Cyclic AMP-elevating agents prolong or inhibit eosinophil survival depending on prior exposure to GM-CSF

1 Purified human eosinophils survived for up to 7 days when cultured in vitro in the presence of 1 ng ml(-1) granulocyte-macrophage colony stimulating factor (GM-CSF) with a viability of 73%. In the absence of GM-CSF, eosinophil viability decreased after one day in culture, and only 4% of cells were viable by day 4. 2 Culture of eosinophils with cholera toxin produced a concentration-dependent decrease in GM-CSF-induced survival at 7 days (IC50=7 ng ml(-1)) which was associated with a 6 fold increase in the intracellular cyclic AMP concentration. This inhibition of cell survival could be prevented by the addition of the protein kinase A inhibitor, H89 (10(-6)M). 3 When eosinophils were cultured with dibutyryl cyclic AMP, there was a concentration-dependent inhibition of GM-CSF-induced survival at 7 days with an IC50 of 200 mu M. The related cyclic nucleotide analogue, dibutyryl cyclic GMP did not inhibit GM-CSF-induced eosinophil survival over the same concentration range. 4 Culture of eosinophils with forskolin, or with the phosphodiesterase inhibitors, rolipram and SK&F94120, had no effect on GM-CSF-induced eosinophil survival at any concentration examined. 5 After 7 days' culture in the absence of GM-CSF, fractionation of eosinophil DNA on agarose gels demonstrated a 'ladder' pattern characteristic of apoptosis. GM-CSF prevented DNA fragmentation and this protection could be overcome by both cholera toxin and dibutyryl cyclic AMP. 6 GM-CSF did not affect intracellular cyclic AMP concentrations in unstimulated eosinophils or in cells stimulated by cholera toxin. Thus, GM-CSF does not apparently increase eosinophil survival by affecting cyclic AMP levels. 7 In the absence of GM-CSF both cholera toxin and dibutyryl cyclic AMP decreased the rate of eosinophil death, when compared to cells cutured with medium alone. The t(1/2) values for cell death were 1.63+/-0.3, 2.46+/-0.3 and 4.62+/-1.0 days for cells cultured in the presence of medium, cholera toxin and dibutyryl cyclic AMP respectively. 8 In conclusion, cyclic AMP exerts opposing effects on eosinophil survival depending on prior exposure of the cells to GM-CSF.