytoplasmic processes increased compared with DMEM (P = 0.017). Alcian blue labeling decreased (P = 0.02) and cartilage thinned (P ≤ 0.05); however, there was no change to surface roughness, chondrocyte density, chondrocyte volume, or picrosirius red labeling (P > 0.05). CONCLUSIONS: In this ex vivo model, chondrocyte viability was maintained in human femoral heads for up to 10 weeks in culture, a novel finding not previously reported. This human model could prove invaluable for the exploration, development, and assessment of preclinical cartilage repair and regenerative therapies.

Effects of Antibiotics on α-Toxin Levels during Staphylococcus aureus Culture: Implications for the Protection of Chondrocytes in a Model of Septic Arthritis.

OBJECTIVE: Septic arthritis results from joint infection by Staphylococcus aureus, which produces potent α-toxin causing cell death, potentially leading to permanent cartilage damage. Treatment is by joint irrigation and antibiotics, although it is unclear if, following treatment with antibiotics which cause bacterial lysis, there is release of additional stored α-toxin. DESIGN: A rabbit erythrocyte hemolysis assay was optimised to assess biologically-active α-toxin from cultured S. aureus α-toxin strain DU5946. Hemoglobin release was measured spectrophotometrically following addition of a bacteriostatic antibiotic (linezolid) or a bacteriolytic antibiotic (penicillin). A bovine cartilage model of septic arthritis was used to test the protective effects of antibiotics against S. aureus infection. RESULTS: During S. aureus culture, α-toxin levels increased rapidly but the rate of rise was quickly (within 20 minutes) suppressed by linezolid (25 µg/mL). Penicillin also reduced the increase in α-toxin levels; however, the time course was relatively slow compared to linezolid even at high concentrations (50,000 U/mL). The efficacy of penicillin (250,000 U/mL) at reducing the rise in α-toxin was approximately 8% less than that of linezolid (P < 0.05) suggesting the presence of additional toxin. This could be due to a delayed action of penicillin, and/or release of a small pool of stored α-toxin from dying bacteria. In a bovine cartilage model, however, there was no difference between the protection of in situ chondrocytes against S. aureus by penicillin or linezolid (P > 0.05). CONCLUSION: The results suggested that equally effective protection of chondrocytes against S. aureus septic arthritis may be obtained by the bacteriostatic or bacteriolytic antibiotics tested.

Editorial Commentary: Raising the Osmolarity of Arthroscopic Irrigating Solutions May Be Chondroprotective: We Must Be Kind to Joints During Arthroscopy!

The irrigation of joints during arthroscopic procedures typically uses a non-physiological solution. This replaces the natural synovial fluid and rapidly subjects the connective tissues to an alien hypo-osmotic environment in which cartilage cells are far more sensitive to iatrogenic injury. Raising the osmolarity of the irrigating solution may be a simple, safe, and effective chondroprotective strategy.

Intraoperative Cochlear Implant Reinsertion Effects Evaluated by Electrode Impedance.

OBJECTIVES: To assess the effect on impedance levels of intraoperative reinsertion of a cochlear implant (CI) array compared with matched controls. STUDY DESIGN: Retrospective patient review. SETTING: Cochlear implant center. PATIENTS: CI recipients in the Sydney Cochlear Implant Centre (SCIC) database who required intraoperative array reinsertion and matched controls. Exclusion criteria; known preceding meningitis or labyrinthitis ossificans; electrode array buckling; incomplete "final" insertion. INTERVENTION: Cochlear implantation. MAIN OUTCOME MEASURES: Impedance values measured intraoperatively, at switch on, 3 months, 6 months, and 12 months postoperatively were analyzed. The Generalized Estimating Equation (GEE) Model was used to compare cases with controls for each device, at each time point, and for each channel. RESULTS: Thirty-one reinsertion cases identified; six CI 422 arrays; 14 CI 24RE (ST) arrays, and 11 CI 512 arrays. No increase in impedance levels was found in the reinsertion cases when compared with their matched controls. The only statistical difference in impedance was seen in the CI 422 cohort at switch on with the reinsertion cases having lower impedances (p = 0.03). CONCLUSION: This is the first study to examine impedance values in patients who underwent intraoperative CI array reinsertion and to compare them with the impedances of matched controls. No significant increase found in impedances between our reinsertion cases and matched controls, suggesting the reinsertion did not result in any additional trauma or inflammation. This has implications for surgery both in routine cases such as a faulty electrode and also for future design of mechanisms for delivery of intracochlear therapies.

