Figure 5.

p70S6K1 signaling results in upregulation of Bcl2 in breast cancer cells. A. Western blot analysis of MDA MB 231 cells treated with the indicated inhibitor (PF‐4708671 10 μM or Temsirolimus 10, 20 and 100 nM) or left untreated. Vinculin expression was used as loading control. B. Western blot analysis of primary tumor lysates derived from injection of MDA MB 231 control cells (2 × 10⁶), in nude mouse thoracic mammary fat pads. When tumors reached a volume of 50–100 mm³, mice were intraperitoneally injected with PF‐4708671 (50 mg/kg, i.e. 1200 μg/mouse) or Temsirolimus (12.5 mg/kg, i.e. 300 μg/mouse or 25 mg/kg, i.e. 600 μg/mouse) or vehicle (Untreated) for three consecutive days and then sacrificed. C. Immunohistochemistry analysis of Bcl2 on tumor section from a Tems‐treated mouse of the experiment described in (B). Nor untreated tumors nor those from PF‐treated mice displayed positivity for Bcl2 (magnification 200×). D. Immunofluorescence analysis on tumor sections from experiment described in (B), acquired by confocal microscopy. The panels show the merge of the immunostaining for Bcl2 (AF‐488, pseudocolored in green) with the nuclear staining with propidium iodide (pseudocolored in red). Bars correspond to 35 μm. The panel at the right shows higher magnification of the dashed area.