The Effect of Different Saline Suspension Height on Injured Articular Cartilage During Arthroscopic Surgery.

Background Osteochondral allografts are currently stored at 4°C for 2 to 6 weeks before implantation. At 4°C, chondrocyte viability, especially in the superficial zone, deteriorates starting at 2 weeks. Alternative storage conditions could maintain chondrocyte viability beyond 2 weeks, and thereby facilitate increased graft availability and enhanced graft quality. Purpose The objective of the study was to determine the effects of prolonged 37°C storage compared with traditional 4°C storage on chondrocyte viability and cartilage matrix content. Study Design Controlled laboratory study. Methods Osteochondral samples from humeral heads of adult goats were analyzed (i) fresh, or after storage in medium for (ii) 14 days at 4°C including 10% fetal bovine serum, (iii) 28 days at 4°C including 10% fetal bovine serum, (iv) 28 days at 37°C without fetal bovine serum, (v) 28 days at 37°C including 2% fetal bovine serum, or (vi) 28 days at 37°C including 10% fetal bovine serum. Portions of samples were analyzed by microscopy after LIVE/DEAD staining to determine chondrocyte viability and density, both en face (to visualize the articular surface) and vertically (overall and in superficial, middle, and deep zones). The remaining cartilage was analyzed for sulfated glycosaminoglycan and collagen. Results The 37°C storage maintained high chondrocyte viability compared with 4°C storage. Viability of samples after 28 days at 37°C was ˜80% at the cartilage surface en face, ˜65% in the superficial zone, and ˜70% in the middle zone, which was much higher than ˜45%, ˜20%, and ˜35%, respectively, in 4°C samples after 28 days, and slightly decreased from ˜100%, ˜85%, and ˜95%, respectively, in fresh controls. Cartilage thickness, glycosaminoglycan content, and collagen content were maintained for 37°C and 4°C samples compared with fresh controls. Conclusion The 37°C storage of osteochondral grafts supports long-term chondrocyte viability, especially at the vulnerable surface and superficial zone of cartilage. Clinical Relevance Storage of allografts at a physiologic temperature of 37°C may prolong storage duration, improve graft availability, and improve treatment outcomes.

Micro-particles From Arthroscopic Tools May Induce A Pathologic Cascade Mediated by Synoviocytes (SS-20)

Grain-size distribution in suspension over a sand-gravel bed in open channel flow

The levitation state of a large magnetic sphere held in equilibrium above a thick superconducting layer in the Meissner state is a single temperature-independent state as long as the maximum magnetic field at the superconducting (SC) surface does not exceed the critical field ${\mathit{H}}_{\mathit{c}}$(T). In contrast, a magnetic microsphere trapped by a superconducting microring exhibits very different behavior. When the radius b of the SC ring is of the same order as the Ginzburg-Landau coherence length \ensuremath{\xi}(T), the system exhibits, in general, a small set of distinct, quantized, temperature-dependent levitation and suspension states. For certain discrete values of b the flux in the ring is quantized, and the levitation and suspension heights are temperature independent. An abrupt temperature induced transition in the suspension height is also found for a special set of parameters. \textcopyright{} 1996 The American Physical Society.

Background To date, the morphological, biochemical, and biomechanical characteristics of articular cartilage in osteochondral allografts that have been stored have not been fully described. Hypothesis Osteochondral allografts procured and stored commercially for a standard period as determined by tissue banking protocol will have compromised chondrocyte viability but preserved extracellular matrix quality. Study Design Controlled laboratory study. Methods Unused cartilage from 16 consecutive osteochondral allografts was sampled during surgery after tissue bank processing and storage. Ten grafts were examined for cell viability and viable cell density using confocal microscopy, proteoglycan synthesis via 35SO4 uptake, and glycosaminoglycan content and compared with fresh cadaveric articular cartilage. Biomechanical assessment was performed on the 6 remaining grafts by measuring the indentation stiffness of the cartilage. Results The mean storage time for the transplanted specimens was 20.3 ± 2.9 days. Chondrocyte viability, viable cell density, and 35SO4 uptake were significantly lower in allografts at implantation when compared to fresh, unstored controls, whereas matrix characteristics, specifically glycosaminoglycan content and biomechanical measures, were unchanged. In addition, chondrocyte viability in the stored allografts was preferentially decreased in the superficial zone of cartilage. Conclusion Human osteochondral allografts stored for a standard period (approximately 3 weeks) before implantation undergo decreases in cell viability, especially in the critically important superficial zone, as well as in cell density and metabolic activity, whereas matrix and biomechanical characteristics appear conserved. The exact clinical significance of these findings, however, is unknown, as there are no prospective studies examining clinical outcomes using grafts stored for extended periods. Clinical Relevance Surgeons who perform this procedure should understand the cartilage characteristics of the graft after 21 days of commercial storage in serum-free media.

