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Plant-derived cellulose biomaterials have recently been utilized in several tissue engineering applications. These naturally-derived cellulose scaffolds have been shown to be highly biocompatible in vivo, possess structural features of relevance to several tissues, and support mammalian cell invasion and proliferation. Recent work utilizing decellularized apple hypanthium tissue has shown that it possesses a pore size similar to trabecular bone and can successfully host osteogenic differentiation. In the present study, we further examined the potential of apple-derived cellulose scaffolds for bone tissue engineering (BTE) and analyzed their mechanical properties in vitro and in vivo. MC3T3-E1 pre-osteoblasts were seeded in cellulose scaffolds. Following chemically-induced osteogenic differentiation, scaffolds were evaluated for mineralization and for their mechanical properties. Alkaline phosphatase and Alizarin Red staining confirmed the osteogenic potential of the scaffolds. Histological analysis of the constructs revealed cell invasion and mineralization throughout the constructs. Furthermore, scanning electron microscopy demonstrated the presence of mineral aggregates on the scaffolds after culture in differentiation medium, and energy-dispersive spectroscopy confirmed the presence of phosphate and calcium. However, although the Young’s modulus significantly increased after cell differentiation, it remained lower than that of healthy bone tissue. Interestingly, mechanical assessment of acellular scaffolds implanted in rat calvaria defects for 8 weeks revealed that the force required to push out the scaffolds from the surrounding bone was similar to that of native calvarial bone. In addition, cell infiltration and extracellular matrix deposition were visible within the implanted scaffolds. Overall, our results confirm that plant-derived cellulose is a promising candidate for BTE applications. However, the discrepancy in mechanical properties between the mineralized scaffolds and healthy bone tissue may limit their use to low load-bearing applications. Further structural re-engineering and optimization to improve the mechanical properties may be required for load-bearing applications.

There is an urgent need in the field of soft tissue engineering (STE) to develop biomaterials exhibiting a high degree of biological and mechanical functionality as well as modularity so that they can be tailored according to patient-specific requirements. Recently, biomimetic soft network composites (SNC) consisting of a water-swollen hydrogel matrix and a reinforcing fibrous network fabricated by melt electrospinning writing technology have demonstrated exceptional mechanical and biological properties, thus becoming strong candidates for STE applications. However, there is a lack of design approaches to tailor and optimize their properties in a non-empirical way. To address this challenge, we propose a numerical model-based approach for the rational design of patient-specific SNC for tissue engineering applications. The approach is rooted in an in silico design library that allows for the selection of biomaterial and architecture combinations for the target application, resulting in reduced time, manpower and costs. To demonstrate the validity of the design strategy, a multiphasic SNC with predefined zone-specific properties that captured the complex zonal mechanical and compositional features of articular cartilage was developed based on the natural design of the native tissue.

Adverse tissue reactions to metal implants, including pseudotumors, can compromise implant functionality and survivorship. The identification of specific proteins in the synovial fluid (SF) of hip arthroplasty patients with a pseudotumor may lead to a better understanding of the underlying pathomechanisms. The objective of the present study was to compare the protein content of SF from patients with a short-term metal-on-metal hip implant associated with a pseudotumor and patients with a long-term metal-on-polyethylene hip implant associated with periprosthetic osteolysis. Discovery proteomics was used to identify differentially abundant proteins in albumin-depleted SF. In toto, 452 distinct proteins (present in at least half of the patients in one or both groups) were identified. Thirty of these 452 proteins were differentially abundant between the two groups, including two potential biomarkers: 6-phosphogluconate dehydrogenase (which plays a major protective role against oxidative stress) for the pseudotumor group, and scavenger receptor cysteine-rich type 1 protein M130 (which is involved in low-grade inflammation) for the periprosthetic osteolysis group. Other differentially abundant proteins identified suggest the presence of an adaptive immune response (particularly a type-IV hypersensitivity reaction), necrosis, and greater oxidative stress in patients with a pseudotumor. They also suggest the presence of an innate immune response, oxidative stress, tissue remodeling, and apoptosis in both patient groups, although differences in the specific proteins identified in each group point to differences in the pathomechanisms. Overall, results provide insights into the molecular mechanisms underlying metal-related pseudotumors and periprosthetic osteolysis, and may ultimately help elucidate pseudotumor etiology and assess the risk that asymptomatic pseudotumors will develop into an aggressive lesion. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:1849-1859, 2018.

Plant-derived cellulose biomaterials have recently been utilized in several tissue engineering applications. Naturally-derived cellulose scaffolds have been shown to be highly biocompatible in vivo, possess structural features of relevance to several tissues, as well as support mammalian cell invasion and proliferation. Recent work utilizing decellularized apple hypanthium tissue has shown that it possesses a pore size and properties similar to trabecular bone. In the present study, we examined the potential of apple-derived cellulose scaffolds for bone tissue engineering (BTE). Confocal microscopy revealed that the scaffolds had a suitable pore size for BTE applications. To analyze their in vitro mineralization potential, MC3T3-E1 pre-osteoblasts were seeded in either bare cellulose scaffolds or in composite scaffolds composed of cellulose and collagen I. Following chemically-induced differentiation, scaffolds were mechanically tested and evaluated for mineralization. The Young’s modulus of both types of scaffolds significantly increased after cell differentiation. Alkaline phosphatase and Alizarin Red staining further highlighted the osteogenic potential of the scaffolds. Histological sectioning of the constructs revealed complete invasion by the cells and mineralization throughout the entire constructs. Finally, scanning electron microscopy demonstrated the presence of mineral aggregates deposited on the scaffolds after differentiation, and energy-dispersive spectroscopy confirmed the presence of phosphate and calcium. In summary, our results indicate that plant-derived cellulose is a promising scaffold candidate for bone tissue engineering applications.

Implant wear and corrosion have been associated with adverse tissue reactions that can lead to implant failure. Wear and corrosion products are therefore of great clinical concern. For example, Co2+ and Cr3+ originating from CoCrMo-based implants have been shown to induce a proinflammatory response in macrophages in vitro. Previous studies have also shown that the polarization of macrophages by some proinflammatory stimuli is associated with a hypoxia-inducible factor-1α (HIF-1α)-dependent metabolic shift from oxidative phosphorylation (OXPHOS) towards glycolysis. However, the potential of Co2+ and Cr3+ to induce this metabolic shift, which plays a determining role in the proinflammatory response of macrophages, remains largely unexplored. We recently demonstrated that Co2+ , but not Cr3+ , increased oxidative stress and decreased OXPHOS in RAW 264.7 murine macrophages. In the present study, we analyzed the effects of Co2+ and Cr3+ on glycolytic flux and HIF-1α stabilization in the same experimental model. Cells were exposed to 6 to 24 ppm Co2+ or 50 to 250 ppm Cr3+ . Glycolytic flux was determined by analyzing extracellular flux and lactate production, while HIF-1α stabilization was analyzed by immunoblotting. Results showed that Co2+ , and to a lesser extent Cr3+ , increased glycolytic flux; however, only Co2+ acted through HIF-1α stabilization. Overall, these results, together with our previous results showing that Co2+ increases oxidative stress and decreases OXPHOS, suggest that Co2+ (but not Cr3+ ) can induce a HIF-1α-dependent metabolic shift from OXPHOS towards glycolysis in macrophages. This metabolic shift may play an early and pivotal role in the inflammatory response induced by Co2+ in the periprosthetic environment.

