Cytokine receptor profile of arthroplasty macrophages, foreign body giant cells and mature osteoclasts

In the arthroplasty pseudomembrane surrounding a loose prosthesis there is a marked macrophage and foreign body giant cell (FBGC) response to implant-derived wear particles. These cells contribute to the osteolysis of loosening by releasing cytokines and growth factors which influence the formation and activity of osteoclasts. Using a panel of monoclonal antibodies directed against known cytokine/growth factor receptors, we have determined by immunohis-tochemistry whether arthroplasty macrophages, FB-GCs and osteoclasts express receptors for cytokines and growth factors that are known to modulate osteolysis.

All these cell types reacted with antibodies directed against the following cytokine/growth factor receptors: gp130, IL-1R type 1, IL-2R, IL-4R, IL-6R, TNFR, M-CSFR, GM-CSFR and SCFR but not with antibodies directed against IL-3R and IL-8R. Arthroplasty macrophages, FBGCs and osteoclasts thus show a similar pattern of cytokine/growth factor receptor expression. This reflects the fact that arthroplasty macrophages are capable of osteoclast differentiation and that these cell types form part of the mononuclear phagocyte system. As regards the osteolysis of aseptic loosening, it also indicates that these cells are targets for numerous cytokines and growth factors which influence osteoclast formation and bone resorption.     

骨痂中多核巨细胞与破骨细胞的组织学及超微结构观察

本文通过光镜与透射电镜对家兔桡骨标准分析模型骨痂内多核巨细胞与破骨细胞形态进行了观察。观察显示，两种细胞均参与骨吸收，但多核细胞主要在分析早期吸收死骨和骨碎屑，吸收方式包括吞噬和细胞外降解；破骨细胞主要吸收钙化软骨骨痂和新生骨小粱，通过细胞外吸收方式完成骨痂改建。作者认为，多核巨细胞表面的丝状伪足和破骨细胞表面的皱折缘与骨矿物质细胞外降解有关，但其降解机理的差别尚有待揭示。     

Role of DC-STAMP in cellular fusion of osteoclasts and macrophage giant cells

Osteoclasts are the only cells that can resorb bone matrix physiologically and maintain the bone content. Osteoclasts are derived from macrophage/monocyte lineage cells; stimulation by macrophage colony stimulating factor and receptor activator of NFκB ligand induces osteoclastogenesis. During osteoclastogenesis, preosteoclasts fuse to form multinuclear mature osteoclasts. Cellular fusion is a unique phenomenon and enables fertilization, myotube formation, and efficient bone resorption in vertebrates. To date, several molecules have been reported to be fusion related in osteoclasts, namely CD44, CD47, ADAM12, MCP-1, and CD9, although the molecules which regulate osteoclast cellular fusion remain unclear. Here, we show that the seven-transmembrane-region receptor dendritic cell-specific transmembrane protein (DC-STAMP) is required for cell–cell fusion of osteoclasts and foreign body giant cells.     

The cells of giant cell tumor of tendon sheath resemble osteoclasts

To determine whether the mononuclear cells (MC) and multinucleated giant cells (GC) of giant cell tumor of tendon sheath (GCTTS) exhibit evidence of monocyte/macrophage lineage, we studied their antigenic features (seven cases, paraffin sections; two cases, frozen sections) and enzymatic features in situ (four cases, plastic sections). Both MC and GC expressed a monocyte/macrophage phenotype: HLA-A,B,C+, HLA-DR+, T200+ (leukocyte common antigen), Leu-M3+ and Leu-3+. MC and GC also expressed similar enzymatic phenotypes which resembled that of osteoclasts. Both were rich in acid phosphatase and contained smaller, variable amounts of ATPase, beta-glucuronidase, alpha-naphthyl acetate esterase, and 5'-nucleotidase. Both lacked alkaline phosphatase. Reactive osteoclasts in plastic and paraffin sections were also T200+, a finding strongly supporting their bone marrow derivation and leukocytic differentiation. In plastic sections, osteoclasts were additionally reactive with macrophage antigen EBM11. In aggregate, these data suggest that GCTTS is a true histiocytic lesion of monocyte/macrophage lineage composed of phenotypically similar MC and GC that most closely resemble osteoclasts. We found no evidence that GCTTS cells resemble osteoblasts, fibroblasts, or synovial sarcoma cells. Furthermore, expression of the Ki-67 nuclear antigen by 1-2% of MC but not by GC suggests that the proliferating cells in GCTTS are restricted to its MC component.     

