The role of NO and superoxide (O2-) in tissue injury during cardiac allograft rejection was investigated by using a rat ex vivo organ perfusion system. Excessive NO production and inducible NO synthase (INOS) expression were observed in cardiac allografts at 5 days after cardiac transplantation, but not in cardiac isografts, as identified by electron spin resonance spectroscopy and Northern blotting. Cardiac Isografts or allografts obtained on Day 5 after transplantation were perfused with Krebs bicarbonate buffer with or without various antidotes for NO or O2-, including Nω-monomethyl-L-arglnine (L-NMMA; 1 mM), 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxlde (PTIO; 100 μ/M), 4-amino-6-hydroxypyrazolo[3,4-d]pyrimldlne (AHPP; a xanthine oxidase Inhibitor; 100 μM), and superoxide dismutase (SOD; 100 units/ml). Treatment of the cardiac allografts with PTIO showed most remarkable Improvement of the cardiac Injury as revealed by significant reduction in aspartate transaminase, lactate dehydrogenase, and creatine Phosphokinase concentrations in the perfusate. Similar but less potent protective effect on the allograft injury was observed by treatment with L-NMMA, AHPP, and SOD. Immunohlstochemlcal analyses for INOS and nitrotyrosine indicated that INOS is mainly expressed by macrophages infiltrating the allograft tissues, and nitrotyrosine formation was demonstrated not only in macrophages but also in cardiac myocytes of the allografts, providing indirect evidence for the generation of peroxynitrite during allograft rejection. Our results suggest that tissue Injury in rat cardiac allografts during acute rejection is mediated by both NO and O2-, possibly through peroxynitrite formation.
LangrehrJM, MuraseN, MarkusPM, CaiX, NeuhausP, SchrautW, SimmonsRL, HoffmanRA. Nitric oxide production in host-versus-graft and graft-versus-host reactions in the rat. J Clin Invest90: 679–683, 1992.
3.
LancasterJRJr.., LangrehrJM, BergoniaHA, MuraseN, SimmonsRL, HoffmanRA. EPR detection of heme and nonheme iron-containing protein nitrosylation by nitric oxide during rejection of rat heart allograft. J Biol Chem267: 10994–10998, 1992.
4.
MacMillian-CrowLA, CrowJP, KerbyJD, BeckmanJS, ThompsonJA. Nitration and inactivation of manganese superoxide dismutase in chronic rejection of human renal allografts. Proc Natl Acad Sci U S A93: 11853–11858, 1996.
BeckmanJS, BeckmanTW, ChenJ, MarshallPA, FreemanBA. Apparent hydroxyl radical production by peroxynitrite: Implications for endothelial injury from nitric oxide and superoxide. Proc Natl Acad Sci U S A87: 1620–1624, 1990.
7.
BeckmanJS, KoppenolWH. Nitric oxide, superoxide, and peroxynitrite: The good, the bad, and the ugly. Am J Physiol271: C1424–C1437, 1996.
8.
FreemanBA. Free radical chemistry of nitric oxide: Looking at the dark side. Chest105: 79S–84S, 1994.
9.
Darley-UsmarV, WisemanH, HalliwellB.Nitric oxide and oxygen radicals: A question of balance. FEBS Lett369: 131–135, 1995.
10.
RubboH, Darley-UsmarV, FreemanBA. Nitric oxide regulation of tissue free radial injury. Chem Res Toxicol9: 809–820, 1996.
11.
RadiR, BeckmanJS, BushK, FreemanBA. Peroxynitrite oxidation of sulfhydryls: The cytotoxic potential of superoxide and nitric oxide. J Biol Chem266: 4244–4250, 1991.
RadiR, BeckmanJS, BushKM, FreemanBA. Peroxynitrite-induced membrane lipid peroxidation: The cytotoxic potential of superoxide and nitric oxide. Arch Biochem Biophys288: 481–487, 1991.
14.
IschiropoulosH.Biological tyrosine nitration: A pathophysiological function of nitric oxide and reactive oxygen species. Arch Biochem Biophys356: 1–11, 1998.
15.
CrowJP, BeckmanJS. Reactions between nitric oxide, superoxide, and peroxynitrite: Footprints of peroxynitrite in vivo.Adv Pharmacol34: 17–43, 1995.
16.
KoglinJ, Glysing-JensenT, MudgettJS, RusselME. NOS2 mediates opposing effects in models of acute and chronic cardiac rejection: Insight from NOS2-knockout mice. Am J Pathol153: 1371–1376, 1998.
17.
