Involvement of p38 Mitogen-activated Protein Kinase Signaling Pathway in Osteoclastogenesis Mediated by Receptor Activator of NF-κB Ligand (RANKL)

Abstract

The receptor activator of NF-B ligand (RANKL) induces osteoclast differentiation from bone marrow cells in the presence of macrophage colony-stimulating factor. We found that treatment of bone marrow cells with SB203580 inhibited osteoclast differentiation via inhibition of the RANKL-mediated signaling pathway. To elucidate the role of p38 mitogen-activated protein (MAP) kinase pathway in osteoclastogenesis, we employed RAW264 cells which could differentiate into osteoclastlike cells following treatment with RANKL. In a dose-dependent manner, SB203580 but not PD98059, inhibited RANKL-induced differentiation. Among three MAP kinase families tested, this inhibition profile coincided only with the activation of p38 MAP kinase. Expression in RAW264 cells of the dominant negative form of either p38␣ MAP kinase or MAP kinase kinase (MKK) 6 significantly inhibited RANKL-induced differentiation of the cells. These results indicate that activation of the p38 MAP kinase pathway plays an important role in RANKLinduced osteoclast differentiation of precursor bone marrow cells.Bone morphogenesis, remodeling, and resorption are controlled in part by osteoclasts. These cells differentiate from hematopoietic myeloid precursors of monocyte/macrophage lineage under control of osteotropic hormones and local factors produced by supporting cells such as osteoblasts and stromal cells (1-9).The receptor activator of NF-B ligand (RANKL) 1 (10), also refereed to as osteoclast differentiation factor (11), tumor necrosis factor-related activation-induced cytokine (12), or osteoprotegerin ligand (13), was shown to be highly expressed in supporting cells and to directly induce the differentiation and maturation of osteoclasts (7,(13)(14)(15). To describe RANKL-induced osteoclastogenesis, the sequential phenotype progression model was proposed. The model includes the appearance of mononuclear osteoclasts, the fusion process prior to multinucleated osteoclast formation, and the osteoclast maturation process (6). Moreover, it has been shown that mutant mice disrupted with either RANKL or its receptor RANK revealed severe osteopetrosis and osteoclast defects (16,17), indicating that the RANKL-RANK signaling system plays an essential role in osteoclast differentiation.It has been shown recently that RANK is associated with tumor necrosis factor receptor-associated factors (TRAFs) (18 -21). The intracellular domain of RANK contains two distinct TRAF-binding domains, each of which recognizes different TRAF proteins specifically (18,19). While the C-terminal region of RANK interacts with TRAF2 and TRAF5, the TRAF6-binding domain resides in the middle of the RANK intracellular region. Overexpression of RANK C-terminal deletion mutants has revealed that activation of the RANK-mediated signaling pathway results in the activation of NF-B and c-Jun N-terminal kinase (JNK) which correlate with the TRAF6 interaction activity of mutants (18,19). In addition, mice with disrupted TRAF6 gene exhibit an osteopetrotic phenotype due to a defect in bone resorptio...

Decoy receptor 3 (DcR3) induces osteoclast formation from monocyte/macrophage lineage precursor cells

“…Although DcR3 can trigger ERK and p38 MAPK signals, it did not activate JNK phosphorylation within 1 h incubation. Consistent with previous reports, 16,[41][42][43] we confirmed the ability of RANKL and M-CSF to activate all three types of MAPKs. The differential effect on JNK activation again suggests distinct signaling mechanisms transduced by DcR3 and M-CSF/RANKL.…”

“…26,[41][42][43] To elucidate the signaling pathways underlying DcR3 action, we examined the activation of MAPKs in RAW264.7 cells treated with DcR3, RANKL, or M-CSF by immunoblotting. Similar to the stimulation by RANKL and M-CSF alone, DcR3 at 3 mg/ml markedly induced the phosphorylation of ERK and p38 MAPK (Figure 4a, b).…”

Inhibitory effect of luteolin on osteoclast differentiation and function

“…Activated p38 MAPK phosphorylates transcription factor ATF2, which, in turn, induces transcription of the target genes (Cobb and Goldsmith 1995;Whitmarsh and Davis 1996). It was shown that the expression of dominant-negative forms of p38 MAPK and MKK6 in RAW264.7 cells inhibited RANKL-induced differentiation of RAW264.7 cells into osteoclasts (Matsumoto et al 2000). Moreover, addition of luteolin to osteoclast cultures strongly inhibited the expression of NFATc1, which is a key transcription factor for the expressions of TRAP and other osteoclastogenesis-associated genes (Ikeda et al 2004;Matsumoto et al 2004;Sharma et al 2007).…”

Osteoclast differentiation requires TAK1 and MKK6 for NFATc1 induction and NF-κB transactivation by RANKL

“…10 The involvement of p38 in osteoclastogenesis has been demonstrated by the treatment of bone marrow precursor cells with pharmacological inhibitors of the kinase. [11][12][13] The expression of dominant-negative (DN) form of p38a into RAW264 cells partially inhibited RANKL-induced osteoclastic differentiation of these cells. 13 Interestingly, the activation of p38 MAPK by RANKL was observed in bone marrow precursor cells but not in fully differentiated Ocs.…”