Antiapoptotic role of Spi-1 in preleukemic proerythroblasts. (A) Correlation between cytotoxic effect of Epo withdrawal and Spi-1 reduction was measured by the trypan blue exclusion test. Cells were grown for 18 hours with dox or without and then seeded for 12 hours (30-hour dox treatment) with or without Epo before being counted. (B) Cytotoxic effects of Spi-1 reduction on cells cultured in the presence of SCF. Cells were incubated with dox (100 ng/mL) for 3 or 4 days in the presence of SCF (100 ng/mL), and the percentage of dead cells was measured by trypan blue exclusion test. Data are the mean ± SD of 3 experiments. (C) Fas/FasL signaling does not participate to the cellular death stimulated by Spi-1 depletion. Cells were grown for 3 days with dox (100 ng/mL) or without. They were labeled with PE-conjugated Fas antibody and examined by flow cytometry. Control cells were labeled with a PE-conjugated isotype control. A representative experiment of 3 performed with either A2B or A2C cells is shown. (Left) Thymocytes stimulated with PMA/ionomycine for 12 hours and labeled with control or PE–anti-Fas; middle panel (dox−) A2B cells without dox; right panel (dox+) A2B cells with dox for 3 days.