Flagellin, a specific ligand for Toll-like receptor 5 (TLR5), is a molecular pattern associated with several bacterial species. Recently, TLR signaling has been intensively studied. However, TLR5-associated signaling in non-transformed colonocytes has not been investigated. Here we studied the expression of cytokines induced by flagellin in non-transformed human colonic NCM460 cells and the signaling mechanisms mediating these responses. Cytokine expression array experiments showed that exposure of the cells to flagellin (100 ng/ml) for 12 h increased the expression of interleukin (IL)-8 and macrophage-inflammatory protein 3alpha (MIP3alpha) in a TLR5-specific manner. Flagellin also activated MAP kinases (ERK1/2, JNK, and p38) and degraded IkappaBalpha. Dominant negative MEK1 (a kinase that activates ERK1/2) blocked flagellin-stimulated IL-8 and MIP3alpha transcriptional activity, while the MEK-specific inhibitors PD98059 and U0126 reduced protein production of these cytokines. Conversely, transfection with a constitutively active MEK1 increased IL-8 and MIP3alpha transcriptional activity in a NFkappaB-independent manner. Furthermore, overexpression of the constitutively active MEK1 induced IL-8 and MIP3alpha protein production. We also demonstrated that C-terminal coiled-coil and TRAF-C domains of TRAF6, unable to mediate NFkappaB activation, are involved in MEK-mediated IL-8 and MIP3alpha expression. Thus, in non-transformed human colonocytes, MEK activation following flagellin/TLR5 engagement is a key modulator for NFkappaB-independent, IL-8 and MIP3alpha expression.

Local Anesthetics Induce Chondrocyte Death in Bovine Articular Cartilage Disks in a Dose- and Duration-Dependent Manner

IL-10 reduces apoptosis and extracellular matrix degradation after injurious compression of mature articular cartilage

ObjectivesStaphylococcus 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.MethodsBovine 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.ResultsHla-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.ConclusionThis 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.

Age-related differences in hepatic ischemia/reperfusion: gene activation, liver injury, and protective effect of melatonin

Prior joint injury predisposes an individual to developing post-traumatic osteoarthritis, for which there is presently no disease modifying treatment. In this condition, articular cartilage degenerates due to cell death and matrix breakdown, resulting in tissue with diminished biomechanical function. P188, a nonionic surfactant, and the growth factor IGF-I have been shown to decrease cell death. Additionally, IGF-I is known to have beneficial effects on cartilage matrix. The objective of this study was to determine the efficacy of P188, IGF-I, and their combination following articular cartilage impact injury with two energy levels, 1.1 J ("low") and 2.8 J ("high"), at 24 h and 1 week. Bovine articular cartilage with attached underlying bone was impacted at the low or high level. Impact sites were explanted and examined immediately, or cultured for 24 h or 1 week in serum-free media supplemented with P188 (8 mgml), IGF-I (100 ngml), or their combination. Gross morphology, cell viability, GAG release to the media, and tissue mechanical properties were assessed. Immediately postimpact, high level impacted tissue had significantly increased gross morphology scores, indicating tissue damage, which were maintained over 1 week. Gross scores following low impact were initially similar to nonimpacted controls, but, at 24 h and 1 week, low impact gross scores significantly increased compared to nonimpacted controls. Additionally, at 24 h, high impact resulted in increased cell death, and both low and high impacts had increased GAG release compared to nonimpacted controls. Furthermore, high impact caused decreased tissue stiffness at 24 h that appeared to worsen over 1 week, evident by the percent decrease from nonimpacted controls increasing from 16% to 26%. No treatment type studied mitigated this loss. The combination did not perform better than either individual treatment; however, following low impact at 1 week, P188 reduced cell death by 75% compared to no treatment and IGF-I decreased GAG release from the tissue by 49%. In conclusion, high impact resulted in immediate tissue changes that worsened over 1 week. Though not causing immediate changes, low impact also resulted in tissue degeneration evident by 24 h. No treatment studied was effective at 24 h, but by 1 week P188 and IGF-I ameliorated established detrimental changes occurring in articular cartilage postimpact. However, further work is needed to optimize treatment strategies to prevent and/or reverse cell death and matrix destruction in a way that maintains tissue mechanical properties, and hence its functionality.

