Evaluation of the effect of seven acrylic bone cements on erythrocytes and plasmatic phase of coagulation

The haemolytic activity and the effect on the plasmatic phase of coagulation of seven bone cements were evaluated (Sulfix-60 from Sulzer Orthopedic Inc., a bone cement at low viscosity from Zimmer, a bone cement dough-type from Zimmer, Palacos R from Merck, CMW1, CMW2 and CMW3 from DePuy International Ltd.). Haemolytic activity was tested by adding the cement extracts in phosphate buffered saline to a suspension of erythrocytes. After 4 h incubation at 37 degrees C, the haemoglobin concentration was determined on the supernatants by colorimetric method. The effect on the plasmatic phase of coagulation was tested by adding the cement extracts in saline to human plasma. After 30 min incubation at room temperature activated partial thromboplastin time (APTT) was determined. All extracts induced non-significant variations of haemoglobin concentration and APTT. It was concluded that the tested cement extracts do not induce haemolysis and do not activate the intrinsic pathway of coagulation, at least in the tests that were performed.     

Comparison of the mechanical properties of Simplex P, Zimmer Regular, and LVC bone cements

The mechanical properties of the three cement preparations most widely used in the United States were compared by conducting tensile and fatigue tests on Simplex P, LVC, and Zimmer Regular bone cements. Specimens of all three cement preparations were prepared for mechanical testing with and without centrifugation of the cement immediately after mixing. Although the results of the tensile testing revealed a few specific instances of significant differences in the tensile properties of the three cement preparations, there was no consistent evidence that one cement was superior in tension to the others. However, the fatigue properties of Simplex P were consistently and significantly superior to the fatigue properties of both LVC and Zimmer Regular bone cements. Centrifugation of the cement immediately after mixing significantly improved both the tensile and fatigue properties of all three bone cements. However, the fatigue strength of centrifuged Simplex P was substantially and significantly superior to the fatigue strength of the centrifuged LVC and Zimmer Regular bone cements. Since in total joint replacements bone cement is subjected to cyclic loading, these data suggest that centrifuged Simplex P is a preferable bone cement to LVC and to Zimmer Regular cement with or without centrifugation.     

Influence of antibiotic impregnation on the fatigue life of Simplex P and Palacos R acrylic bone cements, with and without centrifugation

The fatigue properties of Simplex P and Palacos R bone cements were compared to their antibiotic impregnated counterparts AKZ* and Palacos R with gentamycin. The effect of porosity reduction by centrifugation of all four cement types was also assessed. Fifteen specimens of each cement type were prepared according to manufacturer's instructions and 15 additional specimens of each cement type were prepared by mixing the powder with chilled monomer (0 degrees C) and then centrifuging the cement immediately after mixing. Fifteen fully reversed tension-compression fatigue tests were performed at 15 MPa in stress control for each cement preparation in vitro while simulating the in vivo state (37 degrees C and 100% humidity). The number of cycles to failure were recorded. There was no significant difference in the fatigue life of Palacos R and Simplex P when both cements were prepared in the standard fashion. The addition of 1/2 g of gentamycin to Palacos R did not significantly alter its fatigue properties. The addition of 0.5 g of erythromycin and 0.24 g of colistin did not decrease the fatigue life of Simplex P. Centrifugation significantly improved the fatigue properties of Simplex P and AKZ. The fatigue lives of Palacos R and Palacos R with gentamycin were not improved by centrifugation. The fatigue life of centrifuged Simplex P was significantly greater than the fatigue life of Palacos R and of Palacos R with gentamycin, whether the Palacos R based cements were centrifuged or not.     

Platelet activation after in vitro contact with seven acrylic bone cements

Seven acrylic bone cements were evaluated: Cemex Rx (Tecres S.p.a., Italy), Cemex Isoplastic (Tecres S.p.a., Italy), Zimmer Low Viscosity Cement (L.V.C. , Zimmer, IN, USA), Zimmer bone cement—dough type (Zimmer, IN, USA), CMW 3 (DePuy International Ltd., UK), Cerim LT (Cremascoli S.r.l., Italy), and Palacos R (Merck, Wehreim, Germany). The cements after polymerization were put in contact in vitro with platelet-rich plasma. Plasma in contact only with siliconated glass was used as the negative control. After contact, platelet number, β-thromboglobulin (β-TG), and transforming growth factor-β1 (TGF-β1) were determined. The Wilcoxon signed rank test showed Palacos R and L.V.C. induced a significant decrease of platelet number compared with the negative control. All cements determined a significant increase in β-TG. CMW 3 , Palacos , L.V.C. , and Zimmer dough type determined a significant increase in TGF-β1 compared with the negative control.     

