Osteoclasts are multinucleated cells that are formed by the fusion of mononuclear osteoclasts, which is an essential process in bone resorption leading to bone remodeling. Herein we show that GM-CSF promoted the fusion of prefusion osteoclasts (pOCs). The expression of GM-CSF receptor-α was significantly up-regulated at the fusion stage of pOCs induced by RANKL. GM-CSF induced the expression of dendritic cell-specific transmembrane protein (DC-STAMP), which was mediated by inducing NFATc1 via induction of c-Fos. The expression of c-Fos and NFATc1 was regulated by the ERK signaling pathway. Inhibition of ERK and NFATc1 suppressed the expression of DC-STAMP and led to the fusion inhibition of pOC. However, retrovirus-mediated expression of NFATc1 in pOCs rescued the defect in pOC fusion, despite the presence of U0126 and cyclosporin A. GM-CSF-stimulated pOCs had an intact actin ring and could resorb bone. Importantly, pOCs infected with constitutively active MEK adenovirus expressed c-Fos and NFATc1, followed by the binding of NFATc1 to the DC-STAMP promoter, which enables its transcription and expression. Constitutively active MEK-infected pOCs are able to resorb bone by undergoing cell-cell fusion. Taken together, our results demonstrated that GM-CSF induced fusion of pOCs to form multinucleated osteoclasts, making the osteoclast capable of bone resorption.

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The prevention or therapeutic treatment of loss of bone mass is an important means of improving the quality of life for patients with disorders related to osteoclast-mediated bone loss. Fisetin, a flavonoid dietary ingredient found in the smoke tree (Continus coggygria), exhibits various biological activities, but its effect on osteoclast differentiation is unknown. In this study, fisetin dose-dependently inhibited the RANKL-induced osteoclast differentiation with downregulation of the activity or expression of p38, c-Fos, and NFATc1 signaling molecules. The p38/c-Fos/NFATc1-regulated expression of genes required for cell fusion and bone resorption, such as DC-STAMP and cathepsin K, was also inhibited by fisetin. Considering the rescue of fisetin's inhibitory action by NFATc1 over-expression, the cascade of p38-c-Fos-NFATc1 could be strongly involved in the inhibitory effect of fisetin on osteoclast differentiation. Furthermore, fisetin inhibited the bone-resorbing activity of mature osteoclasts. In conclusion, fisetin may be of use in the treatment of osteoclast-related disorders, including osteoporosis.

Migration and invasion comprise key steps in cancer metastasis. Through the migration and invasion into and out of lymphatic and/or blood vessels, cancer cells can be spread out into the tissues in remote site from the origin. Degradation of extracellular matrix (ECM) must be preceded prior to the metastasis of cancer cells. Matrix metalloproteinases (MMP) can degrade ECM, thus allow cells to migrate from the original site. Among MMPs, two gelatinase MMP-2 and MMP-9 play particularly important roles in ECM degradation. Here, we report that recently developed p21-activated kinase 4 inhibitor PF-3758309 shows anti-metastatic effect in A549 human lung cancer cell. PF-3758309 suppresses CREB, NF-κB, and β-catenin pathways, which are well known to be closely related with cell migration. This leads to the downregulation of MMP-2/MMP-9 expressions and the inhibition of A549 lung cancer metastasis.