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. 2014 Jun 27;155(9):3251–3261. doi: 10.1210/en.2014-1002

Figure 5.

Figure 5.

Depletion of SMRT from MCF-7 cells increases apoptosis. Cells were transfected with nontargeting control (siCtrl) or SMRT-specific (siSMRT) siRNA, and after 48 hours were treated with vehicle, 1 nM E2, 100 nM 4HT, or 1 μM staurosporine (positive control) for 24 hours, as indicated. Cells were harvested and the extent of apoptosis was assessed by (A) Cell Death ELISA and (B) annexin V staining. C, MCF-7 cells transfected with control (-) or SMRT (+) siRNA were harvested 48, 72, or 96 hours after transfection for analysis of PARP cleavage by Western blot analysis (top). Tubulin (bottom) is shown as a loading control. Shown is a representative blot (n = 4). D, ZR-75–1 cells were transfected with siCtrl (-) or siSMRT (+) siRNA and harvested 96 hours thereafter for analysis of PARP cleavage by Western blot. MCF-7 cells were transfected with siCtrl (-) or siSαβ (+) siRNA and collected 96 hours thereafter for assessment of (E) cleaved PARP and (F) knock-down of SMRT expression by Western blot. Shown are representative blots of at least 3. G, MCF-7 cells were transfected with siCtrl or siSαβ siRNA and 24 hours after treated with vehicle or 1 nM E2. After 5 days cells were harvested for evaluation of cell growth. Numerical data are presented as average ± SEM (n = 3). Statistical analyses were performed by Student's t test, #, P < .05 versus siCtrl vehicle and *, P < .05 versus respective siCtrl treatment.