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It is widely recognized that innate macrophage immune reactions to implant debris are central to the inflammatory responses that drive biologic implant failure over the long term. Less common, adaptive lymphocyte immune reactions to implant debris, such as delayed type hypersensitivity (DTH), can also affect implant performance. It is unknown which key patient factors, if any, mediate these adaptive immune responses that potentiate particle/macrophage mediated osteolysis. The objective of this investigation was to determine to what degree known adaptive immune responses to metal implant debris can affect particle-induced osteolysis (PIO); and if this pathomechanism is dependent on: 1) innate immune danger signaling, i.e., NLRP3 inflammasome activity, 2) sex, and/or 3) age. We used an established murine calvaria model of PIO using male and female wild-type C57BL/6 vs. Caspase-1 deficient mice as well as young (12–16 weeks old) vs. aged (18–24 months old) female and male C57BL/6 mice. After induction of metal-DTH, and Cobalt-alloy particle (ASTM F-75, 0.4um median diameter) calvaria challenge, bone resorption was assessed using quantitative micro-computed tomography (micro-CT) analysis and immune responses were assessed by measuring paw inflammation, lymphocyte transformation test (LTT) reactivity and adaptive immune cytokines IFN-gamma and IL-17 (ELISA). Younger aged C57BL/6 female mice exhibited the highest rate and severity of metal sensitivity lymphocyte responses that also translated into higher PIO compared to any other experimental group. The absence of inflammasome/caspase-1 activity significantly suppressed DTH metal-reactivity and osteolysis in both male and female Caspase-1 deficient mice. These murine model results indicate that young female mice are more predisposed to metal-DTH augmented inflammatory responses to wear debris, which is highly influenced by active NLRP3 inflammasome/caspase-1 danger signaling. If these results are clinically meaningful for orthopedic patients, then younger female individuals should be appropriately assessed and followed for DTH derived peri-implant complications.

Despite the success in returning people to health saving mobility and high quality of life, the over 1 million total joint replacements implanted in the US each year are expected to eventually fail after approximately 15–25 years of use, due to slow progressive subtle inflammation to implant debris compromising the bone implant interface. This local inflammatory pseudo disease state is primarily caused by implant debris interaction with innate immune cells, i.e., macrophages. This implant debris can also activate an adaptive immune reaction giving rise to the concept of implant-related metal sensitivity. However, a consensus of studies agree the dominant form of this response is due to innate reactivity by macrophages to implant debris danger signaling (danger-associated molecular pattern) eliciting cytokine-based and chemokine inflammatory responses. This review covers implant debris-induced release of the cytokines and chemokines due to activation of the innate (and the adaptive) immune system and how this leads to subsequent implant failure through loosening and osteolysis, i.e., what is known of central chemokines (e.g., IL-8, monocyte chemotactic protein-1, MIP-1, CCL9, CCL10, CCL17, and CCL22) associated with implant debris reactivity as related to the innate immune system activation/cytokine expression, e.g., danger signaling (e.g., IL-1β, IL-18, IL-33, etc.), toll-like receptor activation (e.g., IL-6, tumor necrosis factor α, etc.), bone catabolism (e.g., TRAP5b), and hypoxia responses (HIF-1α). More study is needed, however, to fully understand these interactions to effectively counter cytokine- and chemokine-based orthopedic implant-related inflammation.

Long-term aseptic failures of joint replacements are generally attributed to implant debris-induced inflammation and osteolysis. This response is largely mediated by immune and bone cells (monocytes/macrophages and osteoclasts, respectively), that in the presence of implant debris (e.g. metal particles and ions), release pro-inflammatory cytokines such as IL-1β, TNF-α, and IL-6. The relative degree to which implant debris can illicit inflammatory response(s) from osteoclasts vs monocytes/macrophages is unknown, i.e. are osteoclasts a viable target for anti-inflammatory therapy for implant debris? We investigated relative monocyte versus osteoclast inflammatory responses in a side-by-side comparison using implant debris from the perspective of both danger signaling (IL-1β) and pathogenic recognition (TNF-α) reactivity (Challenge Agents: Cobalt-alloy, Titanium-alloy, and PMMA particles, 0.9-1.8um-dia ECD and Cobalt, and Nickel-ions 0.01-0.1mM, all with and without LPS priming). Human monocytes/macrophages reacted to implant debris with >100 fold greater production of cytokines compared to osteoclast-like cells. Particulate Co-alloy challenge induced >1000 pg/ml of IL-1β and TNF-α, in monocytes and 3000pg/mL IL-1β and TNF-α in monocytes/macrophages and