There is an urgent clinical need to develop accurate in vitro test procedures to better understand, predict and prevent in vivo implant failures. This study aimed to simulate in vivo fatigue fracture and distraction of modular necks in an in vitro setting, and to assess the effects of neck material (Ti6Al4V vs. CoCrMo) and assembly method (hand vs. impact) on implant failure. Fatigue tests were performed on the PROFEMUR® Modular Neck System under two different loads and number of cycles: 2.3 kN for 5 million cycles, and 7.0 kN for 1.3 million cycles. The developed in vitro simulation setup successfully reproduced in vivo modular neck fracture mode and location. Neck failure occurred at the neck-stem taper and the fracture ran from the distal lateral neck surface to the proximal medial entry point of the neck into the stem. None of the necks failed under the 2.3 kN load. However, all hand-assembled Ti6Al4V necks failed under the 7.0 kN load. In contrast, none of the hand-assembled CoCrMo necks and impact-assembled necks (Ti6Al4V or CoCrMo) failed under this higher load. In conclusion, Ti6Al4V necks were more susceptible to fatigue failure than CoCrMo necks. In addition, impact assembly substantially improved the fatigue life of Ti6Al4V necks and also led to overall higher distraction forces for both neck materials. Overall, this study shows that the material and assembly method can affect the fatigue strength of modular necks. Finally, improper implant assembly during surgery may result in diminished modular neck survivability and increased failure rates. This article is protected by copyright. All rights reserved

Developing multifunctional soft biomaterials capable of addressing all the requirements of the complex tissue regeneration process is a multifaceted problem. In order to tackle the current challenges, recent research efforts are increasingly being directed towards biomimetic design concepts that can be translated into soft biomaterials via advanced manufacturing technologies. Among those, soft network composites consisting of a continuous hydrogel matrix and a reinforcing fibrous network closely resemble native soft biological materials in terms of design and composition as well as physicochemical properties. This article reviews soft network composite systems with a particular emphasis on the design, biomaterial and fabrication aspects within the context of soft tissue engineering and drug delivery applications.

Cobalt and chromium ions released from cobalt-chromium-molybdenum (CoCrMo)-based implants are a potential health concern, especially since both ions have been shown to induce oxidative stress in macrophages, the predominant immune cells in periprosthetic tissues. Ions of other transition metals (Cd, Ni) have been reported to inhibit the activity of mitochondrial enzymes in the electron transport chain. However, the effects of Co and Cr ions on the energy metabolism of macrophages remain largely unknown. The objective of the present study was to analyze the effects of Co

Articular cartilage from a material science point of view is a soft network composite that plays a critical role in load-bearing joints during dynamic loading. Its composite structure, consisting of a collagen fiber network and a hydrated proteoglycan matrix, gives rise to the complex mechanical properties of the tissue including viscoelasticity and stress relaxation. Melt electrospinning writing allows the design and fabrication of medical grade polycaprolactone (mPCL) fibrous networks for the reinforcement of soft hydrogel matrices for cartilage tissue engineering. However, these fiber-reinforced constructs underperformed under dynamic and prolonged loading conditions, suggesting that more targeted design approaches and material selection are required to fully exploit the potential of fibers as reinforcing agents for cartilage tissue engineering. In the present study, we emulated the proteoglycan matrix of articular cartilage by using highly negatively charged star-shaped poly(ethylene glycol)/heparin hydrogel (sPEG/Hep) as the soft matrix. These soft hydrogels combined with mPCL melt electrospun fibrous networks exhibited mechanical anisotropy, nonlinearity, viscoelasticity and morphology analogous to those of their native counterpart, and provided a suitable microenvironment for in vitro human chondrocyte culture and neocartilage formation. In addition, a numerical model using the p-version of the finite element method (p-FEM) was developed in order to gain further insights into the deformation mechanisms of the constructs in silico, as well as to predict compressive moduli. To our knowledge, this is the first study presenting cartilage tissue-engineered constructs that capture the overall transient, equilibrium and dynamic biomechanical properties of human articular cartilage.

Adverse tissue reactions to metal implants, including pseudotumors, can compromise implant functionality and survivorship. The identification of specific proteins in the synovial fluid (SF) of hip arthroplasty patients with a pseudotumor may lead to a better understanding of the underlying pathomechanisms. The objective of the present study was to compare the protein content of SF from patients with a short-term metal-on-metal hip implant associated with a pseudotumor and patients with a long-term metal-on-polyethylene hip implant associated with periprosthetic osteolysis. Discovery proteomics was used to identify differentially abundant proteins in albumin-depleted SF. In toto, 452 distinct proteins (present in at least half of the patients in one or both groups) were identified. Thirty of these 452 proteins were differentially abundant between the two groups, including two potential biomarkers: 6-phosphogluconate dehydrogenase (which plays a major protective role against oxidative stress) for the pseudotumor group, and scavenger receptor cysteine-rich type 1 protein M130 (which is involved in low-grade inflammation) for the periprosthetic osteolysis group. Other differentially abundant proteins identified suggest the presence of an adaptive immune response (particularly a type-IV hypersensitivity reaction), necrosis, and greater oxidative stress in patients with a pseudotumor. They also suggest the presence of an innate immune response, oxidative stress, tissue remodeling, and apoptosis in both patient groups, although differences in the specific proteins identified in each group point to differences in the pathomechanisms. Overall, results provide insights into the molecular mechanisms underlying metal-related pseudotumors and periprosthetic osteolysis, and may ultimately help elucidate pseudotumor etiology and assess the risk that asymptomatic pseudotumors will develop into an aggressive lesion. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:1849-1859, 2018.

An increasing number of studies have reported adverse tissue reactions around metal-on-metal (MM) hip implants. However, the origin and mechanisms of these reactions remain unclear. Moreover, the biological effects of nanometer-size chromium oxide particles, the predominant type of wear particles produced by MM implants, remain mostly unknown. The purpose of this study was to analyze the cytotoxic effects of clinically relevant nanometer-size chromium oxide particles on macrophage response in vitro. J774.A1 macrophages were cultured with either 60 nm or 700 nm commercially available Cr2 O3 particles at different concentrations. Two different particle sizes were analyzed to evaluate potential volume effects. Cell mortality was analyzed by light microscopy, flow cytometry (annexin V-fluorescein isothiocyanate and propidium iodide assay), and using a cell death detection enzyme-linked immunosorbant assay (ELISA). Tumor necrosis factor alpha (TNF-α), monocyte chemotactic protein-1 (MCP-1), and macrophage inflammatory protein-1 alpha (MIP-1α) release was measured by ELISA, and gene expression was analyzed by quantitative real-time PCR. Results showed that, at high concentrations, Cr2 O3 particles of both sizes can be cytotoxic, inducing significant decreases in total cell numbers and increases in necrosis. Results also suggested that these effects were dependent on particle volume. However, TNF-α, MCP-1, and MIP-1α cytokine release and gene expression remained low. Overall, this study demonstrates that nanometer-size particles of Cr2 O3 , a stable form of chromium oxide ceramic, have rather low cytotoxic effects on macrophages. Therefore, these particles may not be the main culprit in the initiation of the inflammatory reaction in MM periprosthetic tissues. However, other parameters (e.g., potential intracellular damage) remain to be investigated.