Optimizing the Composition of Irrigation Fluid to Reduce the Potency of Staphylococcus aureus α-Toxin: Potential Role in the Treatment of Septic Arthritis.

OBJECTIVE: Septic arthritis is commonly caused by Staphylococcus aureus and is a medical emergency requiring antibiotics and joint irrigation. The bacteria produce α-toxin causing rapid cartilage cell (chondrocyte) death. Saline (0.9%NaCl) lavage is normally used to remove bacteria and toxins, however, its composition might be suboptimal to suppress the lethal effects of α-toxin. We utilized rabbit erythrocyte hemolysis as a sensitive, biologically relevant assay of α-toxin levels to determine if changes to osmolarity, temperature, pH, and divalent cation (Mg2+, Ca2+) concentration were protective. DESIGN: Erythrocytes were incubated in the various conditions and then exposed to α-toxin ("chronic" challenge) or incubated with α-toxin and then exposed to experimental conditions ("acute" challenge). RESULTS: Raising osmolarity from 300 mOsm (0.9%NaCl) to 400, 600, or 900 mOsm (sucrose addition) when applied chronically, significantly reduced hemolysis linearly. As an acute challenge, osmotic protection was significant and similar over 400 to 900 mOsm. Reducing temperature chronically from 37°C to 25°C and 4°C significantly reduced hemolysis, however, when applied as an acute challenge although significant, was less marked. Divalent cations (Mg2+, Ca2+ at 5mM) reduced hemolysis. Varying pH (6.5, 7.2, 8.0) applied chronically marginally reduced hemolysis. The optimized saline (0.9% NaCl; 900 mOsm with sucrose, 5 mM MgCl2 (37°C)) rapidly and significantly reduced hemolysis compared with saline and Hank's buffered saline solution applied either chronically or acutely. CONCLUSIONS: These results on the effect of S. aureus α-toxin on erythrocytes showed that optimizing saline could markedly reduce the potency of S. aureus α-toxin. Such modifications to saline could be of benefit during joint irrigation for septic arthritis.

Damage control articular surgery: Maintaining chondrocyte health and minimising iatrogenic injury.

Articular cartilage has limited intrinsic regenerative potential. The maintenance of healthy articular cartilage is essential to prevent joint degeneration and the morbidity associated with arthritis. In this review, we outline the structure and function of healthy articular cartilage. We summarise some of the recent literature outlining the influence of surgical factors on chondrocyte health. These factors include mechanical injury from instrumentation and drilling, drying, and the influence of irrigation fluids, antimicrobial solutions and local anaesthetics. We demonstrate that there is scope for improving cartilage viability at the time of surgery if simple chondroprotective measures are routinely adopted.

Total endoscopic stapes surgery: Systematic review and pooled analysis of audiological outcomes.

OBJECTIVE: This study evaluates the current evidence base for total endoscopic stapes surgery, specifically to establish current efficacy and safety of the technique within clinical practice. DATA SOURCES: A systematic review of the literature on endoscopic stapes surgery was performed according to the Preferred Reporting Items for Systematic Reviews and Meta-Analysis checklist. A comprehensive search of PubMed, Embase, and the Cochrane Central Register of Controlled Trials database for relevant publications for all available dates with appropriate Medical Subject Headings search criteria in January 2018. STUDY SELECTION: Out of the 160 articles identified in the search, 14 met the inclusion criteria for further analysis. Four of these were level III and 10 level IV evidence. DATA EXTRACTION: A pooled patient population of 314 individuals was analyzed. DATA SYNTHESIS: Documented postoperative air bone gap closure to within 20 dB was found in 95.3% of individuals (285 patients). Facial nerve palsy (temporary) occurred in three patients (0.6%), with all recovering. No total sensorineural hearing losses were recorded, but two moderate sensorineural hearing losses occurred (0.6%). Perilymph fistula was noted on four occasions (1.3%). Chorda tympani trauma was documented in 3.5% of cases, with taste disturbance documented in 13 patients (5%). CONCLUSIONS: Our pooled analysis uses the current published evidence to establish the complication rate and audiological outcome for the endoscopic approach to stapes surgery. These outcomes are comparable to those documented in traditional use of the microscope for stapedectomy. Laryngoscope, 130:1282-1286, 2020.

