title: MLO-Y4 osteocyte-like cells support osteoclast formation and activation.
abstract: Osteocytes are terminally differentiated cells of the osteoblast lineage that have become embedded in mineralized matrix and may send signals that regulate bone modeling and remodeling. The hypothesis to be tested in this study is that osteocytes can stimulate and support osteoclast formation and activation. To test this hypothesis, an osteocyte-like cell line called MLO-Y4 and primary murine osteocytes were used in coculture with spleen or marrow cells. MLO-Y4 cells support osteoclast formation in the absence of 1,25-dihydroxyvitamin D3 [1,25(OD)2D3] or any other exogenous osteotropic factor. These cells alone stimulate osteoclast formation to the same extent or greater than adding 1,25(OH)2D3. Coaddition of 1,25(OH)2D3 with MLO-Y4 cells synergistically increased osteoclast formation. Optimal osteoclast formation and pit formation on dentine was observed with 200-1,000 MLO-Y4 cells per 0.75-cm2 well. No osteoclast formation was observed with 2T3, OCT-1, or MC3T3-E1 osteoblast cells (1,000 cells/well). Conditioned media from the MLO-Y4 cells had no effect on osteoclast formation, indicating that cell contact is necessary. Serial digestions of 2-week-old mouse calvaria yielded populations of cells that support osteoclast formation when cocultured with 1,25(OH)2D3 and marrow, but the population that remained in the bone particles supported the greatest number of osteoclasts with or without 1,25(OH)2D3. To examine the mechanism whereby these cells support osteoclast formation, the MLO-Y4 cells were compared with a series of osteoblast and stromal cells for expression of macrophage colony-stimulating factor (M-CSF), RANKL, and osteoprotegerin (OPG). MLO-Y4 cells express and secrete large amounts of M-CSF. MLO-Y4 cells express RANKL on their surface and their dendritic processes. The ratio of RANKL to OPG mRNA is greatest in the MLO-Y4 cells compared with the other cell types. RANK-Fc and OPG-Fc blocked the formation of osteoclasts by MLO-Y4 cells. These studies suggest that both RANKL and OPG may play a role in osteocyte signaling, OPG and M-CSF as soluble factors and RANKL as a surface molecule that is functional in osteocytes or along their exposed dendritic processes.
authors.affiliation: Department of Medicine, University of Texas Health Science Center, San Antonio, USA.
authors.forename: S
authors.initials: S
authors.lastname: Zhao
authors.forename: Y Kato Y
authors.initials: YK
authors.lastname: Zhang
authors.forename: S
authors.initials: S
authors.lastname: Harris
authors.forename: S S
authors.initials: SS
authors.lastname: Ahuja
authors.forename: L F
authors.initials: LF
authors.lastname: Bonewald
mesh_terms: D000818:Animals; D001854:Bone Marrow Cells / Q000187:drug effects; D002117:Calcitriol / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism; D002454:Cell Differentiation / Q000502:physiology; D002478:Cells, Cultured; D018920:Coculture Techniques / Q000379:methods; D017077:Culture Media, Conditioned / Q000494:pharmacology; D006023:Glycoproteins / Q000378:metabolism; D016173:Macrophage Colony-Stimulating Factor / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism; D051379:Mice; D010010:Osteoclasts / Q000166:cytology / Q000502:physiology*; D010011:Osteocytes / Q000166:cytology* / Q000378:metabolism; D053244:Osteoprotegerin; D053245:RANK Ligand; D053246:Receptor Activator of Nuclear Factor-kappa B; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism; D018124:Receptors, Tumor Necrosis Factor; D012886:Skull / Q000166:cytology; D013499:Surface Properties
chemical_list: D002352:Carrier Proteins; D017077:Culture Media, Conditioned; D006023:Glycoproteins; D008562:Membrane Glycoproteins; D053244:Osteoprotegerin; D053245:RANK Ligand; D053246:Receptor Activator of Nuclear Factor-kappa B; D018160:Receptors, Cytoplasmic and Nuclear; D018124:Receptors, Tumor Necrosis Factor; C505687:Tnfrsf11a protein, mouse; C506031:Tnfrsf11b protein, mouse; C505689:Tnfsf11 protein, mouse; D016173:Macrophage Colony-Stimulating Factor; D002117:Calcitriol