Figure 2.

HRT-BU and fHRT-BU inhibit RANKL-induced osteoclast formation in BMMs. BMMs (1 × 10⁴ cells/well in a 96-well plate) were incubated with M-CSF (60 ng/mL), RANKL (150 ng/mL), and the indicated concentrations of HRT-BU or fHRT-BU for 4 days. Cell viability was measured by CCK-8 assay (a). The cells were fixed and stained for TRAP activity (b) and then the number of TRAP-positive multinuclear osteoclast cells (TRAP(+)MNCs) (c) with more than 3 nuclei and larger than 100 μm in diameter were counted. Representative microscopic pictures of multinucleated osteoclasts (d) were shown at a magnification of 100x. Data represents mean ± SD of three independent experiments. ^a P < 0.05, versus vehicle treated control. ^b P < 0.05, versus HRT.