1,720,972 research outputs found
Effects of Metal Ions Released from Implants on Energy Metabolism in Macrophages
No full textCobalt-chromium-molybdenum (CoCrMo) alloys are used extensively in orthopaedic applications. However, they can undergo wear and corrosion in vivo, leading to the release of Co and Cr ions that can limit implant functionality and survivorship because of their biological effects. Indeed, previous studies have shown that Co2+ and Cr3+ can stimulate the production of bone-resorbing cytokines through the activation of redox-dependent mechanisms and induce an inflammatory response in macrophages. However, the effects of Co2+ and Cr3+ on the energy metabolism of macrophages remain largely unknown. The objectives of this thesis were to determine, in macrophages: 1. the effects of Co2+ and Cr3+ on oxidative stress and mitochondrial function; 2. the effects of Co2+ and Cr3+ on glycolytic flux and the stabilization of HIF-1α. RAW 264.7 murine macrophages were exposed to different Co2+ or Cr3+ concentrations for up to 48h. Biochemical, and immunoblotting assays as well as extracellular flux analyses were performed to analyze reactive oxygen species (ROS) production, oxidative damage, mitochondrial function, glycolytic activity, as well as stabilization of HIF-1α after macrophage exposure to the metal ions. Mitochondrial function and glycolytic activity were assessed by measuring oxygen consumption rates (OCR) and extracellular acidification rates (ECAR). Overall, results showed that Co2+ but not Cr3+ induced oxidative stress as well as a decrease in oxidative phosphorylation (OXPHOS). Both Co2+ and Cr3+ induced an increase in glycolytic flux, albeit to a lower level with Cr3+. Moreover, Co2+ but not Cr3+ induced the stabilization of HIF-1α, likely contributing to the Co2+-induced increase in glycolytic flux. Altogether, results suggest that Co2+ can induce a metabolic shift from OXPHOS towards aerobic glycolysis similar to that observed in macrophages upon polarization towards a pro-inflammatory phenotype. Further elucidation of the molecular mechanisms activated by Co2+ and Cr3+ may facilitate the development of therapeutic approaches to modulate the inflammatory response to metal wear and corrosion products and re-establish immune homeostasis in periprosthetic tissues in order to increase implant longevity
In Vitro Macrophage Response to Nanometer-size Particles from Materials Used in Hip Implants
No full textWear particle-induced inflammation leading to periprosthetic osteolysis remains a major cause of hip implant failure. As polyethylene particles from conventional metal-on-polyethylene implants have been associated with these failures, an interest in lower wear metal-on-metal (MM) bearings has emerged. However, the biological effects of nanometer-size chromium oxide particles, predominant type of wear particles produced by MM implants, remain mostly unknown. Therefore, this study aimed to determine the cytotoxicity of nanometer-size Cr2O3 particles on macrophages in vitro, by analyzing their effects on cell mortality and cytokine release and comparing them with those of similarly-sized alumina (Al2O3) particles (known to be relatively bioinert). Results showed that at high concentrations, nanometer-size Cr2O3 particles can be cytotoxic to macrophages, inducing significant decreases in total cell numbers and increases in necrosis. Results also showed that, at high concentrations, the cytotoxicity of Cr2O3 particles was overall higher than that of Al2O3 particles, even though Cr2O3 and Al2O3 are both stable forms of ceramic materials. However, it appeared to be lower than that of previously reported conventional polyethylene and CoCrMo particles. Therefore, chromium oxide particles may not be the main culprit in initiating the inflammatory reaction in MM periprosthetic tissues
Simvastatin Encapsulation in Alginate-Based Microspheres
No full textDespite the great success of hip implant surgeries, wear particle-induced implant aseptic loosening still limits the implant longevity. Simvastatin, an FDA-approved cholesterol lowering statin, is a promising drug candidate for the treatment of implant aseptic loosening due to its anti-inflammatory properties as well as its ability to stimulate bone growth and inhibit bone resorption. In addition, alginate microspheres have been used extensively in drug delivery applications because of alginate properties, including biocompatibility and gelation in mild conditions. However, the hydrophobicity of simvastatin, as well as the large alginate microsphere pore size leading to the leakage of low molecular weight drugs are limiting factors for their use as a delivery system for simvastatin. Therefore, the objectives of this thesis were twofold: 1. To complex simvastatin with 2-hydroxypropyl-β-cyclodextrin (HP-βCD) in order to increase its solubility; and 2. To increase simvastatin encapsulation efficiency in alginate microspheres by coating the microspheres with chitosan, adding dextran sulfate in the alginate solution, and optimizing the gelation conditions used for the synthesis of the microspheres (e.g., volume of gelation medium, curing time, and addition of simvastatin in the gelation medium). Results showed that simvastatin complexation with HP-βCD increased with HP-βCD to simvastatin molar ratio, to a maximum of 97.6% at the molar ratio of 10. Results also showed that chitosan coating of the alginate microspheres increased simvastatin encapsulation efficiency (up to 10.6%), which was further improved (up to 14.0%) when adding 2.0% (w/v) dextran sulfate to the alginate solution. This increase was likely due to electrostatic interactions between dextran sulfate and chitosan in addition to alginate, resulting in a denser coating. Finally, the addition of simvastatin in the gelation medium was shown to also increase simvastatin encapsulation (up to 22.4%), likely because of a decrease in the diffusion of simvastatin out of the microspheres. Overall, this work completed the initial steps for the development of an alginate-based drug delivery system for simvastatin with the long-term goal of providing a local delivery of simvastatin to modulate implant aseptic loosening
Molecular Mechanisms Involved in Interleukin-1β Release by Macrophages Exposed to Metal Ions from Implantable Biomaterials
No full textMetal ions released from implantable biomaterials have been associated with adverse biological reactions that can limit implant longevity. Previous studies have shown that, in macrophages, Co2+, Cr3+, and Ni2+ can activate the NLR family pyrin domain-containing protein 3 (NLPR3) inflammasome, which is responsible for interleukin(IL)-1β production through caspase-1. Furthermore, these ions are known to induce oxidative stress, and inflammasome priming is known to involve nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling. However, the mechanisms of inflammasome activation by metal ions remain largely unknown. The objectives of this thesis were to determine if, in macrophages: 1. IL-1β release induced by metal ions is caspase-1-dependent; 2. caspase-1 activation and IL-1β release induced by metal ions are oxidative stress-dependent; and 3. IL-1β release induced by metal ions is NF-κB signaling pathway-dependent. Lipopolysaccharide (LPS)-primed murine bone-marrow-derived macrophages were exposed to Co2+, Cr3+, or Ni2+, with or without an inhibitor of caspase-1, oxidative stress, or NF-κB. Culture supernatants were analyzed for active caspase-1 (immunoblotting) and/or IL-1β (ELISA). Overall, results showed that while both Cr3+ and Ni2+ may be inducing inflammasome activation, Cr3+ is likely a more potent activator, acting through oxidative stress and the NF-κB signaling pathway. Further elucidation of the activation mechanisms may facilitate the development of therapeutic approaches to modulate the inflammatory response to metal ions, and thereby increase implant longevity
Molecular Mechanisms Leading to Interleukin-1β Release by Macrophages in Response to Wear and Corrosion Products from Metal Implants
No full textWear particles and ions from cobalt-chromium-molybdenum (CoCrMo)-based implants have been shown to cause adverse immune responses, including periprosthetic osteolysis leading to aseptic loosening, the main cause of implant failure. Previous studies have shown that these wear and corrosion products can lead to the release of inflammatory cytokines, including interleukin-1β (IL-1β), suggesting the involvement of the NLRP3 inflammasome. However, the mechanisms leading to IL-1β release have not been fully elucidated. The primary objectives of this thesis were to determine if, in murine macrophages, IL-1β release induced by micrometre-size CoCrMo particles and nanometre-size chromium oxide (Cr2O3) particles is: 1. Caspase-1-dependent; 2. Reduction-oxidation (redox)-dependent; and 3. NLRP3 inflammasome-dependent. Additionally, the effects of metal ions (Co2+, Cr3+, and Ni2+) on NLRP3 inflammasome activation and the effects of matrix metalloproteinase (MMP) inhibition on IL-1β release induced by CoCrMo particles were analyzed. Results showed that IL-1β release induced by CoCrMo particles was partly caspase-1-, redox-, and MMP-dependent, but NLRP3 inflammasome-independent. On the other hand, IL-1β release induced by Cr2O3 particles appeared to be NLRP3 inflammasome-dependent. Finally, IL-1β release induced by Cr3+, but not Co2+, appeared to be NLRP3 inflammasome-dependent, while Ni2+-induced IL-1β release appeared to be only partially NLRP3 inflammasome-dependent, suggesting that other pathways may also be involved. These findings, which provide additional insights into the mechanisms leading to IL-1β release induced by wear particles and ions from CoCrMo-based implants, may help the future development of therapeutic treatments to modulate wear product-induced inflammation and increase implant longevity
In Vitro Macrophage Response to Nanometer-size Particles from Materials Used in Hip Implants
