Fig. 4

Nitazoxanide attenuates Ac-KLF5-induced osteoclast differentiation. a Effects of NTZ at different concentrations on the viability of RAW264.7 cells after 48 h treatment, as determined by the CCK8 assay. b, c NTZ inhibits osteoclast differentiation in a dose-dependent manner, as determined by the TRAP staining of RAW264.7 cells treated with RANKL (50 ng/mL) and various concentrations of NTZ for 5 days. Shown are representative images of TRAP⁺ and multinucleated cells (nuclei >3) (b) and their numbers per well (c). One-way ANOVA analysis was performed on the data. d NTZ inhibited osteoclast differentiation of RAW264.7 cells caused by conditioned medium (CM) from PC-3-KQ cells, as indicated by representative images of TRAP-stained cells (left) and the numbers of TRAP⁺ cells per well (right) after NTZ treatments. One-way ANOVA analysis was performed on the data. e NTZ inhibited osteoclast differentiation of RAW264.7 cells co-cultured with PC-3-KQ cells, as indicated by representative images of TRAP-stained cells (left) and the numbers of TRAP⁺ cells per well (right) after NTZ treatments. One-way ANOVA analysis was performed on the data. f NTZ decreased the number of Ac-KLF5-induced osteoclasts in mouse bones, as indicated by images of bone sections with TRAP⁺ cells (left) and the number of osteoclasts (N.Oc) per mm of bone surface (BS). Scale bar, 200 μm. Student’s t-test was performed. n = 5 femurs in vehicle group, n = 5 femurs in NTZ group. For all bar graphs, data are shown as mean ± SD. **, p < 0.01; ***, p < 0.001. T, tumor cells; B, bone; BM, bone marrow; NTZ, nitazoxanide; RANKL, receptor activator of the nuclear factor-κB ligand; TRAP, tartrate-resistant acid phosphatase