ABSTRACT
Osteoclast and osteoblast are essential for proper bone development and remodeling as well as recovery of bone fracture. In this study, we seek chemical compounds that enhance turnover of bone metabolism for promoting bone healing. First, we screen a chemical library which includes 378 compounds by using murine pre-osteoclastic RAW264.7 cells to identify compounds that promote osteoclastic differentiation. We find that two ROCK (Rho-associated coiled-coil kinase) inhibitors, HA-1077 (Fasudil) and Y-27632, enhance osteoclastogenesis. Subsequently, we identify that these two compounds also increase osteoblastic differentiation of MC3T3-E1 cells. Finally, our in vivo experiment shows that the local administration of ROCK inhibitors accelerate the bone healing of the rat calvarial defect.
Subject(s)
1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine/analogs & derivatives , Amides/pharmacology , Osteogenesis/drug effects , Protein Kinase Inhibitors/pharmacology , Pyridines/pharmacology , rho-Associated Kinases/antagonists & inhibitors , 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine/chemistry , 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine/pharmacology , 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine/therapeutic use , Amides/chemistry , Amides/therapeutic use , Animals , Cell Differentiation , Cell Line , Fracture Healing/drug effects , Male , Mice , Osteoblasts/cytology , Osteoblasts/drug effects , Osteoblasts/metabolism , Osteoclasts/cytology , Osteoclasts/drug effects , Osteoclasts/metabolism , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/therapeutic use , Pyridines/chemistry , Pyridines/therapeutic use , RAW 264.7 Cells , Rats , Rats, Sprague-Dawley , Skull/drug effects , Skull/injuries , Skull/metabolism , Small Molecule Libraries/chemistry , Small Molecule Libraries/pharmacology , rho-Associated Kinases/metabolismABSTRACT
Osteoclasts are multinucleated cells with bone resorption activity that is crucial for bone remodeling. RANK-RANKL (receptor activator of nuclear factor κB ligand) signaling has been shown as a main signal pathway for osteoclast differentiation. However, the molecular mechanism and the factors regulating osteoclastogenesis remain to be fully understood. In this study, we performed a chemical genetic screen, and identified a Cdks/GSK-3ß (cyclin-dependent kinases/glycogen synthase kinase 3ß) inhibitor, kenpaullone, and two Cdks inhibitors, olomoucine and roscovitine, all of which significantly enhance osteoclastogenesis of RAW264.7 cells by upregulating NFATc1 (nuclear factor of activated T cells, cytoplasmic 1) levels. We also determined that the all three compounds increase the number of osteoclast differentiated from murine bone marrow cells. Furthermore, the three inhibitors, especially kenpaullone, promoted maturation of cathepsin K, suggesting that the resorption activity of the resultant osteoclasts is also activated. Our findings indicate that inhibition of GSK-3ß and/or Cdks enhance osteoclastogenesis by modulating the RANK-RANKL signaling pathway.
