Aseptic osteolysis is a major complication to total joint arthroplasty requiring several thousand people a year to have to undergo revisions of their joint prosthesis. The formation of the interfacial membrane has been associated with aseptic osteolysis leading to the failure of all types of total joints. Recent evidence suggests that RANKL, a potent activator of bone reabsorption, is present in the interfacial membrane. Prior research in this laboratory to determine the source of RANKL in the interfacial membrane has revealed the presence of intense areas of RANKL concentration in the membrane. These areas of RANKL concentration correspond to multiple nuclei and a distinct cellular structure in the tissue as determined through light microscopy. This structure either represents a multi-nucleated giant cell or a cluster of cells that express high concentrations of RANKL. These following studies attempted to characterize this cell and determine its lineage.The results of these studies show that the RANIU producing anomaly appears multi-nucleated in all examples with a RANKL staining pattern that makes it appear as a multi-nucleated cell. Furthermore this RANIU positive giant cell (RPGC) stains negative for markers typically seen on myeloid cells, osteoclasts, and osteoblasts. This RPGC does however stain very well for fibroblast markers and the inflammatory cytokines TNF-α, IL-1β, and IL-6. The most interesting result from these studies revealed that this cell was positive for cytomegalovirus and expressed high concentrations of TNF-a converting enzyme (TACE). These data lead to a hypothesis as to how this cell might form and how large an impact it might play in the interfacial membrane with respect to aseptic bone reabsorption.

Polyethylene wear and micromotion of the implant play an important role in multifactorial etiology of osteolysis leading to aseptic loosening of the acetabular components. Despite excellent results in primary total hip arthoplasty in a 10-15 year perspective there are still unsolved problems. The weakest link is the longevity of the actabular component. Young and active patients have a clearly worse outcome than older patients. Consequences of polyehtylene wear and ways to reduce wear have therefore been in focus during recent years. Radiostereometry (RSA) is the golden standard in measuring in vivo micromotions. In 4 clinical studies including 332 patients we used therefore RSA to record the efficacy of fixation of cemented and uncemented cups. The amount of wear of old and newly designed polyethylenes (PE) was related to cup stability and radiological and clinical measures of outcome. This study showed that cementless cups inserted with pressfit technique do not need additional augmentation. Screws and pegs increase the risk for radiolucencies and osteolystic lesions but are helpful tools in cases where primary stability is jeopardized. In the second decade clinically silent osteolysis is common for the porous coated Harris Galante cup with unsealed screw holes. The locking mechanism of the PE liner in this cup is unsatisfactory and an increase of liner dissociations is expected. EtO sterilized PE displayed high in vivo wear and we do not recommend its continued use but close monitoring of patients with earlier inserted EtO sterilized implants. The substantially reduced wear in cemented highly cross-linked PE cups without any negative in vivo tradeoffs might have a substantial impact on choice of material and operating technique in the near future. However, we still recommend its restrained use in controlled series until longer follow-up data is available. Nevertheless, the short term in vivo results of modern highly cross-linked PE look promising and ight improve the outcome of cemented and uncemented hip arthroplasties by reducing complication and revisions.

