American Society of Hematology

Figure 11.

$Figure 11. Effect of dyhydropyridines, nicardipine, and nitrendipine on ST1926- and CD437-dependent apoptosis and cytochrome c release from mitochondria in NB4 cells. NB4 cells (150 000/well) were treated with vehicle, the indicated dihydropyridines, ST1926 or CD437, or the indicated mixtures of calcium blockers and RRMs for 6 hours. (A) A typical cytometric profile of cells treated with vehicle (control), ST1926 (1 μM), nicardipine (100 μM), or the combination of the 2 compounds. The profile shows the level of propidium iodide cell positivity (PI+, ordinate axis) and annexin-V positivity (Annexin+, abscissa). The lower left, lower right, upper left, and upper right quadrants indicate the percentage of viable, apoptotic, necrotic, and necrotic+apoptotic cells, respectively. (B) A quantitative summary of an experiment performed as in panel A. The results are expressed as the mean ± SD of 3 replicate cultures. The data presented are representative of 2 independent experiments. *Significantly lower than the corresponding vehicle-treated group (P < .01 according to the Student t test). °Significantly higher than the corresponding vehicle-treated group (P < .01 according to the Student t test). §Significantly lower than the corresponding ST1926-treated group (P < .01 according to the Student t test). (C) Following treatment with ST1926 and CD437 in the presence and absence of the indicated concentrations of dihydropyridines, aliquots of the cultures were subjected to the determination of the apoptotic index by scoring the percentage of cells showing morphologic signs of nuclear fragmentation upon DAPI staining. The results are expressed as the mean ± SD of 3 replicate cultures. The data presented are representative of 2 independent experiments. *Significantly lower than the corresponding ST1926- or CD437-treated group (P < .01 according to the Student t test). °Significantly higher than the corresponding vehicle-treated group (P < .01 according to the Student t test). (D) Cells were treated for 6 hours with 10 μMor100 μM nicardipine in the presence of medium or medium containing 1 μM ST1926. The level of caspase-3 was measured following incubation of cell extracts with the fluorogenic peptide substrate DEVD-amc. The results are expressed as the mean ± SD of 3 replicate cultures. °Significantly lower than the corresponding ST1926- or CD437-treated group (P < .01 according to the Student t test). In panel E, cells were treated with vehicle (control), ST1926 (1 μM), nitrendipine (100 μM), or the combination of the 2 compounds for 4 hours. Cells were harvested, homogenized, and fractionated in a mitochondrial and a cytosolic fraction or left unfractionated (total). Equivalent aliquots of the various fractions were subjected to Western blot analysis with antibodies directed against the indicated proteins.$

Effect of dyhydropyridines, nicardipine, and nitrendipine on ST1926- and CD437-dependent apoptosis and cytochromec release from mitochondria in NB4 cells. NB4 cells (150 000/well) were treated with vehicle, the indicated dihydropyridines, ST1926 or CD437, or the indicated mixtures of calcium blockers and RRMs for 6 hours. (A) A typical cytometric profile of cells treated with vehicle (control), ST1926 (1 μM), nicardipine (100 μM), or the combination of the 2 compounds. The profile shows the level of propidium iodide cell positivity (PI⁺, ordinate axis) and annexin-V positivity (Annexin⁺, abscissa). The lower left, lower right, upper left, and upper right quadrants indicate the percentage of viable, apoptotic, necrotic, and necrotic+apoptotic cells, respectively. (B) A quantitative summary of an experiment performed as in panel A. The results are expressed as the mean ± SD of 3 replicate cultures. The data presented are representative of 2 independent experiments. *Significantly lower than the corresponding vehicle-treated group (P < .01 according to the Student t test). °Significantly higher than the corresponding vehicle-treated group (P < .01 according to the Student t test). §Significantly lower than the corresponding ST1926-treated group (P < .01 according to the Student t test). (C) Following treatment with ST1926 and CD437 in the presence and absence of the indicated concentrations of dihydropyridines, aliquots of the cultures were subjected to the determination of the apoptotic index by scoring the percentage of cells showing morphologic signs of nuclear fragmentation upon DAPI staining. The results are expressed as the mean ± SD of 3 replicate cultures. The data presented are representative of 2 independent experiments. *Significantly lower than the corresponding ST1926- or CD437-treated group (P < .01 according to the Student t test). °Significantly higher than the corresponding vehicle-treated group (P < .01 according to the Student t test). (D) Cells were treated for 6 hours with 10 μMor100 μM nicardipine in the presence of medium or medium containing 1 μM ST1926. The level of caspase-3 was measured following incubation of cell extracts with the fluorogenic peptide substrate DEVD-amc. The results are expressed as the mean ± SD of 3 replicate cultures. °Significantly lower than the corresponding ST1926- or CD437-treated group (P < .01 according to the Student t test). In panel E, cells were treated with vehicle (control), ST1926 (1 μM), nitrendipine (100 μM), or the combination of the 2 compounds for 4 hours. Cells were harvested, homogenized, and fractionated in a mitochondrial and a cytosolic fraction or left unfractionated (total). Equivalent aliquots of the various fractions were subjected to Western blot analysis with antibodies directed against the indicated proteins.

This Feature Is Available To Subscribers Only