Treatment of circulating human neutrophils with recombinant human granulocyte colony-stimulating factor (rhG-CSF) for 30 min augmented superoxide generation and chemotaxis induced by N-formylmethionyl-leucyl-phenylalanine (fMLP) in a dose dependent manner. When neutrophils were treated with 1 pM of methotrexate (MTX) for 60 rnin after incubation with rhG-CSF (10 ngrnl), the effects of rhG-CSF on superoxide generation and chemotaxis were inhibited by approximately 49 and 29%, respectively. Although inhibitory effects of MTX were also seen in neutrophils not pretreated with rhG-CSF, the degree of inhibition was much less. The addition of either hypoxanthine or guanosine at a concentration of 100 pM to the culture medium significantly attenuated the effects of MTX. However, in neutrophils obtained from a patient with Lesch-Nyhan syndrome, which lacked hypoxanthine-guanine phosphoribosyl transferase activity, neither hypoxanthine nor guanosine had any rescue effect. These results suggest that MTX inhibits superoxide generation and chemotaxis in rhG-CSF-activated neutrophils, at least in part, by disturbing purine nucleotide biosynthesis. 8 1996 WiIey-Liss, Inc.
I. iutrophils play a central role in the defense against bacterial infections. Although circulating neutrophils are usually in a resting state, they are known to be activated following an encounter with invading microorganisms. Several lymphokines including granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF) (Lopez et