Biotribological Tests of Osteochondral Grafts after Treatment with Pro-Inflammatory Cytokines.

OBJECTIVE: During osteoarthritis progression, cartilage degrades in a manner that influences its biomechanical and biotribological properties, while chondrocytes reduce the synthesis of extracellular matrix components and become apoptotic. This study investigates the effects of inflammation on cartilage under biomechanical stress using biotribological tests. METHODS: Bovine osteochondral grafts from five animals were punched out from the medial condyle and treated with or without pro-inflammatory cytokines (interleukin-1ß [IL-1ß], tumor necrosis factor-α [TNF-α], IL-6) for 2 weeks. After incubation, biotribological tests were performed for 2 hours (alternating 10 minutes test and pause respectively at 39°C, 180 N, 1 Hz, and 2 mm stroke). Before and after testing, the cartilage surface was imaged with a 3-dimensional microscope. During testing, the coefficient of friction (COF) was measured, while gene expression analysis and investigation of metabolic activity of chondrocytes were carried out after testing. Histological sections of the tissue and wear debris from the test fluid were also analyzed. RESULTS: After biotribological tests, surface cracks were found in both treated and untreated osteochondral grafts. In treated grafts, the COF increased, and the proteoglycan content in the cartilage tissue decreased, leading to structural changes. Chondrocytes from treated grafts showed increased expression of genes encoding for degradative enzymes, while cartilage-specific gene expression and metabolic activity exhibited no significant differences between treated and untreated groups. No measurable difference in the wear debris in the test fluid was found. CONCLUSIONS: Treatment of osteochondral grafts with cytokines results in a significantly increased COF, while also leading to significant changes in cartilage proteoglycan content and cartilage matrix compression during biotribological tests.

Concentration-Dependent Effects of Cobalt and Chromium Ions on Osteoarthritic Chondrocytes.

OBJECTIVE: Cobalt and chromium (CoCr) ions from metal implants are released into the joint due to biotribocorrosion, inducing apoptosis and altering gene expression in various cell types. Here, we asked whether CoCr ions concentration-dependently changed viability, transcriptional activity, and inflammatory response in human articular chondrocytes. DESIGN: Human articular chondrocytes were exposed to Co (1.02-16.33 ppm) and Cr (0.42-6.66 ppm) ions and cell viability and early/late apoptosis (annexin V and 7-AAD) were assessed in 2-dimensional cell cultures using the XTT assay and flow cytometry, respectively. Changes in chondrocyte morphology were assessed using transmitted light microscopy. The effects of CoCr ions on transcriptional activity of chondrocytes were evaluated by quantitative polymerase chain reaction (qPCR). The inflammatory responses were determined by measuring the levels of released pro-inflammatory cytokines (interleukin-1ß [IL-1ß], IL-6, IL-8, and tumor necrosis factor-α [TNF-α]). RESULTS: CoCr ions concentration-dependently reduced metabolic activity and induced early and late apoptosis after 24 hours in culture. After 72 hours, the majority of chondrocytes (>90%) were apoptotic at the highest concentrations of CoCr ions (16.33/6/66 ppm). SOX9 expression was concentration-dependently enhanced, whereas expression of COL2A1 linearly decreased after 24 hours. IL-8 release was enhanced proportionally to CoCr ions levels, whereas IL-1ß, IL-6, and TNF-α levels were not affected by the treatments. CONCLUSIONS: CoCr ions showed concentration- and time-dependent effects on articular chondrocytes. Fractions of apoptotic articular chondrocytes were proportional to CoCr ion concentrations. In addition, metabolic activity and expression of chondrocyte-specific genes were decreased by CoCr ions. Furthermore, exposure to CoCr ions caused a release of pro-inflammatory cytokines.

Biotribological Testing and Analysis of Articular Cartilage Sliding against Metal for Implants.

