TNF- induces JNK activation but does not induce apoptosis in HCD57 cells.
(A) HCD57(R) cells were deprived of EPO for 18 hours and then treated with 10 ng TNF-α/mL for the times indicated. Whole cell lysates were immunoblotted with an anti-phospho-JNK antibody (upper panel, arrows). The blot was then stripped and reprobed with anti-JNK1 to ensure equal loading of proteins (lower panel, arrow). (B) HCD57 cells were deprived of EPO for 18 hours and then treated with no additional growth factor (lane 1), 1 U EPO/mL (lane 2), 10 ng SCF/mL (lane 3), or 10 ng TNF-α/mL (lane 4) for 24 hours. Whole cell lysates were immunoblotted with an anti-phospho-JNK antibody (upper panel, arrows). The blot was then stripped and reprobed with anti-JNK1 to ensure equal loading of proteins (lower panel, arrow). (C) HCD57 cells were deprived of EPO for 18 hours and then treated with 1 U EPO/mL (+EPO); no additional growth factor (No EPO); or 10 ng, 100 ng, or 1000 ng TNF-α/mL for 24 hours. Cells were stained with propidium iodide as indicated in “Materials and methods” and analyzed by using flow cytometry. The number of apoptotic cells is indicated as a percentage of the total number of cells containing sub-G0/G1DNA and is indicated as M1 on plots.
