Nitric Oxide Donor Increases Osteoprotegerin Production and Osteoclastogenesis Inhibitory Activity in Bone Marrow Stromal Cells from Ovariectomized Rats

Abstract

Nitric oxide (NO) has emerged as a potent regulator useful in alleviating estrogen deficiency bone loss. Osteoprotegerin (OPG) and receptor activator of nuclear factor-kappaB ligand (RANKL) play important roles in regulating osteoclastogenesis. Although recent studies have reported NO donor attenuation of bone loss, the effect of NO donor on OPG and RANKL expression of osteogenic stromal cells and bone microenvironment in ovariectomized rats is not fully understood. Here, we showed that optimal NO donor treatment [2,2'-(hydroxynitrosohydrazino)bis-ethanamine; 15 microm] increased OPG, but not RANKL, levels in bone marrow stromal cells from ovariectomized rats. NO donor augmentation of OPG synthesis was transcriptionally mediated. The stimulatory action of NO donor on OPG expression appeared to be regulated by tyrosine kinase-dependent activation of Cbfa1/Runx2 binding to the OPG promoter, because cell cultures pretreated with tyrosine kinase inhibitor (herbimycin A), but not with protein kinase A inhibitor (calphostain C) or protein kinase C inhibitor [(Rp)-cAMP] significantly reduced NO-augmented Runx2 activation and OPG levels. Conditioned medium from NO donor-treated cells inhibited macrophage-colony-stimulating factor and RANKL-induced osteoclast formation of macrophage-colony-stimulating factor-dependent bone marrow macrophages. Neutralization with anti-OPG antibodies abolished the inhibitory effect of conditioned medium on osteoclastogenesis. Immunohistochemical observa…