Link N suppresses interleukin-1ß-induced biological effects on human osteoarthritic cartilage.

Osteoarthritis (OA) is a disease of diarthrodial joints associated with extracellular matrix proteolytic degradation under inflammatory conditions, pain and disability. Currently, there is no therapy to prevent, reverse or modulate the disease course. The present study aimed at evaluating the regenerative potential of Link N (LN) in human OA cartilage in an inflammatory milieu and determining if LN could affect pain-related behaviour in a knee OA mouse injury model. Osteo-chondro OA explants and OA chondrocytes were treated with LN in the presence of interleukin-1ß (IL-1ß) to simulate an osteoarthritic environment. Quantitative von Frey polymerase chain reaction and Western blotting were performed to determine the effect of LN on matrix protein synthesis, catabolic enzymes, cytokines and nerve growth factor expression. Partial medial meniscectomy (PMM) was performed on the knee of C57BL/6 mice and, 12 weeks post-surgery, mice were given a 5 µg intra-articular injection of LN or phosphate-buffered saline. A von Frey test was conducted over 24 h to measure the mechanical allodynia in the hind paw. LN modulated proteoglycan and collagen synthesis in human OA cartilage through inhibition of IL-1ß-induced biological effects. LN also supressed IL-1ß-induced upregulation of cartilage-degrading enzymes and inflammatory molecules in OA chondrocytes. Upon investigation of the canonical signalling pathways IL-1ß and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), LN resulted to significantly inhibit NF-κB activation in a dose-dependent manner. In addition, LN suppressed mechanical allodynia in an OA PMM mouse model. Results supported the concept that LN administration could provide therapeutic potential in OA.

Induction of macrophage apoptosis by ceramic and polyethylene particles in vitro.

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.

Evidence for a humoral mechanism for enhanced osteogenesis after head injury.

The rate of fracture-healing is accelerated and abundant callus develops in patients who have a head injury and fractures. The mechanism underlying this is unclear. We studied the possibility that increased circulating growth factors or circulating factors that stimulate local release of growth factors mediate the increased osteogenesis. Samples of serum were obtained from thirty-two subjects: patients who had a head injury alone, those who had a head injury and fractures of the lower extremities, those who had only fractures, and control subjects who had neither a head injury nor a fracture. Severe head injury was defined as that producing coma of at least three days' duration. Growth-factor activity was determined by assessing the effect of serum on the incorporation of [3H]thymidine and on cell counts in primary cultures of osteoblastic cells from the calvaria of fetal rats. Samples of serum from the two groups of patients who had a head injury had higher mitogenic activity and produced a greater increase in the number of cells than did the samples from the other two groups. The mean levels of activity were not statistically different between the first two groups or between the patients who had fractures only and the control subjects. Dilution studies showed that increased mitogenic activity in the serum from the patients who had a head injury was dose-dependent. In three patients in whom it was studied, the mitogenic activity peaked approximately thirty-seven days after the head injury was sustained.

Analysis of cementless implants using interface nonlinear friction--experimental and finite element studies.

Measured interface nonlinear friction properties are used to develop models to study the short-term fixation response of smooth- and porous-surfaced posts, bone screws, and plates fixed with and without posts/screws. Experimental studies are carried out to validate the model predictions and identify the relative role of posts and screws in fixation of a plate on a polyurethane block under symmetric/eccentric axial compression loads. The idealized Coulomb's friction is also used for the sake of comparison. The incorporation of measured nonlinear, rather than the idealized Coulomb, friction is essential to compute realistic results. For plate fixation, the experimental and finite element results show that the screw fixation yields the stiffest response followed by the smooth- and then porous-coated post fixation. For example, under 1000 N eccentric axial compression, the edge of the plate opposite the loaded edge is measured to lift by 1147 +/- 72, 244 +/- 38, or 112 +/- 28 microns, respectively, for the cases with no fixation, with smooth-surfaced posts, or with screws. The corresponding models predict, respectively, values of 1538, 347, or 259 microns and also 556 microns for the plate fixed with porous coated posts. The satisfactory agreement between numerical and experimental results confirms the importance of proper interface modelling for the analysis of posts, screws, and complex fixation systems. This becomes further evident when considering cementless implants in which the bone-implant interface exhibits relatively large displacements as the maximum resistance force is reached. The developed models can be used to investigate the post-operative short-term stability of various cementless implant designs.

Fixation pull-out response measurement of bone screws and porous-surfaced posts.

