Comparison of different anti-inflammatory agents in suppressing the monocyte response to orthopedic particles

Three different anti-inflammatory agents--diclofenac, dexamethasone, and N-acetylcysteine--were compared to evaluate their effectiveness in suppressing monocyte-macrophage cell culture activation and mediator release (tumor necrosis factor-alpha [TNF-alpha] and interleukin-1beta [IL-1beta]) in response to polymethylmethacrylate particulate debris. N-acetylcysteine and diclofenac were most effective in suppressing TNF-alpha and IL-1beta expression by the monocyte-macrophages. Dexamethasone reduced TNF-alpha expression but was not as effective suppressing IL-1beta expression. N-acetylcysteine and dexamethasone had no effect on cell viability whereas diclofenac at the highest concentrations decreased cell viabilities. N-acetylcysteine and diclofenac, but less so dexamethasone, are effective in suppressing wear debris-related cell activation and mediator release and thus potentially represent therapeutic or preventive modalities for periprosthetic osteolysis.     

Cement particles containing radio-opacifiers stimulate pro-osteolytic cytokine production from a human monocytic cell line

Proponents of the biological theory of aseptic loosening have in recent years tended to concentrate on the production and distribution of particulate ultra-high-molecular-weight polyethylene (UHMWPE) debris around the potential joint space. However, mechanical loading of cemented implants with the differing elastic moduli of metal stems, polymethylmethacrylate (PMMA) cement and bone can result in relative micromotion, implying the potential for production of metal and PMMA particles from the stem-cement interface by fretting wear. In order to investigate the production and biological reactivity of debris from this interface, PMMA and metal particulate debris was produced by sliding wear of PMMA pins containing barium sulphate and zirconium dioxide against a Vaquasheened stainless steel counterface. This debris was characterised by SEM, energy-dispersive analysis by X-ray (EDAX) and image analysis, then added to cell cultures of a human monocytic cell line, U937, and stimulation of proosteolytic cytokines measured by ELISA. Large quantities of PMMA cement debris were generated by the sliding wear of PMMA pins against Vaquasheened stainless steel plates in the method developed for this study. Both cements stimulated the release of pro-osteolytic TNFalpha from the U937 monocytic cell line, in a dose-dependent fashion. There was a trend towards greater TNFalpha release with Palacos cement than CMW cement at the same dose. Palacos particles also caused significant release of IL-6, another pro-osteolytic cytokine, while CMW did not. The particulate cement debris produced did not stimulate the release of GM-CSF or IL1beta from the U937 cells. These results may explain the cytokine pathway responsible for bone resorption caused by particulate PMMA debris. Radio-opaque additives are of value in surgical practice and clinical studies to quantify the relevance of these in vitro findings are required before the use of cement containing radio-opacifier is constrained.     

Synergistic effect of particles and cyclic pressure on cytokine production in human monocyte/macrophages: proposed role in periprosthetic osteolysis.

Macrophages, activated by particulate wear debris, are important in the process of osteolysis, which occurs during joint implant loosening. We previously found increased levels of interleukin-1beta (IL-1beta), IL-6, and tumor necrosis factor-alpha in cultured macrophages subjected to cyclical pressure of 0.138 MPa, suggesting that cyclic pressure may be another relevant cause of macrophage activation. The current study first investigated the effects of a range of cyclic pressures on cultured macrophages, including an investigation of the time course of cytokine expression. At 0.138 MPa, supernatant levels of TNF-alpha were maximal at 12 h, whereas IL-6 and IL-1beta were maximal at 24 h. All four cyclic pressure levels tested (without particles) resulted in increased production of all three cytokines relative to control. These increases were most marked at 0.069 and 0.035 MPa, and the increase in cytokine production at 0.017 MPa was not statistically significant. Further studies demonstrated that conditioned media from cyclically pressurized macrophages stimulated bone resorption in a neonatal mouse calvarial assay system. There were increased levels of calcium released from calvaria cultured in conditioned media from pressurised monocytes, and an increase in tartate-resistant acid phosphatase-positive osteoclasts was observed microscopically. As particulate wear debris is important in implant loosening, ultra high molecular weight polyethylene particles were also added to the pressurized cell cultures. The experiments compared the effect of atmospheric pressure, cyclic pressure alone, particles alone, and particles and cyclic pressure combined. A combination of ultra high molecular weight polyethylene particles and cyclic pressure at 0.017 MPa resulted in a dramatic synergistic elevation of levels of all three cytokines compared with the levels found with either pressure or particles alone. We propose that monocyte/macrophage activation by cyclic pressure plays a major role in the osteolysis seen in aseptic loosening of implants. The synergistic effect observed between particles and pressure could accelerate implant loosening, and implies that reduction in either cyclic pressure (by improving implant fixation) or wear debris load would reduce osteolysis.     

