In order to better understand the enhancing effects of lowered oxygen (O2) tension on the growth in vitro of granulocyte-macrophage progenitor cells (CFU-GM), the effects of oxidizing species derived from molecular O2 were assessed on CFU-GM. Low density or nonadherent low density normal human bone marrow cells were plated at ambient (20%) or lowered (5%) O2 tension in the presence of a source of colony stimulating factors, and in the absence or presence of superoxide dismutase, catalase, glucose oxidase or horseradish peroxidase, alone or in various combinations. Enhanced colony and cluster formation of CFU-GM was noted when low density cells were grown at 5% O2, or when cells were grown at 20% O2 in the presence of superoxide dismutase or glucose oxidase. Both of these enzymes are capable of generating hydrogen peroxide (H2O2), although by different mechanisms. Low concentrations of glucose oxidase resulted in increased formation of colonies and clusters, but higher concentrations of glucose oxidase were inhibitory. Catalase, which converts H2O2 to H2O, had no effect by itself on cells growing at 20% O2, but it eliminated the superoxide dismutase and glucose oxidase enhancing effects. Catalase decreased colony formation of cells grown at 5% O2. Removal of adherent cells ablated the growth-enhancing effects noted at lowered (5%) O2 tension and also the superoxide dismutase and catalase effects at 20% or 5% O2. Horseradish peroxidase, which converts H2O2 to a more toxic oxidant, hypochlorite, had a suppressive effect on colony and cluster numbers and at 20% O2 converted the glucose oxidase effects from stimulatory to inhibitory. The results suggest that adherent cells and low concentrations of H2O2 may mediate growth-enhancing effects of CFU-GM seen at lowered (5%) O2 tension.