Fibrin, derived from key proteins involved in blood clotting (fibrinogen and thrombin), is a self-assembling biopolymer. The range of clinical applications and types of procedure that incorporate fibrin sealants is expanding. While fibrin sealants were initially, and continue to be, used for their hemostasis and tissue sealing properties, they have also been used more recently as a successful platform in various tissue regeneration applications necessitating cell and/or growth factor delivery. One of the primary reasons for this success is that fibrin provides a biocompatible physical structure and many of the important biological properties needed for successful tissue regeneration. In addition, these inherent biological properties can be enhanced by combining fibrin with other biomaterials (hence improving the overall biological and/or mechanical properties of the composite matrix) or by incorporating bioactive molecules to control cell proliferation, migration, or differentiation during tissue regeneration. This chapter summarizes key aspects of the composition, structure, as well as mechanical, fibrinolytic, and biological properties of fibrin as these characteristics influence its efficacy and performance. Applications of fibrin and fibrin-based biomaterials in tissue engineering and in clinical practice are also highlighted, with an emphasis on the use of fibrin as a delivery system for cells and bioactive molecules.

Hip implant neck fractures and adverse tissue reactions associated with fretting-corrosion damage at modular interfaces are a major source of concern. Therefore, there is an urgent clinical need to develop accurate in vitro test procedures to better understand, predict and prevent in vivo implant failures. This study aimed to simulate in vivo fatigue fracture and distraction of modular necks in an in vitro setting, and to assess the effects of neck material (Ti6Al4V vs. CoCrMo) and assembly method (hand vs. impact) on the fatigue life and distraction of the necks. Fatigue tests were performed on the cementless PROFEMUR® Total Hip Modular Neck System under two different loads and number of cycles: 2.3 kN for 5 million cycles, and 7.0 kN for 1.3 million cycles. The developed in vitro simulation setup successfully reproduced in vivo modular neck fracture mode and location. Neck failure occurred at the neck-stem taper and the fracture ran from the distal lateral neck surface to the proximal medial entry point of the neck into the stem. None of the necks failed under the 2.3 kN load. However, all hand-assembled Ti6Al4V necks failed under the 7.0 kN load. In contrast, none of the hand-assembled CoCrMo necks and impact-assembled necks (Ti6Al4V or CoCrMo) failed under this higher load. In conclusion, Ti6Al4V necks were more susceptible to fatigue failure than CoCrMo necks. In addition, impact assembly substantially improved the fatigue life of Ti6Al4V necks and also led to overall higher distraction forces for both neck materials. Overall, this study shows that the material and assembly method can affect the fatigue strength of modular necks. Finally, improper implant assembly during surgery may result in diminished modular neck survivability and increased failure rates. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:2023-2030, 2017.

Osteoarthritis and lesions to cartilage tissue are diseases that frequently result in impaired joint function and patient disability. The treatment of osteoarthritis, along with local bone defects and systemic skeletal diseases, remains a significant clinical challenge for orthopaedic surgeons. Several bone morphogenetic proteins (BMPs) are known to have osteoinductive effects, whereof BMP-2 and BMP-7 are already approved for clinical applications. There is growing evidence that the metabolism of bone as well as the cartilage damage associated with the above disease processes are strongly inter-related with the interactions of the inflammation-related pathways (in particular prostaglandin E2(PGE2)) and osteogenesis (in particular bone morphogenetic protein-2 (BMP-2)). There is strong evidence that the pathways of prostaglandins and bone morphogenetic proteins are intertwined, and they have recently come into focus in several experimental and clinical studies. This paper focuses on PGE2and BMP-2 intertwining pathways in bone and cartilage metabolism, and summarizes the recent experimental and clinical data.

Biological effects of wear products (particles and metal ions) generated by metal-on-metal (MoM) hip replacements made of CoCrMo alloy remain a major cause of concern. Periprosthetic osteolysis, potential hypersensitivity response and pseudotumour formation are possible reactions that can lead to early revisions. To accurately analyse the biological response to wear particles from MoM implants, the exact nature of these particles needs to be characterized. Most previous studies used energy-dispersive X-ray spectroscopy (EDS) analysis for characterization. The present study used energy filtered transmission electron microscopy (TEM) and electron diffraction pattern analysis to allow for a more precise determination of the chemical composition and to gain knowledge of the crystalline structure of the wear particles.Particles were retrieved from two different test rigs: a reciprocating sliding wear tribometer (CoCrMo cylinder vs. bar) and a hip simulator according to ISO 14242-1 (CoCrMo head vs. CoCrMo cup). All tests were conducted in bovine serum. Particles were retrieved from the test medium using a previously published enzymatic digestion protocol.Particles isolated from tribometer samples had a size of 100 - 500 nm. Diffraction pattern analysis clearly revealed the lattice structure of strain induced hcp ε-martensite. Hip simulator samples revealed numerous particles of 15 - 30 nm and 30 - 80 nm size. Most of the larger particles appeared to be only partially oxidized and exhibited cobalt locally. The smallest particles were Cr(2)O(3) with no trace of cobalt. It optically appeared that these Cr(2)O(3) particles were flaking off the surface of larger particles that depicted a very high intensity of oxygen, as well as chromium, and only background noise of cobalt. The particle size difference between the two test rigs is likely related to the conditions of the two tribosystems, in particular the difference in the sample geometry and in the type of sliding (reciprocating vs. multidirectional).Results suggest that there may be a critical particle size at which chromium oxidation and cobalt ionization is accelerated. Since earlier studies have shown that wear particles are covered by organic residue which may act as a passive layer inhibiting further oxidation, it would suggest that this organic layer may be removed during the particle isolation process, resulting in a change of the particle chemical composition due to their pyrophoric properties. However, prior to being isolated from the serum lubricant, particles remain within the contact area of head and cup as a third-body. It is therefore possible that during that time, particles may undergo significant transformation and changes in chemical composition in the contact area of the head and cup within the tribological interface due to mechanical interaction with surface asperities.

This article was updated on May 13, 2011, because of previous editorial errors. On page 76, the affiliation that had previously read “Departments of Mechanical Engineering Surgery, and Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, Ontario, Canada” now reads “Departments of Mechanical Engineering, Surgery & Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, Ontario, Canada.” On page 79, the line that had previously read “The percentage of these needle-shaped particles was the highest with hip replacements implanted for more than twenty years[38][1],” now reads “The percentage of these needle-shaped particles was the highest with hip replacements implanted for more than twenty years (average [and standard deviation] of 28% ± 9%)[38][1].” On page 80, the previous incorrect version of Figure 3 has been replaced with the correct version. On page 82, the last line in reference 15, which had previously read “Paper no. 214,” now reads “Paper no. 365.” An erratum has been published: J Bone Joint Surg Am 2011;93(12):1158. Background: Despite the renewed interest in metal-on-metal implants in the past two decades, the underlying wear mechanisms and biological effects are still not fully understood. Methods: This paper first reviews the tribology of metal-on-metal bearings, bringing new insights into the interaction of wear and corrosion, and putting the characteristics and the potential origin of wear particles in perspective with the proposed wear mechanisms. It then summarizes the current knowledge on the biological effects of particles and metal ions in relation to these wear mechanisms. Results: Tribochemical reactions play an important role in the wear of metal-on-metal joints. The generated tribomaterial, which progressively forms by mechanical mixing of the uppermost nanocrystalline zone of the metal surface with proteins from the synovial fluid, governs the wear rate and influences the corrosive behavior of the bearing. Nanometer-sized wear particles may initially originate from the passivation layer covering the implant surface and then detach from this tribolayer. The inflammatory response observed surrounding metal-on-metal implants appears to be lower than that around metal-on-polyethylene implants. However, metallic byproducts, which can complex with proteins, may lead to a T lymphocyte-mediated hypersensitivity response. Conclusions: The tribolayer appears to have beneficial effects on the wear rate. Much information has been gained on wear particle characteristics, but the exact mechanisms of particle detachment remain to be further elucidated. Excessive wear along with a hypersensitivity response may be at the origin of the early adverse tissue reactions that have been recently reported in some patients with metal-on-metal implants. Clinical Relevance: Future development of new methods to improve the tribolayer retention and optimize the tribocorrosive properties of the implant may minimize the clinical impact of implant wear and immune responses. [1]: #ref-38

Der Verschleis von Gelenkimplantaten unterliegt multifaktoriellen Einflussgrosen, die in Ihrem Zusammenspiel auch bei nominell identischen Gleitpaarungen hochst unterschiedliche Partikelmengen und -formen generieren konnen. Die biologische Reaktion und damit letztendlich der klinische Erfolg des Implantats wird dabei von den speziellen Partikeleigenschaften beeinflusst. Eine dauerhafte Implantatversorgung bedingt einen optimierten Kompromiss zwischen Material, Design, Operationsmethode und patientenspezifischen Faktoren.