Background: Using a nude rat model of site-specific metastatic bone disease (MBD), we developed a semiquantitative histological score for rapid assessment of lytic lesions in bone. This provides additional information to conventional histological measurement by clarifying the extent and location of metastatic infiltration and the tumor growth pattern. The score can also be used to assess the action of bisphosphonates on bone metastases. Materials and Methods: Male nude rats (n = 12 per group) were inoculated with the human breast cancer cell line MDA-MB-231 via the femoral artery. Following appearance of radiographically visible osteolytic lesions on day 18, the animals received phosphate-buffered saline (PBS; controls) or ibandronate (IBN, 10 µg P/kg) daily until day 30. Whole body radiographs were obtained on days 18 and 30, and osteolytic areas (OA) were determined by radiographic computer-based analysis (CBA). On day 30, MBD was assessed in both tibias using conventional histological CBA and the new scoring system. Results: Metastatic tumor area correlated with the total sum of the new score in both PBS- (r = 0.762) and IBN-treated animals (r = 0.951; p < 0.001). OA correlated well with the total sum in both groups (r = 0.845 and 0.854, respectively; p < 0.001). Conclusion: Significant reduction of bone marrow and cortical infiltration of tumor cells with IBN suggested local control of metastases. / Hintergrund: Mit Hilfe eines etablierten Tiermodells zur Erzeugung lokalisationsspezifischer Knochenmetastasen in der Nacktratte wurde ein semiquantitatives histologisches Graduierungssystem zur schnellen Bewertung osteolytischer Knochenmetastasen entwickelt. Das Graduierungssystem liefert hinsichtlich der Metastasenlokalisation, deren Ausmaß und Infiltrationsmuster wertvolle Zusatzinformationen zu den konventionellen histologischen Untersuchungsmethoden. Damit kann beispielsweise auch die pharmakologische Wirkung von Bisphosphonaten auf die Knochenmetastasierung beurteilt werden. Material und Methoden: Männlichen Nacktratten (n = 12 pro Gruppe) wurden Zellen der humanen Brustkrebszellinie MDA-MB-231 in die Oberschenkelarterie inokuliert. Ab dem Auftreten radiologisch erkennbarer Osteolysen 18 Tage nach Inokulation erhielten die Tiere bis zum Studienende (Tag 30) täglich entweder eine subkutane Applikation einer Phosphat-Puffer-Lösung (Kontrollgruppe) oder Ibandronat (IBN, 10 µg P/kg; Behandlungsgruppe). Konventionelle Röntgenaufnahmen wurden an den Tagen 18 und 30 nach Tumorinokulation angefertigt und die Osteolysenflächen mittels Computerauswertung bestimmt. Nach Studienende wurde der Metastasenbefall in beiden Tibiae sowohl konventionell histologisch als auch mittels des neuen Graduierungssystems ausgewertet. Ergebnisse: Die Metastasenfläche korrelierte mit der kummulativen Punktsumme des Graduierungssystems sowohl in der Kontrollgruppe (r = 0,762; p < 0,001) als auch in der Ibandronat- Gruppe (r = 0,951; p < 0,001). Ebenso war die Osteolysenfläche eng mit der Punktesumme in beiden Gruppen korreliert (r = 0,845 und 0,854; p < 0,001). Schlussfolgerung: Die signifikante Reduktion von Knochenmark- und Kortikalisbefall durch IBN deuten auf eine gute lokale Kontrolle des Metastasenwachstums hin. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

Periprosthetic osteolysis remains the leading complication of total hip arthroplasty. It often results in aseptic loosening of the implant with a requirement for a revision surgery. Wear-generated particular debris is the main cause of initiating this destructive process. The most important cellular target for wear debris is a macrophage, which responds to particle challenge by activatig proinflamatory signals, which contribute to increased bone resorption. The activation of the RANKL/RANK/OPG system is considered to be a likely cause of periprosthetic osteolysis leading to implant failure. The aim of this study was to examine the possible correlation between the clinical extent of osteolysis, the number of wear particles and the expression of the osteoclastic mediator RANKL in the tissues around aseptically loosened cemented and non-cemened total hip replacements. Periprosthetic tissues were harvested from 59 patients undergoing revision hip replacement for aseptic loosening. We had observed RANKL-positive cells in 23 of our 59 patients, their presence was noted predominantly in tissues with a loosened cemented endoprosthesis. We have shown that RANKL is present only in the tissues with a large amount of wear debris and predominantly in the cases involving lacunar type of osteolysis. Key words:...

