Figure 12.

X-linked inhibitor of apoptosis protein (XIAP) as a molecular target of Piper genus-derived compounds. (A) Western blot analysis of XIAP in MCF7 cells both untransfected (CTRL, control) or transfected for 24 h with a XIAP-specific and scrambled targeting (scr) siRNA (50 nM). Lactate dehydrogenase (LDH) was used as internal standard. Low panel: densitometric analysis expressed as fold change of scr siRNA. Images and data are representative of three independent experiments. **P < 0.001 relative to scr siRNA. (B) MCF7 cells were transfected for 24 h with a XIAP-specific or scr siRNA (50 nM) and then cultured in the absence (CTRL) and in the presence of 30 μg/mL gibbilimbol B/eriopodol A or 10 µg/mL erioquinol for 6 h, before 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Results are expressed by setting the absorbance of the reduced MTT in the scr siRNA CTRL, as 100%. Data are representative of three independent experiments. * p < 0.0001 relative to the respective scr siRNA. (C) Western blot analysis of cleaved-caspase 7 in MCF7 cells transfected for 24 h with a XIAP-specific or scr siRNA (50 nM) and then cultured in the presence of 30 μg/mL gibbilimbol B and eriopodol A for 6 h. LDH was used as internal standard. Images are representative of three independent experiments. (D) Real-time PCR and (E) Western blot analysis of XIAP mRNA and protein expression, respectively, in MCF7 cells treated for increasing times in the absence (CTRL) and in the presence of 30 μg/mL gibbilimbol B/eriopodol A or 10 µg/mL erioquinol. β-actin (PCR) and LDH (Western blot) were used as internal standards. PCR results are expressed as fold change of respective CTRL, set as 1. Images and data are representative of three independent experiments.