This study analyzed the ability of fibrin gels to deliver added recombinant transforming growth factor beta-1 (TGF-beta1) in a controlled manner and biologically active form. First, the effects of the amount of TGF-beta1 on the release kinetics were analyzed using a single fibrin gel formulation (fibrinogen complex (FC) at 25 mg/mL, thrombin at 2 IU/mL). Then, the effects of FC and thrombin concentrations were analyzed. Finally, to test the biological activity of the released TGF-beta1 from the gels, medium supernatants taken from gels at day 3 were used as culture medium for human mesenchymal stem cell (HMSC) monolayers. Cell proliferation was analyzed after staining with calcein dye, and changes in cell morphology were observed under fluorescence microscopy at days 1, 4, and 7. At day 7, HMSC chondrogenic differentiation was assessed by Alcian Blue staining and osteogenic differentiation by alkaline phosphatase activity and Alizarin Red staining. Results showed that TGF-beta1 added to fibrin gels was gradually released from the gels and increased with the amount of TGF-beta1 initially seeded, with a total of approximately 50% of the initial amount released by day 10 (with gels containing 25 mg/mL of FC and 2 IU/mL of thrombin). The release was lower with increasing FC concentrations, suggesting a binding affinity of TGF-beta1 with the FC component. Varying the thrombin concentration had a lesser effect. HMSC monolayers cultured with medium supernatants collected from gels at day 3 and containing released TGF-beta1 showed a change in morphology (squared to polygonal), lower cell proliferation, positive Alcian Blue staining but low levels of osteogenic differentiation markers. These results demonstrated that released TGF-beta1 was still bioactive and tended to induce mainly chondrogenic differentiation of the HMSC. Overall, the present study demonstrated that fibrin gels could be used as a carrier matrix for controlled release of bioactive TGF-beta1 by adjusting the concentrations of FC and thrombin in the gels.

A T cell-mediated hypersensitivity reaction has been reported in some patients with CoCrMo-based implants. However, the role of cobalt and chromium ions in this reaction remains unclear. The objective of the present study was to analyze the effects of Co

Early adverse tissue reactions around metal-on-metal (MoM) hip replacements, especially pseudotumors, are a major concern. Because the causes and pathomechanisms of these pseudotumors remain largely unknown, clinical monitoring of patients with MoM bearings is challenging.The purpose of this study was to compare the lymphocyte subpopulations in peripheral blood from patients with a failed MoM hip implant with and without a pseudotumor and patients with a well-functioning MoM hip implant without a pseudotumor. Potential differences in the systemic immune response are expected to reflect local differences in the periprosthetic tissues.Consenting patients who underwent a revision of a failed MoM hip implant at The Ottawa Hospital (TOH) from 2011 to 2014, or presented with a well-functioning MoM hip implant for a postoperative clinical followup at TOH from 2012 to 2013, were recruited for this study, unless they met any of the exclusion criteria (including diagnosed conditions that can affect peripheral blood lymphocyte subpopulations). Patients with a failed implant were divided into two groups: those with a pseudotumor (two hip resurfacings and five total hip arthroplasties [THAs]) and those without a pseudotumor (10 hip resurfacings and two THAs). Patients with a well-functioning MoM hip implant (nine resurfacings and three THAs) at 5 or more years postimplantation and who did not have a pseudotumor as demonstrated sonographically served as the control group. Peripheral blood subpopulations of T cells (specifically T helper [Th] and cytotoxic T [Tc]), B cells, natural killer (NK) cells, memory T and B cells as well as type 1 (expressing interferon-γ) and type 2 (expressing interleukin-4) Th and Tc cells were analyzed by flow cytometry after immunostaining. Serum concentrations of cobalt and chromium were measured by inductively coupled plasma-mass spectrometry.The mean percentages of total memory T cells and, specifically, memory Th and memory Tc cells were lower in patients with a failed MoM hip implant with a pseudotumor than in both patients with a failed implant without a pseudotumor and patients with a well-functioning implant without a pseudotumor (memory Th cells: 29% ± 5% [means ± SD] versus 55% ± 17%, d = 1.8, 95% confidence interval [CI] [1.2, 2.5] and versus 48% ± 14%, d = 1.6, 95% CI [1.0, 2.2], respectively; memory Tc cells: 18% ± 5% versus 45% ± 14%, d = 2.3, 95% CI [1.5, 3.1] and versus 41% ± 12%, d = 2.3, 95% CI [1.5, 3.1], respectively; p0.001 in all cases). The mean percentage of memory B cells was also lower in patients with a failed MoM hip implant with a pseudotumor than in patients with a well-functioning implant without a pseudotumor (12% ± 8% versus 29% ± 16%, d = 1.3, 95% CI [0.7, 1.8], p = 0.025). In addition, patients with a failed MoM hip implant with a pseudotumor had overall lower percentages of type 1 Th cells than both patients with a failed implant without a pseudotumor and patients with a well-functioning implant without a pseudotumor (5.5% [4.9%-5.8%] [median with interquartile range] versus 8.7% [6.5%-10.2%], d = 1.4, 95% CI [0.8, 2.0] and versus 9.6% [6.4%-11.1%], d = 1.6, 95% CI [1.0, 2.2], respectively; p ≤ 0.010 in both cases). Finally, serum cobalt concentrations in patients with a failed MoM hip implant with a pseudotumor were overall higher than those in patients with a well-functioning implant without a pseudotumor (5.8 µg/L [2.9-17.0 µg/L] versus 0.9 µg/L [0.6-1.3 µg/L], d = 2.2, 95% CI [1.4, 2.9], p0.001).Overall, results suggest the presence of a type IV hypersensitivity reaction, with a predominance of type 1 Th cells, in patients with a failed MoM hip implant with a pseudotumor.The lower percentages of memory T cells (specifically Th and Tc) as well as type 1 Th cells in peripheral blood of patients with a failed MoM hip implant with a pseudotumor could potentially become diagnostic biomarkers for the detection of pseudotumors. Although implant design (hip resurfacing or THA) did not seem to affect the results, as suggested by the scatter of the data with respect to this parameter, future studies with additional patients could include the analysis of implant design in addition to correlations with histological analyses of specific Th subsets in periprosthetic tissues.