Screws and posts are used in various implant designs to contribute to the short- and long-term fixation stability of artificial joints. This study was undertaken to measure the detailed pull-out load-displacement response of bone screws, beaded porous coated posts, and smooth-surfaced posts in both proximal tibial cancellous bone and polyurethane material under monotonic static and repetitive cyclic loadings. The effect of a number of parameters such as insertion site on the proximal tibia, rate of displacement, insertion depth, outside diameter, drill size, repetitive loading and boundary conditions were studied. Bone screws resisted significantly larger loads than posts of the same size. Smooth-surfaced posts demonstrated much larger (about twice) pull-out forces than beaded porous coated posts of the same size. The pull-out force in the proximal tibia was markedly larger at the medial region followed by the lateral region. The central region exhibited the least force. The resistance of screws and posts diminished with repetitive cyclic loads/displacements, especially when these were larger than 50% of their respective values at pull-out force evaluated under monotonic static loading conditions. The smooth-surfaced posts showed superior performance in maintaining their resistance in fatigue than did the porous coated posts and bone screws. The pull-out force was found to also depend on the pull-out material arrangement and boundary conditions. Pull-out results measured with a specific test design should not, therefore, be compared with those performed using different design configurations.

Fresh osteochondral allografts for post-traumatic defects in the knee. A survivorship analysis.

Fresh osteochondral allografts were used to repair post-traumatic osteoarticular defects in 92 knees. At the time of grafting, varus or valgus deformities were corrected by upper tibial or supracondylar femoral osteotomies. A survivorship analysis was performed in which failure was defined as the need for a revision operation or the persistence of the pre-operative symptoms. There was a 75% success rate at five years, 64% at ten years and 63% at 14 years. The failure rate was higher for bipolar grafts than for unipolar and the results in patients over the age of 60 years were poor. The outcome did not depend on the sex of the patient and the results of allografts in the medial and lateral compartments of the knee were similar. Careful patient selection, correction of joint malalignment by osteotomy, and rigid fixation of the graft are all mandatory requirements for success. We recommend this method for the treatment of post-traumatic osteochondral defects in the knees of relatively young and active patients.

Cytotoxicity and macrophage cytokine release induced by ceramic and polyethylene particles in vitro.

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 microns). Al2O3 was compared with ZrO2 at 0.6 micron and with HDP at 4.5 microns. 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 micron, 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-alpha also increased with the size and concentration of Al2O3 particles, reaching 195% of control (165 pg/ml v 84 pg/ml) at 2.4 microns and 350 particles per cell (p < 0.05). Release of TNF-alpha was higher with HDP than with Al2O3 particles at 4.5 microns. However, we did not detect any significant difference in the release of TNF-alpha between Al2O3 and ZrO2 at 0.6 micron (p > 0.05). We saw no evidence of release of interleukin-1 alpha or interleukin-1 beta after exposure to ceramic or HDP particles.

Fixation response of two cementless tibial implants under static and fatigue compression loading.

The short- and long-term success of tibial cementless implants depend on the initial fixation stability often provided by posts and screws. Excessive relative interface motions are known to inhibit bone ingrowth and, hence, biologic fixation. In the present study, the stability of two cementless tibial implants under static and cyclic loading was studied. An apparatus was developed to perform compression tests on tibial components fixed by screws or posts onto resurfaced tibiae. Motion transducers were placed to record liftoff and subsidence displacements. A load of 1000 N was eccentrically applied on the medial plateau via the femoral component. The response was measured under static and repetitive loading up to 4000 cycles at 1 Hz. Experimental results showed that the screw fixation yielded the stiffer response and, hence, provided greater initial stability. The tibial component fixed with posts demonstrated larger lift off displacements. Comparison of measured results of static tests before and after fatigue indicated that the load-displacement behavior was not significantly affected by the cyclic fatigue loading under 1000 N up to 4000 cycles at 1 Hz. Nevertheless, much larger permanent displacements were recorded for the post fixation systems. In contrast to screw fixation, posts fixation systems demonstrated marked deterioration under larger compression loads of approximately 2000 N.

Deviations between intra-operative navigation data and post-operative weight-bearing X-rays.

Several studies have shown that computer-navigated TKA reduces the rate of outliers. Thirty-one consecutive patients were operated on by the same surgeon using a computer assisted navigation system. Data collected by the system included the final mechanical axis of the extremity (HKA angle) and the coronal angle of the tibial and femoral implants. These same values were measured using CAD software on full weight-bearing long X-rays taken 6 weeks post-surgery. Deviations were observed when X-ray measurements were compared to intra-operative data collected from the navigation system. A statistically significant difference was found in the tibial cut (1.29 degrees +/- 1.35 degrees; p < 0.0001) and in the HKA (1.59 degrees +/- 2.36 degrees; p = 0.0007). Outliers of more than 3 degrees were observed in the coronal plane of the tibial implant in 9.6% of patients, in the coronal plane of the femoral implant in 6.4% of patients, and in the HKA angle of 29% of patients. Our results indicate that the use of navigation alone is insufficient to prevent outliers beyond an acceptable range of 3 degrees .

