title: The impact of N-nitrosomelatonin as nitric oxide donor in cell culture experiments.
abstract: N-nitrosomelatonin (NOMela) is well-known for its capabilities of transnitrosating nucleophiles such as thiols and ascorbate, thereby generating nitric oxide (NO)-releasing compounds. It is unknown, however, whether NOMela can be successfully applied as a precursor of NO in a complex biological environment like a cell culture system. NO donors may be useful to induce the transcription factor hypoxia inducible factor 1 (HIF-1), which coordinates the protection of cells and tissues from the lack of oxygen (hypoxia). In this study, the effects of NOMela in an in vitro cell-free assay [NO-release, inhibition of prolylhydroxylase1 (PHD1)] and in living cells (upregulation of HIF-1, reduction of HIF-1 hydroxylation, upregulation of the HIF-1-target gene PHD2) were compared with those of the frequently applied NO donor S-nitrosoglutathione (GSNO) under normoxic and hypoxic conditions. In contrast to GSNO, NOMela released NO in a predictable manner and this release in vitro was found to be independent of the composition of the buffer system. The NOMela-mediated effects in oxygenated cells were in all cases comparable to the hypoxic response, whereas unphysiological strong effects were observed with GSNO. Probably, because of the antioxidative power of the NOMela-dependent formation of melatonin, cells were completely protected against the attack of reactive nitrogen oxygen species, which are generated by autoxidation of NO. In conclusion, NOMela had to be an excellent NO precursor for cells in culture and potentially tissues.
authors.affiliation: Institut für Physiologie, Universität Duisburg-Essen, Essen, Germany.
authors.forename: Utta
authors.initials: U
authors.lastname: Berchner-Pfannschmidt
authors.forename: Suzan
authors.initials: S
authors.lastname: Tug
authors.forename: Buena
authors.initials: B
authors.lastname: Trinidad
authors.forename: Maria
authors.initials: M
authors.lastname: Becker
authors.forename: Felix
authors.initials: F
authors.lastname: Oehme
authors.forename: Ingo
authors.initials: I
authors.lastname: Flamme
authors.forename: Joachim
authors.initials: J
authors.lastname: Fandrey
authors.forename: Michael
authors.initials: M
authors.lastname: Kirsch
mesh_terms: D000975:Antioxidants; D015153:Blotting, Western; D002021:Buffers; D018929:Cell Culture Techniques; D015687:Cell Hypoxia; D005978:Glutathione / Q000494:pharmacology; D006801:Humans; D051793:Hypoxia-Inducible Factor 1 / Q000378:metabolism; D008550:Melatonin / Q000031:analogs & derivatives* / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology*; D009603:Nitroso Compounds / Q000494:pharmacology*; D011393:Procollagen-Proline Dioxygenase / Q000378:metabolism; D026361:Reactive Nitrogen Species / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D026422:S-Nitrosoglutathione / Q000494:pharmacology; D014162:Transfection
chemical_list: D000975:Antioxidants; D002021:Buffers; D051793:Hypoxia-Inducible Factor 1; C503522:N1-nitrosomelatonin; D020030:Nitric Oxide Donors; D009603:Nitroso Compounds; D026361:Reactive Nitrogen Species; D017382:Reactive Oxygen Species; D009569:Nitric Oxide; D026422:S-Nitrosoglutathione; D011393:Procollagen-Proline Dioxygenase; C030606:proline, 2-oxoglutarate 3-dioxygenase; D005978:Glutathione; D008550:Melatonin