An in vitro study was performed on the effect of recombinant human granulocyte colony-stimulating factor (rhG-CSF) and adenine on the survival of purified human neutrophils. The addition of rhG-CSF (1 to 100 ng/mL) or adenine (100 microM) enhanced the survival of neutrophils. The maintenance of O2- production in response to N-formylmethionyl-leucyl-phenyl-alanine (FMLP) suggested that these neutrophils were functionally alive. Neutrophils in cultures had shown two distinct biochemical changes during cell death: DNA fragmentation and depletion of cellular adenosine triphosphate (ATP) pools. Treatment with rhG-CSF (10 ng/mL) significantly delayed the appearance of DNA fragmentation as measured quantitatively by diphenylamine or by agarose gel electrophoresis. On the other hand, adenine had no effect on the generation of DNA fragmentation. The decrease of ATP during incubation for 12 hours was similar in control and rhG-CSF-treated neutrophils, while rhG-CSF prevented the further decline of ATP seen in control cultures. In contrast, adenine (100 microM) preserved ATP at levels significantly higher than in controls at both 12 hours and 24 hours of incubation. Our results suggest that rhG-CSF and adenine promote the survival of neutrophils in vitro by different mechanisms.