Platelet activation after in vitro contact with seven acrylic bone cements

Seven acrylic bone cements were evaluated: Cemex Rx (Tecres S.p.a., Italy), Cemex Isoplastic (Tecres S.p.a., Italy), Zimmer Low Viscosity Cement (L.V.C., Zimmer, IN, USA), Zimmer bone cement - dough type (Zimmer, IN, USA), CMW (DePuy International Ltd., UK), Cerim LT (Cremascoli S.r.l., Italy), and Palacos (Merck, Wehreim, Germany). The cements after polymerization were put in contact in vitro with platelet-rich plasma. Plasma in contact only with siliconated glass was used as the negative control. After contact, platelet number, beta-thromboglobulin (beta-TG), and transforming growth factor-beta1 (TGF-beta1) were determined. The Wilcoxon signed rank test showed Palacos R and L.V.C. induced a significant decrease of platelet number compared with the negative control. All cements determined a significant increase in beta-TG. CMW 3, Palacos, L.V.C., and Zimmer dough type determined a significant increase in TGF-beta1 compared with the negative control.     

Production of TNF-alpha and bone resorbing activity by macrophages in response to different types of bone cement particles

We have compared the capacity of clinically relevant wear debris from seven different cement types to activate macrophages to produce TNF-alpha, IL-1beta, IL-6 and bone resorbing activity in vitro. The bone cements were: CMW 1 original (PMMA only); CMW 1RO (1 microm BaSO4; 9.2%); CMW copolymer bone cement 1 (10 microm BaSO4; 10%); CMW copolymer bone cement 2 (1 microm BaSO4; 10%); Palacos R (10 microm ZrO2; 15.6%); CMW Calcium phosphate cement 20% (10 microm tri-calcium phosphate; 20%) and CMW calcium phosphate cement 30% (10 microm tri-calcium phosphate; 30%). Cement debris was produced aseptically using a simple configuration wear test. The majority of particles were in the size range 0.1-0.5 microm for each cement type. The cement particles were co-cultured with the U937 macrophage cell line at ratios of 10 and 100 microm3 particle volumes to macrophage cell numbers for 24 h. At the 10:1 ratio the particles had no effect on the cells. At the 100:1 ratio, the major cytokine produced was TNF-alpha and there were no statistical differences between the different types of cement debris. The bone resorption activity of the co-culture supernatants was significantly greater than the control (U937 cells without particles) for particles of CMW 1RO, CMW copolymer bone cement 1, CMW copolymer bone cement 2 and Palacos R (P < 0.05, ANOVA). However there were no statistical differences between the levels of bone resoprtion evoked by these four cement types. The CMW1 original and CMW calcium phosphate containing cements failed to induce the macrophages to elaborate bone resorption activity at the 100:1 ratio. These data suggest that the addition of radio-opaque additives to bone cement may increase the capacity of the debris to induce osteolysis.     

Platelet release of transforming growth factor-beta and beta-thromboglobulin after in vitro contact with acrylic bone cements

Three methacrylate-based bone cements used for the fixation of joint prostheses were evaluated: Sulfix-60 (Sulzer Orthopedic Inc., Baar, Switzerland). CMW1 (DePuy International Ltd., England). and CMW2 (DePuy International Ltd., England). The cements after polymerization were put in contact in vitro with platelet-rich plasma. Plasma, in contact only with siliconized glass, was used as a negative control. After contact, platelet number. beta-thromboglobulin (beta-TG), and transforming growth factor-beta1 (TGF-beta1) were determined. The Student's paired t test showed that the ccments induced no significant modifications of platelet number. CMWI and Sulfix-60 determined a significant increase in beta-TG compared with the negative control. All cements determined a significant increase in TGF-beta1. Significant differences were also seen in the levels of beta-TG and TGF-beta1 between cements with a content of benzoyl peroxide < 1 (Sulfix-60) and those with a content > 1 (CMW1 and CMW2). The cement with zirconium dioxide (Sulfix-60) produced higher levels of beta-TG and TGF-beta1, compared to those with barium sulphate (CMW1 and CMW2). In conclusion, all the cements induced the secretion of TGF-beta1 CMW1 and Sulfix-60 determined also a significant release of beta-TG. Platelet activation induced by the cements from one side could contribute to the pathogenesis of deep venous thrombosis, that often occurs after prosthetic implant and is caused also by other factors, including surgical trauma and venous stasis. From the other side, activated platelets can release growth factors favoring bone formation.     