Orientador: Dagmar Ruth Stach Machado / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-21T19:21:12Z (GMT). No. of bitstreams: 1 Obando-Pereda_GustavoAlberto_D.pdf: 1140735 bytes, checksum: 2f2e2b586d91ad80806eab9cb7b24bf8 (MD5) Previous issue date: 2012 / Resumo: A reação biológica as partículas de desgaste é crítica para indução da perda asséptica do implante mediante a osteólise. Deste modo, foi objetivo deste trabalho avaliar o envolvimento dos receptores Toll-like receptors 2 e 4 no reconhecimento das partículas de titânio e zircônia. Mensurados em cultura de macrófagos murinos desafiados com as partículas de zircônia ou de titânio comparando a expressão de TLRs, seus adaptadores intracelulares e citocinas pró-inflamatórias. Em in vivo foram estudados a indução da osteolise utilizando o modelo de calvária e a geração da resposta inflamatória através da indução do edema e hiperalgesia. As partículas são prontamente fagocitadas pelos macrófagos em cultura, e resultam no aumento da expressão de RNAm para TLRs 2, 3, 4 e 9, os seus adaptadores MyD88 e NF-kB e das citocinas TNF-?, IL-1? e IL-6. Contudo, o padrão de expressão de RNAm para TLRs entre as partículas é distinto, enquanto a zircônia induz um aumento significativamente na expressão de TLR2, o titânio modula a expressão significativamente maior de TLR3, TL4 e TLR9, respectivamente. Todavia, a expressão do RNAm para a molécula adaptadora MyD88 envolvida na sinalização intracelular de TLR é estimulada em ambas as partículas e com uma cinética de expressão semelhante. O fator de transcrição NF-kB necessário para efetuar a expressão gênica das citocinas envolvidas na resposta inflamatória apresenta uma cinética de expressão distinta entre as partículas, na zircônia a expressão é imediata e alcança o máximo da expressão após duas horas, enquanto as partículas de titânio induzem um aumento exponencial do fator de transcrição. A expressão de RNAm das citocinas inflamatórias TNF-?, IL-1? e IL-6 induzidas pelas partículas de zircônia é significativamente menor em comparação com as partículas de ix titânio. Contudo a expressão protéica da citocina TNF-? é maior nas em cultura de macrófagos expostas as partículas de zircônia, enquanto as partículas de titânio induzem a expressão protéica das citocinas inflamatória IL- 6. Ambas as partículas são capazes de induzir osteólise no modelo da calvária, contudo, a osteolise induzida pela zircônia assim como a perda óssea foi significativamente menor em comparação com as partículas de titânio. Assim embora ambas as partículas induzam edema e hiperalgesia nos animais de experimentação contudo as partículas de titânio uma maior sensação de hiperalgesia. Com base nos nossos resultados sugerimos que a biocompatibilidade da zircônia é maior em comparação com o titânio, e a perda asséptica é modulada pelo reconhecimento mediado pelos TLRs os quais ativam as vias de sinalização intracelular como os fatores de transcrição NF-kB levando a expressão de citocinas inflamatórias / Abstract: The biological reaction to wear debris is critical to the osteolysis underlying aseptic loosening of prosthetic implants Therefore was the aim of this study was to evaluate the involvement of Toll-like receptors 2 and 4 in the recognition of titanium and zirconia particles. Measured in cultured murine macrophages challenged with particles of zirconia or titanium by comparing the expression of TLRs, their intracellular adaptors and the proinflammatory cytokines. Particle-induced osteolysis was evaluated in mice calvaria model, whereas the inflammatory responses through induction of hind paw edema and hyperalgesia. The particles are readily phagocytized by macrophages in culture and result in increased expression of mRNA for TLRs 2, 3, 4 and 9, its adaptors MyD88 and NF-kB, TNF-?, IL- 1? and IL-6 . However, the pattern of mRNA expression TLRs is distinct for the particles, while the zirconia induces a significant increase in expression of TLR2; titanium modulates the expression significantly greater TLR3, TLR9 and TL4, respectively. However, the mRNA for the adaptor molecule MyD88 involved in intracellular signaling of TLR is stimulated in both particles and with a similar kinetic. The transcription factor NF-kB is needed to carry the gene expression of cytokines involved in the inflammatory response has a different kinetics expression between the particles, whereas zirconia induces the expression with the maximum after two hours of incubation, the titanium particles induce an exponential increase. The mRNA expression of the inflammatory cytokines TNF-?, IL- 1? and IL-6 induced by the particles of zirconia is significantly lower compared with the particles of titanium. However, the protein expression of TNF-? is greater in cultured xi macrophages exposed to the particles of zirconia, while the titanium particles to induce protein expression of inflammatory cytokine IL-6. Both the particles induce osteolysis in the calvaria model however; the induced osteolysis by zirconia as well as the bone loss was significantly lower compared with titanium particles. So although both particles induce edema and hyperalgesia in animal models, titanium particles induced greater sense of hyperalgesia. Based on our results, we suggest that the biocompatibility of the zirconia is greater in comparison with the titanium and the loss is modulated by aseptic recognition mediated by TLRs which activate the intracellular signaling pathways as transcription factors NF-kB and leads to expression of inflammatory cytokines / Doutorado / Imunologia / Doutor em Genetica e Biologia Molecular

Periprosthetic osteolysis remains the leading complication of total hip arthroplasty. It often results in aseptic loosening of the implant with a requirement for a revision surgery. Wear-generated particular debris is the main cause of initiating this destructive process. The most important cellular target for wear debris is a macrophage, which responds to particle challenge by activatig proinflamatory signals, which contribute to increased bone resorption. The activation of the RANKL/RANK/OPG system is considered to be a likely cause of periprosthetic osteolysis leading to implant failure. The aim of this study was to examine the possible correlation between the clinical extent of osteolysis, the number of wear particles and the expression of the osteoclastic mediator RANKL in the tissues around aseptically loosened cemented and non-cemened total hip replacements. Periprosthetic tissues were harvested from 59 patients undergoing revision hip replacement for aseptic loosening. We had observed RANKL-positive cells in 23 of our 59 patients, their presence was noted predominantly in tissues with a loosened cemented endoprosthesis. We have shown that RANKL is present only in the tissues with a large amount of wear debris and predominantly in the cases involving lacunar type of osteolysis. Key words:...

