C. P. Rader

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.

Effects of polyethylene and TiAIV wear particles on expression of RANK, RANKL and OPG mRNA

Background Wear debris has been associated with periprosthetic osteolysis and loosening of total joint arthroplasties. RANKL (receptor activator of NF-κB ligand), RANK (receptor activator of NF-κB) and OPG (osteoprotegerin) are three key molecules which regulate differentiation, survival, fusion, and activation of osteoclasts.Material and methods We evaluated the effect of TiAlV and polyethylene particles on expression of RANK, RANKL and OPG mRNA. We used a human monocytic leukemic cell line (THP-1) in this in vitro study. THP-1 monocytes were differentiated into macrophage-like cells and exposed to polyethylene particles and prosthesis-derived TiAlV particles. The supernantant was used for measurement of TNFα protein and total RNA was extracted from the cells. Expression of the genes coding for OPG, RANKL and RANK was analysed at the mRNA level using a semiquantitative RT-PCR method.Results Both polyethylene and TiAlV particles induced a significant release of TNFα after 6 h of exposure and a significant upregulation of RANK mRNA. OPG mRNA expression was transiently upregulated after exposure to polyethylene and TiAlV particles. These effects were dependent on particle dose. RANKL mRNA was not detectable in our THP-1 model. In contrast, LPS exhibited a different pattern of RANK/RANKL/OPG mRNA expression.Interpretation Our findings provide evidence that both polyethylene and TiAlV particles induce upregulation of RANK expression in cells of the monocytic lineage, which may be important for periprosthetic osteoclastogenesis.

Heterotopic ossification after total knee arthroplasty : 54/615 cases after 1-6 years'follow-up

We found heterotopic ossifications in 54 (9%) of 615 cases after total knee arthroplasty. The largest ossifications were located in the anterior distal femur. In 12 cases smaller ossifications were found in other knee regions. The development of heterotopic ossification showed a positive correlation with hypertrophic arthrosis and a negative correlation with rheumatoid arthritis. We propose a new 3-grade classification which refers only to the anterior distal femoral region. Grade III heterotopic ossifications occurred in 4 patients (4 knees) who had clinical symptoms; 2 were successfully reoperated with removal of the ossifications. Prophylaxis should be considered in patients with marked hypertrophic arthrosis or marked periosteal damage to the anterior distal femur.

Results of total knee arthroplasty with a metal-backed patellar component. A 6-year follow-up study

This study reviews the clinical outcome of knee arthroplasty (Microloc, Johnson & Johnson Orthopedics, Norderstedt, Germany) with a resurfacing metal-backed patellar component. Thirty patients were available for clinical and radiographic reexamination with follow-up periods of 36 and 72 months. There were 21 women and 9 men, with an average age of 71.1 years. At the follow-up examination, seven patients demonstrated a metallic friction noise, and three had only mild crepitation at the patellofemoral joint level. In the group complaining of knee pain, the average knee score dropped from 76.9 +/- 9.9 at 36 months to 44.7 +/- 15.3 (P < .05) prior to revision surgery on average at 66 months. The function score dropped during the same time from 72.0 +/- 19.7 to 53.5 +/- 14.9. In contrast, the pain-free group did not demonstrate any significant changes in either knee or function score. Radiographic evaluation showed a patellar tilt of 8.1 degrees +/- 4.0 degrees in the group complaining of pain, compared with 2.5 degrees +/- 2.3 degrees in the pain-free patient collective. All 10 patients complaining of pain and increasing disability underwent revision surgery. At the time of surgery, an obvious polyethylene particle-induced synovitis was found, and in 7 patients it was associated with a metallosis. One of the main reasons for failure of metal-backed polyethylene components is thought to be the undue high stresses forced onto the insert. High loading of only a small fraction of the surface will lead to increased polyethylene creep and particle formation, resulting in synovitis and finally metallosis. Early revision seems to be the best solution to prevent progressive destruction of the entire joint. Previously reported results on metal-backed patellar components show a failure rate of 8.4% after 12 to 24 months. In this study, this rate had already increased to 33.3% after 6 years. One might therefore speculate that at 10 years, revision surgery might become necessary in more than 50% of the surviving patients with this type of implant.

