The role of phosphoinositide 3-kinase in spreading osteoclasts induced by colony-stimulating factor-1

Abstract

Background: Colony-stimulating factor-1 (CSF-1), a growth and survival factor for osteoclasts, stimulates these cells to spread and migrate towards a gradient of CSF-1. This may support the translocation of osteoclasts to new sites on the bone surface to be resorbed. Phosphoinositide 3-kinase (PI 3-K) is a lipid kinase participating in various signal transduction pathways. Objective: To investigate the role of PI 3-K in the CSF-1-induced spreading of osteoclasts. Methods: In isolated rat osteoclasts treated with or without CSF-1, the distribution of PI 3-K and proteins phosphorylated on tyrosine were investigated using immunofluorescence. In murine osteoclast-like cells grown from bone marrow cells co-cultured with osteoblasts, the activation of the PI 3-K by CSF-1 was determined both in vivo and in vitro. In vivo, the enzyme product in the cell was determined after extraction and separation with thin layer chromatography; in vitro, PI 3-K activity was measured in the pellet immunoprecipitated from the cell lysate. Results: Inhibition of PI 3-K blocked the CSF-1-induced spreading of osteoclasts. In spreading osteoclasts, a portion of PI 3-K was translocated to the periphery where proteins phosphorylated on tyrosine appeared simultaneously. In osteoclast-like cells, CSF-1 stimulated PI 3-K activity. This activity could be immunoprecipitated with antibody against phophotyrosine residues. Conclusion: PI 3-K participates in the CSF-1-induced spreading of osteoclasts. The activated PI 3-K may induce the reorganization of the cytoskeleton resulting in spreading and migration.

Molecular regulation of osteoclast activity

“…7). Osteoclasts stimulated with M-CSF undergo Srcdependent cytoskeletal reorganization, cell spreading and migration [187,188] and M-CSF treatment induces the membrane relocation of PI3-K [189] and decreases bone resorption [190]. Consistent with this, the migration of PI3-K null macrophages is disrupted in response to M-CSF [191,192].…”

M-CSF Regulates the Cytoskeleton via Recruitment of a Multimeric Signaling Complex to c-Fms Tyr-559/697/721

“…1C). These data, coupled with the information that two specific inhibitors of p85, wortmannin or LY294002, inhibited cytoskeletal organization (31), suggested that recruitment of PI3K to c-Fms could be critical in mediating M-CSF-dependent cytoskeletal organization. To test this hypothesis, we generated OCs from primary BMMs transduced with the WT or the Phe-721-mutated chimera in the presence of RANKL and M-CSF and then treated these cells with Epo to examine subsequent cytoskeletal reorganization dependent on the chimeric EpoR/ c-Fms receptor.…”

“…Although PI3K mediates M-CSF-dependent cytoskeletal organization (31,32), the c-Fms tyrosine residue/s that regulate the cytoskeletal reorganization in response to M-CSF in OCs have not been identified. To address this issue, we generated a chimeric construct comprising the external domain of the EpoR linked to the transmembrane and cytoplasmic domain of c-Fms.…”

Regulation of Embryonic and Postnatal Development by the CSF-1 Receptor

“…In combination with RANKL, CSF-1 drives OC differentiation (Arai et al, 1999; Nakagawa et al, 1998), sustains the survival of mature OC (Bouyer et al, 2007; Fuller et al, 1993), and stimulates their spreading (Palacio & Felix, 2001; Sakai et al, 2006) and migration (Fuller et al, 1993). In vitro, the effects of CSF-1 during OC differentiation are biphasic, with low concentrations (in the normal physiological range) enhancing OC formation and high concentrations promoting macrophage differentiation (Hodge, Kirkland, & Nicholson, 2007; Perkins & Kling, 1995).…”