title: Induction of apoptosis and reactive oxygen species production by N-nitrosopiperidine and N-nitrosodibutylamine in human leukemia cells.
abstract: N-nitrosopiperidine (NPIP) and N-nitrosodibutylamine (NDBA) belong to a group of N-nitrosamines that are widely distributed in foodstuffs and the occupational environment. In the present study, the human promyelocytic leukemia cell line HL-60, was used to characterize the apoptotic effects of N-nitrosamines, and to examine the production of reactive oxygen species (ROS). Apoptotic cells were identified by (i) chromatin condensation (ii) flow cytometry analysis and (iii) poly(ADP-ribose) polymerase (PARP) cleavage. NPIP and NDBA induced morphological changes consistent with apoptotic events in HL-60 cells. Flow cytometry analysis showed that both N-nitrosamines induced apoptotic cell death in a concentration and time dependent-manner. It was observed that NDBA was stronger than NPIP, since it induced a significant apoptotic cell death after 18 h starting from a concentration of 2 mm, whereas NPIP was effective at 10 mm. Furthermore, PARP was markedly cleaved with 0.5 mm of NDBA and 5 mm of NPIP after treatments for 3 and 18 h, respectively. Finally, the ROS level was found to be elevated after 0.5 h of treatment with both N-nitrosamines. Antioxidant N-acetylcysteine (NAC) completely inhibited the ROS production induced by NPIP and NDBA. However, this action seems not to be associated with the apoptosis because NAC did not block N-nitrosamines-induced apoptosis. The data demonstrate that NPIP and NDBA induce apoptosis and ROS production in HL-60 cells.
authors.affiliation: Departamento de Nutrición, Bromatología y Tecnología de los Alimentos, Facultad de Veterinaria, Universidad Complutense de Madrid, 28040 Madrid, Spain.
authors.forename: Almudena
authors.initials: A
authors.lastname: García
authors.forename: Paloma
authors.initials: P
authors.lastname: Morales
authors.forename: Nuria
authors.initials: N
authors.lastname: Arranz
authors.forename: Eugenia
authors.initials: E
authors.lastname: Delgado
authors.forename: Joseph
authors.initials: J
authors.lastname: Rafter
authors.forename: Ana I
authors.initials: AI
authors.lastname: Haza
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology; D000975:Antioxidants / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D002273:Carcinogens / Q000633:toxicity*; D048430:Cell Shape / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D042002:Chromatin Assembly and Disassembly / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D018922:HL-60 Cells; D006801:Humans; D020287:In Situ Nick-End Labeling; D007938:Leukemia / Q000378:metabolism / Q000473:pathology*; D009602:Nitrosamines / Q000633:toxicity*; D018384:Oxidative Stress / Q000187:drug effects*; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013997:Time Factors
chemical_list: D000975:Antioxidants; D002273:Carcinogens; D009602:Nitrosamines; D017382:Reactive Oxygen Species; C002743:N-nitrosopiperidine; C012279:dibutylnitrosamine; D011065:Poly(ADP-ribose) Polymerases; D000111:Acetylcysteine