N-acetylcysteine (NAC), often used as an antioxidant-scavenging reactive oxygen species (ROS) in vitro, was recently shown to increase the cytotoxicity of other compounds through ROS-dependent and ROS-independent mechanisms. In this study, NAC itself was found to induce extensive ROS production in human leukemia HL-60 and U937 cells. The cytotoxicity depends on ROS-modulating enzyme expression. In HL-60 cells, NAC activated NOX2 to produce superoxide (O2•-). Its subsequent conversion into H2O2 by superoxide dismutase 1 and 3 (SOD1, SOD3) and production of ClO- from H2O2 by myeloperoxidase (MPO) was necessary for cell death induction. While the addition of extracellular SOD potentiated NAC-induced cell death, extracellular catalase (CAT) prevented cell death in HL-60 cells. The MPO inhibitor partially reduced the number of dying HL-60 cells. In U937 cells, the weak cytotoxicity of NAC is probably caused by lower expression of NOX2, SOD1, SOD3, and by the absence of MOP expression. However, even here, the addition of extracellular SOD induced cell death in U937 cells, and this effect could be reversed by extracellular CAT. NAC-induced cell death exhibited predominantly apoptotic features in both cell lines. Conclusions: NAC itself can induce extensive production of O2•- in HL-60 and U937 cell lines. The fate of the cells then depends on the expression of enzymes that control the formation and conversion of ROS: NOX, SOD, and MPO. The mode of cell death in response to NAC treatment bears apoptotic and apoptotic-like features in both cell lines.

• Keywords
• HL-60 cells, MPO, N-acetylcysteine, NOX, SOD, U937 cells, oxidative stress,
• MeSH
• Acetylcysteine pharmacology   MeSH
• HL-60 Cells MeSH
• Catalase genetics   MeSH
• Leukemia drug therapy   genetics   metabolism   MeSH
• Humans MeSH
• NADPH Oxidase 2 genetics   MeSH
• Oxidative Stress drug effects   MeSH
• Peroxidase genetics   MeSH
• Cell Proliferation drug effects   MeSH
• Reactive Oxygen Species metabolism   MeSH
• Gene Expression Regulation, Neoplastic drug effects   MeSH
• Gene Expression Profiling MeSH
• Superoxide Dismutase genetics   MeSH
• U937 Cells MeSH
• Cell Survival drug effects   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Acetylcysteine MeSH
• CYBB protein, human MeSH Browser
• Catalase MeSH
• MPO protein, human MeSH Browser
• NADPH Oxidase 2 MeSH
• Peroxidase MeSH
• Reactive Oxygen Species MeSH
• Superoxide Dismutase MeSH

The question as to whether A3 adenosine receptor (A3AR) agonists, N (6)-(3-iodobenzyl)-adenosine-5'-N- methyluronamide (IB-MECA) and 2-chloro-N (6)-(3-iodobenzyl)-adenosine-5'-N-methyluronamide (Cl-IB-MECA), could exert cytotoxic effects at high concentrations with or without the involvement of A3AR has been a controversial issue for a long time. The initial findings suggesting that A3AR plays a crucial role in the induction of cell death upon treatment with micromolar concentrations of IB-MECA or Cl-IB-MECA were revised, however, the direct and unequivocal evidence is still missing. Therefore, the sensitivity of Chinese hamster ovary (CHO) cells transfected with human recombinant A3AR (A3-CHO) and their counter partner wild-type CHO cells, which do not express any of adenosine receptors, to micromolar concentrations of IB-MECA and Cl-IB-MECA was studied. We observed that IB-MECA and Cl-IB-MECA exhibited a strong inhibitory effect on cell proliferation due to the blockage of cell cycle progression at G1/S and G2/M transitions in both A3-CHO and CHO cells. Further analysis revealed that IB-MECA and Cl-IB-MECA attenuated the Erk1/2 signalling irrespectively to A3AR expression. In addition, Cl-IB-MECA induced massive cell death mainly with hallmarks of a necrosis in both cell lines. In contrast, IB-MECA affected cell viability only slightly independently of A3AR expression. IB-MECA induced cell death that exhibited apoptotic hallmarks. In general, the sensitivity of A3-CHO cells to micromolar concentrations of IB-MECA and Cl-IB-MECA was somewhat, but not significantly, higher than that observed in the CHO cells. These results strongly suggest that IB-MECA and Cl-IB-MECA exert cytotoxic effects at micromolar concentrations independently of A3AR expression.

• MeSH
• Adenosine analogs & derivatives   pharmacology   MeSH
• Adenosine A3 Receptor Agonists pharmacology   MeSH
• CHO Cells MeSH
• Cricetulus MeSH
• Cytotoxins pharmacology   MeSH
• Cell Cycle Checkpoints drug effects   MeSH
• Humans MeSH
• Mitogen-Activated Protein Kinase 1 antagonists & inhibitors   genetics   metabolism   MeSH
• Mitogen-Activated Protein Kinase 3 antagonists & inhibitors   genetics   metabolism   MeSH
• Cell Proliferation drug effects   MeSH
• Proto-Oncogene Proteins c-akt genetics   metabolism   MeSH
• Receptor, Adenosine A3 genetics   metabolism   MeSH
• Gene Expression Regulation MeSH
• Signal Transduction drug effects   MeSH
• Transfection MeSH
• Cell Survival drug effects   MeSH
• Dose-Response Relationship, Drug MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• 2-chloro-N(6)-(3-iodobenzyl)adenosine-5'-N-methyluronamide MeSH Browser
• Adenosine MeSH
• Adenosine A3 Receptor Agonists MeSH
• Cytotoxins MeSH
• MAPK1 protein, human MeSH Browser
• Mitogen-Activated Protein Kinase 1 MeSH
• Mitogen-Activated Protein Kinase 3 MeSH
• N(6)-(3-iodobenzyl)-5'-N-methylcarboxamidoadenosine MeSH Browser
• Proto-Oncogene Proteins c-akt MeSH
• Receptor, Adenosine A3 MeSH