Overexpression of Akt1 results in increased intratibial tumor growth. A) Cells were cultured in presence or absence of bone conditioned medium (upper panel) and co-cultured with human bone stromal cells (lower panel), then analyzed by Western blot for pAkt(S473). B) Cells were injected into the tibiae of mice intratibially. After harvesting at 8 weeks, bones were analyzed by x-ray. C) H&E was performed on tissue sections from DU145‒Neo and DU145‒HA‒Akt1 intratibial tumors. Images were taken at 5X and digitally merged. D) Trabecular bone and tumor growth was analyzed at 8 weeks. E) Tissue sections from DU145-Neo and DU145-HA-Akt1 intratibial tumors were immunostained with antibodies directed against pAkt(S473) or CXCR4. Images were taken at 20X and 40X (insert). Bar represents 50 μM.