Fig. 4. PA induces apoptosis in MCF-7 cells. (A) MCF-7 cells were grown in coverslips and were treated with PA with three different concentrations (10, 15 and 20 μM) for 24 h. Next, the medium was removed and washed with PBS. Additionally, the cells were fixed with ice-cold acetone:methanol for 20 min. After air drying, the cells were incubated with Annexin V (FITC conjugated) for 30 min in the dark. After counter staining, the coverslips were placed in a slide with anti-fading agent. PA dose-dependently increased the expression of Annexin V-FITC at 24 h. (B) Flow cytometric analysis of Annexin V-FITC/PI was represented. PA administration, dose-dependently increased the apoptosis (both early and late). (C) Western blot analysis of mitochondrial cytochrome C, Bax and Bcl-2. Treatment with PA for 24 h increased the expression of mitochondrial cytochrome C, Bax and decreased the expression of Bcl-2. (D, E) Quantification of the respective western blots (Cytochrome C, Bax and Bcl-2) was represented in graphs. The results are from 3 independent experiments and are presented as the means ± SD. *Indicates a significant difference by Tukey’s test (p<0.05) relative to their respective control.
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