Detailed characterization of wear particles is necessary to understand better the implant wear mechanisms and the periprosthetic tissue response. The purposes of the present study were to compare particle characteristics of current with older designs of metal-on-metal (MM) total hip replacements (THRs), and to determine the effect of implantation time on wear particle characteristics. Metal wear particles isolated from periprosthetic tissues from 19 patients with MM THRs of current and older designs and at different implantation times (very short, longer, and very long) were studied using transmission electron microscopy and energy dispersive X-ray analysis. The particles from the current design implants with implantation times of not more than 15 months (very short term) were almost exclusively round to oval chromium oxide particles. In all other cases, although the predominance was still round to oval chromium oxide particles, greater proportions of cobalt-chromium-molybdenum (Co-Cr-Mo) particles, mainly needle-shaped, were detected. Very long term THRs implanted for more than 20 years had the highest percentage of needle-shaped Co-Cr-Mo particles. Particle lengths were not markedly different between the different designs and implantation times except for the current design implants of not more than 15 months, which had a significantly smaller mean length of 39 nm. In conclusion, the implant design did not seem to have a significant influence on particle characteristics whereas the implantation time appeared to have the most effect on the particles. It should be noted that, because of the limited number of tissue retrievals available, some uncertainty remains regarding the generality of these findings.

The potential toxicity of metal ions in tissues surrounding metal-metal hip replacements is a cause for concern. Previous studies conducted in our laboratory demonstrated that Co(2+) and Cr(3+) induce TNF-alpha secretion in macrophages, as well as cell mortality. However, the degree of apoptosis and necrosis remained to be investigated. The aim of the present study was to quantify the rate of macrophage mortality by apoptosis vs. necrosis induced by Co(2+) and Cr(3+). J774 mouse macrophages were incubated in growth medium containing 0-10 ppm Co(2+) and 0-500 ppm Cr(3+) for 24 and 48 h under conventional cell culture conditions. Transmission electron microscopy, flow cytometry (Annexin-V fluorescein isothiocyanate/propidium iodide assay) and a specific cell death detection ELISA were used to illustrate cell death and differentiate between apoptotic and necrotic cells. Cell culture exposed to low concentrations of Co(2+) (0-6 ppm) revealed a low degree of mortality. In contrast, at the highest concentrations (8-10 ppm), late apoptosis occurred within 24 h. After 48 h, however, there was a clear evidence for an increase in the rate of necrosis while apoptosis occurred at much lower rate. Macrophages exposed to Cr(3+) demonstrated a predominance of apoptosis after 24h. At concentrations lower than 250 ppm, early and late apoptosis occurred at the same rate. At higher concentrations (250-500 ppm), the number of early apoptotic cells decreased in favor of late apoptosis. After 48 h, lower concentrations of Cr(3+) (150 ppm) induced a higher degree of early apoptosis than after 24 h, and some necrosis. At higher concentrations, the percentage of early apoptotic cells decreased, while necrosis became predominant over late apoptosis. In conclusion, this study demonstrates that macrophage mortality induced by metal ions depends on the type and concentration of metal ions as well as the duration of their exposure. Overall, apoptosis was predominant after 24 h with both Co(2+) and Cr(3+) ions, but high concentrations induced mainly necrosis at 48 h. These results point to the potential for these ions of inducing tissue damage by necrosis if present in large concentrations in vivo.

There has been a renewed interest in the use of the metal-on-metal (MOM) implants for total hip arthroplasty (THA). It is well known, however, that the MOM articulation generates both metal particles and ions. The physiologic effects of these ions are poorly understood and their potential toxicity remains a cause for concern. In the present study, murine J774 macrophages were incubated with Co2+ and Cr3+ ions and the mode of cell death (apoptosis/necrosis) was evaluated in vitro by transmission electron microscopy and cell death ELISA. Overall, results demonstrated that the mode of cell death was dependent on the ion concentration and the incubation time. Indeed, at short incubation times (24 h), the noninflammatory process of apoptosis was predominant. At longer incubation times (48 h), however, necrosis was predominant at higher ion concentrations.

The purpose of the present study was to compare wear particles isolated from metal-metal (MM) hip implants worn in an orbital bearing simulator with particles from similar MM total hip replacement (THR) implants worn in vivo. Comparison of these particles is important because it will help to assess the overall suitability of this type of hip simulator for reproducing in vivo wear and for producing physiological wear particles suitable for biological studies of in vitro cellular response. Commercial grade components made of ASTM F75 (cast) alloy were evaluated. Simulator tests were performed in 95% bovine calf serum with a 28-mm-diameter implant. Wear particles were collected from 0 to 0.25 million cycles (run-in wear period) and 1.75 to 2 million cycles (steady-state wear period). Tissues from seven patients with MM implants (surface replacement or stem type) were harvested at revision surgeries (after 1-43 months). Metal wear particles were isolated from serum lubricant or tissues using an enzymatic protocol that was previously optimized to minimize particle changes due to reagents. After isolation, particles were centrifuged, embedded in epoxy resin, and characterized by transmission electron microscopy (TEM) and energy dispersive X-ray analysis (EDXA). Results of EDXA on particles from the hip simulator primarily indicated a predominance of particles containing Cr and O but no Co (most likely chromium oxide particles), and fewer CoCrMo particles presenting varying ratios of Co and Cr. Image analysis of TEM micrographs demonstrated that the majority of the particles from the simulator were round to oval, but a substantial number of needle-shaped particles were also found, especially from 0 to 0.25 Mc. The particles generated from 0 to 0.25 Mc had an average length of 53 nm, whereas those generated from 1.75 to 2 Mc had an average length of 43 nm. In vivo, EDXA and TEM analysis of particles that were retrieved from two patients at 23 and 43 months respectively, revealed that they were the most comparable in composition, average length (57 nm), and shape to particles generated in the hip simulator during the run-in wear period. Because a large clinical retrieval study in the literature suggested that a run-in wear regime might occur in vivo for some 6-36 months, the fidelity of the simulator of the present study was strongly supported. However, some uncertainties existed, including the finding that the particles isolated from the other five patients generated from 1 month up to 15 months (shorter implantation times than the other two patients) were smaller and mostly contained only Cr and O (no Co). In the opinion of the authors, this particular very short term patient group was somewhat atypical. Therefore, despite these uncertainties, the present study was deemed to support the ability of the orbital bearing hip simulator to produce physiological wear particles.

There has been a revived interest in metal-metal total hip replacements because of their potential for improved wear performance compared with conventional metal-polyethylene implants. The aim of the present study was to characterize metal wear particles isolated from metal-metal hip simulator testing of various clinically relevant alloys and to analyze the effects of these alloys and the number of loading cycles on wear particle characteristics. Implants were manufactured using medical-grade cobalt-chromium-molybdenum (CoCrMo) alloys that were high-carbon wrought, low-carbon wrought, or cast (with solution annealing). Testing was performed in a MATCO orbital bearing hip simulator in 95% bovine calf serum. The wear particles were isolated from the serum at test periods of 0-0.25 million cycles (Mc) (run-in wear) and 1.75-2 Mc (steady-state wear) using an enzymatic protocol previously optimized to minimize particle changes due to reagents. Isolated particles embedded in epoxy resin were characterized by transmission electron microscopy (TEM) and energy dispersive X-ray analysis (EDXA). The EDXA results revealed the predominance of "lighter" particles containing Cr and O (most likely chromium oxide particles from the passivation layer) and fewer darker CoCrMo particles, with varying ratios of Co and Cr (possibly from carbides and from implant matrix material). More CoCrMo particles were observed with the low-carbon wrought alloy, but the majority of the particles for all three alloys was chromium oxides, especially for the 1.75-2 Mc test period. Image analysis of TEM micrographs revealed that for 0-0.25 Mc, there was up to 21% needle-shaped particles but that the majority remained round to oval in shape, reflecting the predominance of chromium oxide particles. Particle length averaged about 52 +/- 4 nm, with only small differences due to the alloy. For 1.75-2 Mc, most particles were round to oval in shape. They were even less needle-shaped than at 0.25 Mc, and they had a slightly smaller length, averaging 46 +/- 3 nm. In addition to characterizing the size and shape of particles from a MATCO simulator, this study is the first to demonstrate that particles that do not contain Co (presumably chromium oxides) can be predominant in the wear of metal-metal hip implants. It is therefore recommended that future in vitro and in vivo studies include the effects of these particles rather than just the effects of CoCrMo particles on the overall tissue response.