The osteolytic response of macrophages to challenge with particles of Simplex P, Endurance, Palacos R, and Vertebroplastic bone cement particles in vitro

The capacity of clinically relevant wear particles from Simplex P, Endurance, Vertebroplastic and Palacos R bone cements to activate macrophages to produce cytokines and bone resorbing activity in vitro was compared. Cement particles were generated aseptically by using a pin on plate rig. The particles were irregular in shape, and there were no significant differences in the particle characteristics of the different bone cement types (mean equivalent circle diameter range 0.225--0.36 mum, mean area range 0.048--0.063 microm(2), mean aspect ratio range 1.481--1.593, and mean length 0.412--0.523 microm). The volumetric concentration of particles in the 0.1- to 1.0-microm size range was 85% Palacos R, 82% Endurance, 80% Simplex P, and 77% Vertebroplastic. Particles were cultured with C3H macrophages at 100 microm(3) per cell for 24 h. Cytokines were determined by enzyme-linked immunosorbent assay and bone resorption (BR) measured by Ca(45) release from murine calvarias. Particles of Palacos R and Endurance stimulated enhanced production of TNF-alpha, IL-1-beta, and IL-6 (p<0.05; ANOVA). Simplex P particles only stimulated IL-1-beta (p<0.05; ANOVA). Vertebroplastic particles did not stimulate production of any of the cytokines. Particles of Palacos R generated the highest BR (1.38), but this did not reach statistical significance. The BRs for the other bone cements were no greater than the control. Hence, compared with the same volumetric concentrations, particles of Palacos R and Endurance were the most, and particles of Vertebroplastic were the least biologically reactive.     

Staphylococcus aureus biofilm formation on different gentamicin-loaded polymethylmethacrylate bone cements

In this in vitro study, the formation of a Staphylococcus aureus biofilm on six gentamicin-loaded bone cements (CMW1, CMW3, CMW Endurance, CMW2000, Palacos, and Palamed) was determined in a modified Robbins device over a 3 days time span and related with previously (Van de Belt et al., Biomaterials 21 (2000) 1981) measured kinetics of antibiotic release by these cement brands. The influence of gentamicin release on biofilm formation was quantified by expressing the number of colony-forming units on gentamicin-loaded cement relative to the number of viable organisms on unloaded cement of the same brand. Biofilms formed on all gentamicin-loaded cements, despite the release of antibiotics, followed a consistent pattern in time with a maximum number of colony-forming units per unit cement area found between 24 and 30 h after inoculation. None of the gentamicin-loaded cements showed a reduction in biofilm formation relative to unloaded cements within 6 h after inoculation, whereas only gentamicin-loaded CMW1 and Palacos reduced biofilm formation 24 h after inoculation. Alternatively, CMW Endurance, CMW2000, and Palamed did not exhibit any initial reductions in biofilm formation, but effects started after 72, 48, and 72 h, respectively. Biofilm reduction by gentamicin-loaded CMW3 lasted the longest from 24 to 72 h. Interestingly, each cement seemed to have a different "window-of-effectiveness" with regard to reduction in biofilm formation that did not relate with the gentamicin-release kinetics. Summarising, this study demonstrates that although gentamicin loading of bone cements yields reductions in biofilm formation, S. aureus is able to grow on gentamicin-loaded bone cements.     