Cochlear Implant Outcomes in Large Vestibular Aqueduct Syndrome-Should We Provide Cochlear Implants Earlier?

OBJECTIVE: Examine postoperative speech perception outcomes in a large vestibular aqueduct syndrome (LVAS) patients at a major cochlear implantation center. STUDY DESIGN: Retrospective analysis of the Sydney Cochlear Implant Centre (SCIC) database and medical records from January 1994 to December 2015 was performed. SETTING: Tertiary referral center. PATIENTS: Patients with a diagnosis of LVAS who received a cochlear implant (CI). Only those with speech perception outcomes recorded at least 12 months post implant were included in our analysis. INTERVENTION(S): Therapeutic. MAIN OUTCOME MEASURE(S): Postoperative speech perception scores. RESULTS: Between 1994 and 2015, 176 adult and pediatric patients with a diagnosis of LVAS underwent cochlear implantation at SCIC. Postoperative Bamford-Kowal Bench (BKB) sentence test scores were obtained for 97 patients. The postoperative median BKB score was 93% with a lower quartile score of 85% and an upper quartile score of 98%. Smaller numbers were available for post-CI City University of New York (CUNY) and Consonant-Nucleus-Consonant (CNC) word scores yet similar excellent results were seen. CONCLUSIONS: Our study results suggest the CI should be considered when BKB scores have dropped to 85%. We suggest that rather than LVAS cases representing a challenge to cochlear implantation, they are amongst the best candidates for surgery, and should receive a CI at an earlier stage in hearing loss, when they have better speech perception. This allows stable hearing to be established earlier along with excellent speech perception outcomes.

The Role of Chondrocyte Morphology and Volume in Controlling Phenotype-Implications for Osteoarthritis, Cartilage Repair, and Cartilage Engineering.

PURPOSE OF REVIEW: Articular chondrocytes are exclusively responsible for the turnover of the extracellular matrix (ECM) of hyaline cartilage. However, chondrocytes are phenotypically unstable and, if they de-differentiate into hypertrophic or fibroblastic forms, will produce a defective and weak matrix. Chondrocyte volume and morphology exert a strong influence over phenotype and a full appreciation of the factors controlling chondrocyte phenotype stability is central to understanding (a) the mechanisms underlying the cartilage failure in osteoarthritis (OA), (b) the rationale for hyaline cartilage repair, and (c) the strategies for improving the engineering of resilient cartilage. The focus of this review is on the factors involved in, and the importance of regulating, chondrocyte morphology and volume as key controllers of chondrocyte phenotype. RECENT FINDINGS: The visualisation of fluorescently-labelled in situ chondrocytes within non-degenerate and mildly degenerate cartilage, by confocal scanning laser microscopy (CLSM) and imaging software, has identified the marked heterogeneity of chondrocyte volume and morphology. The presence of chondrocytes with cytoplasmic processes, increased volume, and clustering suggests important early changes to their phenotype. Results from experiments more closely aligned to the normal physico-chemical environment of in situ chondrocytes are emphasising the importance of understanding the factors controlling chondrocyte morphology and volume that ultimately affect phenotype. An appreciation of the importance of chondrocyte volume and morphology for controlling the chondrocyte phenotype is advancing at a rapid pace and holds particular promise for developing strategies for protecting the chondrocytes against deleterious changes and thereby maintaining healthy and resilient cartilage.

The clustering and morphology of chondrocytes in normal and mildly degenerate human femoral head cartilage studied by confocal laser scanning microscopy.