With the introduction of second generation metal-on-metal (MM) hip implants in the mid-1990s, increased interest has arisen regarding the local biological effects of metal wear particles and ions on soft tissues, which have been grouped under the term adverse reactions to metal debris (ARMD). When these reactions constitute a soft tissue mass, they are usually referred to as a pseudotumor [15]. Histological studies of these masses have shown the presence of features characteristic of a non-specific inflammatory reaction (macrophages and wear particles) but also the presence of perivascular infiltrates of lymphocytes that are suggestive of a hypersensitivity reaction [3, 7, 13]. The first description of a hip implant-related pseudotumor was made in 1988 with a metal-on-polyethylene (MPE) bearing. The mass was attributed to a local response to metal ion release from corrosion at the Morse taper head-neck junction [16]. In 2008, a report of a multisurgeon experience with MM hip resurfacing arthroplasties (HRA) estimated that pseudotumors occurred in approximately 1% of patients within 5 years, but acknowledged that with time, the incidence of these pseudotumors may increase [15]. A year later, the same group reported on 16 cases of pseudotumors that went to revision surgery [6]. Five cases required re-revision for instability and loosening, and three of these cases had evidence of pseudotumor recurrence at the time of revision. We report on a patient who, after a hip resurfacing, presented with a pseudotumor that recurred after conversion to a conventional MPE total hip replacement (THR). Our hypothesis is that the recurrence was caused by ongoing corrosion at the Morse taper junction between the titanium stem and the cobalt-chromium-molybdenum (CoCrMo) head. The patient consented for the publication of this case.

We recently identified apoptosis in in vitro wear particle-stimulated macrophages. The recent explosion of interest in apoptosis lies in the fact that it is under positive and negative regulation through evolutionary conserved biochemical pathways. It may also be possible to modulate macrophage apoptosis in the treatment of periprosthetic osteolysis. The purpose of this study was to compare the macrophage response to identically sized particles of alumina ceramic (Al2O3) and ultra-high-molecular-weight polyethylene (UHMWPE) in terms of TNF-alpha release and induction of apoptosis. J774 mouse macrophages were incubated for 0-24 h in the presence of Al2O3 and UHMWPE particles. TNF-alpha release was measured by ELISA; Poly(ADP-ribose)polymerase (PARP) and caspase-3 expression was analyzed by Western blot; DNA fragmentation (DNA laddering) was visualized on agarose gel containing ethidium bromide. Al2O3 particles induced TNF-alpha release after 4 h incubation with concentrations reaching 483 and 800 pg/ml after 24 h with 125 and 250 particles/macrophage, respectively (control = 161 pg/ml) (P0.05 vs. control). The same concentrations of UHMWPE particles induced a much larger and significant TNF-alpha release after only 1 h incubation, increasing up to 6250 pg/ml after 24 h (P0.05 vs. control). Western blot analysis demonstrated that the active caspase-3 fragment (17 kDa) and the proteolytic PARP fragment (85 kDa) were expressed after 2 h incubation with 125 and 250 Al2O3 particles/macrophage. The active caspase-3 and the PARP fragment had lower expression and appeared after a longer incubation time (8 h) with 125 and 250 UHMWPE particles/macrophage. Finally, DNA fragmentation (DNA laddering) was observed after 16 h with 125 and 250 particles of Al2O3 per macrophage whereas no laddering was induced by UHMWPE particles even after 24 h incubation. This study shows that although both Al2O3 and UHMWPE particles induce TNF-alpha release, this stimulation was much greater (8-10 times higher) with UHMWPE than Al2O3 (P0.05 vs. control). As well, the induction of apoptosis, as measured by activation of caspase-3, PARP cleavage and DNA laddering, is different for these two particles, being faster and more important with Al2O3 than UHMWPE. We hypothesize that the ability of Al2O3 to induce macrophage apoptosis may explain the lower TNF-alpha release observed with these particles and explain the differences seen in osteolysis patterns of ceramic-ceramic (CC) vs. metal-polyethylene (Mpe) articulations. In conclusion, apoptosis may be a major internal mechanism to decrease macrophage activity and may be a desired therapeutic endpoint. The identification of an apoptosis-related pathway in the macrophage response to ceramic particles provides crucial data for a rational approach in the treatment and/or prevention of periprosthetic osteolysis.

The aim of this study was to evaluate the cytotoxic and apoptotic effects of cobalt and chromium ions on macrophages in vitro, and analyze the implication of caspase-3 in the apoptotic pathway. J774 mouse macrophages (5 x 10(5) cells/ml) were exposed for up to 24 h to 0-10 ppm Co2+ and 0-500 ppm Cr3+. The cytotoxic effect of ions was measured by Trypan blue exclusion. DNA analysis on agarose gel was used as a specific test for detection of DNA fragmentation into oligonucleosomes that occurs in apoptotic cells. The proteolytic cleavage of poly(ADP-ribose)polymerase (PARP), closely associated with the induction of apoptosis, was also analyzed along with the appearance of the active fragment of caspase-3, implicated in several apoptosis pathways. Results demonstrated that both Co2+ and Cr3+ ions induce macrophage mortality in a dose-dependent manner. However, Co2+ is more toxic inducing a cell mortality up to 28% with only 10 ppm vs. 37% with 500 ppm of Cr3+. DNA analysis demonstrated that both Co2+ and Cr3+ ions induce DNA fragmentation, between 6-10 ppm Co2+ and 250-500 ppm Cr3+ after 24 h incubation. PARP cleavage and the appearance of caspase-3 active fragment were observed after 6 h with both Co+ and Cr3+ ions, with a stronger signal after 24 h and 10 ppm of Co2+ or 500 ppm of Cr3+. In conclusion, this study demonstrates that after 24 h incubation, both Co2+ and Cr3+ ions can induce macrophage mortality, and more specifically apoptosis. The results also suggest that apoptosis occurs via a caspase-3 pathway. However, the relative importance of necrosis and apoptosis and the effects of longer exposure times on the induction of macrophage death by these metal ions remain to be investigated.