Background The increasing complexity of articulating spinal implants prohibits the use of serum-supplemented simulator fluid testing because multicomponent interfaces retain residual protein and preclude gravimetric measurement. Our original hypothesis was that simulator testing of a posterior dynamic stabilization implant that has metal-on-metal articulating bearings will not produce dramatically different wear debris when tested using pure saline versus testing in saline supplemented with 20% serum. Methods This hypothesis was tested using simulator testing of 12 dynamic stabilization spinal implants, 6 in 100% saline and 6 in 20%-serum saline. Gravimetric and particle analysis were performed after every million cycles up to 10 million cycles, with flexion of 11.3°/extension of 5.6° coupled with axial rotation of ± 4°. Results The mean gravimetric weight loss was approximately 200 mg over 10 million cycles for the implants tested in 100% saline, while the mean weight loss for those tested in 20%-serum saline was below the method detection limits (< 10 mg over 10 million cycles). For the 100%-saline and 20%-serum simulator fluids, the average particle size over the course of 0 to 10 million cycles remained relatively constant at 0.2 µm-dia (saline) and 3.2 µm-dia (20%-serum saline). Testing in 100% saline generated > 1000-fold more particles, compared to testing in 20% serum-supplemented saline. Energy-dispersive X-ray (EDAX) analyses of particles demonstrated that the 100% saline debris was composed of Co-Cr-P-O (Cr-Co metal oxides), and for the 20%-serum saline debris only bulk metal Co-Cr was detected. Conclusion Our initial hypothesis was not supported. There were significant differences in gravimetric wear, average size, and type of wear debris that were mechanistically attributable to the type of simulator fluid used. The over-protective effect of serum proteins appears to underscore the importance of using both saline and serum when establishing upper and lower bounds of predictive implant debris generation modeling, where saline represents a worst-case scenario and as little as 20% serum masks all weight loss completely in highly modular articulating implants. Clinical Relevance Clinical Relevance = 5 (Oxford Centre for Evidence-based Medicine Levels of Evidence). Study findings are limited to a greater understanding of the science associated with predictive wear testing of articulating spinal implants.

Background Metal-on-metal prostheses undergo wear and corrosion, releasing soluble ions and wear particles into the surrounding environment. Reports described early failures of the metal-on-metal prostheses, with histologic features similar to a Type IV immune response. Mechanisms by which metal wear products and metal ion causing this reaction are not completely understood, and the effects of metal ions on osteocytes, which represent more than 95% of all the bone cells, have not been also studied. We hypothesized that soluble metal ions released from the cobalt-chromium-molybdenum (Co-Cr-Mo) prosthesis may have cytotoxic effect on osteocytes. Methods MLO-Y4 osteocytes were treated with various metal ion solutions for 24 and 48 h. The effect of ion treatment on cytotoxicity was assessed by WST-1 reagents and cell death ELISA. Morphological changes were analyzed by a phase-contrast microscope or fluorescent microscope using Hoechst 33342 and propidium iodine staining. Results Cr and Mo ions did not cause cell death under 0.50 mM, highest concentration studied, whereas Co and Ni ions had significant cytotoxic effect on MLO-Y4 cells at concentrations grater than 0.10 mM and at 0.50 mM, respectively, in a dose-dependent manner. According to the ELISA data, osteocytes treated with Co ions were more susceptible to necrotic than apoptotic cell death, while Ni ions caused osteocyte apoptosis. The morphological assays show that cells treated with Co and Ni ions at high concentration were fewer in number and rounded. In addition, fluorescent images showed a marked reduction in live cells and an increase in dead osteocytes treated with Co and Ni ions at high concentration. Conclusions Metal ions released from metal-on-metal bearing surfaces have potentially cytotoxic effects on MLO-Y4 osteocytes, in vitro.

The historical success of orthopedic implants has been recently tempered by unexpected pathologies and early failures of some types of Cobalt-Chromium-Molybdenum alloy containing artificial hip implants. Hypoxia-associated responses to Cobalt-alloy metal debris were suspected as mediating this untoward reactivity at least in part. Hypoxia Inducible Factor-1α is a major transcription factor involved in hypoxia, and is a potent coping mechanism for cells to rapidly respond to changing metabolic demands. We measured signature hypoxia associated responses (i.e. HIF-1α, VEGF and TNF-α) to Cobalt-alloy implant debris both in vitro (using a human THP-1 macrophage cell line and primary human monocytes/macrophages) and in vivo. HIF-1α in peri-implant tissues of failed metal-on-metal implants were compared to similar tissues from people with metal-on-polymer hip arthroplasties, immunohistochemically. Increasing concentrations of cobalt ions significantly up-regulated HIF-1α with a maximal response at 0.3 mM. Cobalt-alloy particles (1 um-diameter, 10 particles/cell) induced significantly elevated HIF-1α, VEGF, TNF-α and ROS expression in human primary macrophages whereas Titanium-alloy particles did not. Elevated expression of HIF-1α was found in peri-implant tissues and synovial fluid of people with failing Metal-on-Metal hips (n = 5) compared to failed Metal-on-Polymer articulating hip arthroplasties (n = 10). This evidence suggests that Cobalt-alloy, more than other metal implant debris (e.g. Titanium alloy), can elicit hypoxia-like responses that if unchecked can lead to unusual peri-implant pathologies, such as lymphocyte infiltration, necrosis and excessive fibrous tissue growths.