Biological and mechanical properties of PMMA-based bioactive bone cements

We reported previously that a bioactive PMMA-based cement was obtained by using a dry method of silanation of apatite-wollastonite glass ceramic (AW-GC) particles, and using high molecular weight PMMA particles. But handling and mechanical properties of the cement were poor (Mousa et al., J Biomed Mater Res 1999;47:336-44). In the present study, we investigated the effect of the characteristics of PMMA powder on the cement. Different cements containing different PMMA powders (CMW1, Surgical Simplex, Palacos-R and other two types of PMMA powders with Mw 270,000 and 1,200,000) and AW-GC filler in 70 wt% ratio except Palacos-R (abbreviated as B-CMW1 and B-Surg Simp, B-Palacos 50 [50 wt% AW-GC filler] and B-Palacos 70 [70 wt% AW-GC filler], B-270 and B-1200) were made. Dough and setting times of B-CMW1, B-Surg Simp B-270 and B-1200 were similar to the commercial CMW1 cement which did not contain bioactive powder (C-CMW1), but B-palacos which contained large PMMA beads with high Mw had delayed setting time. B-270 had the highest bending strength among the tested cements. After 4 and 8 weeks of implantation in the medullary canals of rat tibiae, the bone-cement interface was examined using SEM. The affinity index of B-1200 was significantly higher than the other types of cements. B-270 showed good combination of handling properties, high mechanical properties and showed higher bioactivity with minimal soft tissue interposition between bone and cement compared with commercial PMMA bone cement. This may increase the strength of the bone-cement interface and increase the longevity of cemented arthroplasties.     

Antibiotic bone cement for the treatment of Pseudomonas aeruginosa in joint arthroplasty: comparison of tobramycin and gentamicin-loaded cements

One hundred clinical isolates of Pseudomonas aeruginosa were collected from 22 medical centers throughout Europe and were challenged with two aminoglycoside-loaded bone cements, employing a modified in vitro Kirby-Bauer susceptibility model. The results of this study show that Simplex P with tobramycin exhibits antibacterial activity against 98% of the strains tested, compared to 93% for Palacos with gentamicin. Additionally, for strains that were susceptible to the antibiotic bone cement formulations, the average zone of inhibition produced around the tobramycin-loaded cement disks was approximately 25% greater than that seen around the gentamicin-loaded cement disks. This difference was statistically significant (p < 0.01). Tobramycin-loaded bone cement is therefore the preferred formulation when addressing Pseudomonas aeruginosa in septic joint arthroplasty.     

Comparison of the response of human peripheral blood mononuclear cells to challenge with particles of three bone cements in vitro

This study compared the effects of different sizes of three clinically relevant endotoxin free bone cement particles on primary human macrophage TNF-alpha production in vitro. The bone cements used were CMW original, CMW1RO and Palacos R. The cement wear debris was generated aseptically and then sequentially filtered to produce the size ranges 0.1-1 microm, 0.1-10 microm, 1-10 microm and >10 microm. The debris was cultured with human peripheral blood mononuclear cells at particle volume (microm(3)) per cell ratios of 100:1, 10:1 and 1:1. TNF-alpha production was determined by ELISA and cell viability by MTT conversion. CMW1RO particles induced increased TNF-alpha production by PBMNCs when tested in the size range 0.1-1 microm, and also to a lesser degree in the sizes 0.1-10 microm and 1-10 microm at the particle volume (microm(3)) to cell number ratios of 100:1 and 10:1.The increase in TNF-alpha production induced by Palacos R debris was only observed with the particle size ranges less than 10 microm at the ratio of 100:1. This study demonstrated that bone cement particles are capable of inducing raised TNF-alpha production in vitro. This is dependent upon cement particle size, volume and cement particle type, with cement particles containing radio-opaque additives being the most active.     

Effect of four acrylic bone cements on transforming growth factor-beta1 expression by osteoblast-like cells MG63.