The effect of spinal instrumentation particulate wear debris. an in vivo rabbit model and applied clinical study of retrieved instrumentation cases.

STUDY DESIGN: The current study was undertaken to determine if the presence of spinal instrumentation wear particulate debris deleteriously influences early osseointegration of posterolateral bone graft or disrupts an established posterolateral fusion mass.OBJECTIVES: Using an in vivo animal model, the first phase (basic science) of this study was to evaluate the effect(s) of titanium wear particulate on a posterolateral spinal arthrodesis based on serological, histological and immunocytochemical analyses. The second phase (clinical) was to perform the same analysis of soft tissue surrounding spinal instrumentation in 12 symptomatic clinical patients. SUMMARY OF BACKGROUND DATA: The effect of unintended wear particulate resulting from micromotion between the interconnection mechanisms in spinal instrumentation remains a clinical concern. METHODS: Thirty-four New Zealand White rabbits were randomized into two groups based on postoperative time periods of 2 months (Group 1, n=14) and 4 months (Group II, n=20). Group I underwent a posterolateral arthrodesis (PLF) at L5-L6 using tricortical iliac autograft or tricortical iliac autograft plus titanium particulate. Group 2 all received iliac autograft at the initial surgery and were reoperated on after 8 weeks and treated with PLF exposure alone or titanium particulate. Postoperative analysis included serological quantification of systemic cytokines. Postmortem microradiographic, immunocytochemical and histopathological assessment of the intertransverse fusion mass quantified the extent of osteolysis, local proinflammatory cytokines, osteoclasts and inflammatory infiltrates. Clinical aspect of study: Over the last 2 years, 12 patients more than 0.4 years after spinal instrumentation presented with painful paraspinal inflammation. At surgical exploration, the cultures were negative for infection and the surrounding soft tissue was examined for cytokine reactions. There was loosening of implants and osteolysis in the location of the wear debris in 8 of 12 patients. RESULTS: Basic science phase: serological analysis of systemic cytokines indicated no significant differences in cytokine levels (p>.05) between the titanium or autograft treatments. Immunocytochemistry indicated increased levels of local cytokines: TNF-alpha at the titanium-treated PLF sites at both time periods (p<.05). Osteoclast cell counts and regions of osteolytic resorption lacunae were higher in the titanium-treated versus autograft-alone groups (p<.05), and the extent of cellular apoptosis was markedly higher in the titanium-treated sites at both time intervals. Electron microscopy indicated definitive evidence of phagocytized titanium particles and foci of local, chronic inflammatory changes in the titanium-treated sites. Clinical aspect: Eleven of 12 clinical cases demonstrated elevated TNF-alpha levels and an increased osteoclastic response in the vicinity of wear debris caused by dry frictional wear particles of titanium or stainless steel. Osteolysis most commonly involved loose transverse connectors. Resection of the wear debris and surrounding fibroinflammatory glycocalyx resulted in resolution of clinical symptoms in all 12 cases. CONCLUSIONS: Titanium particulate debris introduced at the level of a spinal arthrodesis elicits a cytokine-mediated particulate-induced response favoring proinflammatory infiltrates, increased expression of intracellular TNF-alpha, increased osteoclastic activity and cellular apoptosis. This is the first basic scientific study and the first clinical study demonstrating associations of spinal instrumentation particulates wear debris and increased cytokines and increased osteoclastic activity. Osteolysis is the number one cause of failure of orthopedic implants in the appendicular skeleton. Spinal surgeons need to increase their awareness of this destructive process.     

Accumulation of LPS by polyethylene particles decreases bone attachment to implants.

