Resistance to nitric oxide-mediated apoptosis in HL-60 variant cells is associated with increased activities of Cu,Zn-superoxide dismutase and catalase

Nitric oxide (NO) released from (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl)a mino]diazen-1-ium-1,2-diolate (DETA/NO or NOC-18) induces apoptosis in huma n leukemia HL-60 cells. In this study, we isolated a HL-60 variant cell lin e, HL-NR6, that is resistant to DETA/NO toxicity as assessed by DNA fragmen tation, morphology, and colony forming ability. The variant cells also show ed resistance to reactive oxygen species (ROS) such as superoxide and hydro gen peroxide as well as NO donors, but not to anti-tumor drugs. We found th at HL-NR6 cells when compared with HL-60 cells possessed twice the activiti es of Cu,Zn-superoxide dismutase (Cu,Zn-SOD) and catalase, but no change in Mn-SOD nor in glutathione peroxidase. Immunoblotting confirmed the high le vels of both enzymes in the variant cell. We also observed that ROS generat ion following DETA/NO exposure was substantially higher in HL-60 cells than in HL-NR6 cells, using the 2',7'-dichlorofluorescein fluorometric method. Moreover, the SOD mimetic Mn(III) tetrakis(1-methyl-4-pyridyl) porphyrin an d exogenous catalase effectively attenuated DETA/NO-elicited DNA fragmentat ion in HL-60 cells. Taken together, these data suggested that the NO resist ance in HL-NR6 cells is associated with the increased Cu,Zn-SOD/catalase an d that NO-mediated apoptosis in HL-60 cells is correlated with the generati on of ROS and derived molecules like peroxynitrite. (C) 1998 Elsevier Scien ce Inc.