Standardisiertes Testen von Skelett-Implantatoberflächen mit einem Osteoblasten-Zellkultursystem. II. Titanoberflächen unterschiedlicher Rauhigkeit - Standardised Testing of Bone-implant Surfaces using an Osteoblast Gell Culture System. II. Titanium Surfaces of varying Degrees of Roughness

Der Effekt von Titanoberflächen unterschiedlicher Oberflächenrauhigkeit auf die Proliferation und Differenzierung von Osteoblasten wurde mit einem standardisierten Zellkultursystem untersucht. Humane fetale Osteoblasten (hFOB 1.19) wurden auf polierten (Ti pol), sandgestrahlten (Ti sb) und sandgestrahlt-wärmebehandelten Titanoberflächen (Ti sb-ht) über einen Zeitraum von 17 Tagen kultiviert. Als Kontrolle wurde Zellkultur-Polystyrol (Ps) verwendet. Zellzahl und -Vitalität wurden als Proliferationsparameter bestimmt. Als Parameter der osteoblastären Differenzierung wurden die Aktivität der alkalischen Phosphatase, die Kollagen Iund die Osteocalcinproduktion gemessen. Auf Ti pol wurden in der Frühphase zunächst höhere Proliferationsparameter gemessen. Dagegen zeigten sich in der Spätphase gesteigerte Werte auf Ti sb und Ti sb-ht. Die Aktivität des frühen Differenzierungsparameters alkalische Phosphatase war auf Ti pol erhöht. Für die späten Differenzierungsparameter Kollagen I und Osteocalcin bestanden keine Unterschiede zwischen den drei Oberflächen. Mit dem vorliegenden Testsystem kann der Einfluß der Oberflächengestaltung auf die Dynamik des osteoblastären Entwicklungszyklus dargestellt werden. Die vergrößerte Oberfläche der rauhen Materialien führt zu einem später einsetzenden und länger anhaltenden Wachstum. Das Modell korreliert mit entsprechenden In-vivo-Befunden und bestätigt den Einsatz rauher Oberflächen für Implantate im direkten Knochenkontakt auch auf zellulärer Ebene.

[Effects of clinically relevant alumina ceramic, zirconia ceramic and titanium particles of different sizes and concentrations on TNF-alpha release in a human macrophage cell line].

Abstract Einleitung: Als großes Problem der modernen Endoprothetik gilt die aseptische Lockerung von Prothesenkomponenten. Als Initiatorzytokin des Particle Disease wird derzeit der Tumor Nekrose Faktor α (TNFα) vermutet. Ziel der Studie war es die TNFα-Ausschüttung von Makrophagen-ähnlichen Zellen (MäZ) zu untersuchen, die durch Partikel, wie sie typischerweise während Revisionsoperationen gefunden werden stimuliert wurden. Hierzu verwendeten wir Aluminiumkeramik- (Al2O3), Zirkoniumoxid- (ZrO2) und Titanpartikel (Ti) unterschiedlicher Konzentration und Größe wie sie typischerweise bei Revisionsoperationen gefunden werden. Insbesondere sollte die Frage beantwortet werden, ob unterschiedliche Partikelgrößen und Materialgruppen (Titan und Keramik) eine differente TNFα-Ausschüttung bewirken. Methode: Um ein TNFα-Profil der einzelnen Materialien zu erhalten, verwendeten wir ein etabliertes Makrophagenmodell (Rader et al. 1999) mit THP-1 Zellen (humane Leukämiezellen). Die verschiedenen Partikelgrößen wurden in unterschiedlichen Konzentrationen für 6h mit 106 MäZ inkubiert. Der Überstand wurde entnommen und durch ELISA-Technik auf die TNFα-Konzentration untersucht. Ergebnis: die Verwendung von Ti-Partikel löste in beiden verwendeten Größen (0,2 μm und 2 μm) mit jeweils 8facher und 17facher TNFα-Sekretion gegenüber der Leerprobe den stärksten Anstieg aus. Es waren jedoch deutlich höhere Mengen an Ti-Partikel der Größe 0,2 μm notwendig, um o.g. Werte zu erreichen. Al2O3-Partikel zeigten ebenfalls eine signifikante Erhöhung der TNFα-Ausschüttung, lagen jedoch mit einer 4fachen Steigerung gegenüber der Leerprobe deutlich unter den Ti-Werten. In der maximalen TNFα-Sekretion fand sich kein Unterschied beider Partikelgrößen (0,6 μm und 2,5 μm), jedoch war eine nahezu 1000fach höhere Konzentration an Al2O3-Partikel der Größe 0,6 μm notwendig. Beim direkten Vergleich von Al2O3- und Ti-Partikeln gleicher Größe und Konzentration stimulierte Ti signifikant höhere TNFα-Ausschüttungen. ZrO2 konnte keine signifikante TNFα-Sekretion hervorrufen. Schlussfolgerung: Aufgrund der vorliegenden Ergebnisse empfiehlt sich die Verwendung von Keramikgleitpaarungen, die geringere biologische Potenz aufweisen als Metall- oder PE-Gleitpaarungen. Zusätzlich sollten größere Abriebpartikel vermieden werden. Frühzeitige Revisionsoperationen vermeiden große Mengen an Abriebpartikeln, somit könnten ausgedehnte lokale Osteolysen verhindert werden. Introduction: Aseptic loosening is considered to be the main problem of modern endoprothesis. Tumor necrosis factor α (TNFα) seems to be the initiator protein of particle disease. The aim of our study was to investigate the TNFα response of macrophage like cells (MLC) after stimulation with periprosthetic particles, typically found during revision surgery. For this purpose alumina ceramic (Al2O3), circonia ceramic (ZrO2) and titanium (Ti) particles of different sizes and concentrations were used. Important was to study the effects of different sizes due to TNFα secretion and the comparison of the biological effects of alumina ceramic and titanium. Method: To obtain an TNFα profile we used an established macrophage model (Rader et al.) with THP-1 cells (human monocytic cell line). Therefore 106 MLC were incubated with different particle concentrations and sizes for 6 h. The supernatent was then investigated for TNF·using ELISA assay. Results: Ti-particles provoked in both sizes (0,2 μm and 2,5 μm) the greatest TNFα response, 8 times and 17 times as high in comparison with control. But substantially more 0,2 μm sized Ti-particles were necessary to get the above mentioned results. Al2O3-particles were not as effective as Ti, but they released fourfold more TNFα compared to control. There was no difference in TNFα-secretion comparing Al2O3- particles of different sizes (0,6 μm and 2 μm), but a 1000times greater concentration of the 0,6Ìm sized particles were needed. Using Al2O3- and Ti-particles of the same size and concentration, Ti provoked a significant higher TNFα response. ZrO2 showed no effects on TNFα release. Conclusion: Because of our results we recommend ceramic articulating surfaces, which are superior to metal on metal matings ion term of biological reactions. Additionally bigger wear particles should be avoided. Revisionoperation should be done early to avoid huge amount of wear particles and to minimize local osteolysis

