Analysis of differentiation associated apoptosis of M1, M1myc, M1Egr-1, and M1Egr-1/myc cells following treatment with IL-6. Cells were seeded at 0.05 × 106 cells/mL with or without IL-6 (50 ng/mL). At indicated time points cells were analyzed as indicated. Because M1myc cells treated with IL-6 continue to proliferate, it was necessary to split the cell culture on days 3, 5, and 7. (A) Cell viability was assessed by trypan blue dye exclusion using a hemocytometer, as described in “Materials and methods.” Results are the average of 3 independent experiments yielding similar results, with an SD up to 10%. (B) DNA fragmentation was determined as indicated in “Materials and methods.” DNA samples were resolved on a 2% agarose gel and assessed for fragmentation. Results shown are representative of 3 independent experiments. (C) Analysis of PARP cleavage was determined as indicated in “Materials and methods.” This is a representative experiment that was carried out 3 times. (D) Caspase 8 activity was determined using the ApoAlert FLICE assay (Clontech), as described in “Materials and methods.” Results are the average of 3 independent experiments, with SDs less than 10% for each sample.