Molecules absorbed on the surface of particulate wear debris may contribute to inflammatory reactions that lead to aseptic loosening of implants. Lipopolysaccharide (LPS), a bacterial endotoxin, can attach to many biomaterials and stimulate macrophages to secrete osteoclast-activating cytokines. We tested the adsorption of LPS by polyethylene particles in vitro and examined the biological effects of LPS absorption on bone remodeling around implants in vivo. Polyethylene particles were incubated in radiolabeled LPS solutions, and adsorption of LPS by the particles was quantified by radioassay. Because polyethylene particles are hydrophobic and less dense than water, they floated and clumped when incubated in a water solution of LPS, resulting in low adsorption of LPS. However, when particles were incubated in an ethanol solution of LPS, most of the LPS was adsorbed by the particles, and was resistant to washing with water. Triton X-100 (10%), however, effectively washed the LPS off the particles. In a rat model, the presence of polyethylene particles around the implant in the femoral canal decreased bone attachment to the implant at 6 weeks. Incubating the particles with LPS before implantation, or intermittent administration of LPS systemically, further decreased bone-implant attachment to similar extents, but had no effect on the bone density of the control side femurs. Our data indicate that polyethylene particles have high affinity for LPS, depending on many factors, especially the solvents of the LPS. Intermittent systemic administration of LPS affects bone remodeling but only occurs in the area containing polyethylene particles and titanium implants, supporting the hypothesis that the presence of polyethylene particles around implants can result in accumulation of LPS from exogenous sources. This may cause local levels of LPS that are high enough to affect bone remodeling around implants.     

Pathophysiological aspects of hip implants

Osteolysis and aseptic loosening are the major causes of failure of total hip replacements (THRs). These processes are induced by wear debris released from components of implants over time. Wear particles are a clinically relevant size (0.1-10 microm) that activates macrophages and induces a foreign-body response. Activated macrophages stimulate the production of proinflammatory cytokines (IL-1, IL-6, and TNF-alpha) to the synovial capsule, leading to chronic inflammation and bone resorption. Wear mechanics, including characteristics of wear particles from metal, polyethylene, and ceramic articulating surfaces, fixation materials, and surface coatings, have been reported in the literature and are compiled here. Data concerning the pathophysiological effects of wear debris are reported in this article, whereas data on the metal ions released from implants are presented in the following article (see pp. 113-114). Clinical data obtained from in vivo use and laboratory studies conducted in vitro are summarized.     

Inhibition of the PI3K-Akt signaling pathway reduces tumor necrosis factor-alpha production in response to titanium particles in vitro

BACKGROUND: Wear debris contributes to implant loosening after total joint arthroplasty, and few advances have been made in our ability to inhibit the biological response to wear particles. Bacterial endotoxins augment the effects of wear particles in vitro and in vivo. The cytokine, tumor necrosis factor-alpha (TNF-alpha), is produced by macrophages in response to bacterial endotoxins and wear particles, and it increases osteoclast activity resulting in bone resorption and implant loosening. The phosphoinositol-3-kinase (PI3K)-Akt intracellular signal transduction pathway contributes to cytokine production in response to soluble endotoxin. We investigated the role of the PI3K-Akt pathway in the production of TNF-alpha in response to wear particles with adherent endotoxin and so-called endotoxin-free wear particles. METHODS: Cultured RAW264.7 murine macrophages were incubated with titanium particles with adherent endotoxin or with endotoxin-free titanium particles in the presence and absence of specific inhibitors of PI3K (LY294002) or Akt (SH-5). Akt activation was assessed with use of Western blot. TNF-alpha production was measured with use of enzyme-linked immunosorbent assay. Cytotoxicity was determined by measuring lactic dehydrogenase release. RESULTS: Titanium particles with adherent endotoxin increased Akt activation, whereas endotoxin-free titanium particles did not. The PI3K inhibitor reduced TNF-alpha production by 70% in response to titanium with adherent endotoxin without increasing cytotoxicity. Similarly, the Akt inhibitor reduced TNF-alpha production by 83% in response to titanium particles with adherent endotoxin without increasing cytotoxicity. High concentrations of endotoxin-free titanium particles resulted in a small delayed increase in TNF-alpha production that was completely blocked by the PI3K inhibitor. CONCLUSIONS: Inhibition of the PI3K-Akt pathway reduces macrophage TNF-alpha production in response to titanium particles with adherent endotoxin and endotoxin-free particles in vitro.     

