Ikarisoside A inhibits osteoclastogenesis by reducing iNOS and NF-kB

Objectives:The present study was undertaken to elucidate the effect of Ikarisoside A on osteoclastogenesis in a murine RAW 264.7 cell line, for the first time.

Methods:Cell transfection and luciferase reporter assay-

RAW 264.7 cells were seeded at 5 x 104/well in a 24-well dish and grown to 90–95 % confluence in complete growth media. For each well, 1.6 ìg of luciferase reporter plasmid construct harboring the NF-êB binding site (pGL2-NF-êB) and 0.5 ìg of pCMV-â-galactosidase control vector were co-transfected into cells with Lipofectamine 2000 (Invitrogen) according to the manufacturer's instructions. The activities of firefly luciferase in the cellular extracts were measured using the luciferase reporter assay system.

Results:The results indicated that Ikarisoside A inhibited the enzyme activaties of cathepsin K, JNK and MMP9, which were known to be involved in osteoclastogenesis. Ikarisoside A also reduced transcriptional activity of NF-kB and NFATc as demonstrated by luciferase assay in a concentration-dependent manner. Furthermore, Ikarisoside A inhibited the differentiation of osteoclasts as shown by TRAP staining and pit formation assay.

Conclusion:The present results suggest that Ikarisoside A has an inhibitory effect on RANKL-mediated osteoclastogenesis in RAW cells through the inhibition of JNK/NF-kB activity, and thus could be used as a therapeutic agent for bone-lytic diseases.