kt-5720 and 2-phenyl-4-4-5-5-tetramethylimidazoline-1-oxyl-3-oxide

Incubation with TNF-alpha (50 ng/ml) for 72 hours markedly reduced viability of endothelial cells. A 6-hour preincubation with S-nitroso-N-acetyl-D,L-penicillamine (SNAP, 3-100 microM) protected cells in a concentration-dependent manner and decreased TNF-alpha-mediated toxicity by up to 70%. Cytoprotection by SNAP was completely abolished by the adenylyl cyclase inhibitor 2', 5'-dideoxyadenosine and mimicked by 8-bromo cyclic AMP or forskolin. SNAP produced significant increases in cyclic GMP and cyclic AMP, both being abrogated in the presence of the NO scavenger 2-phenyl-4, 4,5,5,-tetramethylimidazoline-1-oxyl-3-oxide (PTIO). Moreover, no endothelial protection by SNAP was detected in the presence of the protein kinase A inhibitor KT5720, whereas the protein kinase G inhibitor KT5823 left cytoprotection virtually unaltered. Our results demonstrate a crucial role for cyclic AMP in mediating NO-induced endothelial protection against TNF-alpha, possibly through cyclic GMP-dependent inhibition of cyclic AMP breakdown. NO-dependent endothelial protection may ultimately result from cyclic AMP-induced up-regulation of antioxidant proteins or down-regulation of cytotoxic processes.

Topics: 8-Bromo Cyclic Adenosine Monophosphate; Adenylyl Cyclase Inhibitors; Alkaloids; Animals; Carbazoles; Cattle; Cell Line; Cell Survival; Colforsin; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Cyclic GMP; Cyclic N-Oxides; Dideoxyadenosine; Endothelium, Vascular; Enzyme Inhibitors; Free Radical Scavengers; Imidazoles; Indoles; Kinetics; Nitric Oxide; Nitric Oxide Donors; Oxadiazoles; Penicillamine; Pulmonary Artery; Pyrroles; Quinoxalines; S-Nitroso-N-Acetylpenicillamine; Tumor Necrosis Factor-alpha