Effect of a Type II Collagen Fragment on the Expression of Genes of the Extracellular Matrix in Cells of the Intervertebral Disc.

Knowledge of factors regulating the turnover, repair, and degeneration of the intervertebral disc (IVD) is lacking. Although type II collagen (CII) fragments accumulate in the degenerative IVD, little is known of how they affect the degenerative process. A better understanding of the cellular interactions with fragments of matrix molecules are a key factor in promoting therapies for degenerative disc diseases. In the present study, we have investigated the effect of the CII (245-270) peptide on the expression of matrix molecules, proteinases, and interleukin genes in cells of the IVD. Cells isolated from the nucleus pulposus (NP) and annulus fibrosus (AF) of adult bovine tails were cultured up to 8 days in the absence (control) or presence of the CII (245-270) peptide. RT-PCR was used to analyze the expression of the different genes. Exposure of these cells to the CII (245-270) peptide led to a transient up-regulation of the aggrecan gene in AF cells while this up-regulation was maintained for a longer time in NP cells. The fragment also enhanced a transient up-regulation of the type II collagen gene in AF cells but had no effect in NP cells. The peptide enhanced transiently the expression of matrix metalloproteinase (MMP)-1 and cathepsin K genes in both AF and NP cells whereas it increased MMP-13 expression only in NP cells. The peptide up-regulated tissue inhibitor of metalloproteinase (TIMP)-1, TIMP-2, and TIMP-3 gene expression on day 1 in AF cells but had very little effect on their expression in NP cells. Finally, the CII (245-270) peptide had no effect on IL-6 expression while IL-1alpha was not expressed in these cells. In conclusion, our results showed that the CII (245-270) peptide differentially alter the expression of genes in bovine AF and NP cells and suggest that degradation products of collagen may be involved in the regulation of IVD homeostasis.

Experience of total hip replacement and its effect on recovery after second hip replacement.

OBJECTIVE: To determine whether the knowledge gained through total hip replacement (THR) influences the time to recovery after a second THR. DESIGN: A retrospective study of data gathered prospectively on the recovery course of patients from their preoperative visit to 1 year after surgery. SETTING: A university teaching hospital. PATIENTS: Sixty-nine patients underwent bilateral hip replacements at the Royal Victoria Hospital, Montreal, between May 1985 and May 1992. The inclusion criteria were: (a) similar diagnoses and prostheses bilaterally, (b) time between the two THRs of less than 3 years, (c) both replacements carried out by the same surgeon and (d) attendance at the arthroplasty clinic. Forty-one patients did not fulfil the criteria for inclusion, leaving 28 patients having 56 THRs for the study. An additional 28 patients who had unilateral hip arthroplasty were randomly chosen on the basis of sex and age to act as the control group. INTERVENTION: Bilateral cementless THR. MAIN OUTCOME MEASURES: Harris hip score at one preoperative visit and postoperatively at 6 weeks, 3 months, 6 months and 1 year. RESULTS: There were no significant differences in measures of recovery between the initial and the subsequent hip replacement at any of the time intervals. Significant differences were found only between the various time periods. CONCLUSIONS: Time and not familiarity dictated progress toward recovery for patients who underwent bilateral THR with cementless prostheses. Randomized clinical trials, assessing whether time to recovery, ultimate outcome and cost-effectiveness are affected by preoperative teaching programs, are necessary to further elucidate this important issue.

Biomechanical response of the passive human knee joint under anterior-posterior forces.