Polyethylene content and cell size of foreign body giant cells in aseptic hip prosthesis loosening--a histomorphometric study

INTRODUCTION: In aseptic loosening of cementless hip arthroplasty polyethylene particles are phagocytized by macrophages and foreign-body giant cells. This initiates an osteolytic cascade. In this study, the authors investigated if there are correlations between the size of foreign-body giant cells and the polyethylene loading and number of intracellular particles as determined by light microscopy. MATERIAL AND METHOD: Histological specimens were processed from tissues retrieved during revision surgery of 7 cases of cementless hip arthroplasty which had become aseptically loose. The specimens were analyzed by histolomorphology and histomorphometry. The cell size, polyethylene loading and intracellular particle number of 111 foreign-body giant cells were determined. A regression analysis was performed to investigate if there was a correlation between these variables. RESULTS: The mean cell size was 1417 +/- 487 micron 2, the mean polyethylene loading was 49 +/- 42 micron 2 and the mean intracellular particle number was 10.4 +/- 5.4. The cell size correlated with both the intracellular particle number (r = 0.25) and the polyethylene loading (r = 0.39). CONCLUSIONS: In the cellular reaction to polyethylene particles in aseptic loosening of cementless total hip arthroplasty, the size of foreign-body giant cells correlates with intracellular polyethylene loading and intracellular polyethylene particle number as determined by light microscopy. The presence of large foreign body giant cells might be associated with a high particle load of the tissue and a high osteolytic activity.     

The role of macrophages and giant cells in loosening of joint replacement

Summary Macrophages and foreign-body giant cells from the bone-cement interface of loosened joint replacement were assayed for direct bone resorption in an in vitro experimental model. Human osteoclasts from giant cell tumors and experimental animal osteoclasts were used as controls. Macrophages and giant cells were not capable of resorbing the bone under experimental conditions, but osteoclasts are. It is suggested that macrophages and giant cells in loosened implants may initiate bone resorption, removing the extra-cellular barrier that normally protects mineral crystals from osteoclastic recognition.

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Zusammenfassung Makrophagen und Fremdkörperriesenzellen von der Knochen/Zementgrenze gelokkerter Gelenkendoprothesen wurden hinsichtlich der direkten Knochenresorption in einem experimentellen in vitro-Modell untersucht. Als Kontrollen dienten Osteoklasten aus menschlichen Riesenzelltumoren und von embryonalen Hühnerknochen. Während Osteoklasten unter den experimentellen Bedingungen Knochen resorbierten waren Makrophagen und Riesenzellen dazu nicht in der Lage. Es wird angenommen, daß Makrophagen and Riesenzellen bei gelockerten Implantaten die Knochenresorption anbahnen, indem sie die extrazelluläre Barriere entfernen, welche normalerweise die Mineralkristalle dagegen schützen von den Osteoklasten als solche erkannt zu werden.

     

The role of macrophages and giant cells in loosening of joint replacement

Macrophages and foreign-body giant cells from the bone-cement interface of loosened joint replacement were assayed for direct bone resorption in an in vitro experimental model. Human osteoclasts from giant cell tumors and experimental animal osteoclasts were not capable of resorbing the bone under experimental conditions, but osteoclasts are. It is suggested that macrophages and giant cells in loosened implants may initiate bone resorption, removing the extra-cellular barrier that normally protects mineral crystals from osteoclastic recognition.     

Identification,characterization, and isolation of a common progenitor for osteoclasts,macrophages, and dendritic cells from murine bone marrow and periphery