AkaikeT, NoguchiY, IjiriS, SetoguchiK, SugaM, ZhengYM, DietzscholdB, MaedaH.Pathogenesis of influenza virus-induced pneumonia: Involvement of both nitric oxide and oxygen radicals. Proc Natl Acad Sci U S A93: 2448–2453, 1996.
18.
WrayG, ThiemermannC.NO in sepsis. In: FangFC Ed. Nitric Oxide and Infection. New York: Kluwer Academic/Plenum Publishers, pp 265–280, 1999.
19.
TownsendG, ScheidM.Nitric oxide in bacterial meningitis. In: FangFC Ed. Nitric Oxide and Infection. New York: Kluwer Academic/Plenum Publishers, pp 417–428, 1999.
20.
AkaikeT, YoshidaM, MiyamotoY, SatoK, KohnoM, SasamotoK, MiyazakiK, UedaS, MaedaH.Antagonistic action of imidazolineoxyl N-oxides against endothelium-derived relaxing factor/-153537020022500207NO through a radical reaction. Biochemistry32: 827–832, 1993.
21.
YoshidaM, AkaikeT, WadaY, SatoK, IkedaK, UedaS, MaedaH.Therapeutic effects of imidazolineoxyl N-oxide against endotoxin shock through its direct nitric oxide-scavenging activity. Biochem Biophys Res Commun202: 923–930, 1994.
22.
WhiteCR, Darley-UsmarV, BerringtonWR, McAdamsM, GoreJZ, ThompsonJA, ParksDA, TarpeyMM, FreemanBA. Circulating plasma xanthine oxidase contributes to vascular dysfunction in hyper-cholesterolemic rabbits. Proc Natl Acad Sci U S A93: 8745–8749, 1996.
YamaguchiY, HarlandRC, WybleC, MoriK, BollingerRR. Variable allograft responses to pretreatment with donor splenocytes treated with mitomycin C in the rat. Transplantation47: 360–363, 1989.
25.
ZweierJL. Measurement of superoxide-derived free radicals in the reperfused heart: Evidence for a free radical mechanism of reperfusion injury. J Biol Chem263: 1353–1357, 1988.
26.
AkaikeT, InoueK, OkamotoT, NishinoH, OtagiriM, FujiiS, MaedaH.Nanomolar quantification and identification of various nitrosothiols by high performance liquid chromatography coupled with flow reactors of metals and Griess reagent. J Biochem122: 459–466, 1997.
27.
SetoguchiK, TakeyaM, AkaikeT, SugaM, HattoriR, MaedaH, AndoM, TakahashiK.Expression of inducible nitric oxide synthase and its involvement in pulmonary granulomatous inflammation in rats. Am J Pathol149: 2005–2022, 1996.
28.
FujiiS, AkaikeT, MaedaH.Role of nitric oxide in pathogenesis of herpes simplex virus encephalitis. Virology256: 203–212, 1999.
29.
IyonagaK, TakeyaM, YamamotoT, AndoM, TakahashiK.A novel monoclonal antibody, RM-4, specifically recognizes rat macrophages and dendritic cells in formalin-fixed, paraffin-embedded tissues. Histochem J29: 105–116, 1997.
30.
WorrallNK, MiskoTP, SullivanPM, HuiJJ, FergusonTBJr.Inhibition of inducible nitric oxide synthase attenuates established acute cardiac allograft rejection. Ann Thorac Surg62: 378–385, 1996.
PieperGM, CooperM, JohnsonCP, AdamsMB, FelixCC, RozaAM. Reduction of myocardial nitrosyl complex formation by a nitric oxide scavenger prolongs cardiac allograft survival. J Cardiovasc Pharmacol35: 114–120, 2000.
33.
NathanC.Inducible nitric oxide synthase: What difference does it make?J Clin Invest100: 2417–2423, 1997.
34.
AkaikeT, MaedaH.Pathophysiological actions of high-output production of nitric oxide. In: IgnarroL Ed. Nitric Oxide: Biology and Pathobiology. San Diego: Academic Press, pp 733–745, 2000.
AkaikeT.Mechanism of biological S-nitrosation and its measurement. Free Radic Res (in press) 2000.
37.
InoueK, AkaikeT, MiyamotoY, OkamotoT, SawaT, OtagiriM, SuzukiS, YoshimuraT, MaedaH.Nitrosothiol formation catalyzed by ceruloplasmin: Implication for cytoprotective mechanism in vivo.J Biol Chem274: 27069–27075, 1999.
38.
RauhalaP, LinAM, ChiuehCC. Neuroprotection by S-nitrosoglutathione of brain dopamine neurons from oxidative stress. FASEB J12: 165–173, 1998.
39.