OBJECTIVE: To investigate the detailed biomechanics of the passive tibiofemoral knee joints in full extension under anterior/posterior drawer forces of up to 400 N. DESIGN: A nonlinear three-dimensional finite element model of the entire human tibiofemoral joint consisting of bony structures, their articular cartilage layers, menisci, and four principal ligaments was utilized. BACKGROUND: The mechanics of the knee joint, specially under drawer forces, have extensively been investigated. Despite all these works, the detailed joint biomechanics, specially the role of boundary conditions, load transmission through menisci/articular cartilage layers, and coupling between menisci and cruciate ligaments are not yet quantified. METHODS: Nonlinear elastostatic analyses were carried out considering the tibiofemoral joint at full extension under anterior and posterior loads of up to 400 N applied either to the tibial or the femoral shaft. Cases with various boundary conditions, cruciate ligament deficiency (anterior or posterior), and total unilateral meniscectomy (medial or lateral) were analysed. RESULTS: In addition to the total primary anterior-posterior motion of about 9 mm at +/-400 N, significant coupled external tibial rotations of about 9 degrees and 10 degrees were computed under 400 N femoral posterior and anterior forces, respectively. The response was influenced by the manner of loading and boundary conditions. The anterior cruciate ligament and posterior cruciate ligament were the primary restraints to femoral posterior and anterior drawer forces, respectively. Section of either of these ligaments drastically increased the joint anterior-posterior motion. In the absence of cruciates, the collaterals became the primary restraints in both anterior-posterior forces. In this case, the tibial plateaus, specially the medial one in the anterior cruciate ligament-deficient joint, experienced much larger compressive forces. In addition to causing an increase in joint primary anterior-posterior laxity and anterior cruciate ligament forces, medial meniscectomy substantially increased coupled tibial external rotation, forces on the lateral plateau, and stresses in the articular cartilage of the lateral plateau. RELEVANCE: Our results suggest an increased role for the medial meniscus in the anterior cruciate ligament-deficient joint. Lateral meniscectomy had much smaller effects on results than the medial one. The success of any anterior cruciate ligament replacement or meniscal transplantation in the restoration of the joint stability and the protection of the articular cartilage against excessive stresses depends on the coupling between the anterior cruciate ligament and medial meniscus. Absence of any of these components would drastically influence the joint response.

Finite element analysis of human knee joint in varus-valgus.

OBJECTIVE: The overall response, load transmission, role of ligaments, and state of stress in various components under varus-valgus moments in the intact and collateral-deficient tibiofemoral joint are investigated. DESIGN: A non-linear finite element model consisting of bony structures (tibia and femur), their articular cartilage layers, medial and lateral menisci and four primary ligaments (cruciates and collaterals) is utilized. BACKGROUND: Valgus and varus stresses are among the primary mechanisms of injury to knee ligaments. Several in vitro studies have investigated the role of ligaments in resisting such loads and on the way deficiency in either collateral may affect the response. METHODS: Cartilage layers are isotropic while menisci are non-homogeneous composite. The articulation of cartilage layers with each other and with the intervening menisci and the wrapping of the medial collateral ligament around the tibial edge are treated as large displacement frictionless contact problems. The non-linear elastostatic response of the joint at full extension is computed under varus-valgus moments applied to the femur with the tibia fixed. Cases simulating deficiency in collaterals and constraint on femoral axial rotation are also studied. RESULTS: The response is non-linear with large coupled axial rotations, internal in varus and external in valgus. In intact and collateral-deficient states, the joint shows varus or valgus openings so that the articulation occurs at one plateau only, medial in varus and lateral in valgus. Large tensile forces in cruciates in collateral-cut models generate higher compression penalty on the loaded plateau. CONCLUSIONS: Collaterals are the primary load-bearing structures; their absence would substantially increase primary laxities, coupled axial rotations, forces in cruciates, and articular contact forces. Good agreement with measurements is found. RELEVANCE: Detailed knowledge of joint biomechanics is essential in the diagnosis, prevention and treatment of observed disorders. Absence of collateral ligaments increases the loads in cruciates and contact stresses transmitted through cartilage layers and menisci, and thus places the affected components at more risk, especially when varus-valgus is accompanied by other modes of loading as well.

Flow cytometric analysis of macrophage response to ceramic and polyethylene particles: effects of size, concentration, and composition.

Using the J774 macrophage cell line, we designed an in vitro model to analyze by flow cytometry the effects of size, concentration, and composition of ceramic (Al2O3 and ZrO2) and high density polyethylene (HDP) particles on phagocytosis and cell mortality. Inflammatory mediator (TNF-alpha) also was measured by ELISA. Kinetic studies revealed that phagocytosis of the particles begins very early after cell exposure, increasing with time and particle concentration and reaching a plateau after 15 h. This implies that the optimum period to evaluate cellular response to particulate debris is between 15 and 24 h of incubation. Results also showed that phagocytosis increases with concentration for particles up to 2 microns. For larger particles (up to 4.5 microns), phagocytosis seems to reach a plateau independent of size and concentration, which suggests a saturation of phagocytosis that is most likely dependent on overall particle volume ingested. We did not detect any significant difference in phagocytosis between Al2O3 and ZrO2 at 0.6 microns. Al2O3 seems to be more easily phagocytosed than HDP at the same size (4.5 microns) and concentrations. Cytotoxicity studies revealed that macrophage mortality increases with particle size and concentration for sizes greater than 2 microns. Smaller particles (0.6 microns) cause cell mortality only at higher concentrations (from 1,250 particles per cell), but the mortality is still very low (10%). No significant difference in cell mortality and TNF-alpha release was found between Al2O3 and ZrO2. Effects of Al2O3 and HDP at 4.5 microns were compared by measuring TNF-alpha release. Results showed that TNF-alpha release increases with particle concentrations and is higher with HDP than with Al2O3.

