PED/PEA-15 Inhibits Hydrogen Peroxide-Induced Apoptosis in Ins-1E Pancreatic Beta-Cells via PLD-1
Figure 5
PLD-1 role in PED/PEA-15 anti-apoptotic action.
A) PKC alpha phosphorylation. Cells were treated or not for 48 hours with 150 µM propranolol, as indicated. Whole-cell extracts (30 µg) were analyzed by Western blot with anti-phospho PKC alpha antibody. Tubulin was used as loading control (n = 3). A representative image is shown. B) Cell viability. Cells were cultured in 96-well cell culture plates, pretreated with 150 µM propranolol and then incubated with 70 µM hydrogen peroxide as indicated. After 48 hours, cell viability was estimated by use of the sulforhodamine B assay. Cell survival is expressed as the percent of control (untreated Ins-1ECTRL cells). Each point is the mean value from eight identical wells. Values represent the mean ±SD of three independent experiments. *p<0.05, **p<0.01 and ***p<0.001. C) Apoptosis. Cells were pretreated with 150 µM propranolol and then incubated with 70 µM hydrogen peroxide as indicated. After 48 hours, cells were harvested and apoptosis was quantitated by evaluating the level of DNA fragmentation using the Roche Cell Death Detection ELISAPLUS. Data are the mean value of four identical wells. Values represent the mean ±SD of three independent experiments. *p<0.05, ***p<0.001.