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M. Ladli et al.
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Figure 6. AMPK activation by GSK621 provokes the death of mature GPAhigh 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 mM GSK621 and 2x10-7 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 ery- throblasts 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 mM GSK621 for 24 or 48 h. Cells were incu- bated 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.
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haematologica | 2019; 104(5)