The osteoclastogenic molecules RANKL and RANK are associated with periprosthetic osteolysis

Extensive osteolysis adjacent to implants is often associated with wear particles of prosthetic material. We have investigated if RANKL, also known as osteoprotegerin ligand, osteoclast differentiation factor or TRANCE, and its natural inhibitor, osteoprotegerin (OPG), may be important in controlling this bone loss. Cells isolated from periprosthetic tissues containing wear particles expressed mRNA encoding for the pro-osteoclastogenic molecules, RANKL, its receptor RANK, monocyte colony-stimulating factor (M-CSF), interleukin (IL)-1beta, tumour necrosis factor (TNF)alpha, IL-6, and soluble IL-6 receptor, as well as OPG. Osteoclasts formed from cells isolated from periprosthetic tissues in the presence and absence of human osteoblastic cells. When osteoclasts formed in the absence of osteoblastic cells, markedly higher levels of RANKL mRNA relative to OPG mRNA were expressed. Particles of prosthetic materials also stimulated human monocytes to express osteoclastogenic molecules in vitro. Our results suggest that ingestion of prosthetic wear particles by macrophages results in expression of osteoclast-differentiating molecules and the stimulation of macrophage differentiation into osteoclasts.     

Effect of PMMA particles and movement on an implant interface in a canine model

The pathogenesis of aseptic loosening of total joint prostheses is not clearly understood. Two features are associated with loosened prostheses, namely, particulate debris and movement of the implant. While numerous studies have evaluated the cellular response to particulate biomaterials, few have investigated the influence of movement of the implant on the biological response to particles. Our aim was therefore to test the hypothesis that excessive mechanical stimulation of the periprosthetic tissues induces an inflammatory response and that the addition of particulate biomaterials intensifies this. We allocated 66 adult Beagle dogs to four groups as follows: stable implants with (I) and without (II) particulate polymethylmethacrylate (PMMA) and moving implants with (III) and without (IV) particulate PMMA. They were then evaluated at 2, 4, 6, 12 and 24 weeks. The stable implants were well tolerated and a thin, fibrous membrane of connective tissue was observed. There was evidence of positive staining in some cells for interleukin-6 (IL-6). Addition of particulate PMMA around the stable implants resulted in an increase in the fibroblastic response and positive staining for IL-6 and tumour necrosis factor-alpha (TNF-alpha). By contrast, movement of the implant resulted in an immediate inflammatory response characterised by large numbers of histiocytes and cytokine staining for IL-1beta, TNF-alpha and IL-6. Introduction of particulate PMMA aggravated this response. Animals with particulate PMMA and movement of the implant have an intense inflammatory response associated with accelerated bone loss. Our results indicate that the initiation of the inflammatory response to biomaterial particles was much slower than that to gross mechanical instability. Furthermore, when there was both particulate debris and movement, there was an amplification of the adverse tissue response as evidenced by the presence of osteolysis and increases in the presence of inflammatory cells and their associated cytokines.     

Third-body wear of highly cross-linked polyethylene in a hip simulator

The wear performance of a radiation cross-linked melted ultrahigh-molecular-weight polyethylene (UHMWPE) articulating against 28-mm cobalt chrome femoral heads in the presence of third-body particulate debris was investigated in a hip simulator and compared with the wear of conventional UHMWPE. Particles of aluminum oxide or bone cement containing barium sulfate were added to the serum. In the presence of aluminum oxide particles, the incremental wear rates of conventional UHMWPE averaged as high as 149 +/- 116 mg/million cycles compared with 37 +/- 38 mg/million cycles for the highly cross-linked components. The difference in the average weight loss was statistically significant at P <.01. With bone cement particles, the conventional UHMWPE components had an average incremental wear rate of 19 +/- 5mg/million cycles, and the wear rate of the highly cross-linked UHMWPE components was 0.5 +/- 0.7 mg/million cycles.     

Cytokine and prostaglandin E2 release from leukocytes in response to metal ions derived from different prosthetic materials: an in vitro study

Cytokines produced by leukocytes in the periprosthetic membranes surrounding joint replacements have been implicated as causal agents in osteolysis and prosthetic loosening. In this study, we used an in vitro leukocyte culture system to monitor the response of leukocytes to various metal ions and their possible roles in the mechanism of aseptic loosening. Human peripheral leukocytes were isolated and incubated with various concentrations of Co2+, Cr3+, and Ti3+ ions. Leukocyte cell counts and the levels of the tumor necrosis factor-alpha (TNF-alpha), interleukin-1 (IL-1), interleukin-6 (IL-6) and prostaglandin E2 (PGE2) released into the media were analyzed at 1 h, 3 h, and 1, 3, and 7 day intervals. The results showed that adding different metal ions into leukocyte cultures did not affect the cell counts. Exposure of leukocytes to Co2+ ion increased the release of TNF-alpha, IL-6, and PGE2. Exposure of leukocytes to Cr3+ ion did not increase the release of TNF-alpha but increased the secretion of IL-6 and PGE2. In contrast, exposure of the leukocytes to Ti3+ ions was associated with a decrease in the release of TNF-alpha and PGE2 and a minimal change in IL-6 noted after 7 days' culture. The present study elucidated the possible mechanisms involved in periprosthetic osteolysis and the inflammatory response of human leukocytes to metal ions. We found that cobalt ion is the most potent stimulant for cytokines and prostaglandin secretion by leukocytes. This elucidation, in combination with other efforts to reduce the generation of wear debris and metal ions, may improve the longevity of orthopedic implants.     

