, 31:344 | Cite as

Glutamine Reduces TNF-α by Enhancing Glutathione Synthesis in Lipopolysaccharide-Stimulated Alveolar Epithelial Cells of Rats

  • Feng Zhang
  • Xinying Wang
  • Weiya Wang
  • Ning Li
  • Jieshou Li


To investigate the role of glutathione (GSH) synthesis in the regulation on nuclear factor (NF)-κB activity and tumor necrosis factor-alpha (TNF-α) release by glutamine (GLN) in lipopolysaccharide (LPS)-stimulated alveolar type II (AT-II) epithelial cells of rat lungs. Primary cultured AT-II cells were pre-treated with various doses of GLN for 2, 8, 16, 24 h. At the 8 h time point before LPS stimulation, various doses of l-buthionine-(S,R)-sulfoximine (BSO), an inhibitor of GSH synthesis, were added with 10 mM GLN. Then the cells were stimulated with 1 μg/ml LPS for 24 h. The cells were obtained for GSH measurement. TNF-α level in the supernatant was determined by enzyme-linked immunosorbent assay. NF-κB activity was assessed by electrophoretic mobility shift assay. Eight hours before LPS exposure was the best time point for GLN’s enhancing GSH synthesis. LPS could significantly decrease the GSH level, increase NF-κB activation and TNF-α release in AT-II cells. Supplementation of GLN could increase the GSH level and attenuate the release of TNF-α in LPS-stimulated AT-II cells in a dose-dependant manner. And NF-κB activation also could be prevented by GLN. BSO could block the effect of GLN. As a precursor of GSH, glutamine could prevent the NF-κB activation and attenuate the release of TNF-α in LPS-stimulated AT-II cells and the effect may be mediated via GSH synthesis.


glutamine glutathione NF-κB tumor necrosis factor lipopolysaccharide 



This work was supported by the National Natural Science Foundation (30500404) and the Jiangsu Provincial Natural Science Foundation (BK2006530).


