Bone morphogenic protein 2 directly enhances differentiation of murine osteoclast precursors

Abstract

Previous studies found that BMPs support osteoclast formation, but it is not clear whether this is a direct effect on osteoclasts or mediated indirectly through osteoblasts. We have shown that a mouse deficient for the BMP antagonist Twisted gastrulation suggested a direct positive role for BMPs on osteoclastogenesis. In this report, we further determine the significance of BMP signaling on osteoclast formation in vitro. We find that BMP2 synergizes with suboptimal levels of RANKL to enhance in vitro differentiation of osteoclast-like cells. The enhancement by BMP2 is not a result of changes in the rate of proliferation or survival of the bone marrow derived cultures, but is accompanied by an increase in expression of genes involved in osteoclast differentiation and fusion. Treatment with BMP2 did not significantly alter expression of RANKL or OPG in our osteoclast cultures, suggesting that the enhancement of osteoclastogenesis is not mediated indirectly through osteoblasts or stromal cells. Consistent with this, we detected phosphorylated SMAD1,5, in the nuclei of mononuclear and multinucleated cells in osteoclast cultures. Levels of p-SMAD, BMP2 and BMP receptors increased during differentiation. RNAi suppression of Type II BMP receptor inhibited RANKL-stimulated formation of multinuclear TRAP positive cells. The BMP antagonist noggin inhibited RANKL-mediated osteoclast differentiation when added prior to day 3, while addition of noggin on day 3 or later failed to inhibit their differentiation. Taken together, these data indicate that osteoclasts express BMP2 and BMP receptors, and that autocrine BMP signaling directly promotes the differentiation of osteoclasts-like cells. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author ManuscriptBone is a highly dynamic tissue, characterized by a continuous cycle of bone formation by osteoblasts and bone resorption by osteoclasts [Henriksen et al., 2009;Sims and Gooi, 2008]. This cycle permits physiological bone growth, repair of damaged bone, and is important for regulation of systemic calcium and phosphate levels. Dysregulation of the cycle results in numerous pathological conditions such as osteoporosis, Paget's disease, and arthritis [Rodan and Martin, 2000]. It also contributes to the progression and morbidity of osteolytic cancers such as myeloma, osteosarcoma and metastatic breast, lung and prostate tumors [Guise et al., 2006;Roodman, 2009].Osteoclasts are large, multinucleated cells formed by fusion of mononuclear cells that derive from the monocyte/macrophage lineage [Vaananen and Laitala-Leinonen, 2008]. They produce proteases and other factors to degrade the inorganic mineral and organic protein components of bone, thereby facilitating repair and remodeling [Teitelbaum, 2000]. Two factors that are necessary and sufficient for osteoclast formation are M-CSF [Cecchini et al., 1997] and Receptor Activator of NF-κB Ligand (RANKL) [Wada et al., 2006], both of which are expressed by osteoblasts. M-CSF is required for survival and proliferation of...

TGF-β Family Signaling in Connective Tissue and Skeletal Diseases

“…The high-bone-mass phenotype in mice with osteoblast-specific Bmpr1a silencing was found to be triggered by impaired expression of sclerostin (SOST) (Kamiya et al 2008), an antagonist of Wnt signaling that acts by sequestering the co-receptor low-density lipoprotein receptor-related protein 5 (LRP5) into inactive complexes (van Bezooijen et al 2007;Kamiya et al 2010;Krause et al 2010), as also discussed below in the context of sclerosteosis and Van Buchem disease. In addition, treatment of osteoclastic precursors with BMP induces differentiation along the osteoclast cell lineage, and further stimulates expression of key enzymes for bone degradation (Kaneko et al 2000;Jensen et al 2010). The observation that BMP can regulate both bone formation and bone resorption is consistent with the notion that, with the exception of fibrodysplasia ossificans progressiva (FOP), mutations in BMP signaling components do not generally cause anabolic bone diseases.…”

“…This effect is in part mediated by the BMP-dependent increase of OPG expression in osteoblasts, and consequent suppression of osteoclast differentiation (Kaneko et al 2000;Wan et al 2001;Jensen et al 2010;Kamiya and Mishina 2011). However, BMP signaling also regulates bone resorption, as is apparent by the high bone mass phenotype when Bmpr1a expression is specifically inactivated in osteoblasts, which results in failed osteoclastogenesis (Kamiya et al 2008).…”

Modulation of Stromal Cell-Derived Factor-1/CXC Chemokine Receptor 4 Axis Enhances rhBMP-2-Induced Ectopic Bone Formation

“…45 Osteoclastogenesis could then be occurring in the ectopically implanted ACS site due to differentiation of HPCs (monocyte-macrophage lineage) to osteoclasts, particularly in the presence of rhBMP-2 and rhBMP-2 induced osteoblastic differentiation of recruited MSCs. 55 Reports by other groups have previously shown evidence that SDF-1 plays a crucial regulatory role in BMP-2 induced osteogenic differentiation of MSCs. Hosogane et al showed that BMP-2-induced alkaline phosphatase (ALP) activity, osteocalcin (OC) synthesis, and calcium deposition was significantly decreased by blocking the SDF-1/CXCR4 signal axis with AMD3100.…”

Molecular Variations Related to the Regional Differences in Periosteal Growth at the Mandibular Ramus

“…We found the opposite in this study, which suggests that BMP2 may also be involved in regulating periosteal bone resorption. In recent years, studies have in fact found that BMP2 can enhance osteoclastogenesis indirectly by stimulating osteoblasts or stromal cells to produce osteoclast-promoting factors, such as RANKL (Abe et al, 2000;Otsuka et al, 2003), or directly by activating osteoclast differentiation in the presence of RANKL (Jensen et al, 2009). It is hence likely that stronger expression of BMP2 at the anterior periosteal region plays an assisting role in promoting osteoclastogenesis by synergizing with RANKL.…”