Based on the hypothesis that bone cements cause changes in the production of transforming growth factor-beta 1 (TGF-beta1) by bone cells, the effects of four acrylic bone cements (Sulfix-60, CMW 1, CMW 2 and CMW 3) were examined using the osteoblast-like cell line MG63. The extracts in MEM of the cements were tested, following 1 h- and 7 day-curing. MG63 cells seldom expressed mRNA specific for TGF-beta1 in basal conditions. The cultures expressed mRNA constantly after incubation with the extract of CMW 1 cured for 1 h. TGF-beta1 specific mRNA was seldom expressed after incubation with the other cement extracts. The release of TGF-beta1 into the conditioned medium was increased significantly by CMW 1 extract at 1 h-curing, but was not changed significantly by CMW 1 extract at 7 day-curing and by the extracts of the other cements, at both curing times. The stimulating effect of CMW 1 on the secretion of TGF-beta1, even with all the restrictions of an in vitro study of continuous cell lines, if confirmed in vivo, might favor the development of the synovial-like membrane around the implant, and therefore impair the chance of success of the prosthesis.     

Gene expression of bone-associated cytokines in MG63 osteoblast-like cells incubated with acrylic bone cement extracts in minimum essential medium

This study examined the effects of in vitro challenge with four polymerized acrylic bone cements (Sulfix-60, CMW 1, CMW 2, and CMW 3) on the expression of interleukin-1beta (IL-1beta), interleukin-6 (IL-6), and transforming growth factor-beta1 (TGF-beta1) mRNAs in the osteoblast-like cell line MG63. The extracts of the cements in minimal essential medium (MEM) were tested following 1-h and 7-day curing. A semi-quantitative analysis of the cytokine-specific mRNAs was carried out by agarose gel densitometry and expression was compared with the GAPDH housekeeping gene. The ratio between cytokine gene expression and GAPDH expression was calculated. The mRNA specific for the bone-resorbing cytokines IL-1beta and IL-6 was low in basal conditions. IL-1beta mRNA increased only after incubation with the extract of CMW 1 following 1-h curing. The mRNA specific for the bone-resorbing cytokine IL-6 also increased after contact with CMW 1 at both curing times. Sulfix-60 and CMW 3 following 7-day curing, but not after 1 h, induced higher levels of IL-6 mRNA than the control. CMW 2 after 1-h curing constantly determined the expression of IL-6 mRNA, but at low levels. The mRNA specific for TGF-beta1 was also expressed by the MG63 osteoblast-like cells in basal conditions. The levels increased after contact with Sulfix-60 after 7-day curing and with CMW 1 after 1-h curing. CMW 2 after 7-day curing decreased TGF-beta1 mRNA. In conclusion, the highest expression of the cytokines IL-1beta, IL-6, and TGF-beta1 mRNA was determined by CMW 1. If the results are confirmed in vivo, the increased expression of the osteolytic cytokines induced by the bone cement might result in loosening of the prosthesis, even with all the restrictions of an in vitro study on continuous cell lines.     

Interleukin-6 expression by osteoblast-like MG63 cells challenged with four acrylic bone cements

Periprosthetic osteolysis is a major clinical problem in total hip and total knee arthroplasty and polymethylmethacrylate (PMMA) is a possible etiologic factor. Recently, increasing importance was ascribed to interleukin-6 (IL-6) as an agent favouring bone resorption. The aim of the present study was to investigate the role of bone cements on IL-6 production by MG63. The effect of four acrylic bone cements (Sulfix-60, CMW 1, CMW 2, and CMW 3) on the protein release and mRNA expression of IL-6 in osteoblast-like cell line MG63 was examined using IL-1beta (0.2 microg ml(-1)) as the positive control. The extracts in minimum essential medium (MEM) of the cements were tested, following 1-h and 7-day curing. CMW 1 and CMW 2 significantly increased the IL-6 release into the culture media (p < 0.01). The cells incubated with Sulfix-60 and CMW 3 produced no significantly different levels of IL-6 than the basal production. A positive correlation was found between the concentration of IL-6 and the contents of benzoylperoxide (p = 0.0003) and barium sulphate (p < 0.0001). MG63 expressed IL-6 mRNA constitutively, as demonstrated by the positivity of the negative controls too. We conclude that CMW 1 and CMW 2 increase the production of IL-6 in MG63 cells. The response to Sulfix-60 and CMW 3 is not significantly greater than the negative control.     

