This contribution discusses photoinitiated crosslinking of
multifunctional acrylic esters in polymeric binders based on digital
imaging using the Computer to Plate (CtP) technology applying laser
exposure at 830 nm in the near infrared (NIR). All coating
components (NIR photoinitiator system, monomers, binder, adhesion
promoter, contrast dye, and oxygen barrier material) were applied as
thin double layer film (1–2 μm dry film thickness)
on Al-plates with an anodized surface. Materials exposed exhibit
a sensitivity between 30–200 mJ/cm2 depending on the
NIR-photoinitiator composition. This was processed in a weak aqueous
alkaline bath to obtain the image. Generation of initiating radicals
occurs by electron transfer from the excited state of the
NIR-sensitizer to the radical generator; that is either an onium salt
or neutral electron deficient compound. Redox potentials were
determined for the NIR-sensitizers and the radical generator. These
data allow a rough estimate regarding the free energy of electron
transfer of the excited state of the sensitizer and radical generator
in NIR-photosensitive imaging material. Photoinduced electron transfer
plays a major function to generate initiating radicals by a sensitized
mechanism but thermal events also influence sensitivity of the
coating. Particular the non-radiative deactivation of the
NIR-sensitizer possesses a major function to release selectively the
heat. Absorption data of many NIR-sensitizers used exhibit molar
extinction coefficient of more than
200000 M–1 cm–1.
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.
The patch clamp technique has been applied to isolated vacuoles from green suspension cells of Chenopodium rubrum to record electrical parameters of the tonoplast. In a symmetrical K+ solution of 46 mᴍ, the membrane displays a near zero voltage, whereas 2 mᴍ ATP will hyper-polarize it to 15 or 20 mV (vacuole positive). The conductance amounts to about one S · m-2. Fluctuations of the clamp current are explained by an unknown channel species having opening times of 5 - 10 ms. Together with previous work on a tonoplast vesicle preparation and unpublished data on vacuoles from our laboratory, the present results indicate an electrogenic membrane ATPase pumping protons from the cytoplasm to the vacuole.
Tumor necrosis factor α (TNFα) plays a fundamental role in the pathogenesis of wear particle-induced periprosthetic osteolysis. However, particle-induced mechanisms that control TNFα gene expression are not yet well characterized. LITAF [lipopolysaccharide (LPS)-induced TNFα factor] is a novel transcription factor that regulates expression of the TNFα gene, but nothing is known about its role in wear particle-induced osteolysis. We evaluated the effect of titanium aluminum vanadium (TiAlV) and polyethylene particles on mRNA expression of LITAF. A human monocytic leukemia cell line (THP-1) was used in this in vitro study. THP-1 monocytes were differentiated to macrophage-like cells and exposed to LPS-detoxified polyethylene particles and prosthesis-derived TiAlV particles. Supernatant was used for TNFα protein measurement and total RNA was extracted from cells. LITAF was analyzed at the mRNA level using semiquantitative RT-PCR. Both polyethylene and TiAlV particles induced significant upregulation of LITAF mRNA that was followed by a significant TNFα response. These effects were dependent on the particle dose. Low particle concentrations exhibited no significant effect on expression of TNFα and LITAF mRNA. In comparison to exposure to polyethylene and TiAlV particles, LPS stimulation exhibited similar upregulation of LITAF mRNA, but led to an overwhelming TNFα response. Our findings provide evidence that LITAF is implicated in the pathogenesis of wear particle-induced osteolysis.
In the recent years the number of commercially available immunoassays for the detection of human cytomegalovirus (HCMV)–specific immunoglobulin M (IgM) antibodies has rapidly increased. The aim of the present study was to evaluate five commercial immunoassays for the serological diagnosis of HCMV-infection. These methods, namely the IMx CMV IgM assay, the AxSYM CMV IgM assay (both Abbott), the Gull CMV IgM, the CMV-IgM-ELA test PCS Medac and the Biotest Anti-HCMV recombinant IgM ELISA, were compared for their diagnostic effectiveness and interference with substances eventually producing cross-reactions with HCMV-IgM (Epstein-Barr-virus (EBV)-IgM, rheumatoid factor (RF)). In addition, repeated measurements on samples from kidney and heart transplant recipients with active HCMV infection were examined to compare the temporal development of the HCMV-IgM measured with the five assay systems. Since there is no commercially available gold standard, it was assumed that the true classification, of whether the patient sample is HCMV-IgM positive or negative, was unknown. Hence sensitivity and specificity were assessed based on a maximum likelihood approach using a “latent class” model. The cross-reactions were quantified by a Bayesian statistical model using prior information for the expected prevalences in the EBV-IgM and rheumatoid factor sample groups. The results of the study demonstrated that there are great differences in sensitivity and specificity as well as in cross-reactions with EBV-IgM and RF between the tested ELISAs.