Chondrocytes are the major cell type present in hyaline cartilage and they play a crucial role in maintaining the mechanical resilience of the tissue through a balance of the synthesis and breakdown of extracellular matrix macromolecules. Histological assessment of cartilage suggests that articular chondrocytes in situ typically occur singly and demonstrate a rounded/elliptical morphology. However, there are suggestions that their grouping and fine shape is more complex and that these change with cartilage degeneration as occurs in osteoarthritis. In the present study we have used confocal laser scanning microscopy and fluorescently labelled in situ human chondrocytes and advanced imaging software to visualise chondrocyte clustering and detailed morphology within grade-0 (non-degenerate) and grade-1 (mildly degenerate) cartilage from human femoral heads. Graded human cartilage explants were incubated with 5-chloromethylfluorescein diacetate and propidium iodide to identify the morphology and viability, respectively, of in situ chondrocytes within superficial, mid- and deep zones. In grade-0 cartilage, the analysis of confocal microscope images showed that although the majority of chondrocytes were single and morphologically normal, clusters (i.e. three or more chondrocytes within the enclosed lacunar space) were occasionally observed in the superficial zone, and 15-25% of the cell population exhibited at least one cytoplasmic process of ~ 5 µm in length. With degeneration, cluster number increased (~ 50%) but not significantly; however, the number of cells/cluster (P < 0.001) and the percentage of cells forming clusters increased (P = 0.0013). In the superficial zone but not the mid- or deep zones, the volume of clusters and average volume of chondrocytes in clusters increased (P < 0.001 and P < 0.05, respectively). The percentage of chondrocytes with processes, the number of processes/cell and the length of processes/cell increased in the superficial zone of grade-1 cartilage (P = 0.0098, P = 0.02 and P < 0.001, respectively). Processes were categorised based on length (L0 - no cytoplasmic processes; L1 < 5 µm; 5 < L2 ≤ 10 µm; 10 < L3 ≤ 15 µm; L4 > 15 µm). With cartilage degeneration, for chondrocytes in all zones, there was a significant decrease (P = 0.015) in the percentage of chondrocytes with 'normal' morphology (i.e. L0), with no change in the percentage of cells with L1 processes; however, there were significant increases in the other categories. In grade-0 cartilage, chondrocyte clustering and morphological abnormalities occurred and with degeneration these were exacerbated, particularly in the superficial zone. Chondrocyte clustering and abnormal morphology are associated with aberrant matrix metabolism, suggesting that these early changes to chondrocyte properties may be associated with cartilage degeneration.

Chondroprotection in Models of Cartilage Injury by Raising the Temperature and Osmolarity of Irrigation Solutions.

Objectives During arthroscopic or open joint surgery, articular cartilage may be subjected to mechanical insults by accident or design. These may lead to chondrocyte death, cartilage breakdown and posttraumatic osteoarthritis. We have shown that increasing osmolarity of routinely used normal saline protected chondrocytes against injuries that may occur during orthopedic surgery. Often several liters of irrigation fluid are used during an orthopedic procedure, which is usually kept at room temperature, but is sometimes chilled. Here, we compared the effect of normal and hyperosmolar saline solution at different temperatures on chondrocyte viability following cartilage injury using in vitro and in vivo models of scalpel-induced injury. Design Cartilage injury was induced in bovine osteochondral explants and the patellar groove of rats in vivo by a single pass of a scalpel blade in the presence of normal saline (300 mOsm) or hyperosmolar saline solution (600 mOsm, sucrose addition) at 4°C, 21°C, or 37°C. Chondrocytes were fluorescently labeled and visualized by confocal microscopy to assess cell death. Results Hyperosmolar saline reduced scalpel-induced chondrocyte death in both bovine and rat cartilage by ~50% at all temperatures studied (4°C, 21°C, 37°C; P < 0.05). Raising temperature of both irrigation solutions to 37°C reduced scalpel-induced cell death ( P < 0.05). Conclusions Increasing the osmolarity of normal saline and raising the temperature of the irrigation solutions to 37°C reduced chondrocyte death associated with scalpel-induced injury in both in vitro and in vivo cartilage injury models. A hyperosmolar saline irrigation solution at 37°C may protect cartilage by decreasing the risk of chondrocyte death during mechanical injury.