The isolation of metal wear particles from hip simulator lubricants is important for understanding wear mechanisms and the tissue response to particulate material. Part I of this study demonstrated that isolation protocols involving digestion reagents can chemically attack metal-metal wear particles, reducing their size and changing their shape. In part II of this study, Co and Cr ion concentrations in solution after each digestion protocol were measured by flame atomic absorption spectrometry, and wear particle composition was determined by X-ray analysis spectra. The exposure of wear particles in water to alkaline solutions caused an increasing release of Cr ions in solution with alkaline concentration and time, and a corresponding decrease in particle Cr peak intensity on X-ray spectra. As a result, particles exposed to 12N KOH for 48 h displayed Co peaks and no Cr. In contrast, enzymatic protocols caused a release of Co ions in solution and a corresponding decrease in particle Co peak intensity on X-ray spectra, especially with sodium phosphate as a buffer. However, when isolating particles from 95% serum, there was an initial protective effect of serum proteins, presumably because of their binding to Co and Cr. As a result, the extent of Cr ion release from metal wear particles in 95% serum after alkaline treatments was diminished, although still present, whereas both enzymatic protocols resulted in a negligible release of Co and Cr ions into solution. Particle composition analysis after enzymatic treatments revealed the presence of chromium oxide particles and CoCrMo particles with variable Co/Cr ratios. After alkaline treatments, the chromium oxide particles increasingly disappeared with time and alkaline concentration, demonstrating a change in particle composition after these treatments. This study demonstrated that digestion reagents can induce chemical changes that affect particle composition. Of all the protocols tested, the enzymatic protocols were the least damaging to the particles and appeared to be the best compromise for isolation and characterization of metal particles, especially in 95% serum. Special care on the choice of buffers should be taken when isolating particles from a lower concentration of serum.

Isolation of metal wear particles from hip simulator lubricants or tissues surrounding implants is a challenging problem because of small particle size, their tendency to agglomerate, and their potential for chemical degradation by digestion reagents. To provide realistic measurements of size, shape, and composition of metal wear particles, it is important to optimize particle isolation and minimize particle changes due to the effects of the reagents. In this study (Part I of II), transmission electron microscopy (TEM) was used to examine and compare the effects of different isolation protocols, using enzymes or alkaline solutions, on the size and shape of three different types of cobalt-based alloy particles produced from metal-metal bearings. The effect on particle composition was examined in a subsequent study (Part II). Large particles (

Although the response of macrophages to polyethylene debris has been widely studied, it has never been compared with the cellular response to ceramic debris. Our aim was to investigate the cytotoxicity of ceramic particles (Al2O3 and ZrO2) and to analyse their ability to stimulate the release of inflammatory mediators compared with that of high-density polyethylene particles (HDP). We analysed the effects of particle size, concentration and composition using an in vitro model. The J774 mouse macrophage cell line was exposed to commercial particles in the phagocytosable range (up to 4.5 μm). Al2O3 was compared with ZrO2 at 0.6 μm and with HDP at 4.5 μm. Cytotoxicity tests were performed using flow cytometry and macrophage cytokine release was measured by ELISA. Cell mortality increased with the size and concentration of Al2O3 particles. When comparing Al2O3 and ZrO2 at 0.6 μm, we did not detect any significant difference at the concentrations analysed (up to 2500 particles per macrophage), and mortality remained very low (less than 10%). Release of TNF-α also increased with the size and concentration of Al2O3 particles, reaching 195% of control (165 pg/ml v 84 pg/ml) at 2.4 μm and 350 particles per cell (p < 0.05). Release of TNF-α was higher with HDP than with Al2O3 particles at 4.5 μm. However, we did not detect any significant difference in the release of TNF-α between Al2O3 and ZrO2 at 0.6 μm (p > 0.05). We saw no evidence of release of interleukin-1α or interleukin-1ß after exposure to ceramic or HDP particles.

The purpose of this study was to investigate in vitro the presence of apoptotic cell death after macrophage stimulation with different ceramic (Al2O3 and ZrO2) and high density polyethylene (HDP) particles. We also analyzed the effects of particle size, concentration, and composition. The J774 mouse macrophage cell line was exposed to commercial particles of different sizes (up to 4.5 microm) and concentrations (up to 500 particles per macrophage). Fluorescence microscopy and DNA laddering were used to investigate the presence of apoptosis in cell cultures after 24 h of incubation. Fluorescence microscopy of propidium iodide stained cells showed two characteristic morphological features that occur in apoptotic cells, namely nuclear condensation and heterogeneity of stain uptake. The effect of ceramic particles on apoptotic nuclear morphology was size- and concentration-dependent and reached a plateau above 150 particles per macrophage at 1.3 microm. With regards to composition, we did not find any difference in cell morphology between Al2O3 and ZrO2. Ceramic and HDP particles induced DNA fragmentation into oligonucleosomes as evidenced by DNA laddering, another characteristic of apoptosis. The induction of DNA laddering was size- and concentration-dependent whereas particle composition (Al2O3 vs. ZrO2 and Al2O3 vs. HDP) had no effect. In conclusion, our results demonstrated that ceramic and HDP particles induce macrophage apoptotic cell death in vitro and open doors for possible modulation of debris-induced periprosthetic osteolysis.

Osteoarthritis and lesions to cartilage tissue are diseases that frequently result in impaired joint function and patient disability. The treatment of osteoarthritis, along with local bone defects and systemic skeletal diseases, remains a significant clinical challenge for orthopaedic surgeons. Several bone morphogenetic proteins (BMPs) are known to have osteoinductive effects, whereof BMP-2 and BMP-7 are already approved for clinical applications. There is growing evidence that the metabolism of bone as well as the cartilage damage associated with the above disease processes are strongly inter-related with the interactions of the inflammation-related pathways (in particular prostaglandin E₂ (PGE₂)) and osteogenesis (in particular bone morphogenetic protein-2 (BMP-2)). There is strong evidence that the pathways of prostaglandins and bone morphogenetic proteins are intertwined, and they have recently come into focus in several experimental and clinical studies. This paper focuses on PGE₂ and BMP-2 intertwining pathways in bone and cartilage metabolism, and summarizes the recent experimental and clinical data.

An increasing number of studies have reported adverse tissue reactions around metal-on-metal (MM) hip implants. However, the origin and mechanisms of these reactions remain unclear. Moreover, the biological effects of nanometer-size chromium oxide particles, the predominant type of wear particles produced by MM implants, remain mostly unknown. The purpose of this study was to analyze the cytotoxic effects of clinically relevant nanometer-size chromium oxide particles on macrophage response in vitro. J774.A1 macrophages were cultured with either 60 nm or 700 nm commercially available Cr2 O3 particles at different concentrations. Two different particle sizes were analyzed to evaluate potential volume effects. Cell mortality was analyzed by light microscopy, flow cytometry (annexin V-fluorescein isothiocyanate and propidium iodide assay), and using a cell death detection enzyme-linked immunosorbant assay (ELISA). Tumor necrosis factor alpha (TNF-α), monocyte chemotactic protein-1 (MCP-1), and macrophage inflammatory protein-1 alpha (MIP-1α) release was measured by ELISA, and gene expression was analyzed by quantitative real-time PCR. Results showed that, at high concentrations, Cr2 O3 particles of both sizes can be cytotoxic, inducing significant decreases in total cell numbers and increases in necrosis. Results also suggested that these effects were dependent on particle volume. However, TNF-α, MCP-1, and MIP-1α cytokine release and gene expression remained low. Overall, this study demonstrates that nanometer-size particles of Cr2 O3 , a stable form of chromium oxide ceramic, have rather low cytotoxic effects on macrophages. Therefore, these particles may not be the main culprit in the initiation of the inflammatory reaction in MM periprosthetic tissues. However, other parameters (e.g., potential intracellular damage) remain to be investigated.

