Effects of Various Antioxidants on Endotoxin-Induced Lung Injury and Gene Expression: mRNA Expressions of MnSOD, Interleukin-1β and iNOS

Nan-Hsiung Feng；Shi-Jye Chu；David Wang；Kang Hsu；Chun-Hsiu Lin；Hen-I Lin

lung injury ； lipopolysaccharide ； interleukin-1β ； manganese superoxide dismutase ； catalase ； inducible nitric oxide synthase ； oxygen radicals ； antioxidants ； vitamin C

The Chinese Journal of Physiology

47卷3期（2004 / 09 / 30）

111 - 120

英文

Antioxidants have been shown to be effective in attenuating acute lung injury. In this study, we determine the effects of various antioxidants by different mechanisms on the lipopolysaccharide (LPS)-induced changes. LPS was administered intravenously at a dose of 10 mg/kg to anesthetized rats. LPS induced a significant decrease in blood pressure (P＜0.01) and increased exhaled nitric oxide (NO) from 3.60±0.18 to 35.53±3.23 ppb (P＜0.01) during an observation period of 4 b. Plasma nitrate concentrations also increased from 0.61±0.06 to 1.54 ±0.22 μmol/l (P＜0.05). LPS-induced oxygen radical release from white blood cells isolated from rat peripheral blood also increased significantly (P＜0.001). After the experiment, the lung weight was obtained and lung tissues were taken for the determination of mRNA expression of inducible nitric oxide synthase (iNOS), tumor necrosis factor α (TNF-α), interleukin-1β(IL-1β) and manganese superoxide dismutase (MnSOD). Histological examination of the lungs was also performed. In the control group injected with saline solution, mRNA expressions of iNOS, IL-1β, TNF-α and MnSOD were absent. Four hours after LPS administration, mRNA expressions of iNOS, IL-1, and MnSOD were significantly enhanced, but TNF-αwas not discernibly expressed. LPS also caused a twofold increase in lung weight. Pathological examination revealed endothelial cell damage and interstitial edema. Various antioxidants were given 1h after LPS administration. These agents include SOD, catalase (CAT), SOD+CAT or vitamin C (ascorbic acid). These antioxidants effectively reversed the systemic hypotension, reduced the quantity of exhaled NO and plasma nitrate concentration, and prevented acute lung injury. Administration of various antioxidants also significantly attenuated LPS. induced oxygen radical release by rat white blood cells. EPS induced mRNA expressions of MnSOD and iNOS were significantly depressed by these antioxidants. However, only SOD+CAT and vitamin C inhibited the mRNA expression of IL-1β. These results suggest that oxygen radicals are responsible for LPS-induced lung injury. Antioxidants can attenuate the lung injury by inhibiting mRNA expressions of iNOS and IL-1β.

