title: DNA-protein crosslinks induced by nickel compounds in isolated rat lymphocytes: role of reactive oxygen species and specific amino acids.
abstract: Isolated rat lymphocytes in salts-glucose medium (pH 7.2) were incubated with nickel chloride, nickel acetate, nickel sulfate, and a soluble form of nickel subsulfide (0-2 mM) at 37 degrees C for 2 h. The soluble form of nickel subsulfide induced a significant increase in DNA-protein crosslinks (DPXLs) (111%) beginning at 0.5 mM and a maximum increase of 700% from that of the control value was reached at a 2 mM concentration, whereas nickel sulfate produced only a 65% increase of such crosslinks at the 2 mM concentration only. No significant reduction in viability of rat lymphocytes (as measured by trypan blue exclusion) due to these nickel compounds was observed at any concentration used. Time-course studies of DPXLs and cellular viability due to 2 mM nickel subsulfide indicate that DPXL formation may not be due in part to cellular necrosis. Coincubation of nickel subsulfide (2 mM) with l-histidine (16 mM), l-cysteine (4 or 8 mM), or l-aspartic acid (24 mM) significantly reduced the DPXLs induced by 2 mM nickel subsulfide. But Mg(2+) even at 24 mM failed to antagonize nickel subsulfide-induced increase in DPXLs. High concentrations of these amino acids significantly decreased the accumulation of Ni(2+) from nickel subsulfide in lymphocytes, suggesting that such reduction of cellular uptake of Ni(2+) by these amino acids is partly responsible for the potent protective effects of these amino acids against such genotoxicity of nickel subsulfide. In vitro exposure of lymphocytes to nickel subsulfide (0-2 mM) increased the formation of reactive oxygen species (ROS) in a concentration-dependent manner. Furthermore, coincubation of 2 mM nickel subsulfide with catalase, dimethylthiourea, mannitol, or vitamin C at 37 degrees C for 2 h resulted in a significant decrease of nickel subsulfide-induced formation of DPXLs, suggesting that nickel subsulfide-induced DPXLs formation in isolated rat lymphocytes is caused by the formation of ROS. The amino acid treatment also abrogated Ni(3)S(2)-induced generation of ROS. Deferoxamine (a highly specific iron chelator) treatment prevented nickel subsulfide-induced DNA-protein crosslink formation, suggesting that Ni(2+)-induced DPXL formation in rat lymphocytes is caused by the induction of Fenton/Haber-Weiss reaction, generating hydroxyl radicals. The potent protective effects of these specific amino acids against nickel subsulfide-induced DPXL formation in isolated rat lymphocytes may be due in part to impaired cellular uptake of Ni(2+), inhibition of the binding of Ni(2+) to deproteinized DNA, and a reduction in reactive oxygen species.
authors.affiliation: Département de santé environnementale et santé au travail, Faculté de médecine, Université de Montréal, Québec, Quebec, H3C 3J7, Canada.
authors.forename: S K
authors.initials: SK
authors.lastname: Chakrabarti
authors.forename: C
authors.initials: C
authors.lastname: Bai
authors.forename: K S
authors.initials: KS
authors.lastname: Subramanian
mesh_terms: D000085:Acetates / Q000633:toxicity; D000818:Animals; D003432:Cross-Linking Reagents / Q000633:toxicity; D003545:Cysteine / Q000494:pharmacology; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004249:DNA Damage; D003676:Deferoxamine / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D016166:Free Radical Scavengers / Q000494:pharmacology; D006639:Histidine / Q000494:pharmacology; D008214:Lymphocytes / Q000187:drug effects* / Q000378:metabolism; D008274:Magnesium / Q000494:pharmacology; D009532:Nickel / Q000378:metabolism / Q000633:toxicity; D016273:Occupational Exposure; D009942:Organometallic Compounds / Q000633:toxicity; D018384:Oxidative Stress; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism*
chemical_list: D000085:Acetates; D003432:Cross-Linking Reagents; D016166:Free Radical Scavengers; D009942:Organometallic Compounds; D011506:Proteins; D017382:Reactive Oxygen Species; C029938:nickel sulfate; D006639:Histidine; C022838:nickel chloride; D009532:Nickel; D004247:DNA; C119536:nickel acetate; D008274:Magnesium; D003676:Deferoxamine; D003545:Cysteine; C017557:nickel subsulfide