title: Cellular and humoral mechanisms of osteoclast formation in Ewing's sarcoma.
abstract: Cellular mechanisms that account for tumour osteolysis associated with Ewing's sarcoma are uncertain. Osteoclasts are marrow-derived multinucleated cells (MNCs) that effect tumour osteolysis. Osteoclasts are known to form from macrophages by both receptor activator for nuclear factor-kappaB (RANK) ligand (RANKL)-dependent and -independent mechanisms. In this study, our aim has been to determine whether tumour-associated macrophages (TAMs) isolated from Ewing's sarcoma are capable of differentiating into osteoclasts and to characterise the cellular and humoral mechanisms whereby this occurs. Tumour-associated macrophages were isolated from two Ewing's sarcomas and cultured on both coverslips and dentine slices for up to 21 days with soluble RANKL and macrophage colony stimulating factor (M-CSF). Osteoclast formation from TAMs (CD14+) was evidenced by the formation of tartrate-resistant acid phosphatase (TRAP) and vitronectin receptor (VNR)-positive MNCs, which were capable of carrying out lacunar resorption. This osteoclast formation was inhibited by the addition of bisphosphonates. Both Ewing's sarcoma-derived fibroblasts and some bone stromal cells expressed RANKL and supported osteoclast formation by a contact-dependent mechanism. We also found that osteoclast differentiation occurred when Ewing's TAMs were cultured with tumour necrosis factor (TNF)-alpha in the presence of M-CSF and that TC71 Ewing's sarcoma cells stimulated osteoclast formation through the release of a soluble factor, the action of which was abolished by an antibody to TNF-alpha. These results indicate that TAMs in Ewing's sarcoma are capable of osteoclast differentiation by both RANKL-dependent and TNF-alpha-dependent mechanisms and that Ewing's sarcoma cells produce osteoclastogenic factor(s). Our findings suggest that anti-resorptive and anti-osteoclastogenic therapies may be useful in inhibiting the osteolysis of Ewing's sarcoma.
authors.affiliation: Department of Pathology, Nuffield Department of Orthopaedic Surgery, Nuffield Orthopaedic Centre, University of Oxford, Oxford OX3 7LD, UK.
authors.forename: Y S
authors.initials: YS
authors.lastname: Lau
authors.forename: I E
authors.initials: IE
authors.lastname: Adamopoulos
authors.forename: A
authors.initials: A
authors.lastname: Sabokbar
authors.forename: H
authors.initials: H
authors.lastname: Giele
authors.forename: C L M H
authors.initials: CL
authors.lastname: Gibbons
authors.forename: N A
authors.initials: NA
authors.lastname: Athanasou
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D001859:Bone Neoplasms / Q000235:genetics / Q000473:pathology*; D001862:Bone Resorption / Q000473:pathology; D001842:Bone and Bones / Q000166:cytology / Q000473:pathology; D002454:Cell Differentiation; D005260:Female; D006801:Humans; D008264:Macrophages / Q000502:physiology; D008297:Male; D008875:Middle Aged; D010010:Osteoclasts / Q000378:metabolism / Q000502:physiology*; D053244:Osteoprotegerin / Q000378:metabolism / Q000494:pharmacology; D053245:RANK Ligand / Q000378:metabolism / Q000494:pharmacology; D012512:Sarcoma, Ewing / Q000235:genetics / Q000473:pathology*; D017154:Stromal Cells / Q000166:cytology / Q000473:pathology; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000378:metabolism; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
chemical_list: D053244:Osteoprotegerin; D053245:RANK Ligand; D053221:TNF-Related Apoptosis-Inducing Ligand; C506030:TNFRSF11B protein, human; C505653:TNFSF10 protein, human; C505688:TNFSF11 protein, human; D014409:Tumor Necrosis Factor-alpha