a b s t r a c t In biomedical research, because of the critical nature of corrosion within the environment of motion and interactions of weight bearing surfaces, scientists and engineers have shown a growing interest in the area of "tribocorrosion". Fundamentally, tribocorrosion combines the disciplines of tribology and corrosion and their concurrent exchanges. Principles of tribocorrosion are applied in a range of industries including the offshore, space and biomedical (for example, dental and orthopedic) industries. One of the challenges in tribocorrosion research is the lack of standard testing equipment and method- ologies, particularly in its bio-medical applications. Many investigators and research laboratories are engaged in modifying their existing classical tribology apparatus, whether they are commercially avail- able or customized tribometers, to integrate electrochemical monitoring. The current work is focused on developing a tribocorrosion test apparatus to model the contact situation of artificial hip joints with particular attention on possible erroneous issues that could arise in such a system. The test system facilitates studies on the tribocorrosion behavior of implant metals or coatings as a function of diverse tribological and electrochemical parameters, and allows monitoring temperature, pH and dissolved oxygen level during testing. Initial results on the HC CoCrMo alloy in bovine calf serum (BCS), indicate that the proposed system is reliable and useful for the investigation of tribocorrosion behavior of implant materials. Electrochemical impedance spectroscopy tests (EIS) are employed before and after the sliding tests. The usefulness of such tests in understanding the changes in the surface chemistry and corrosion kinetics are explained, with the help of optimum EIS models. An attempt has also been made to highlight the relevance on wear quantification methods and possible ways to analyse the synergistic interactions of wear and corrosion for the biomedical tribosystems in orthopedics and

We have previously described a unique gene expression signature exhibited by synovial fluid leukocytes in response to bacterial infection, identifying a number of potential biomarkers for infection. However, the diagnostic performance of these potential biomarkers in an immunoassay format is unknown. We therefore evaluated the sensitivity, specificity, and accuracy of several potential synovial fluid biomarkers for infection, and compared them to current standards of testing for periprosthetic infection. We prospectively collected synovial fluid from 14 patients classified as having a periprosthetic infection and 37 patients classified as having an aseptic failure. The synovial fluid samples were tested for 23 potential biomarkers for periprosthetic infection. We then determined differences in biomarker levels between infected and aseptic groups, then computed the sensitivity, specificity, positive predictive value, negative predictive value and accuracy for select biomarkers, and finally compared those to current standard tests for infection. Twelve synovial fluid biomarkers had substantially higher average levels in the synovial fluid of infected versus aseptic patients. Synovial fluid levels of IL-1 were a mean of 258 times higher in patients with a periprosthetic infection compared to patients having revision for aseptic diagnoses. Synovial fluid IL-1 and IL-6 levels correctly classified all patients in this study with a sensitivity, specificity, positive predictive value, negative predictive value and accuracy equal to 1. Several markers tested in this study outperformed the ESR and CRP tests. Patients with a periprosthetic infection have elevated levels of numerous synovial fluid biomarkers, when compared to patients with aseptic diagnoses. Several of these biomarkers exhibited nearly ideal sensitivity, specificity, and accuracy in this study, suggesting that synovial fluid biomarkers could be a valuable tool for diagnosing periprosthetic infection. Level III, prognostic study. See Guidelines for Authors for a complete description of levels of evidence.

