Figure 1

HuR knockdown prevents bone metastasis of breast cancer cells in mice. (a) Bioluminescence images taken 6 weeks after luciferase-transfected HuR-knockdown (shHuR) and control (shNC) breast cancer cells were inoculated into the left cardiac ventricles of nude mice (n = 10). (b,c) X-ray and 3D images of mandibles (b), femora, and tibiae (c) derived from μCT scans performed on week 6. (c) Goldner’s trichrome and TRAP staining of femoral tissues. Arrowheads: TRAP-positive osteoclasts; T: tumor; B: bone; BM: bone marrow. Scale bar: 0.5 mm for Goldner’s trichrome staining and 10 μm for TRAP staining. (d,e) Tumor areas (d) and osteoclast surface per bone surface (Oc.S/BS) (e) values for stained femoral sections. (f) Bone morphometric parameters BV/TV (%), Tb.Th (mm), Tb.N (1/mm), Tb.Sp (mm), and SMI of the mouse femora analyzed by μCT. (g) Serum levels of the bone resorption markers TRAP 5b and CTX quantified using commercial kits. The data are expressed as the mean ± standard error (s.e.m.). ^# P < 0.05 versus control mice without cancer cells; *P < 0.05, **P < 0.01 versus mice injected with shNC MDA-MB-231 cells. (h) Viabilities of shNC and shHuR MDA-MB-231 cells. The cells were cultured for 24, 48, and 72 h, and cell viability was assessed by MTT assay. (i) Cell migration. Movement of cells in scratched areas was observed under an inverted optical microscope (original magnification, × 40) and measured using ImageJ software at 0 h and 24 h. (j) Cell invasion. Cell invasion was measured by transwell invasion assay. Representative images of cells that invaded the lower surfaces of the filters were captured using an optical microscope (original magnification, × 200). The data are expressed as the mean ± s.e.m. *P < 0.01 versus shNC.