Osteochondral defects in middle-aged patients might be treated with focal metallic implants. First developed for defects in the knee joint, implants are now available for the shoulder, hip, ankle and the first metatarsalphalangeal joint. While providing pain reduction and clinical improvement, progressive degenerative changes of the opposing cartilage are observed in many patients. The mechanisms leading to this damage are not fully understood. This protocol describes a tribological experiment to simulate a metal-on-cartilage pairing and comprehensive analysis of the articular cartilage. Metal implant material is tested against bovine osteochondral cylinders as a model for human articular cartilage. By applying different loads and sliding speeds, physiological loading conditions can be imitated. To provide a comprehensive analysis of the effects on the articular cartilage, histology, metabolic activity and gene expression analysis are described in this protocol. The main advantage of tribological testing is that loading parameters can be adjusted freely to simulate in vivo conditions. Furthermore, different testing solutions might be used to investigate the influence of lubrication or pro-inflammatory agents. By using gene expression analysis for cartilage-specific genes and catabolic genes, early changes in the metabolism of articular chondrocytes in response to mechanical loading might be detected.

Effects of Loading Conditions on Articular Cartilage in a Metal-on-Cartilage Pairing.

The aim of this in vitro study was to investigate the response of articular cartilage to frictional load when sliding against a metal implant, and identify potential mechanisms of damage to articular cartilage in a metal-on-cartilage pairing. Bovine osteochondral cylinders were reciprocally slid against metal cylinders (cobalt-chromium-molybdenum alloy) with several variations of load and sliding velocity using a microtribometer. The effects of different loads and velocities, and the resulting friction coefficients on articular cartilage, were evaluated by measuring histological and metabolic outcomes. Moreover, the biotribocorrosion of the metal was determined. Chondrocytes stimulated with high load and velocity showed increased metabolic activity and cartilage-specific gene expression. In addition, higher load and velocity resulted in biotribocorrosion of the metal implant and damage to the surface of the articular cartilage, whereas low velocity and a high coefficient of friction increased the expression of catabolic genes. Articular cartilage showed particular responses to load and velocity when sliding against a metal implant. Moreover, metal implants showed tribocorrosion. Therefore, corrosion particles may play a role in the mechano-biochemical wear of articular cartilage after implantation of a metal implant. These findings may be useful to surgeons performing resurfacing procedures and total knee arthroplasty. © 2019 The Authors. Journal of Orthopaedic Research® published by Wiley Periodicals, Inc. on behalf of Orthopaedic Research Society J Orthop Res 37:2531-2539, 2019.

Effect of osteochondral graft orientation in a biotribological test system.

Autologous osteochondral transplantation (AOT) utilizing autografts is a widely used technique for the treatment of small-to-medium cartilage defects occurring in knee and ankle joints. The application of viable cartilage and bone ensures proper integration, early weight bearing, as well as restoration of biomechanical and biotribological properties. However, alignment of the autografts onto the defect site remains a pivotal aspect of reinstating the properties of the joint toward successful autograft integration. This is the first study to perform tests with different orientations of osteochondral grafts in a cartilage-on-cartilage test system. The objective was to estimate if there are differences between aligned and 90°-rotated grafts concerning molecular biological and biomechanical parameters. Tissue viability, assessed by XTT assay indicated lower metabolic activity in tested osteochondral grafts (aligned, p = 0.0148 and 90°-rotated, p = 0.0760) in favor of a higher anabolic gene expression (aligned, p = 0.0030 and 90°-rotated, 0.0027). Tissue structure was evaluated by Safranin O histology and microscopic images of the surface. Aligned and 90°-rotated grafts revealed no apparent differences between proteoglycan content or cracks and fissures on the cartilage surface. Test medium analyzed after tribological tests for their sulfated glycosaminoglycan content revealed no differences (p = 0.3282). During the tests, both the friction coefficient and the relative displacement between the two cartilage surfaces were measured, with no significant difference in both parameters (COF, p = 0.2232 and relative displacement, p = 0.3185). From the methods we deployed, this study can infer that there are no differences between aligned and 90°-rotated osteochondral grafts after tribological tests in the used ex vivo tissue model. © 2019 The Authors. Journal of Orthopaedic Research® Published by Wiley Periodicals, Inc. on behalf of Orthopaedic Research Society. J Orthop Res.