Figure 5.

Effects of AIRE expression on cell cycle and apoptosis of luminal breast cancer cell lines. (A) AIRE protein expression was evaluated in MCF-7 and MDA-MB-361 luminal breast cancer cells transiently transfected with AIRE-expressing or empty vector after 72 h by Western blot with anti-AIRE mouse monoclonal antibody. Vinculin was used to normalize total protein loading using anti-vinculin. The distribution of AIRE-transfected cells in phases of the cell cycle was evaluated by staining with anti-AIRE antibody and propidium iodide. In MCF-7 and MDA-MB-361 cells 72 h after transfection with AIRE, the percentage of AIRE-positive cells in G1 (B) and S phases (C), as assessed by propidium iodide assay, was determined as compared to AIRE-negative cells. Data represent the mean ± s.e.m. of at least 3 independent experiments. *p < 0.05, **p < 0.01. D) Adherent cells were trypsinized and the catalytic activity of caspase-9, −3, and −8 was measured as described in Methods. Data are expressed as fold-increase with respect to cells transfected with empty vector. E) MDA-MB-361 cells transiently transfected with AIRE-expressing or empty vector were analyzed for cleaved caspase-7 and p21 protein expression after 72 h by Western blot by using anti-cleaved caspase-7 rabbit polyclonal antibody and anti-p21 polyclonal antibody. Vinculin was used to normalize total protein loading.