The pA20-36 peptide induces apoptosis of A20 cells in a caspase-dependent manner. A20 cells (2 × 107/mL) were incubated with monomeric pA20-36 or pCNT peptide (20 μg/mL), or with the streptavidin-coated pA20-36 or pCNT peptide, and analyzed for viability, cell-cycle profile, and apoptosis. As positive control of apoptosis, cells were cultured in the presence of goat anti–mouse IgG F(ab)′2 (20 μg/mL). (A) Cell viability is strongly affected by streptavidin-coated pA20-36. Number of viable A20 cells was measured by trypan blue. Values are the mean ± SEM of 3 independent experiments; statistical analysis was performed by Student t test. (B) The pA20-36 treatment increases sub-G1 (apoptotic) cell population. Histograms for cell-cycle profiles of 3 independent experiments with similar results, as measured by propidium iodide staining and flow cytometry. (C) The pA20-36 treatment induces apoptosis of A20 target cells. Dot plots for annexin V binding of 3 independent experiments with similar results. Numbers in the quadrants indicate the percentage of cells. (D) The pA20-36 peptide induces activation of caspases. The activity of caspase-8, caspase-9, and effector caspase 3/7 was analyzed by luminometric assay of A20 cell extracts. Values are the mean ± SEM of 3 independent experiments.