Background Proximal femoral bone loss during revision hip arthroplasty often requires bypassing the deficient metaphyseal bone to obtain distal fixation. The purpose of this study was to determine the effect of stem diameter and length of diaphyseal contact in achieving rotational stability in revision total hip arthroplasty. Methods Twenty-four cadaveric femoral specimens were implanted with a fully porous-coated stem. Two different diameters were tested and the stems were implanted at multiple contact lengths without proximal bone support. Each specimen underwent torsional testing to failure and rotational micromotion was measured at the implant-bone interface. Results The larger stem diameter demonstrated a greater torsional stability for a given length of cortical contact (p ≤ 0.05). Decreasing length of diaphyseal contact length was associated with less torsional stability. Torsional resistance was inconsistent at 2 cm of depth. Conclusion Larger stem diameters frequently used in revisions may be associated with less diaphyseal contact length to achieve equivalent rotational stability compared to smaller diameter stems. Furthermore, a minimum of 3 cm or 4 cm of diaphyseal contact with a porous-coated stem should be achieved in proximal femoral bone deficiency and will likely be dependent on the stem diameter utilized at the time of surgery.

Wear debris-induced osteolysis remains the greatest limitation of long-term success for total joint replacements with ultra-high molecular weight polyethylene (UHMWPE) bearings. To address oxidative degradation post-gamma irradiation, manufacturers are investigating the incorporation of antioxidants into PE resins. Similarly, larger molecular weight monomers have been developed to increase crosslinking and decrease wear debris, and ultimately osteolysis. However, the effects of modifying monomer size, crosslink density, and antioxidant incorporation on UHMWPE particle-induced osteoclastic bone resorption and coupled osteoblastic bone formation have never been tested. Here, we review the field of antioxidant-containing UHMWPE, and present an illustrative pilot study evaluating the osteolytic and osteogenic potential of wear debris generated from three chemically distinct particles (MARATHON®, XLK, and AOX™) as determined by a novel 3D micro-CT algorithm designed for the murine calvaria model. The results demonstrate an approach by which the potential osteoprotective effects of antioxidants in UHMWPE can be evaluated.

Background All prostheses with metallic components release metal debris that can potentially activate the immune system. However, implant-related metal hyper-reactivity has not been well characterized. In this study, we hypothesized that adaptive immunity reaction(s), particularly T-helper type 1 (Th1) responses, will be dominant in any metal-reactivity responses of patients with total joint replacements (TJAs). We tested this hypothesis by evaluating lymphocyte reactivity to metal "ions" in subjects with and without total hip replacements, using proliferation assays and cytokine analysis. Methods Lymphocytes from young healthy individuals without an implant or a history of metal allergy (Group 1: n = 8) were used to assess lymphocyte responses to metal challenge agents. In addition, individuals (Group 2: n = 15) with well functioning total hip arthroplasties (average Harris Hip Score = 91, average time in-situ 158 months) were studied. Age matched controls with no implants were also used for comparison (Group 3, n = 8, 4 male, 4 female average age 70, range 49–80). Group 1 subjects' lymphocyte proliferation response to Aluminum+3, Cobalt+2, Chromium+3, Copper+2, Iron+3, Molybdenum+5, Manganeese+2, Nickel+2, Vanadium+3 and Sodium+2 chloride solutions at a variety of concentrations (0.0, 0.05, 0.1, 0.5, 1.0 and 10.0 mM) was studied to establish toxicity thresholds. Mononuclear cells from Group 2 and 3 subjects were challenged with 0.1 mM CrCl3, 0.1 mM NiCl2, 0.1 mM CoCl2 and approx. 0.001 mM titanium and the reactions measured with proliferation assays and cytokine analysis to determine T-cell subtype prominence. Results Primary lymphocytes from patients with well functioning total hip replacements demonstrated a higher incidence and greater magnitude of reactivity to chromium than young healthy controls (p < 0.03). Of the 15 metal ion-challenged subjects with well functioning total hip arthroplasties, 7 demonstrated a proliferative response to Chromium, Nickel, Cobalt and/or Titanium (as defined by a statistically significant >2 fold stimulation index response, p < 0.05) and were designated as metal-reactive. Metals such as Cobalt, Copper, Manganese, and Vanadium were toxic at concentrations as low as 0.5 mM while other metals, such as Aluminum, Chromium, Iron, Molybdenum, and Nickel, became toxic at much higher concentrations (>10 mM). The differential secretion of signature T-cell subsets' cytokines (Th1 and Th2 lymphocytes releasing IFN-gamma and IL-4, respectively) between those total hip arthroplasty subjects which demonstrated metal-reactivity and those that did not, indicated a Th1 type (IFN-gamma) pro-inflammatory response. Conclusion Elevated proliferation and production of IFN-gamma to metals in hip arthroplasty subjects' lymphocytes indicates that a Th1 (vs. Th2) type response is likely associated with any metal induced reactivity. The involvement of an elevated and specific lymphocyte response suggests an adaptive (macrophage recruiting) immunity response to metallic implant debris rather than an innate (nonspecific) immune response.