Figure 4. Clodronate liposome treated mice had increased bone volume in an intratibial tumor model in vivo.

Four-six week old C57Bl6 male mice were treated with 3 consecutive injections of clodronate loaded liposomes (CLOD) (10μl/g) or vehicle (VEH). RM-1 murine prostate cancer cells were injected intratibially on the fourth day and booster injections were given every third day for depletion maintenance. Fourteen days later mice were euthanized and evaluated. A. Representative image (4X) of intratibial RM-1 tumor of C57Bl6 mice treated with vehicle (VEH) or clodronate (CLOD) and quantification of total bone volume (BV/TV) by histomorphometric analyses at day 14. B. Total tumor volume (Tm V/TV) and tumor volume per bone surface (Tm V/BS) were quantified by histomorphometric analyses in intratibial tumors of C57Bl6 mice treated with vehicle (VEH) or clodronate (CLOD) at day 14. C. Serum analyses of TRAP5b (units per liter) in C57Bl6 mice with intratibial RM-1 tumors treated with vehicle (VEH) or clodronate (CLOD) after 14 days. D. Representative image (20X) of TRAP positive osteoclast surface per bone surface (Oc.S/BS), osteoclast numbers per total tissue area (N.Oc/T.Ar) (P = 0.052), and osteoclast numbers per bone perimeter (N.Oc/B. Pm) were quantified by histomorphometric analyses in intratibial tumors of C57Bl6 mice treated with vehicle (VEH) or clodronate (CLOD) at day 14. E. Representative 2D and 3D μCT images of femurs without tumors from mice with intratibial tumors show increased trabecular bone volume in clodronate-treated mice. Trabecular total bone volume (BV/TV), trabecular thickness (Tb.Th), number (Tb.N) and spacing (Tb.Sp) were quantified. Data for figures A, B and D are mean ± SEM; n = 10-13 per group. Data for figures C is mean ± SEM; n = 18 per group. Data for figure E is mean ± SEM; n = 7-10 per group. Statistically significant differences were calculated using unpaired t tests between treatments. *P < 0.05, ***P < 0.001 vs. VEH.