Figure 6.

AMPK activation by GSK621 provokes the death of mature GPA^high erythroblasts. (A) GSK621 does not increase the percentage of annexin V-positive cells in an immature cell population. Erythroid cells were incubated in the absence (vehicle) or presence of 20 μM GSK621 and 2×10⁻⁷ M dexamethasone (Dexa)/vehicle or Dexa/GSK621 from day 0 to the indicated days. Cells were labeled with anti-GPA antibodies and annexin V before analysis by flow cytometry. A representative experiment and the percentage of annexin V-positive cells from three experiments are shown. (B) GSK621 does not induce cell death when the population of erythroblasts is immature. The proportion of dead cells was determined by trypan blue exclusion dye assay. (C) GSK621-mediated AMPK activation specifically induces the death of mature day 9 erythroblasts. Mature erythroblasts were incubated in the absence (vehicle) or presence of 20 μM GSK621 for 24 or 48 h. Cells were incubated with anti-GPA antibodies and annexin V before analysis by flow cytometry. A representative experiment and the percentage of annexin V-positive cells from three experiments are shown (lower panel). (D) The proportion of dead cells was determined by a trypan blue exclusion dye assay. Results are expressed as the mean ± SD. ^*P<0.05, ^**P<0.01, ^***P<0.001. d: day; Dexa: dexamethasone; GPA: glycophorin A; h: hours.