BACKGROUND:Tumor cell lines that can be tracked in vivo during tumorigenesis and metastasis provide vital tools for studying the specific cellular mechanisms that mediate these processes as well as investigating therapeutic targets to inhibit them. The goal of this study was to engineer imageable mouse mammary tumor cell lines with discrete propensities to metastasize to bone in vivo. Two novel luciferase expressing cell lines were developed and characterized for use in the study of breast cancer metastasis to bone in a syngeneic mouse model.RESULTS:The 4 T1.2 luc3 and 66c14 luc2 cell lines were shown to have high levels of bioluminescence intensity in vitro and in vivo after orthotopic injection into mouse mammary fat pads. The 4 T1.2 luc3 cell line was found to closely model the sites of metastases seen in human patients including lung, liver, and bone. Specifically, 4 T1.2 luc3 cells demonstrated a high incidence of metastasis to spine, with an ex-vivo BLI intensity three orders of magnitude above the commercially available 4 T1 luc2 cells. 66c14 luc2 cells also demonstrated metastasis to spine, which was lower than that of 4 T1.2 luc3 cells but higher than 4 T1 luc2 cells, in addition to previously unreported metastases in the liver. High osteolytic activity of the 4 T1.2 luc3 cells in vivo in the bone microenvironment was also detected.CONCLUSIONS:The engineered 4 T1.2 luc3 and 66c14 luc2 cell lines described in this study are valuable tools for studying the cellular events moderating the metastasis of breast tumor cells to bone.

Background: Using a nude rat model of site-specific metastatic bone disease (MBD), we developed a semiquantitative histological score for rapid assessment of lytic lesions in bone. This provides additional information to conventional histological measurement by clarifying the extent and location of metastatic infiltration and the tumor growth pattern. The score can also be used to assess the action of bisphosphonates on bone metastases. Materials and Methods: Male nude rats (n = 12 per group) were inoculated with the human breast cancer cell line MDA-MB-231 via the femoral artery. Following appearance of radiographically visible osteolytic lesions on day 18, the animals received phosphate-buffered saline (PBS; controls) or ibandronate (IBN, 10 µg P/kg) daily until day 30. Whole body radiographs were obtained on days 18 and 30, and osteolytic areas (OA) were determined by radiographic computer-based analysis (CBA). On day 30, MBD was assessed in both tibias using conventional histological CBA and the new scoring system. Results: Metastatic tumor area correlated with the total sum of the new score in both PBS- (r = 0.762) and IBN-treated animals (r = 0.951; p < 0.001). OA correlated well with the total sum in both groups (r = 0.845 and 0.854, respectively; p < 0.001). Conclusion: Significant reduction of bone marrow and cortical infiltration of tumor cells with IBN suggested local control of metastases. / Hintergrund: Mit Hilfe eines etablierten Tiermodells zur Erzeugung lokalisationsspezifischer Knochenmetastasen in der Nacktratte wurde ein semiquantitatives histologisches Graduierungssystem zur schnellen Bewertung osteolytischer Knochenmetastasen entwickelt. Das Graduierungssystem liefert hinsichtlich der Metastasenlokalisation, deren Ausmaß und Infiltrationsmuster wertvolle Zusatzinformationen zu den konventionellen histologischen Untersuchungsmethoden. Damit kann beispielsweise auch die pharmakologische Wirkung von Bisphosphonaten auf die Knochenmetastasierung beurteilt werden. Material und Methoden: Männlichen Nacktratten (n = 12 pro Gruppe) wurden Zellen der humanen Brustkrebszellinie MDA-MB-231 in die Oberschenkelarterie inokuliert. Ab dem Auftreten radiologisch erkennbarer Osteolysen 18 Tage nach Inokulation erhielten die Tiere bis zum Studienende (Tag 30) täglich entweder eine subkutane Applikation einer Phosphat-Puffer-Lösung (Kontrollgruppe) oder Ibandronat (IBN, 10 µg P/kg; Behandlungsgruppe). Konventionelle Röntgenaufnahmen wurden an den Tagen 18 und 30 nach Tumorinokulation angefertigt und die Osteolysenflächen mittels Computerauswertung bestimmt. Nach Studienende wurde der Metastasenbefall in beiden Tibiae sowohl konventionell histologisch als auch mittels des neuen Graduierungssystems ausgewertet. Ergebnisse: Die Metastasenfläche korrelierte mit der kummulativen Punktsumme des Graduierungssystems sowohl in der Kontrollgruppe (r = 0,762; p < 0,001) als auch in der Ibandronat- Gruppe (r = 0,951; p < 0,001). Ebenso war die Osteolysenfläche eng mit der Punktesumme in beiden Gruppen korreliert (r = 0,845 und 0,854; p < 0,001). Schlussfolgerung: Die signifikante Reduktion von Knochenmark- und Kortikalisbefall durch IBN deuten auf eine gute lokale Kontrolle des Metastasenwachstums hin. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