No full textWear particle-induced inflammation leading to periprosthetic osteolysis remains a major cause of hip implant failure. As polyethylene particles from conventional metal-on-polyethylene implants have been associated with these failures, an interest in lower wear metal-on-metal (MM) bearings has emerged. However, the biological effects of nanometer-size chromium oxide particles, predominant type of wear particles produced by MM implants, remain mostly unknown. Therefore, this study aimed to determine the cytotoxicity of nanometer-size Cr2O3 particles on macrophages in vitro, by analyzing their effects on cell mortality and cytokine release and comparing them with those of similarly-sized alumina (Al2O3) particles (known to be relatively bioinert). Results showed that at high concentrations, nanometer-size Cr2O3 particles can be cytotoxic to macrophages, inducing significant decreases in total cell numbers and increases in necrosis. Results also showed that, at high concentrations, the cytotoxicity of Cr2O3 particles was overall higher than that of Al2O3 particles, even though Cr2O3 and Al2O3 are both stable forms of ceramic materials. However, it appeared to be lower than that of previously reported conventional polyethylene and CoCrMo particles. Therefore, chromium oxide particles may not be the main culprit in initiating the inflammatory reaction in MM periprosthetic tissues
Analysis and Modulation of In Vitro Cell Response to Metal Ions From CoCrMo Alloys Used in Orthopaedic Applications
No full textDespite the high success rates of hip replacements, implant-wear mediated periprosthetic osteolysis remains the most prominent cause of long-term implant failure. Other adverse tissue reactions including hypersensitivity reactions and pseudotumors have also recently been reported as a cause for short-term implant failures. The objectives of this thesis were: 1.) To analyze the effects of Co2+ and Cr3+ released from CoCrMo alloys used in hip implants on macrophage chemokine release; 2.) To determine if Co2+, Cr3+, and the chemokines in cultures of macrophages exposed to Co2+ and Cr3+ can induce migration of T and B lymphocytes; and 3) To analyze the potential modulation of macrophage response to Cr3+ using simvastatin as an anti-inflammatory agent. Results showed that the release of TNF–α and CC chemokines were ion-specific and dose-dependent. Results also suggested that Co2+ and Cr3+ may be capable of directly stimulating the migration of T cells, but not that of B cells, suggesting the potential of these ions to create a micro-environment that would favour a T cell-mediated response in vivo. Results also showed that simvastatin was capable of decreasing chemokine release in macrophages exposed to Cr3+, suggesting its potential to modulate the Cr3+-induced inflammatory response. Together, these studies improve the understanding of the role metal ions play in ion-mediated adverse tissue reactions and potential therapies that may modulate the immune response to metal ions
Immunophenotypic Analysis of Peripheral Blood and Synovial Fluid Lympocytes from Patients with Failed Hip Implants
No full textMetal-on-metal (MM) bearings have been considered as an alternative to conventional metal-on-polyethylene (MPE) bearings because of their lower volumetric wear, but concern exists due to potential metal hypersensitivity. Metal hypersensitivity reactions have been thought to be T cell-mediated delayed type hypersensitivity (DTH) reaction. However some of the MM periprosthetic tissues show the presence of B- and plasma cells, as well as massive fibrin exudation, which are not characteristic of a DTH reaction. Therefore, the exact nature of the hypersensitivity reaction(s) MM implants remains unclear. The present study aimed to compare the phenotypes of lymphocytes from the peripheral blood and synovial fluid of patients with failed MM and MPE implants, and from volunteers with no implant (peripheral blood only). Results in peripheral blood showed differences in the T-cell populations depending on the implant type. This included differences in the proportions of T-helper and T-cytotoxic cells, and T-cells expressing IFN-g. Results in synovial fluid showed a significant difference between MM and MPE groups for the B-cells. Both groups depicted a predominance of T-cell lymphocytes in synovial fluid and overall larger proportions of memory cells than in peripheral blood, but group sizes were rather small. Overall, T-cell cytokine expression (analyzed in peripheral blood only because of the limited number of synovial fluid samples) did not exhibit characteristics of a DTH reaction and the proportions of memory lymphocytes did not indicate activation of a specific subset in the MM group. Nevertheless, group sizes still remain to be increased
In Vitro Simulation of Modular Neck Fracture, Wear, Corrosion, and Distraction in Total Hip Replacements
No full textTotal hip replacements are being used to relieve pain and restore the hip function of unhealthy hip joints. The various sizes and geometries of the modular femoral neck implants allow the surgeon to optimize the range of motion and patient’s leg length. However, some in vivo modular femoral neck retrievals have shown early fatigue and advanced wear-corrosion at the neck-stem taper interface, which can lead to adverse tissue reactions and failure of the implant.
