Fig. 5

MDM2 knockdown in ERα⁺ T47D orthotopic transplant reduces tumor volume. a T47D cells with inducible shmdm2 or mir30 shRNA-expressing control vector were treated with 4 μg/ml doxycycline (Dox) for 10 days to induce and maintain shRNA expression. Western blot shows the levels of MDM2, MDMX, E-cadherin, and mtp53 with Dox treatment (lanes 1 and 2) prior to mammary fat pat implantation. Actin was used as a loading control. b Animals were provided with 2 mg/ml Dox and 8 μg/ml E₂ in their drinking water during the entire experiment. Primary tumor growth was measured over a period of 60 days using calipers *** p < 0.001 calculated by two-tailed unpaired t test. c The experimental endpoint tumor volume was determined at the time of necropsy. d mdm2 mRNA expression in primary tumors was determined by real-time qRT-PCR. The p value was determined by two-tailed unpaired t test. e E-cadherin, MDM2, MDMX, and mtp53 protein levels from primary tumors were determined by Western blot analysis. Actin was used as the loading control. f Representative H&E staining images of T47D.vector and T47D.shmdm2 under 200× and 1000× magnification. T represents Tumor; nm represents normal mammary fat pad; M represents muscle; arrowhead depicts tumor cells infiltrating into muscle layer