Molecular actions of a Mn(III)Porphyrin superoxide dismutase mimetic and peroxynitrite scavenger: reaction with nitric oxide and direct inhibition of NO synthase and soluble guanylyl cyclase.
Mn(III)tetrakis(1-methyl-4-pyridyl)porphyrin (MnTMPyP), described as a superoxide dismutase mimetic and peroxynitrite scavenger, has been used previously to investigate the cytotoxic potential of superoxide and peroxynitrite in several pathological models. Here we report on the interference of MnTMPyP with NO/cGMP signaling using cultured endothelial cells as well as purified soluble guanylyl cyclase (sGC) either activated by the NO donor 2,2-diethyl-1-nitroso-oxyhydrazine sodium salt (DEA/NO) or reconstituted with nitric oxide synthase (NOS). MnTMPyP inhibited endothelial cGMP accumulation induced by A23187 (0.3 microM) with an IC50 of 75.0 +/- 10.4 microM but had no significant effect on the potency of the Ca2+ ionophore. Purified NOS was inhibited by MnTMPyP (IC50 = 5.5 +/- 0.8 microM) because of an interference of the Mn-porphyrin with the reductase domain of the enzyme. The most pronounced actions of MnTMPyP were direct inhibition of sGC and scavenging of NO. Purified sGC stimulated with either Ca2+/calmodulin-activated NOS (in the presence of GSH) or DEA/NO (in the absence of GSH) was inhibited with IC50 values of 0.8 +/- 0.09 microM and 0.6 +/- 0.2 microM, respectively. In the presence of GSH, MnTMPyP was reduced to the Mn(II) complex, resulting in efficient scavenging of NO under these conditions. Our data demonstrate that MnTMPyP (i) interferes with the reductase domain of NOS, (ii) scavenges NO in the presence of GSH, and (iii) is a potent direct inhibitor of sGC. These results cast doubt on the usefulness of MnTMPyP and related Mn-porphyrin complexes as probes to study the involvement of peroxynitrite/superoxide in biological systems.
Pfeiffer
S
S
Institut für Pharmakologie und Toxikologie, Karl-Franzens Universität Graz, Austria.
Schrammel
A
A
Koesling
D
D
Schmidt
K
K
Mayer
B
B
D000818:Animals; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000201:enzymology / Q000378:metabolism; D016166:Free Radical Scavengers / Q000737:chemistry / Q000494:pharmacology*; D005978:Glutathione / Q000378:metabolism; D006162:Guanylate Cyclase / Q000037:antagonists & inhibitors*; D006801:Humans; D008345:Manganese / Q000737:chemistry; D008665:Metalloporphyrins / Q000737:chemistry / Q000494:pharmacology*; D009566:Nitrates / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors*; D051381:Rats; D012995:Solubility; D013053:Spectrophotometry; D013482:Superoxide Dismutase / Q000378:metabolism*; D013552:Swine
D016166:Free Radical Scavengers; D008665:Metalloporphyrins; D009566:Nitrates; C039642:peroxynitric acid; D009569:Nitric Oxide; D008345:Manganese; C050950:tetrakis(N-methyl-4-pyridiniumyl)porphine manganese(III) complex; D019001:Nitric Oxide Synthase; D013482:Superoxide Dismutase; D006162:Guanylate Cyclase; D005978:Glutathione