1. Bagby, G.J.,Plessala, K.J.,Wilson, L.A.,Thompson, J.J.,Nelson, S.(1991).Divergent efficacy of antibody to tumor necrosis factor-alpha in intravascular and peritonitis models of sepsis.J. Infect. Dis.,163,83-88.
2. Barnes, P.J.(1997).Nuclear factor-kB.Int. J. Biochem. Cell Biol.,29,867-870.
3. Blackwell, T.S.,Blackwell, T.R.,Holden, E.P.,Christman, B.W.,Christman, J.W.(1996).In viva antioxidant treatment suppresses nuclear factor-kB activation and neutrophilic lung inflammation.J Immunol.,157,1630-1637.
4. Cadenas, S.,Cadenas, A.M.(2002).Fighting the stranger-antioxidant protection against endotoxicity.Toxicology,180,45-63.
5. Cadenas, S.,Rojas, C.,Barja, G.(1998).Endotoxin increases oxidative injury to proteins in guinea pig liver: protection by dietary vitamin.C. Pharmacol. Toxicol.,82,11-18.
6. Chabot, E.,Mitchell, J.A.,Gutteridge, J.M.C.,Evans, T.W.(1998).Reactive oxygen species in acute lung injury.Eur. Respir. J.,11,745-757.
7. Chu, E.K.,Ribeiro, S.P.,Slutsky, A.S.(1997).Heat stress increase survival rates in lipopolysaccharide-stimulated rats.Crit. Care Med.,25,1727-1732.
8. Dinarello, C.A.,Wolff, S.M.(1993).The role of interleukin-1 in disease.N. Engl. J. Med.,328,106-113.
9. Dwenger, A.,Funck, M.,Leuken, B.,Schweitzer, G,Lehmann, U.(1992).Effect of ascorbic acid on neutrophil functions and hypoxanthine/xanthine oxidase-generated, oxygen-derived radicals.Eur. J. Clin. Chem. Clin. Biochem.,30,187-191.
10. Dwenger, A.,Pape, H.C.,Bantel, C.,Schweitzer, G.,Krumm, K.,Grotz, M.,Lueken, B.,Funck, M.,Regel, G.(1994).Ascorbic acid reduced the endotoxin-induced lung injury in awake sheep.Eur. J. Clin. Invest.,24,229-235.
11. Galey, H. F.,Davies, M.J.,Webster, N.R.(1996).Xanthine oxidase activity and free radical generation in patients with sepsis syndrome.Crit. Care Med.,24,1649-1653.
12. Godde, H.F.,Webster, N.R.(1993).Free radicals and antioxidants in sepsis.Crit. Care Med.,21,1770-1776.
13. Han, Y.J.,Kwon, Y.G.,Chung, H.T.,Lee, S.K.,Simmons, R.L.,Billiar, T.R.,Kim, Y.M.(2001).Antioxidant enzymes suppress nitric oxide production through the inhibition of NF-kB activation: role of H2O2 and nitric oxide in inducible nitric oxide synthase expression in macrophages.Nitric Oxide,5,504-513.
14. Hotchkiss, R.S.,Karl I. E.(2002).The pathophysiology and treatment of sepsis.N. Engl. J. Med.,348,138-150.
15. Hsu, K.,Wang, D.,Shen, C.Y.,Hsu, Y.S.,Shieh S.D.(1988).Protective effect of ketotifen on chemiluminescence of neutrophils.J. Formosan Med. Assoc.,87,939-943.
16. Koyama, S.,Kobayashi, T.,Kubo, K.,Sekiguchi, M.,Ueda, G.(1992).Recombinant human superoxide dismutase attenuates endotoxin-induced lung injury in awake sheep.Am. Rev. Respir. Dis.,145,1404-1409.
17. Levy, M.M.,Fink, M.P.,Marshall, J.C.,Abraham, E.,Angus, D.,Cook, D.,Cohen, J.,Opal, S.M.,Vincent, J.L.,Ramsay, G.(2003).SCCM/ESICM/ACCP/ATS/SIS. 2001 SCCM/ESICM/ACCP/ATS/SIS International sepsis definitions conference.Crit. Care Med.,31,1250-1256.
18. Liu, S.,Adcock, I.M.,Old, R.W.,Barnes P.J.,Evans, T.W.(1993).Lipopolysaccharide treatment in viva induces widespread tissue expression of inducible nitric oxide synthase mRNA.Biochem. Biophys. Res. Common.,196,1208-1213.
19. Liu, S.F.,Ye, X.,Malik, A.B.(1997).In viva inhibition of nuclear factor-KB activation prevents inducible nitric oxide synthase expression and systemic hypotension in a rat model of septic shock.J. Immunol.,159,3976-3983.
20. Lung, H.L.,Ip, W.K.,Wang, C.K.,Mak, N.K.,Chen, Z.Y.,Leung, K.N.(2002).Anti-proliferative and differentiation-inducing activities of the green tea catechin epigallocatechin-3-gallate (EGCG) on the human eosinophilic leukemia EoL-1 cell line.Life Sci.,72,257-268.