Polymyxin B antagonizing biological activity of lipopolysaccharide

Objective ： To investigate the mechanism of polymyxin B （ PMB ） antagonizing the biological activity of Hipopolysaccharide （LPS）. Methods： The affinity of PMB for LPS and lipid A was assayed by biosensor, and the neutralization of PMB for LPS （2 ng/ml ） was detected by kinetic turbidimetric limnins test. The releases of TNF-α and IL-6 in murine peritoneal macrophages （PMφ） after exposure to LPS （ 100 ng/ml） were detected, and the expression levels of TLR4, TNF-α and IL-6 mRNA in PMφ induced by LPS （100 ng/ml） were measured by RT-PCR. Results： PMB had high-affinity to LPS and lipid A with dissociation equilibrium constants of 18.9 nmol/L and 11.1 nmol/L, respectively, and neutralized LPS in a dosedependent manner. Furthermore, PMB could markedly inhibit the expressions of TLR4, TNF-α and IL-6 mRNA and the release of cycokines in LPS-stimniated murine PMφ. Conclusions： PMB neutralizes LPS and inhibites the expression and release of cycokines in macrophages, in which the affinity of PMB for lipid A plays an important role.     

Adherent endotoxin on orthopedic wear particles stimulates cytokine production and osteoclast differentiation.

Aseptic loosening of orthopedic implants is thought to be caused primarily by osteoclast differentiation induced by bone resorptive cytokines produced in response to phagocytosis of implant-derived wear particles. This study examined whether adherent endotoxin on the wear particles is responsible for inducing osteoclast differentiation as well as production of interleukin-1beta (IL-1beta), IL-6, and tumor necrosis factor a (TNF-alpha). Removal of adherent endotoxin almost completely inhibited the responses to titanium (Ti) particles by both murine marrow cells and human peripheral blood monocytes. In vivo experiments showed that endotoxin removal reduced particle-induced osteolysis by 50-70%. Addition of lipopolysaccharide (LPS) to the "endotoxin-free" particles restored their ability to induce cytokine production and osteoclast differentiation in vitro. Moreover, marrow cells from mice that are hyporesponsive to endotoxin because of mutation of Toll-like receptor 4 induced significantly less cytokine production and osteoclast differentiation in response to Ti particles with adherent endotoxin than did marrow cells from normoresponsive mice. This mutation also resulted in significantly less particle-induced osteolysis in vivo. Taken together, these results show that adherent endotoxin is involved in many of the biological responses induced by orthopedic wear particles and should stimulate development of new approaches designed to reduce the activity of adherent endotoxin in patients with orthopedic implants.     

Activation of cytokine synthesis by systemic infusions of lipopolysaccharide and peptidoglycan in a porcine model in vivo and in vitro

