Figure 5. A role for Ac45 in osteoclast formation and fusion.

(A) RANKL-stimulated pre-osteoclasts transfected with control or Ac45 siRNA for 48hrs were stained for TRAP activity to demonstrate multinucleated osteoclast formation. The number (B) and average size (µm²) (C) of TRAP+ve multinucleated osteoclasts (???3 nuclei) were quantified. (D) The number of nuclei per osteoclast from control and Ac45 siRNA treated groups. (E–G) BMM precursors were silenced for Ac45 expression prior to commission to the osteoclast lineage and sustained following RANKL stimulation for osteoclast formation. The number (F) and size (G) of resulting TRAP+ve osteoclasts were quantified. (H) Western blot analysis of protein expression of V-ATPase V0 domain subunits a1, a3, d1, and d2 in osteoclasts following Ac45 gene silencing. Protein expression of α-tubulin was used as loading control. Data representative of three independent experiments (mean ± SEM). *P-values<0.05.