Age-related differences in hepatic ischemia/reperfusion: Gene activation, liver injury, and protective effect of melatonin

BackgroundPrior research has highlighted the involvement of a transcriptional complex comprising C-terminal binding protein 2 (CtBP2), histone acetyltransferase p300, and nuclear factor kappa B (NF-κB) in the transactivation of proinflammatory cytokine genes, contributing to inflammation in mice with acute respiratory distress syndrome (ARDS). Nonetheless, it remains uncertain whether the therapeutic targeting of the CtBP2-p300-NF-κB complex holds potential for ARDS suppression.MethodsAn ARDS mouse model was established using lipopolysaccharide (LPS) exposure. RNA-Sequencing (RNA-Seq) was performed on ARDS mice and LPS-treated cells with CtBP2, p300, and p65 knockdown. Small molecules inhibiting the CtBP2-p300 interaction were identified through AlphaScreen. Gene and protein expression levels were quantified using RT-qPCR and immunoblots. Tissue damage was assessed via histological staining.Key findingsWe elucidated the specific role of the CtBP2-p300-NF-κB complex in proinflammatory gene regulation. RNA-seq analysis in LPS-challenged ARDS mice and LPS-treated CtBP2-knockdown (CtBP2KD), p300KD, and p65KD cells revealed its significant impact on proinflammatory genes with minimal effects on other NF-κB targets. Commercial inhibitors for CtBP2, p300, or NF-κB exhibited moderate cytotoxicity in vitro and in vivo, affecting both proinflammatory genes and other targets. We identified a potent inhibitor, PNSC928, for the CtBP2-p300 interaction using AlphaScreen. PNSC928 treatment hindered the assembly of the CtBP2-p300-NF-κB complex, substantially downregulating proinflammatory cytokine gene expression without observable cytotoxicity in normal cells. In vivo administration of PNSC928 significantly reduced CtBP2-driven proinflammatory gene expression in ARDS mice, alleviating inflammation and lung injury, ultimately improving ARDS prognosis.ConclusionOur results position PNSC928 as a promising therapeutic candidate to specifically target the CtBP2-p300 interaction and mitigate inflammation in ARDS management.

Introduction: Shoulder impingement syndrome is a common diagnosis leading to shoulder disabilities. Subacromial decompression surgery either arthroscopic or mini open is indicated in patients who failed conservative therapy. Arthroscopic surgery is introduced to reduce soft tissue dissection hence less soft tissue damage and fasten rehabilitation. This study is conducted to compare the outcome of mini open and arthroscopic surgery. Materials and Methods: A cross sectional study was conducted in patients who were diagnosed with shoulder impingement syndrome with or without rotator cuff tear who underwent subacromial decompression surgery, with mini open or arthroscopic surgery at Hospital Tengku Ampuan Afzan, Kuantan. Functional outcome was assessed using American Shoulder and Elbow Surgeon (ASES) score and Constant score. Results: Six patients were recruited in the mini open group while 11 in the arthroscopic group. Of these, all were female in mini open group, aged 56.7 ± 7.9 while seven female and four male in the arthroscopic group aged 55.1 ± 9.4 (p-value 0.733). They were evaluated at 3 to 5 years after surgery in the mini open, and at six months in the arthroscopic group. There was no significant different in the ASES score in the mini open group (89.8 ± 11.7) and arthroscopic group (89.8 ±17.9), with p-value of 0.998. Constant score in the mini open was 18.67 ± 7.61, while in the arthroscopic was 16.18 ± 15.03; and there was no significant difference in this two groups (p-value 0.712). Three patients each has good and fair outcome in the mini open surgery, while six patients has excellent, two each has good and fair; and one has poor outcome in the arthroscopic surgery. Conclusion: There were no significant differences in the functional outcomes between mini open and arthroscopic subacromial decompression surgery.

Recent studies have suggested that cartilage progenitor cells (CPCs) could be activated and differentiated into chondrocytes to produce matrix and to restore the integrity of damaged cartilage after injury. However, the mechanism involved in CPC activation upon damage is still unclear. This study aims to investigate the role of high mobility group box chromosomal protein 1 (HMGB1) in both activation and migration of CPCs during cartilage injury. Explants harvested from mature bovine stifle joints were used for impact injury. The proliferation and migration of CPCs were examined via confocal imaging. Gene and protein expression of Hmbg1, Cxcl12, and Cxcr4 was also examined by quantitative polymerase chain reaction (qPCR), ELISA, and western blot. Each experiment was repeated 3 times. ANOVA and Student's t-test were performed for statistical analysis. HMGB1 released from dead and damaged chondrocytes after an impact injury could activate CPCs in the superficial zone of cartilage and promote their migration and proliferation to injury sites. However, the block of HMGB1 activation with its specific binding inhibitor glycyrrhizin inhibits the proliferation and migration of CPCs. Further investigations demonstrate that HMGB1 promotes CPCs migration through the pathway of C-X-C motif chemokine 12 (CXCL12) and its receptor CXCR4. Quantitative analysis of HMGB1 in cell culture medium also indicates that CPCs may have a self-activation property after the HMGB1 released from dead cells has been exhausted. HMGB1 is a pivotal factor that could enhance the migration and proliferation of CPCs through the CXCL12/CXCR4 pathway after cartilage injury, which could provide useful information for cartilage repair and osteoarthritis treatment.