Nitric Oxide and Lipid Peroxidation Valerie B. O Donnell a, Neil Hogg b and Victor M. Darley Usmar c a Dept of Med. Biochem. University of Wales College of Medicine, Heath Park, Cardiff CF14 4XN, UK. Ph: 0044-2920-748447 Fax: 0044-2920-744905 Email: o-donnellvb@cardiff.ac.uk b Biophysics Research Institute, Medical College of Wisconsin, Milwaukee, WI USA. c Department of Pathology, University of Alabama at Birmingham, AL USA NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 1
Nitric Oxide is a Free Radical N=O NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 2
Nitric oxide is not particularly oxidizing. Nitric oxide is (infinitely) stable*. Nitric oxide does not rapidly react with organic molecules. *In the absence of dioxygen. NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 3
The Nitric Oxide Radical and some of its stable end-products N=O nitric oxide - O-N=O nitrite RO-N=O alkyl nitrite RS-N=O nitrosothiol NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 4
Reactions of NO with Other Free Radicals Oxidation of NO to nitrite: 4 NO+ O=O +2H 2 O 4HNO 2 Nitric oxide reaction with organic peroxyl radicals: NO+ ROO ROONO?? NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 5
Lipid Peroxidation LOO + LH + O 2 LOO + LOOH LOO is the central and rate-limiting species of lipid peroxidation NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 6
LOOH Non-enzymatic lipid oxidation Metal or enzyme AOO O 2 LOO LH L LOOH O 2 LOO LH L LOOH O 2 LOO LH GPx GPx Initiation LOH Propagation LOH Propagation NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 7
Lipid peroxidation products in an atherosclerotic lesion are both enzymatically and nonenzymatically produced. Foam cell 15-LOX Necrotic core lipid hydroxides, hydroperoxides, aldehydes, epoxides, ketones, ceroid, isoprostanes Fibrous cap NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 8
Location of lipid oxidation enzymes in the vasculature LOX: lipoxygenase PGHS: prostaglandin H synthase Smooth muscle: 12-LOX Endothelium PGHS-1 Monocyte: PGHS-1,2 5, 15-LOX Platelet: PGHS-1 12-LOX Neutrophil: PGHS-1,2 5,12-LOX NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 9
Lipid oxidation enzymes are upregulated during vascular disease. * Inhibition of 12-LOX restores blood pressure in Angiotensin- II dependent hypertensive rats. * Increased products of PGHS in hypertensive rats, and PGHS inhibition restores blood pressure in humans and rats. * Smooth muscle 12-LOX in hypertension * Platelet 12-LOX in hypertension * Foam cell/macrophage 15-LOX in atherosclerosis. NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 10
Nitric oxide reaction with lipid peroxyl radicals LOO NO LOONO H 2 O [LO NO 2 ] LOOH + NO 2 - LO NO 2 NO [L(O) NO 2 ] NO LONO NO - 2 L(O)NO 2 NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 11
Reactions between NO-derived reactive nitrogen species and unsaturated lipid NO LOONO L(O)NO 2 O 2 O 2 - NO 2 ONOO - NO HONO LH O L 2 LOO e -, H + LOOH NO 2, ONOO - LNO 2 NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 12
Nitric oxide inhibition of lipid oxidation coincides with fast rates of NO uptake during ABAP-dependent linoleate oxidation. (A) (B) 200 0.6 Oxygen (µm) 100 ABAP 0.2 NO (µm) -0.2 0 [ NO] = 0.9 µm added at vertical lines to 7.3 mm linoleate in 0.5% cholate, 11 mm ABAP -0.6 0 400 800 1200 Time/s NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 13
Mechanism of Inhibition of LDL Oxidation by NO LOO + NO LOONO LO + NO 2 LONO 2 LOOH OLOO + NO OLOONO OLONO 2 Two NO consumed for each LOO 4 NO + 2LOO + H 2 O 2HNO 2 + LONO 2 + LONO NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 14
Cholesteryl Ester Triglyceride Phospholipid Apolipoprotein B Oxidant LH NO 2 O 2 O 2 L NO 2 LH LOO NO LNO 2 2 L α-tocopherol LOOH NO L(O)NO 2 Cholesterol NO NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 15
500 NO and LDL Oxidation: The effect of HPODE Lag time of LDL (125 µg/ml) oxidation by Cu(II) (20 µm) with and without HPODE (5 µm) length of lag time (min) 400 300 200 100 0 200 150 100 50 A SNN SNN + HpODE 0 4 8 12 SNN (µm) B BHT BHT + HpODE 0 0 2 4 6 8 10 BHT (µm) NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 16
Comparison of nitric oxide and tocopherol inhibition of linoleate oxidation. 300 ABAP α -tocopherol (1 µm) [Oxygen]/µM 200 100 T inh NO (1 µm) T inh 0 0 200 400 600 800 1000 1200 Time/s Oxidation of linoleate (7.3 mm) by ABAP (11 mm) NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 17
Nitric oxide is consumed by rabbit 15-lipoxygenase during oxidation of linoleate. 2 LOX LOX LOX NO (µm) 1 Add figure caption 0 (i) (ii) (iii) 0 500 1000 1500 Time (secs) NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 18
Site of lipoxygenase inhibition by NO E-Fe 2+ LOOH E-Fe 3+ NO E-Fe 3+ LOO E-Fe 2+ L O 2 E-Fe 2+ LOO NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 19
Activation of LOX inhibits activation of sgc by NO NO Lox 100 sgc cgmp cgmp (% control) 50 ** 0 control AA, 5 µm LOX AA, 5 µm, LOX NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 20
Further Reading: Darley-Usmar, VM et al., (1992) Free Radic. Res. Commun. 17, 9-20 Graham, A et al., (1993) FEBS Lett. 330, 181-185 Rubbo, H et al., (1994) J. Biol. Chem. 265, 26066-26075 Hogg, N et al., (1995) J. Lipid Res. 36, 1756-1762 Struck, N et al., (1995) FEBS Lett 361, 291-294 O Donnell, VB et al., (1997) Biochemistry 36, 15216-15223 O Donnell, VB et al., (1999) Chem. Res. Toxicol. 12, 83-92 O Donnell, VB et al., (1999) J. Biol. Chem. 274, 20083-20091 Hogg, N & Kalyanaraman, B (1999) Biochim. Biophys. Acta. 1411, 378-384 Rubbo, H et al., (2000) J. Biol. Chem. 275, 10812-10818 O Donnell, VB & Freeman, BA (2001) Circ. Res. 88, 16-25 NO & LPO Society For Free Radical Biology and Medicine O Donnell et al. 21