Informations about wear particles in metallosis (mode II wear) and their effects in vitro and in vivo are limited. The aim of this study was to characterize wear particles obtained intraoperatively and to analyse their effects on cytokine response in an established human macrophage-like cell culture model .
Method: Wear particles were obtained intraoperatively from four patients with metallosis resulting from CrCoMo/PE/TiAlV-implants (mode II wear) (3 knee, 1hip prosthesis). After purification, particles were characterized regarding to their composition and size (particle size analyser, electron microscopy, edx-analysis, histological slices). The effects of particles on the release of cytokines (PDGF, IL-1β, IL-8, TNF α) were determined in an established human macrophage-like cell culture system by ELISA-assays.
Results: The metal wear particles consisted of TiAlV with a mean size of 0,1± 0,15 μm, independent of the prosthesis location. CrCoMo particles could not be detected.
In the cell culture model 1456 × 108 particles per 1 × 106 macrophages released maximum amounts of TNFα (8-fold) and IL-8 and IL-1ß (5-fold) while the survival rate of the cells was more than 90 percent. A particle-dependent increase of PDGF-levels could not be detected. Conclusion: As already shown for mode I wear particles (contact between primary bearing surfaces), also mode II wear particles cause release of bone resorbing cytokines in a macrophage-like cell culture model. Because their local and systemic effects in vivo are still not completely understood, we recommend a complete removal of wear particles in cases of metallosis to avoid possible immunological reactions of the body as well as periprosthetic osteolysis.
Informationen über Abriebpartikel in Fällen von Metallose (Mode IIAbrieb) und über deren Effekte in vitro und in vivo sind limitiert. Ziel dieser Studie war die Charakterisierung von intraoperativ gewonnenen Abriebpartikeln und die Analyse ihrer Effekte auf die Zytokinausschüttung in einem etablierten, humanen Makrophagen-ähnlichen Zellkulturmodell .
Methode: Bei vier Patienten resultierte eine Metallose aufgrund pathologischen Abriebs von CrCoMo/PE/TiAlV – Implantaten (Mode IIAbrieb, 3 Knie, 1 Hüftprothese). Die intraoperativ gewonnenen Abriebpartikel wurden aufgereinigt und bezüglich ihrer Zusammensetzung und Größe charakterisiert (particle size analyser, Elektronenmikroskopie, Edx-Analyse, Histologie). Die Effekte der Partikel auf die Freisetzung von Zytokinen (PDGF, IL-1β, IL-8, TNF α) wurden mittels ELISA in einem etablierten, humanen Makrophagen-ähnlichen Zellkulturmodell untersucht.
Ergebnisse: Unabhängig von der Lokalisation der Prothese bestanden die metallernen Abriebpartikel aus TiAlV einer durchschnittlichen Größe von 0,1± 0,15 μm. CrCoMo-Partikel konnten nicht nachgewiesen werden. Im Zellkulturmodell verursachten 1456 × 108 Partikel pro 1 × 106 Makrophagen maximale Zytokinausschüttungen (8-fache TNFα -, 5-fache IL-8 und IL-1ß Ausschüttung) bei einer Überlebensrate der Zellen von mehr als 90 Prozent. Ein Partikel-abhängiger PDGF-Anstieg konnte nicht beobachtet werden.
Schlussfolgerung: Wie bereits für Mode I- Abriebpartikel gezeigt (Kontakt zwischen primär belasteten Oberflächen), verursachen auch Mode II- Abriebpartikel die Freisetzung von Knochenresorption auslösenden Zytokinen in einem Makrophagen-ähnlichen Zellkulturmodell. Weil ihre lokalen und systemischen Effekte in vivo nicht komplett verstanden sind, empfehlen wir die vollständige Entfernung von Abriebpartikeln bei Patienten mit Metallose, um mögliche immunologische Reaktionen des Körpers sowie periprothetische Osteolysen zu vermeiden.