Fig. 4.
Down-regulation of LRP6 significantly inhibits breast tumor growth in vivo. MDA-MB-231 cells (pooled clones) stably expressing control or LRP6 shRNA were injected s.c. into female BNX immunocompromised mice. Tumors initiated from 5 × 105 or 2 × 106 cancer cells were indicated as Low # or High #, respectively. Low #, thoracic pair; High #, caudal pair; control tumors, left; LRP6-KD tumors, right. Tumor growth was monitored over time for 3 weeks by in vivo bioluminescent imaging and caliper measurements. (A) Representative bioluminescence images over time from the same mouse bearing MDA-MB-231 xenografts. (B and D) Growth of tumors over time for control and LRP6-KD xenografts is shown as fold changes of bioluminescence photon flux values over initial value (1 day postinjection). Data are mean ± SEM (six animals, four tumors each) from two independent experiments. (C and E) Tumor volume was monitored for 3 weeks by caliper measurement. (LxWxD). *P < 0.05; **P < 0.01. (F) Gross examination of xenograft tumors. (G) Levels of LRP6 and Wnt target gene expressions (Cyclin D1, c-Myc, and Axin2) in control and LRP6-KD xenograft tumors detected by Western blot analysis. (H) Immunohistochemical analysis of LRP6 level in control and LRP6-KD xenograft tumors with anti-LRP6 antibody (Abgent). (Scale bars, 50 μm.) (I) Western blot analysis showing total and free β-catenin in LRP6-KD tumors were decreased.