Osteoclasts are specialized bone‐resorbing cells that derive from monocyte precursors. We have identified three populations of cells with high osteoclastogenic potential in murine bone marrow, which expressed the phenotype B220^‐CD3^‐CD11b^?/low CD115⁺ and either CD117^hi, CD117^intermediate, or CD117^low. We have evaluated these populations for their ability to also generate macrophages and dendritic cells. At a single‐cell level, the population expressing higher CD117 levels was able to generate bone‐resorbing osteoclasts, phagocytic macrophages, and antigen‐presenting dendritic cells in vitro with efficiencies of more than 90%, indicating that there exists a common developmental pathway for these cell types. Cells with osteoclastogenic potential also exist in blood and peripheral hematopoietic organs. Their functional meaning and/or their relationship with bone marrow progenitors is not well established. Hence, we characterized murine peripheral cell populations for their ability to form osteoclasts, macrophages, and dendritic cells in vitro. The spleen and peripheral blood monocyte progenitors share phenotypic markers with bone marrow progenitors but differ in their expression of CD11b, which was low in bone marrow but high in periphery. We propose that circulating monocyte progenitors are derived from a common bone marrow osteoclasts/macrophage/dendritic cell progenitor (OcMDC), which we have now characterized at a clonal level. However, the lineage relationship between the bone marrow and peripheral monocyte progenitors has yet to be defined. © 2013 American Society for Bone and Mineral Research.     

The origin of osteoclasts: An immunohistochemical study on macrophages and osteoclasts in embryonic rat bone

Summary The origin of osteoclasts was studied in embryonic rat bone primordia using a set of monoclonal antibodies (ED1, ED2, and ED3) that exclusively recognize monocytes and macrophages. ED1 recognizes monocytes and macrophages. Mononuclear phagocytes which were ED1 positive were found in the perichondrium/periosteum of developing bone. These cells started to infiltrate the primordia when the cartilage became hypertrophic. During bone formation, multinucleated ED1-positive cells with the morphological characteristics of osteoclasts were found in the developing bone marrow cavity and against the bone collar. The present findings support the notion that osteoclasts arise by fusion of mononuclear phagocytes derived from blood monocytes.     

Development of an in vitro culture method for stepwise differentiation of mouse embryonic stem cells and induced pluripotent stem cells into mature osteoclasts

The development of methods for differentiation of embryonic stem cells (ESCs) and induced pluripotent stem cell (iPSCs) into functional cells have helped to analyze the mechanism regulating cellular processes and to explore cell-based assays for drug discovery. Although several reports have demonstrated methods for differentiation of mouse ESCs into osteoclast-like cells, it remains unclear whether these methods are applicable for differentiation of iPSCs to osteoclasts. In this study, we developed a simple method for stepwise differentiation of mouse ESCs and iPSCs into bone-resorbing osteoclasts based upon a monoculture approach consisting of three steps. First, based on conventional hanging-drop methods, embryoid bodies (EBs) were produced from mouse ESCs or iPSCs. Second, EBs were cultured in medium supplemented with macrophage colony-stimulating factor (M-CSF), and differentiated to osteoclast precursors, which expressed CD11b. Finally, ESC- or iPSC-derived osteoclast precursors stimulated with receptor activator of nuclear factor-B ligand (RANKL) and M-CSF formed large multinucleated osteoclast-like cells that expressed tartrate-resistant acid phosphatase and were capable of bone resorption. Molecular analysis showed that the expression of osteoclast marker genes such as Nfatc1, Ctsk, and Acp5 are increased in a RANKL-dependent manner. Thus, our procedure is simple and easy and would be helpful for stem cell-based bone research.     

Rapamycin-treated mature dendritic cells have a unique cytokine secretion profile and impaired allostimulatory capacity

Rapamycin (RAPA, sirolimus) is a recently introduced immunosuppressive agent. Its effect on the differentiation and antigen uptake of immature dendritic cells (iDCs) has been studied. However, whether it can also modulate the function of mature DCs (mDCs) is unknown. We investigated the effects of RAPA on rat bone marrow-derived DCs at different stages of maturation. RAPA affected maturation, increased apoptosis and reduced lipopolysaccharide (LPS)-induced IL-12 and IL-10 production in iDCs. However, mDCs were resistant to RAPA-induced apoptosis. RAPA-mDCs produced significantly less IL-10 and TNF-α when compared with mature DCs but similar amounts of IL-12. RAPA did not affect constitutive NF-κB activity, but inhibited allostimulatory activity in mature DCs. In conclusion, mDCs treated with RAPA are reprogrammed to produce a unique cytokine secretion profile and exhibit low allostimulatory capacity, which may play an important role in rapamycin-based immunomodulation.     

