Poly(ADP-ribose) synthetase activation mediates increased permeability induced by peroxynitrite in Caco-2BBe cells.
OBJECTIVE
Peroxynitrite induces cytotoxicity by generating DNA single-strand breaks and activating poly(ADP-ribose) synthetase (PARS), a nuclear enzyme that consumes oxidized nicotinamide adenine dinucleotide (NAD+) and depletes cellular adenosine triphosphate (ATP). The aim of this study was to examine this mechanism of injury in an intestinal epithelial cell model after exposure to exogenous peroxynitrite (ONOO-) and nitric oxide (NO).


METHODS
Caco-2BBe cell monolayers exposed to donors of peroxynitrite (3-morpholino-sydnonimine [SIN-1], 3 mmol/L) or NO (S-nitroso-N-acetyl penicillamine [SNAP]; 3 mmol/L) were analyzed for DNA strand breaks, [NAD+], [ATP], and transepithelial flux of fluorescein sulfonic acid.


RESULTS
SIN-1 but not SNAP induced DNA single-strand breakage. Both SIN-1 and SNAP reduced [ATP], but only SIN-1 reduced [NAD+]. Inhibition of PARS activity by the PARS inhibitors 5-iodo-6-amino 1,2-benzopyrone or 3-aminobenzamide prevented the SIN-1-induced reduction in [NAD+] and [ATP] but had no effect on the SNAP-induced reduction in [ATP]. PARS inhibition reduced SIN-1-but not SNAP-induced hyperpermeability.


CONCLUSIONS
Peroxynitrite but not NO increases transepithelial permeability by inducing DNA strand breaks that activate the PARS pathway and cause the depletion of intracellular energy stores. Inhibition of PARS activity may represent a novel strategy in ameliorating peroxynitrite-mediated epithelial injury during intestinal inflammation.
Kennedy
M
M
Division of Gastroenterology, Children's Hospital Medical Center, Cincinnati, Ohio 45229, USA.
Denenberg
A G
AG
Szabó
C
C
Salzman
A L
AL
D000255:Adenosine Triphosphate / Q000032:analysis; D018938:Caco-2 Cells; D004249:DNA Damage; D006801:Humans; D015212:Inflammatory Bowel Diseases / Q000188:drug therapy; D007413:Intestinal Mucosa / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008981:Molsidomine / Q000031:analogs & derivatives / Q000494:pharmacology; D009566:Nitrates / Q000633:toxicity*; D009569:Nitric Oxide / Q000502:physiology; D010396:Penicillamine / Q000031:analogs & derivatives / Q000494:pharmacology; D010539:Permeability; D011065:Poly(ADP-ribose) Polymerases / Q000502:physiology*; D026423:S-Nitroso-N-Acetylpenicillamine
D009566:Nitrates; C039642:peroxynitric acid; D009569:Nitric Oxide; C002385:linsidomine; D026423:S-Nitroso-N-Acetylpenicillamine; D000255:Adenosine Triphosphate; D008981:Molsidomine; D011065:Poly(ADP-ribose) Polymerases; D010396:Penicillamine