Creep properties of three low temperature-curing bone cements: a preclinical assessment

The mechanical characteristics of new bone cements should be assessed before these cements are released on the orthopedic market in great quantities. In this study, we present the deformational response of 3 relatively new, low-curing temperature bone cements (Cemex RX, Cemex System, and Cemex Isoplastic) to a dynamic compressive force in comparison to Simplex P bone cement. For this purpose, dynamic compressive creep tests were performed on cylindrical shaped specimens at a maximal load level of 20 MPa for a period of 250,000 cycles. The results showed that Cemex System and Cemex RX produced creep rates that were higher (20% and 30%, respectively) as compared to Simplex P bone cement. The creep behavior of Cemex Isoplastic was very similar to that of Simplex P. It was concluded that although Cemex RX and Cemex System produced higher creep rates than Simplex P, these differences were not considered excessive. Hence, although other tests are required to assess the safety and efficacy of these new cements, the dynamic creep properties under compression can be considered adequate for clinical use.     

Effect of choice of surgical gloves on dough time measurements of acrylic bone cement

In this study we investigated the effect of the brand of surgical gloves on the dough time determination for acrylic bone cements. Four different brands of powder-free latex surgical gloves were tested. Two commercial bone cements, Surgical Simplex P and Palacos R, were used for dough time measurement following standard test methods for acrylic bone cements (ASTM F-451 and ISO 5833). The results show that the measured dough time depended largely on the brand of gloves used, and could vary by nearly 250%. The surface morphological structures of gloves, determined by SEM, probably contribute to the differences in the measured dough time. This study provides experimental evidence that supports the need to describe the type of gloves used, in detail, when dough time is reported. It also illustrates the importance of the glove brand, when bone cement is to be handled as a dough in the clinical setting.     

Structural degradation of acrylic bone cements due to in vivo and simulated aging

Acrylic bone cement is the primary load-bearing material used for the attachment of orthopedic devices to adjoining bone. Degradation of acrylic-based cements in vivo results in a loss of structural integrity of the bone-cement-prosthesis interface and limits the longevity of cemented orthopedic implants. The purpose of this study is to investigate the effect of in vivo aging on the structure of the acrylic bone cement and to develop an in vitro artificial aging protocol that mimics the observed degradation. Three sets of retrievals are examined in this study: Palacos brand cement retrieved from hip replacements, and Simplex brand cement retrieved from both hip and knee replacement surgeries. In vitro aging is performed using oxidative and acidic environments on three acrylic-based cements: Palacos, Simplex, and CORE. Gel permeation chromatography (GPC) and Fourier transform infrared spectroscopy (FTIR) are used to examine the evolution of molecular weight and chemical species within the acrylic cements due to both in vivo and simulated aging. GPC analysis indicates that molecular weight is degraded in the hip retrievals but not in the knee retrievals. Artificial aging in an oxidative environment best reproduces this degradation mechanism. FTIR analysis indicates that there exists a chemical evolution within the cement due to in vivo and in vitro aging. These findings are consistent with scission-based degradation schemes in the cement. Based on the results of this study, a pathway for structural degradation of acrylic bone cement is proposed. The findings from this investigation have broad applicability to acrylic-based cements and may provide guidance for the development of new bone cements that resist degradation in the body.     

Relationship between fracture toughness and impact strength of acrylic bone cement

This work involved determining the fracture toughness, KIc (in MPa(square root)m) (using rectangular compact tension specimens) and impact strength, IS (in kJ/m2) (Charpy type specimens) of Surgical Simplex P and three variants of Palacos R acrylic bone cements. The best fit to these results yielded a power relationship KIc = 0.795(IS)0.59. The usefulness of this relationship is detailed, especially for the purpose of performing quality control checks on bone cements.     

VersaBond bone cement prospective randomized study of the clinical properties of a new bone cement in total knee replacement

VersaBond is a newly developed bone cement. To investigate its clinical performance, VersaBond was compared to Palacos R in a prospective randomized study in total knee replacement. Fifty-nine patients (61 knees) undergoing total knee replacement were randomized to either VersaBond or Palacos R bone cement and followed for 24 months using radiostereometric analysis (RSA). Up to 2 years there were no significant differences in clinical performance between the two cements. The mean/median values for implant migration were very similar for the two bone cements, as were the dispersion, and distribution of outliers. Also the proportion "stable" and "continuously migrating" implants was similar between the two cements. The result of this study indicates that VersaBond bone cement will perform at least equally as well as Palacos R in total knee replacement as regards as aseptic loosening.