Figure 3.
5-HT6R increases RANKL-induced osteoclastogenesis via the small RhoA GTPase pathway. (A-B) BMMs were stimulated with RANKL plus 1 μM ST1936 in the absence or presence of 1 μM Rhosin (RhoA GTPase inhibitor), 1 μM NSC23766 (Rac1 GTPase inhibitor), or 1 μM ZCL278 (Cdc42 GTPase inhibitor). F-actin-positive cells (red, (A)), and actin ring formation were measured in actin ring-positive MNCs (B) by confocal microscopy following staining with TRITC-phalloidin (red) and DAPI (blue). Scale bar: 200 μm. Rho: Rhosin; NSC: NSC23766; ZCL: ZCL278. *, p < 0.05 compared to RANKL. #, p < 0.05 compared to RANKL + ST1936. (C-E) BMMs (left) or OC (right) were stimulated with 1 μM ST1936 for the indicated time after being serum-starved for 6 h. RhoA (C), Rac1 (D), and Cdc42 (E) activities were assessed by pull-down assay to detect the GTP-bound form (active form). Total-RhoA, -Rac1, and -Cdc42 were detected as loading controls by western blotting. *, p < 0.05 compared to control. Data shown represent mean ± SEM.