title: Nitrite-mediated modulation of HL-60 cell cycle and proliferation: involvement of cyclin-dependent kinase 2 activation.
abstract: Recent research suggests the vivid possibility of using nitrite therapy against various pathological conditions. Moreover, chronic nitrite therapy offers protection against ischemia and augments endothelial cell proliferation through unknown mechanisms. Nitrite-mediated augmentation in the number of circulating neutrophils has also been reported; however, the exact mechanism is not known. In the present study, we have investigated the effect of nitrite (0.5-10 mM) on the proliferation of the neutrophilic cell line HL-60 and also explored the underlying mechanism. Treatment of HL-60 cells with sodium nitrite (0.5-5 mM) led to an increase in cell proliferation, which was confirmed by cell cycle analysis and 5-bromo-2-deoxyuridine and thymidine incorporation, whereas cells accumulated in the G(0)/G(1) phase after treatment with 10 mM nitrite. Experiments on the synchronized cells exhibited similar effect, which seems to be nitric oxide (NO)-dependent, because carboxyl-1H-imidazol-1-yloxy,2-(4-carboxyphenyl)-4,5-dihydro 4,4,5,5-tetramethyl-3-oxide abolished nitrite-mediated proliferative effect. Moreover, the NO donor sodium nitroprusside at micromolar concentrations also exhibited similar effects. Nitrite induced augmentation in S phase, and intracellular reactive oxygen species (ROS) generation was prevented by ROS scavenger/inhibitors. Moreover, mitochondrial blockers, rotenone and antimycin A, also reduced nitrite-mediated cell proliferation. Assessment of the cell cycle regulators cyclin-dependent kinase 2 (Cdk2), Cdk4, cyclin A, cyclin D, cyclin E, and p21 suggested augmentation in the expression and interaction of Cdk2/cyclin E and Cdk2 activity, whereas p21 was down-regulated. Indeed proliferative effect of nitrite was blocked by roscovitine, a Cdk2 inhibitor. The results obtained demonstrate that the proliferative effect of nitrite on HL-60 cells seems to be NO-mediated, redox-sensitive, and Cdk2 activation-dependent, warranting detailed studies before initiating its clinical use.
authors.affiliation: Pharmacology Division, Central Drug Research Institute, Lucknow 226 001, India.
authors.forename: Sachin
authors.initials: S
authors.lastname: Kumar
authors.forename: Manoj Kumar
authors.initials: MK
authors.lastname: Barthwal
authors.forename: Madhu
authors.initials: M
authors.lastname: Dikshit
mesh_terms: D002453:Cell Cycle / Q000187:drug effects* / Q000235:genetics; D049109:Cell Proliferation / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects*; D003497:Cyclic N-Oxides / Q000494:pharmacology; D051357:Cyclin-Dependent Kinase 2 / Q000378:metabolism*; D018844:Cyclin-Dependent Kinases / Q000378:metabolism; D016213:Cyclins / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation; D016166:Free Radical Scavengers / Q000494:pharmacology; D016193:G1 Phase / Q000187:drug effects; D018922:HL-60 Cells; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism; D009599:Nitroprusside / Q000494:pharmacology; D010084:Oxidation-Reduction; D017382:Reactive Oxygen Species / Q000378:metabolism; D012977:Sodium Nitrite / Q000494:pharmacology*
chemical_list: D003497:Cyclic N-Oxides; D016213:Cyclins; D016166:Free Radical Scavengers; D007093:Imidazoles; D017382:Reactive Oxygen Species; D009599:Nitroprusside; C079391:2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide; D009569:Nitric Oxide; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases; D012977:Sodium Nitrite