Low-molecular-weight heparin and partial thromboplastin time-adjusted unfractionated heparin in thromboprophylaxis after total knee and total hip arthroplasty

Thromboprophylaxis with heparins after total hip arthroplasty (THA) and total knee arthroplasty (TKA) is well established. The aim of this study was to compare low-molecular-weight heparin (enoxaparin) with partial thromboplastin time (PTT)-adjusted, unfractionated heparin (heparin sodium). In a prospective study of THA and TKA 246 patients, physical examination and compression and duplex ultrasound were performed 1 day before and 7 and 14 days after surgery. One hundred thirty patients received 40 mg enoxaparin subcutaneously once per day. One hundred sixteen patients received 5,000 IU heparin sodium subcutaneously 3 times daily. As the PTT did not reach 40 seconds, the heparin sodium dosage was increased to 7,500 IU 3 times daily. The overall thrombosis rate was 4% (n = 10). In the enoxaparin group, the rate was 2.9% of the 70 THAs and 10% of the 60 TKAs. Thrombosis also occurred in the group that received heparin sodium: 1.8% of the THAs and 1.7% of the TKAs. For TKA, the difference between the 2 heparin groups was statistically significant. In the thromboprophylaxis of TKA, PTT-adjusted unfractionated heparins are superior to fixed doses of low-molecular-weight heparins.

9- to 11-Year Results of Cemented Titanium Mueller Straight Stem in Total Hip Arthroplasty

This retrospective study reviewed 9- to 11-year results after total hip arthroplasty (THA) with cemented titanium stems (Mueller-Straight-Stem). Ninety-one patients (110 hips) were examined clinically and radiologically at an average 9.5-year follow-up. Revisions for aseptic loosening were performed in 4 (4%) patients. Subsidence or varus position could only be observed in one of these patients. Radiolucent lines were found in 37 patients, mainly located around the proximal zones of the stem (zone 1, 7, 8, and 14). Harris scores were good or excellent in 78% and satisfactory in 20% of patients. The 9.5-year survival rate of the cemented titanium stem with regard to aseptic loosening was 96.4%. Body weight was significantly higher (88 +/- 5.4 kg) in the 4 patients with aseptic loosening, compared to patients without radiolucent lines (75 +/- 15 kg). The body weight to stem surface ratio showed a significant difference (1.5 kg/cm2 versus 1 kg/cm2; P < .05). No significant differences were found in other factors, including sex, size or type of stem, Harris score, heterotopic ossification, or body mass index. Good long-term results can be achieved with cemented titanium stem implants. This titanium implant is recommended for patients with hypersensitivity to chrome, cobalt, and nickel. mplanting the biggest possible stem seems to be most beneficial.