Chondrocyte Morphology in Stiff and Soft Agarose Gels and the Influence of Fetal Calf Serum.

Changes to chondrocyte volume/morphology may have deleterious effects on extracellular matrix (ECM) metabolism potentially leading to cartilage deterioration and osteoarthritis (OA). The factors controlling chondrocyte properties are poorly understood, however, pericellular matrix (PCM) weakening may be involved. We have studied the density, volume, morphology, and clustering of cultured bovine articular chondrocytes within stiff (2% w/v) and soft (0.2% w/v) three-dimensional agarose gels. Gels with encapsulated chondrocytes were cultured in Dulbecco's Modified Eagle's Medium (DMEM; fetal calf serum (FCS) 1-10%;380 mOsm) for up to 7 days. Chondrocytes were fluorescently labeled after 1, 3, and 7 days with 5-chloromethylfluorescein-diacetate (CMFDA) and propidium iodide (PI) or 1,5-bis{[2-(di-methylamino)ethyl]amino}-4,8-dihydroxyanthracene-9,10-dione (DRAQ5) to identify cytoplasmic space or DNA and imaged by confocal laser scanning microscopy (CLSM). Chondrocyte density, volume, morphology, and clustering were quantified using Volocity™ software. In stiff gels after 7 d with 10% FCS, chondrocyte density remained unaffected and morphology was relatively normal with occasional cytoplasmic processes. However, in soft gels by day 1, chondrocyte volume increased (P = 0.0058) and by day 7, density increased (P = 0.0080), along with the percentage of chondrocytes of abnormal morphology (P < 0.0001) and enhanced clustering (P < 0.05), compared to stiff gels. FCS exacerbated changes to density (P < 0.01), abnormal morphology (P < 0.001) and clustering (P < 0.01) compared to lower concentrations at the same gel strength. Reduced gel stiffness and/or increased FCS concentrations promoted chondrocyte proliferation and clustering, increased cell volume, and stimulated abnormal morphology, producing similar changes to those occurring in OA. The increased penetration of factors in FCS into soft gels may be important in the development of these abnormal chondrocyte properties. J. Cell. Physiol. 232: 1041-1052, 2017. © 2016 Wiley Periodicals, Inc.

Mapping Chondrocyte Viability, Matrix Glycosaminoglycan, and Water Content on the Surface of a Bovine Metatarsophalangeal Joint.

OBJECTIVE: The purpose of this study was to determine if there were variations in chondrocyte viability, matrix glycosaminoglycan (GAG), and water content between different areas of the articular surface of a bovine metatarsophalangeal joint, a common and reliable source of articular cartilage for experimental study, which may compromise the validity of using multiple samples from different sites within the joint. METHODS: Nine fresh cadaveric bovine metatarsophalangeal joints were obtained. From each joint, 16 osteochondral explants were taken from 4 facets, yielding a total of 144 cartilage specimens for evaluation of chondrocyte viability, matrix GAG, and water content. A less invasive method for harvesting osteochondral explants and for processing the biopsy for the assessment of chondrocyte viability was developed, which maintained maximal viability within each cartilage explant. RESULTS: There was no significant difference between the 16 biopsy sites from the different areas of the joint surface with respect to chondrocyte viability, matrix GAG and water content. Pooled data of all samples from each joint established the baseline values of chondrocyte viability to be 89.4% ± 3.8%, 94.4% ± 2.2%, and 77.9% ± 7.8%, in the superficial quarter, central half, and deep quarter (with regard to depth from the articular surface), respectively. The matrix GAG content of bovine articular cartilage was 6.06 ± 0.41 µg/mg cartilage, and the cartilage water content was 72.4% ± 1.5%. There were also no significant differences of these 3 variables between the different joints. CONCLUSION: It is thus reasonable to compare biopsies obtained from different sites, as a biopsy from one site would be considered representative of the whole joint.