No description available.

Le desserrage des tiges est une complication fréquente des plâtres avec tiges transcorticales (TP) chez les grands animaux, nécessitant souvent leur retrait prématuré avant la guérison des fractures. Les charges excessives centrées sur le cortex à l’interface os-tige proximo-externe et disto-interne causent de l'ostéolyse. En utilisant un modèle de veau nouveau-né, ce projet a évalué un nouveau système de tige-manchon et anneau integré dans un plâtre (PS) optimisé pour réduire la contrainte péri-implant et le stress à l'interface os-implant. On a émis l'hypothèse que les PS se traduiraient par une ostéolyse péri-implant moindre par rapport aux TP. Dix veaux en bonne santé, de 3 semaines d'âge, ont été implantés avec les TP ou PS dans le métacarpe droit, à raison de 2 implants par veau. Les veaux ont été observés quotidiennement pour le confort et la boiterie et ont été euthanasiés à 28 jours. Les données recueillies comprenaient les radiographies à la chirurgie et à l'euthanasie et les mesures histomorphométriques de contact os-implant sur des échantillons non-décalcifiés avec les implants in situ. Les données ont été analysées en utilisant le test de Cochran-Mantel-Haenszel, une valeur de P <0,05 a été considérée comme significative. L'épaisseur corticale était plus importante pour les implants distaux que proximaux pour les deux groupes lors de la chirurgie (P = 0,03), mais était similaire entre les groupes (P > 0,3). Les veaux avec TP ont développé une boiterie plus tôt (au jour 21) que les veaux avec PS (P = 0,04). Histologiquement, il y avait plus de contact direct os-implant cortical pour les implants PS distaux que les implants TP (P = 0,04). La jonction métaphyso-diaphysaire osseuse où les implants proximaux étaient situés est impropre aux deux systèmes; chacun a un minimum de contact os-implant et de l'ostéolyse extensive. Le système PS n'ayant pas causé une ostéolyse importante lorsque implantés dans l'os diaphysaire et peut-être une alternative convenable aux TP pour des fractures comminutives des membres distaux. / Pin loosening is a common complication of transfixation pincasts (TP) in large animals, often necessitating premature removal before fracture healing. The excessive loads centered on the proximo-external and disto-internal cortices of the bone-pin interface cause osteolysis. Using a neonatal calf model, this project evaluated a novel pin-sleeve and ring cast system (PS) optimized to decrease peri-implant strain and evenly share stress at the bone-implant interface. It was hypothesized that PS would result in less peri-implant osteolysis compared to TP. Ten, 3-week-old, healthy calves were implanted with either TP or PS in the right metacarpus, 2 implants per calf. Calves were scored daily for lameness and were euthanized at day 28. Collected data included radiographs at surgery and euthanasia and histomorphometric measures of bone-implant contact on non-decalcified specimens with the implants in situ. Data was analyzed using Cochran-Mantel-Haenszel test; a P-value <.05 was considered significant. The cortical thickness was larger for distal implants than proximal implants for both groups at surgery (P = 0.03), but were similar between groups (P > 0.3). TP calves developed lameness sooner, at day 21, than PS calves (P = 0.04). Histologically, there was more direct cortical bone-implant contact for PS distal implants than TP implants (P = 0.04). The metaphyseal-diaphyseal junction where the proximal implants were situated is unsuitable bone for either system; each had minimal bone-implant contact and extensive osteolysis. The PS system did not cause significant osteolysis when instrumented in diaphyseal bone and is a suitable alternative to TP for comminuted distal limb fractures.