Synergistic effects of mixed TiAlV and polyethylene wear particles on TNFalpha response in THP-1 macrophages.

Abstract TNFα is a potent osteoclastogenic cytokine that has a fundamental role in the pathogenesis of wear particle-induced osteolysis. Wear particles of one composition and their biological effects are well characterised. In contrast, little is known about the effects of mixed particles with respect to mix ratio and particle concentration. We evaluated the effects of different mix ratios of polyethylene and TiAlV particles on TNFα response. We used a human monocytic cell line (THP-1) in this in vitro study. THP-1 monocytes were differentiated to macrophage-like cells and exposed to different mixtures of lipopolysaccharide-detoxified polyethylene and TiAlV particles. TNFα was analysed in culture supernatants using ELISAs. Both polyethylene and TiAlV particles induced a dose- and time-related release of TNFα, with maximum levels after 6 h. A PE/TiAlV mix ratio of 36:1 at 108 particles/ml induced significantly higher TNFα concentrations compared to equal particle concentrations of isolated TiAlV (p=0.047) or PE (p=0.044), indicating the synergistic effect of mixed particles. These results provide evidence that TiAlV and polyethylene particles have significant synergistic effects, depending on the mix ratio and particle concentrations. This supra-additive effect can contribute substantially to the pathogenesis of implant particle-induced osteolysis.TNFalpha is a potent osteoclastogenic cytokine that has a fundamental role in the pathogenesis of wear particle-induced osteolysis. Wear particles of one composition and their biological effects are well characterised. In contrast, little is known about the effects of mixed particles with respect to mix ratio and particle concentration. We evaluated the effects of different mix ratios of polyethylene and TiAlV particles on TNFalpha response. We used a human monocytic cell line (THP-1) in this in vitro study. THP-1 monocytes were differentiated to macrophage-like cells and exposed to different mixtures of lipopolysaccharide-detoxified polyethylene and TiAlV particles. TNFalpha was analysed in culture supernatants using ELISAs. Both polyethylene and TiAlV particles induced a dose- and time-related release of TNFalpha, with maximum levels after 6 h. A PE/TiAlV mix ratio of 36:1 at 10(8) particles/ml induced significantly higher TNFalpha concentrations compared to equal particle concentrations of isolated TiAlV (p=0.047) or PE (p=0.044), indicating the synergistic effect of mixed particles. These results provide evidence that TiAlV and polyethylene particles have significant synergistic effects, depending on the mix ratio and particle concentrations. This supra-additive effect can contribute substantially to the pathogenesis of implant particle-induced osteolysis.

Standardisiertes Testen von Skelett-lmplantatoberflächen mit einem Osteoblasten-Zellkultursystem. I. Orthopädische Standardmaterialien. Standardised Testing of Bone-implant Surfaces Using an Osteoblast Cell Culture System. Part I: Standard Materials in Orthopaedic Surgery

Der Effekt orthopädischer Standardmaterialien auf Proliferation und Differenzierung von Osteoblasten wurde mit einem standardisierten Zellkultursystem untersucht. Osteoblasten der Zellinie hFOB 1.19 wurden über 12 Tage auf rostfreiem Stahl (Stainless Steel, SS), einer Chrom-Kobalt-Molybdän-Legierung (CrCoMb) und Reintitan (commercially pure Titan, cpTi) kultiviert. Als Referenzoberfläche wurde Zellkultur-Polystyrol (PS) eingesetzt. Zellzahl und -Vitalität wirkten als Parameter der Zellproliferation, als Differenzierungsparameter wurden die alkalische Phosphatase-Aktivität, die Kollagen Iund die Osteocalcinsynthese bestimmt. Die Proliferationsparameter zeigten auf PS und cpTi früher maximale Werte. Auf SS und CrCoMb war die Proliferation demgegenüber deutlich verzögert. Die höchsten Werte der Differenzierungsparameter wurden jeweils auf cpTi gemessen. Die Aktivität der alkalischen Phosphatase zeigte einen zweigipfligen Verlauf als Hinweis auf eine Apoptose mit nachfolgender Redifferenzierung. Das Zellkultursystem hFOB 1.19 ist damit geeignet, das unterschiedliche Proliferationsund Differenzierungsverhalten von Osteoblasten auf orthopädischen Standardmaterialien nachzuweisen. Die Resultate konrelieren mit bisherigen In-vivo-Befunden. Mit diesem Testsystem wird erstmals ein dynamischer Einfluß der Materialoberfläche auf den Entwicklungsprozeß von Osteoblasten dargestellt.