Radio-opaque marker of a surgical sponge appearing as an intra-articular foreign body after total hip arthroplasty

We report a 73-year-old female patient who underwent total hip arthroplasty for osteoarthritis of the right hip. An anteroposterior radiograph obtained 1 week after surgery showed an intra-articular striated foreign body. This was subsequently removed and found to be a radio-opaque marker of a surgical sponge that had been used during the operation. The patient made an uneventful recovery. In recent years, cases of retained surgical sponge after surgery have been reported occasionally. Counting surgical sponges and using sponges with radio-opaque markers are methods for preventing them from being accidentally left in situ. However, to our knowledge, there has been no report of retention of a surgical sponge radio-opaque marker at the operation site, appearing as a foreign body on an X-ray film.     

The giant cells recruited by subcutaneous implants of mineralized bone particles and slices in rabbits are not osteoclasts.

We have compared structural and functional characteristics of native osteoclasts and the multinucleated giant cells (MNGC) recruited by subcutaneous implants of mineralized bone particles and slices in normal rabbits. Weekly evaluation of the implants for 5 weeks showed distinct differences between MNGC and osteoclasts in the host with respect to morphology and the ability to stain for tartrate-resistant acid phosphatase and acid ATPase. An osteoclast-specific monoclonal antibody bound strongly to osteoclasts but not MNGC. Ground bone slices similarly implanted were surrounded by MNGC but did not show resorption pits by scanning electron microscopy. These data show that the MNGC recruited to subcutaneous implants of mineralized bone particles and slices lack the enzymatic, cell surface, and functional features of osteoclasts.     

Antagonistic effects of different classes of bisphosphonates in osteoclasts and macrophages in vitro.

Nitrogen-containing bisphosphonates, such as alendronate and ibandronate, inhibit bone resorption by preventing protein prenylation in osteoclasts, whereas non-nitrogen-containing bisphosphonates, such as clodronate, are metabolized to nonhydrolyzable analogs of ATP, resulting in osteoclast apoptosis. Because these two classes of bisphosphonates have different molecular mechanisms of action, we examined in vitro whether combined treatment with clodronate and alendronate would alter antiresorptive effectiveness. Although, in cultures of rabbit osteoclasts, the antiresorptive effect of 10 microM alendronate was increased by the addition of clodronate, the effect of higher concentrations of alendronate was not altered by addition of clodronate. Furthermore, the inhibition of protein prenylation in osteoclasts caused by higher alendronate concentrations was partially prevented by cotreatment with clodronate. As in osteoclasts, the inhibition of protein prenylation in J774 cells caused by alendronate or ibandronate treatment was dose-dependently prevented by cotreatment with clodronate. Furthermore, alendronate-induced J774 apoptosis was significantly inhibited in the presence of clodronate. The presence of clodronate also decreased the short-term cellular uptake of [14C]ibandronate. These observations suggest that combined treatment with clodronate could enhance the antiresorptive effect of a low concentration of nitrogen-containing bisphosphonate, but clodronate can also antagonize some of the molecular actions and effects of higher concentrations of nitrogen-containing bisphosphonates. The exact molecular basis for the antagonistic effects between bisphosphonates remain to be determined, but could involve competition for cellular uptake by a membrane-bound transport protein.     

Migration and phenotypic transformation of osteoclast precursors into mature osteoclasts: the effect of a bisphosphonate

Osteoclast-devoid bone explants were cultured together with embryonic liver as a source of osteoclast precursors, but separated from each other by a filter. Cells migrated through the filter toward the calcified matrix and acquired the characteristics of mature, tartrate-resistant acid phosphatase-positive (TRAP+) osteoclasts upon contact with the bone explant. Migration and attachment could be visualized separately. Progressive reduction of filter pore size resulted in progressive reduction of resorption because the use of smaller pores made it increasingly difficult for cells to pass. Indeed, the use of 0.22-micron filters, through which no cells can pass, but which still allow full passage of medium, completely blocked the resorption. When migrating cells from fetal liver were arrested for 10 days by using a combination of filters with different pore sizes, the arrested cells showed a tendency to fuse just opposite the mineralized matrix. Furthermore, a great number of the arrested cells expressed the macrophage-specific cell-surface antigen F4/80 and showed acid phosphatase activity, but none of these cells were tartrate resistant. The acquisition of tartrate-resistant acid phosphatase activity upon contact with the bone explant and subsequent resorption of this explant could be prevented by exposure of the system to the bisphosphonate dimethyl-APD (Me2-APD), whereas migration of cells through the filter was not affected. We suggest that the bisphosphonate interferes with a matrix factor that is essential for the attachment and subsequent transformation of the osteoclast precursor into the mature phenotype.