RESUMO
Previous researches have proved that Pueraria lobata up-regulates bone mineral contents and bone mineral density in bone-loss model, ovariectomized mice and orchidectomized rats. However, the precise effects and mechanisms of Pueraria lobata on osteoclast differentiation and bone resorbing activity of mature osteoclasts still remains unknown. Therefore, we investigated the effect and mechanism of Pueraria lobata on receptor activator of nuclear factor-kappaB ligand (RANKL) and macrophage colony stimulation factor (M-CSF)-induced osteoclast differentiation in bone marrow macro-phages (BMMs). First of all, we treated BMMs derived from mice with various concentrations of Pueraria lobata in order to perform screening by tartrate-resistant acid phosphatase (TRAP) staining. Also, we conducted western blotting and RT-PCR for the purpose of verifying the treatment mechanism of Pueraria lobata and lastly, we used hydroxyapatite-coated plate to evaluate the effects of Pueraria lobata on bone resorbing activity of mature osteoclasts. As a result, Pueraria lobata has inhibitory effect on phosphorylation of p38, Akt, c-Jun N-terminal kinase (JNK), and IkappaB which are essential early signaling pathway of osteoclastogenesis. Also, the inactivation of nuclear factor of activated T cells (NFAT)c1, and c-Fos which is caused by Pueraria lobata is followed by the suppression effects of Pueraria lobata on osteoclast-related various genes, osteoclast-associated receptor (OSCAR), TRAP, Integrin beta3, osteoclast stimulatory transmembrane protein (OC-STAMP), and dendritic cell-specific transmembrane protein (DC-STAMP). Particularly, Pueraria lobata blocks the formation of pit area on hydroxyapatite-coated plate in a dose-dependent manner as well as the mRNA expression of Cathepsin K, which is associated with bone resorbing activity. These results demonstrate the molecular mechanism relating to anti-osteoclastogenesis effect of Pueraria lobata as well as the inhibitory effect of Pueraria lobata on mature osteoclast formation and bone resorbing activity.