21. Macarthur, H.,Westfall, T.C.,Riley, D.P.,Misko, T.P.,Salvemini, D.(2000).Inactivation of catecholamines by superoxide gives new insights on the pathogenesis of septic shock.Proc. Natl. Acad. Sci. USA,97,9753-9758.
22. Macdonald, J.,Galley H.F.,Webster, N.R.(2003).Oxidative Stress and gene expression in sepsis.Br. J. Anaesth.,90,221-232.
23. Manna, S.K.,Zhang, H.J.,Yan, T.,Oberley, L.W.,Aggarwal, B.B.(1998).Overexpression of manganese superoxide dismutase Suppresses tumor necrosis factor-induced apoptosis and activation of nuclear transcription factor-kappaB and activated protein-1.J. Biol Chem.,273,13245-13254.
24. May, J.M.(2000).How does ascorbic acid prevent endothelial dysfunction?.Free Radic. Biol. Med.,28,1421-1429.
25. Metnitz, P.G.,Bartens, C.,Fischer, M.,Fridrich, P.,Steltzer, H.,Druml, W.(1999).Antioxidant status in patients with acute respiratory distress syndrome.Intensive Care Med.,25,180-185.
26. Meyer, M.,Schreck, R.,Baeuerle, P.A.(1993).H2O2 and antioxidants have opposite effects on activation of NF-kappa B and AP-1 in intact cells: AP-1 as secondary antioxidant-responsive factor.EMBO J.,12,2005-2015.
27. Milligan, S.A.,Heffel, J.M.,Goldstein, I.M.,Flick, M.R.(1988).Effect of catalase on endotoxin-induced acute lung injury in unanesthetized sheep.Am. Rev. Respir. Dis.,137,420-426.
28. Nonaka, A.,Manabe, T.,Tobe, T.(1990).Effect of a new synthetic free radical scavenger, 2-octadecyl ascorbic acid, on the mortality in mouse endotoxemia.Life Sci.,47,1933-1939.
29. O`Reilly, M.,Newcomb, D.E.,Remick, D.(1999).Endotoxin, sepsis, and the primrose path.Shock,12,411-420.
30. Okusawa, S.,Gelfand, J.A.,Ikejima, T.,Connally, R.J.,Dinarello, C.A.(1988).Interleukin-l induced a shock like state in rabbits: Synergism with tumor necrosis factor and the effect of cycloxygnease inhabitation.J. Clin. Invest.,81,1162-1172.
31. Padayatty, S.J.,Levine, M.(2001).New insights into the physiology and pharmacology of vitamin.C. C. A. M. J.,164,353-355.
32. Pugin, J.,Ricou, B.,Steinberg, K.P.,Suter, P.M.,Martin, T.R.(1996).Proinflammatory activity in bronchoalveolar lavage fluids from patients with ARDS, a prominent role for interleukin-l.Am. J. Respir. Crit. Care Med.,153,1850-1856.
33. Rojas, C.,Cadenas, S.,Herrero, A.,Mendez, J.,Barja, G.(1996).Endotoxin depletes ascorbate in the guinea pig heart. Protective effects of vitamins C and E against oxidative stress.Life Sci.,59,649-657.
34. Schreck, R.,Rieber, P.,Baeuerle, P.A.(1991).Reactive oxygen intermediates as apparently widely used messengers in the activation of NF-kappa B transcription factor and HIV-1.EMBO J.,10,2247-2258.
35. Zelko, I.N.,Mariani, T.J.,Folz, R. J.(2002).Superoxide dismutase multigene family: a comparison of the CuZn-SOD (SOD1), MnSOD (SOD2), and EC-SOD (SOD3) gene structures, evolution, and expression.Free Radic. Biol. Med.,33,337-349.
36. Zuckerman, S.H.,Shellhaas, J.,Butler, L.D.(1989).Differential regulation of lipopolysaccharide-induced interleukin 1 and tumor necrosis factor synthesis: effects of endogenous and exogenous glucocorticoids and the role of the pituitary-adrenal axis.Eur. J. Immunol.,19,301-305.

1. Yang, Victoria-Hui,Shen, Amy-Ru,Liou, Teh-Ling,Lee, Chia-Wei,Hwu, Chii-Min,Huang, Zih-Ling,Chien, Eileen Jea,Chien, Chau-Heng(2006).Inducible Nitric Oxide Synthase Expression and Plasma Bilirubin Changes in Rats under Intermittent Hypoxia Treatment.Chinese Journal of Physiology,49(5),275-280.