The overall objective of this study was to simulate in vivo fatigue fracture, wear, and corrosion of modular necks at the neck-stem taper interface in a laboratory setting (in vitro) to better predict the failure mechanisms and implant limitations. More specifically, after optimizing the laboratory setup and the testing conditions, this study aimed to compare the effects of the modular neck material (Ti6Al4V and CoCrMo) and the implant assembly technique (hand and impact assembly) on fatigue life, wear-corrosion resistance, and distraction force. The PROFEMUR® Modular Neck System with CoCrMo femoral heads and Ti6Al4V stems was used in this study. The in vitro simulation was divided into two types of tests: fatigue tests (high compression load for a short cyclic loading duration) and corrosion tests (low compression load for a long cyclic loading duration). The neck-stem interface was submersed in a phosphate buffered saline solution, which was maintained at a temperature of 80 ºC to accelerate the corrosion reaction.
The simulation results showed that the Ti6Al4V necks were more vulnerable to fatigue fracture than CoCrMo necks. In addition, impact assembly of the components resulted in an increased implant fatigue life compared to hand assembly, but also increased the distraction force. The observed wear-corrosion damage was higher in fatigue tests than corrosion tests, suggesting that the level of mechanical load was a major factor influencing implant surface damage and fatigue fracture. On the other hand, corrosion tests showed that longer exposure resulted in more fluid accumulation in the stem pocket. This may lead to the formation of a corrosion cell with strongly acidic conditions in the stem pocket, as well as the potential for larger metal ion release. Overall, the in vitro simulation was successful in reproducing femoral modular neck fracture and wear-corrosion damage similar to retrieved in vivo specimens. Results may play a major role in the future development of total hip replacements and international standards for implant testing
Immunophenotypic Analysis of Peripheral Blood and Synovial Fluid Lympocytes from Patients with Failed Hip Implants
No full textMetal-on-metal (MM) bearings have been considered as an alternative to conventional metal-on-polyethylene (MPE) bearings because of their lower volumetric wear, but concern exists due to potential metal hypersensitivity. Metal hypersensitivity reactions have been thought to be T cell-mediated delayed type hypersensitivity (DTH) reaction. However some of the MM periprosthetic tissues show the presence of B- and plasma cells, as well as massive fibrin exudation, which are not characteristic of a DTH reaction. Therefore, the exact nature of the hypersensitivity reaction(s) MM implants remains unclear. The present study aimed to compare the phenotypes of lymphocytes from the peripheral blood and synovial fluid of patients with failed MM and MPE implants, and from volunteers with no implant (peripheral blood only). Results in peripheral blood showed differences in the T-cell populations depending on the implant type. This included differences in the proportions of T-helper and T-cytotoxic cells, and T-cells expressing IFN-g. Results in synovial fluid showed a significant difference between MM and MPE groups for the B-cells. Both groups depicted a predominance of T-cell lymphocytes in synovial fluid and overall larger proportions of memory cells than in peripheral blood, but group sizes were rather small. Overall, T-cell cytokine expression (analyzed in peripheral blood only because of the limited number of synovial fluid samples) did not exhibit characteristics of a DTH reaction and the proportions of memory lymphocytes did not indicate activation of a specific subset in the MM group. Nevertheless, group sizes still remain to be increased
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