BACKGROUND: The incidence of gram-positive and mixed bacterial infections in surgical patients has increased, and there has been an alarming rise in the number of drug-resistant bacteria. Peptidoglycan (PepG) is a cell wall component of gram-positive bacteria that stimulates inflammatory responses both ex vivo and in vivo. The systemic effects of PepG on inflammation have not been studied in a large animal model. METHODS: Anesthetized pigs were subjected to 8-h continuous intravenous infusions of lipopolysaccharide (LPS) (4 mcg/kg/h), PepG (40 mcg/kg/h), LPS plus PepG, or saline. The concentrations of tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, IL-6, IL-8, and IL-10 were measured in the plasma prior to infusion (time 0) and thereafter every second hour until the end of the experiments. Heparinized whole blood samples drawn at time 0 and after a 6-h infusion of LPS or PepG were incubated ex vivo with PepG (10 mcg/mL), LPS (10 ng/mL), or a combination of PepG and LPS to study the immunologic consequences of systemic inflammation. Concentrations of TNF-alpha, IL-8, and IL-1beta were measured in the supernatant liquids. RESULTS: In vivo, there was transient upregulation of TNF-alpha after infusion of LPS, PepG, or the combination. Interleukin-6 and IL-8 were upregulated by LPS but not by PepG. In vitro studies of whole blood obtained at time 0 revealed a synergistic effect of LPS and PepG on the release of TNF-alpha. Incubation of whole blood obtained after 6 h of infusion of LPS or PepG revealed tolerance and cross-tolerance between the two bacterial components in the induction of TNF-alpha, IL-8, and IL-1beta. CONCLUSIONS: Peptidoglycan is a potent inducer of TNF-alpha in this large animal model. Peptidoglycan and LPS synergized to increase the formation of the proinflammatory cytokine TNF-alpha. The study demonstrates for the first time the development of tolerance and cross-tolerance between LPS and PepG in a large animal model. These phenomena could be of importance for the signs and symptoms of sepsis.     

Polyethylene particles from a hip simulator cause (45)Ca release from cultured bone

Periprosthetic osteolysis is a dominant factor in the success or failure of total hip prostheses. Polyethylene wear debris has been implicated in the process of bone resorption and subsequent implant loosening. The present study is the first to examine the effect of ultra high molecular weight polyethylene (UHMWPE) wear debris produced by a hip simulator on calvarial bone resorption in vitro. (45)Ca release was measured in cultured mouse calvarial bone samples. Although short-term exposure to UHMWPE particles (2 h) decreased (45)Ca release, longer-term exposure for 1-2 days increased release in a dose-dependent manner. After one-day exposure to 7.5 x 10(6) particles per mL, 18% more (45)Ca was released from cultured calvarial bone than from control samples. It was concluded that UHMWPE wear particles either directly or indirectly stimulated osteoclasts to activate bone resorption. Polyethylene wear debris contributes to the osteolytic process at the bone-implant interface.     

Cytokine response of human macrophage-like cells after contact with polyethylene and pure titanium particles.

The aim of this study was to establish a human macrophage cell culture system to examine the effect of polyethylene (PE) and titanium particles on cytokine release by macrophage-like cells (MLC) and to quantify this response with respect to the nature and concentration of particles. Human monocytic leukemia cells were differentiated under standard conditions with vitamin D3 and granulocyte macrophage-colony-stimulating factor. Cells were characterized by fluorescence-activated cell-sorter Scan of CD 14 expression analysis as well as a phagocytosis test exploiting fluorescence-labeled particles of bacteria] walls. To achieve a relevant contact between the floating PE particles (approximately 1 microm in size) and MLC, a rotation device was used (15 rotations/min) during incubation. The same was done with the titanium particles. Cell culture supernatants were then analyzed for interleukin (IL)-1beta, IL-8, and tumor necrosis factor (TNF)-alpha using the enzyme-linked immunosorbent assay technique in the absence or presence of particles. Rotation of incubated MLC alone did not influence the secretion of TNF-alpha, but it enhanced secretion of IL-1beta and IL-8 about 30-fold compared to background levels. Both PE and titanium particles significantly enhanced MLC cytokine release, the amount of which depended on the concentration of particles. Using 40 X 10(8) PE particles (0.7 x 10(8) titanium particles) and 10(6) MLC, the maximal release of IL-1beta was about 20-fold (7-fold titanium particles) higher than that of the rotating control sample. The stimulation of IL-8 release was 4-fold (3-fold titanium particles) and of TNF-alpha. 300-fold (170-fold titanium particles) compared to controls. MLC were viable (>90% cell survival) at concentrations less than 108 x 10(8) polyethylene particles per 10(6) MLC and 16 x 10(8) titanium particles per 10(6) MLC. Rotation per se as well as exposure to increasing concentrations of PE and titanium particles stimulates cytokine release (TNF-alpha, IL-1beta, IL-8) by macrophages in vitro. This in vitro model resembles the in vivo situation near arthroplasties, where implant particles make contact with inflammatory cells, such as macrophages. Cytokine release by macrophages may impair osteoblast function as well as stimulate bone resorption by osteoclasts and macrophages, thereby causing aseptic loosening of arthroplasties. Our in vitro model provides a reproducible human cell system that might shed light on the pathogenesis of particle disease and might serve as a reproducible in vitro test system for the biocompatibility of foreign materials.