Apoptosis in interface membranes of aseptically loose total hip arthroplasty.

The terminal events leading to periprosthetic osteolysis are multifactorial in nature and modulation of this process after the stage of osteolytic mediator release has been futile. Recently, the demonstration of the ability of bisphosphonates to inhibit bone resorption that is mediated by particle-stimulated macrophages and their induction of osteoclast apoptosis suggests a potent area for modulation of osteolysis at the prosthesis-bone interface. The purpose of this study was to determine the mode of cell death that occurs at the osteolytic interface of failed total hip arthroplasty (THA). TUNEL staining, DNA laddering, and immunodetection of poly(ADP-ribose)polymerase (PARP) protein were used to identify the presence of apoptosis in interface membranes from 25 patients aged 28-88 years old (mean, 58 years) harvested at the time of hip revision surgery. Our results demonstrated positive TUNEL stain in 100% of specimens with an average 37% of cells (range 12-60%) positively stained for TUNEL whereas less than 8% of control tissue cells showed positive staining. DNA laddering, a characteristic feature of apoptotic cells, was observed in 82% (28/34) of specimens studied at both the acetabular and femoral side of aseptically loose THAs. No laddering was observed in control tissues. Finally, using Western blot analysis, we observed the appearance of the 89 kDa PARP fragment associated with apoptosis in 92% of specimens (30/33). Our results demonstrate the presence of apoptotic cell death in interface membranes of THAs suggesting that apoptosis-related events are indeed associated with periprosthetic osteolysis and could serve as a specific target point for therapeutic modulation.

Cytotoxic and apoptotic effects of cobalt and chromium ions on J774 macrophages - Implication of caspase-3 in the apoptotic pathway.

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.

Hip pinning, past and present: Richards' compression-screw fixation versus Ender's nailing.

The authors trace the operative treatment of intertrochanteric hip fractures from the earliest to the most modern methods and describe how current fixation devices evolved. The operative and perioperative morbidity, mortality and outcome in 57 patients treated by Ender's nailing were compared with those of 51 patients treated by Richards' compression-screw fixation. Results were comparable to those found in an extensive review of the literature except for a higher mortality in the group managed by Ender's nailing. The authors conclude that both methods of treatment are acceptable and that the ultimate choice of fixation device should be based on the surgeon's experience tailored to the individual patient's needs.

Revision total hip arthroplasty with osseous allograft reconstruction. A clinical and roentgenographic analysis.

One hundred twelve irradiation-sterilized, -70 degrees frozen allografts have been used in 72 patients who were available for follow-up study in a prospective analysis of allograft revision total hip arthroplasty. Clinical objectives were achieved in 85% of patients with a follow-up period ranging from six to 72 months. Based on clinical and roentgenographic analysis, specific recommendations are made to further improve the success rate of revision total hip arthroplasty associated with protrusio, shelf, acetabular, calcar, and large fragment femoral allografts.

Effects of digestion protocols on the isolation and characterization of metal-metal wear particles. II. Analysis of ion release and particle composition.

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.

Effects of digestion protocols on the isolation and characterization of metal-metal wear particles. I. Analysis of particle size and shape.

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 (<1200 nm) were generated by dry abrasion of CoCrMo alloy against itself and small particles (<300 nm) were generated by hip simulator testing of a metal-metal implant pair in the presence of either distilled-deionized water or a 95% bovine serum solution. The reagents changed particle size and to a lesser extent particle shape. For both large particles and small particles generated in water, the changes in size were more extensive after alkaline than after enzymatic protocols and increased with alkaline concentration and time in solution, up to twofold at 2 h and threefold at 48 h. However, when isolating particles from 95% serum, an initial protective effect of serum proteins and/or lipids was observed. Because of this protective effect, there was no significant difference in particle size and shape for both oval and needle-shaped particles after 2 h in 2N KOH and after enzymatic treatments. However, round particles were significantly smaller after 2 h in 2N KOH than after enzymatic treatments. Particle composition may also have been affected by the 2N KOH treatment, as suggested by a difference in particle contrast under TEM, an issue examined in detail in Part II.