KonorevEA, TarpeyMM, JosephJ, BakerJE, KalyanaramanB.S-nitrosoglutathione improves functional recovery in the isolated rat heart after cardioplegic ischemic arrest: Evidence for a cardioprotective effect of nitric oxide. J Pharmacol Exp Ther274: 200–206, 1995.
40.
CastroL, RodriguezM, RadiR.Aconitase is readily inactivated by peroxynitrite, but not by its precursor, nitric oxide. J Biol Chem269: 29409–29415, 1994.
41.
HausladenA, FridovichI.Superoxide and peroxynitrite inactivate aconitase, but nitric oxide does not. J Biol Chem269: 29405–29408, 1994.
42.
SzabóC, CuzzocreaS, ZingarelliB, O'connorM, SalzmanAL. Endothelial dysfunction in a rat model of endotoxic shock: Importance of the activation of poly (ADP-ribose) synthetase by peroxynitrite. J Clin Invest100: 723–735, 1997.
43.
SzabóC, ZingarelliB, O'connorM, SalzmanAL. DNA strand breakage, activation of poly (ADP-ribose) synthetase, and cellular energy depletion are involved in the cytotoxicity of macrophages and smooth muscle cells exposed to peroxynitrite. Proc Natl Acad Sci U S A93: 1753–1758, 1996.
44.
OkamotoT, AkaikeT, NaganoT, MiyajimaS, SugaM, AndoM, IchimoriK, MaedaH.Activation of human neutrophil procollagenase by nitrogen dioxides and peroxynitrite: A novel mechanism for procollagenase activation involving nitric oxide. Arch Biochem Biophys342: 261–274, 1997.
45.
FrearsER, ZhangZ, BlakeDR, O'connellJP, WinyardPG. Inactivation of tissue inhibitor of metalloproteinase-1 by peroxynitrite. FEBS Lett381: 21–24, 1996.
46.
BeckmanJS, YeYZ, AndersonP, ChenJ, AccavettiMA, TarpeyMM, WhiteCR. Extensive nitration of protein tyrosines observed in human atherosclerosis detected by immunohistochemistry. Biol Chem Hoppe Seyler375: 81–88, 1994.
47.
MillerMJS, ThompsonJH, ZhangX-J, Sadowska-KrowickaH, KakkisJL, MunshiUK, SandovalM, RossiJL, Eloby-ChildressS, BeckmanJS, YeYZ, RodiCP, ManningPT, CurrieMG, ClarkDA. Role of inducible nitric oxide synthase expression and peroxynitrite formation in guinea pig ileitis. Gastroenterology109: 1475–1483, 1995.
48.
KooyNW, RoyallJA, YeYZ, KelleyDR, BeckmanJS. Evidence for in vivo peroxynitrite production in human acute lung injury. Am Rev Respir Dis151: 1250–1254, 1995.
49.
EstévezAG, CrowJP, SampsonJB, ReiterC, ZhuangY, RichardsonGJ, TarpeyMM, BarbeitoL, BeckmanJ.Induction of nitric oxide-dependent apoptosis in motor neurons by zinc-deficient superoxide dismutase. Science286: 2498–2500, 1999.
50.
EiserichJP, HristovaM, CrossCE, JonesAD, FreemanBA, HalliwellB, van der VlietA.Formation of nitric oxide-derived inflammatory oxidants by myeloperoxidase in neutrophils. Nature391: 393–3971998.
51.
PfeifferS, MayerB.Lack of tyrosine nitration by peroxynitrite generated at physiological pH. J Biol Chem273: 27280–27285, 1998.
52.
SawaT, AkaikeT, MaedaH.Tyrosine nitration by peroxynitrite formed from nitric oxide and superoxide generated by xanthine oxidase. J Biol Chem275 (in press) 2000.
53.
ReiterCD, TengRJ, BeckmanJS. Superoxide reacts with nitric oxide to nitrate tyrosine at physiological pH via peroxynitrite. J Biol Chem275 (in press) 2000.
54.
GoldsteinS, CzapskiG.Kinetics of nitric oxide autooxidation in aqueous solution in the absence and presence of various reductants: The nature of the oxidizing intermediates. J Am Chem Soc117: 12078–12084, 1995.
55.
AmanoF, NodaT.Improved detection of nitric oxide radical (NO) production in an activated macrophage culture with a radical scavenger, carboxy PTIO, and Griess reagent. FEBS Lett368: 425–428, 1995.
56.
AkaikeT, MaedaH.Quantitation of nitric oxide using 2-phenyl-4,4,5.5-tetramethylimidazoline-1-oxyl 3-oxide (PTIO). Meth Enzymol268: 211–221, 1996.