A theoretical clinical advantage of hip resurfacing (HR) is the preservation of femoral bone. HR femoral component revision reportedly yields postoperative function comparable to that of primary THA. However, few studies have looked at the outcome of both HR femoral and acetabular side revisions.We determined whether (1) patients undergoing HR revision to THA have perioperative measures and outcome scores comparable to those of patients undergoing primary THA or revision of primary THA and (2) patients undergoing HR revision of both components have perioperative measures and outcome scores comparable to those of patients undergoing HR revision of the femoral component only.We retrospectively reviewed and compared 22 patients undergoing revision HR to a THA to a matched (age, sex, BMI) group of 23 patients undergoing primary THA and 12 patients undergoing primary THA revision. Patients completed the WOMAC and SF-12 questionnaires before surgery and at latest followup (range, 24-84 months for HR revision, 28-48 months for primary THA, and 24-48 months for revision THA). Blood loss, days in hospital, complications, and outcome scores were compared among groups.We observed no differences in SF-12 scores but observed lower WOMAC stiffness, function, and total scores in the HR revision group than in the primary THA group. Patients undergoing HR revision of both components had comparable SF-12 and WOMAC stiffness, function, and total scores but overall lower WOMAC pain scores compared to patients undergoing HR revision of the femoral side only. The HR revision group had greater intraoperative blood loss compared to the primary THA group but not the revision THA group.The perioperative measures and outcome scores of HR revision are comparable to those of revision THA but not primary THA. Longer followup is required to determine whether these differences persist. Patients undergoing HR revision of one or both components can expect comparable stiffness and function.Level IV, therapeutic study. See the Instructions for Authors for a complete description of levels of evidence.

This paper first presents a brief overview about the mechanism of wear particle formation as well as wear particle characteristics in metal-on-polyethylene and metal-on-metal artificial hip joints. The biological effects of such particles are then described, focusing on the inflammatory response induced by each type of particles as well as on how metal wear products may be the source of a T lymphocyte-mediated specific immune response, early adverse tissue responses, and genotoxicity. Finally, some of the current in vivo models used for the analysis of tissue response to various wear particles are presented.

Aseptic loosening resulting from periprosthetic osteolysis continues to be an important cause of hip implant failure. Wear particles from the bearing surfaces play a major role in initiating periprosthetic osteolysis, which is also potentiated by mechanical factors such as increased synovial fluid pressure. The precise mechanisms by which wear particles induce periprosthetic osteolysis have not been fully elucidated and remain an active subject of research. Particle characteristics such as composition, size, shape, and number (especially for particles in the most biologically active, submicrometer-size range) are recognized to significantly affect the overall cell and tissue response. The production of corrosion products, especially from metal-on-metal implants, also is a clinically significant issue, and individual variability in innate and adaptive immune responses is important but not yet completely defined. Because of the increasing need to implant hip prostheses in younger and more active patients, a better understanding of the biologic activity of wear particles from bearing couples is critical in the attempt to modulate the clinical effects of these particles and to develop materials with improved wear and corrosion resistance.

Metal-on-metal bearings have demonstrated low wear after many years in vivo1,2. Consequently, there has been a resurgence of interest in metal-on-metal bearings. There are, however, concerns about potential adverse effects due to local accumulation and systemic distribution of cobalt-chromium wear moieties and ions, especially after several decades in situ3-5. Cobalt and chromium particles have been shown to induce carcinoma in animal models6, giving rise to the concern that such alloys could have the same effect in human tissue, if present in sufficient amounts for a sufficient length of time. Autopsy retrievals provide the opportunity to assess the distribution of wear products in the tissues immediately surrounding as well as at a distance from the implant7. The aim of this study was to assess the distribution and histological effects of wear particles from a McKee-Farrar metal-on-metal-bearing total hip replacement after nearly thirty years in situ. Amoderately obese, forty-seven-year-old woman fell and sustained a transcervical fracture of the right femur, which was treated initially with internal fixation and a spica cast. A subsequent nonunion was treated with a Judet prosthesis, which was eventually removed, leaving the patient with a Girdlestone resection of the right hip. In 1970, at the age of sixty-eight years, the patient underwent total hip arthroplasty of the left hip for the treatment of extensive osteoarthritis, and a metal-on-metal McKee-Farrar total hip prosthesis was implanted. In 1995, the patient was contacted by our research group, and she agreed to be evaluated as part of a long-term study of McKee-Farrar total hip replacements8,9. She also consented to join the “Willed Joint” program of the Joint Replacement Institute, thereby agreeing to donate her hip joints and to permit postmortem biopsies of her organs. The patient …

Cytokines associated with osteolysis have been demonstrated in tissues surrounding failed metal-metal (MM) total hip replacements (THRs). The objective of the present study was to semi-quantify the amounts of inflammatory cytokines in tissues from 28 failed MM THRs, and determine their relationship with the quantity of metal particles. Paraffin sections were immunohistochemically stained with monoclonal antibodies: anti-IL-1-beta, anti-IL-6 and anti-TNF-alpha. Cytokines and metal particles were rated in 10 fields per tissue using standard light microscopy. Because of the use of light microscopy, only relatively large particles or agglomerations of particles were visible. Therefore, a polarized light and a semi-quantitative scheme based on the discoloration of cell cytoplasms induced by the presence of particles were used to evaluate the quantity of metal particles. Results showed an overall higher amount of IL-6 than IL-1beta while TNF-alpha remained at very low levels. For each patient, the average IL-1beta and IL-6 ratings decreased when the average particle rating increased, following a linear regression, with relatively high correlation factors (r=-0.69 for IL-1beta and r=-0.57 for IL-6). IL-1beta decreased about twice as fast as IL-6. TNF-alpha, remaining at very low levels, did not demonstrate any correlation with particle rating. When multiple tissues were available for the same patient, the correlation factors between the average cytokine and particle ratings were highly variable between samples, demonstrating the heterogeneity between the tissues from the same patient. At the cellular level, there was an even higher correlation between the quantity of metal particles and the production of IL-1beta and IL-6 (r=-0.99), while TNF-alpha did not demonstrate any correlation, remaining at very low levels. In conclusion, this study showed that tissues surrounding failed MM THRs with low to moderate quantities of metal particles can induce the production of potentially osteolytic cytokines. However, the overall number of cells producing these cytokines tended to be lower than that typically seen in tissues surrounding metal-polyethylene THRs.

This study analyzed human mesenchymal stem cell (hMSC) behavior in a fibrin sealant. hMSC morphology, proliferation, and osteogenic differentiation were analyzed after up to 28 days of incubation in eight different formulations of fibrin gels (Tisseel) prepared with various concentrations of fibrinogen complex (FC) and thrombin. Cell morphology and distribution within the gels were observed by fluorescence microscopy after cell staining with calcein dye. Cell proliferation was assessed by measuring the fluorescence intensity of the cell suspension stained with calcein dye after dissolution of the gels. A standard alkaline phosphatase (ALP) assay, von Kossa staining, and real-time reverse transcriptase-polymerase chain reaction (RT-PCR) were used to analyze hMSC osteogenic differentiation. Cell behavior varied depending on the gel formulation. Proliferation was higher in the formulations containing a low FC concentration, but ALP activity was higher in the formulations containing a high FC concentration. Variations in thrombin concentration had a lesser effect. Small nodules of mineralization were observed at days 21 and 28 in a formulation containing a high FC concentration, in addition to a marked increase in bone sialoprotein (BSP) gene expression level as well as a lower increase in ALP and osteopontin (OPN) levels. However, there was no significant increase in osteocalcin (OCN) expression, a late marker of osteogenic differentiation, up to day 28. In conclusion, this study demonstrated that hMSC morphology, proliferation, and osteogenic differentiation in fibrin gels depended on the FC/thrombin ratio. hMSCs appeared to undergo osteogenic differentiation when seeded in Tisseel fibrin sealant containing a high FC concentration, but they did not fully differentiate into mature osteoblasts.