  1. 1.
    Rahman, I., S. K. Biswas, L. A. Jimenez, M. Torres, and H. J. Forman. 2005. Glutathione, stress responses, and redox signaling in Lung Inflammation. Antioxid. Redox Signal 7:42–59 doi: 10.1089/ars.2005.7.42.PubMedCrossRefGoogle Scholar
  2. 2.
    Rahman, I. 2005. Redox signaling in the lungs. Antioxid. Redox Signal 7:1–5 doi: 10.1089/ars.2005.7.1.PubMedCrossRefGoogle Scholar
  3. 3.
    Cantin, A. M., S. L. North, R. C. Hubbard, and R. G. Crystal. 1987. Normal alveolar epithelial lining fluid contains high levels of glutathione. Am. J. Physiol. 63:152–157.Google Scholar
  4. 4.
    Rahman, I., B. Mulier, P. S. Gilmour, T. Watchorn, K. Donaldson, P. K. Jeffery et al. 2001. Oxidant-mediated lung epithelial cell tolerance: the role of intracellular glutathione and nuclear factor-kappaB. Biochem. Pharmacol. 62:787–794 doi: 10.1016/S0006-2952(01)00702-X.PubMedCrossRefGoogle Scholar
  5. 5.
    Coeffier, M., O. Miralles-Barrachina, F. Le Pessot, O. Lalaude, M. Daveau, A. Lavoinne et al. 2001. Influence of glutamine on cytokine production by human gut in vitro. Cytokine 13:148–154 doi: 10.1006/cyto.2000.0813.PubMedCrossRefGoogle Scholar
  6. 6.
    Wischmeyer, P. E., J. Riehm, K. D. Singleton, H. Ren, M. W. Musch, M. Kahana et al. 2003. Glutamine attenuates tumor necrosis factor-α release and enhances heat shock protein 72 in human peripheral blood mononuclear cells. Nutrition 19:1–6 doi: 10.1016/S0899-9007(02)00839-0.PubMedCrossRefGoogle Scholar
  7. 7.
    Singleton, K. D., V. E. Beckey, and P. E. Wischmeyer. 2005. Glutamine prevents activation of NF-κB and stress kinase pathways, attenuates inflammatory cytokine release, and prevents acute respiratory distress syndrome (ARDS). Shock 24:583–589 doi: 10.1097/01.shk.0000185795.96964.71.PubMedCrossRefGoogle Scholar
  8. 8.
    Cao, Y. H., Z. L. Feng, A. Hoos, and V. S. Klimberg. 1998. Glutamine enhances gut glutathione production. J. Parenter. Enter. Nutr. 22:224–227.CrossRefGoogle Scholar
  9. 9.
    Moon, M., T. Pritts, A. Salzman, J. Fischer, and P. O. Hasselgren. 1998. Glutamine prevents LPS-induced NF-κB activation in human intestinal epithelial cells. Gastroenterology 114:A1410–A1411 doi: 10.1016/S0016-5085(98)85735-1.CrossRefGoogle Scholar
  10. 10.
    Babu, R., S. Eaton, D. P. Drake, L. Spitz, and A. Pierro. 2001. Glutamine and glutathione counteract the inhibitory effects of mediators of sepsis in neonatal hepatocytes. J. Pediatr. Surg. 36:282–286 doi: 10.1053/jpsu.2001.20690.PubMedCrossRefGoogle Scholar
  11. 11.
    Dobbs, L. G., R. Gonzalez, and M. C. Williams. 1986. An improved method for isolating type II cells in high yield and purity. Am. Rev. Respir. Dis. 134:141–145.PubMedGoogle Scholar
  12. 12.
    Schreiber, E., P. Matthias, M. M. Müller, and W. Schaffner. 1989. Rapid detection of nuclear binding proteins with mini-extracts prepared from a small number of cell. Nucleic Acids Res. 17:6419 doi: 10.1093/nar/17.15.6419.PubMedCrossRefGoogle Scholar
  13. 13.
    Haddad, J. J. 2001. L-Buthionine-(S, R)-sulfoximine, an irreversible inhibitor of γ-glutamylcysteine sythetase, augments LPS-mediated pro-inflammatory cytokine biosynthesis: evidence for the implication of an IκB-α/NF-κB insensitive pathway. Eur. Cytokine Netw. 12:614–624.PubMedGoogle Scholar
  14. 14.
    Lesur, O., K. Arsalane, and D. Lane. 1996. Lung alveolar epithelial cell migration in vitro: modulators and regulation processes. Am. J. Physiol. 270:L311–L319.PubMedGoogle Scholar
  15. 15.
    Warshamana, G. S., M. Corti, and A. R. Brody. 2001. TNF-alpha, PDGF, and TGF-beta(1) expression by primary mouse bronchiolar-alveolar epithelial and mesenchymal cells: tnf-alpha induces TGF-beta(1). Exp. Mol. Pathol. 71:13–33 doi: 10.1006/exmp.2001.2376.PubMedCrossRefGoogle Scholar
  16. 16.
    Rahman, I. 2002. Oxidative stress, transcription factors and chromatin remodelling in lung inflammation. Biochem. Pharmacol. 64:935–942 doi: 10.1016/S0006-2952(02)01153-X.PubMedCrossRefGoogle Scholar
  17. 17.
    Hammarqvist, F., J. Luo, I. A. Cotgreave, K. Anderson, and J. Wernerman. 1997. Skeletal muscle glutathione is depleted in critically ill patients. Crit. Care Med. 25:78–84 doi: 10.1097/00003246-199701000-00016.PubMedCrossRefGoogle Scholar
  18. 18.
    Manhart, N., K. Vierlinger, A. Spittler, H. Bergmeister, T. Sautner, and E. Roth. 2000. Oral feeding with glutamine prevents lymphocyte and glutathione depletion of peyer’s patches in endotoxemic mice. Ann. Surg. 234:92–97 doi: 10.1097/00000658-200107000-00014.CrossRefGoogle Scholar
  19. 19.
    Bunnel, E., and E. R. Pacht. 1993. Oxidised glutathione is increased in the alveolar fluid of patients with the adult respiratory distress syndrome. Am. Rev. Respir. Dis. 148:1174–1178.Google Scholar
  20. 20.
    Luthen, R., J. H. Grendel, C. Niederau, and D. Haussinger. 1998. Trypsinogen activation and glutathione content are linked to pancreatic injury in models of bilary acute pancreatitis. Int. J. Pancreatol. 24:193–202.PubMedGoogle Scholar
  21. 21.
    Biswas, S. K., D. Mcclure, L. A. Jimenez, I. L. Megson, and I. Rahman. 2005. Curcumin induces glutathione biosynthesis and inhibits NF-κB activation and interleukin-8 release in alveolar epithelial cells: mechanism of free radicals scavenging activity. Antioxid. Redox Signal 7:32–41 doi: 10.1089/ars.2005.7.32.PubMedCrossRefGoogle Scholar
  22. 22.
    Haddad, J. J., and S. C. Land. 2002. Redox signaling-mediated regulation of lipopolysaccharide-induced proinflammatory cytokine biosynthesis in alveolar epithelial cells. Antioxid. Redox Signal 4:179–193 doi: 10.1089/152308602753625942.PubMedCrossRefGoogle Scholar
  23. 23.
    Rouse, K., E. Nwokedi, J. E. Woodliff, J. Epstein, and V. S. Klimberg. 1995. Glutamine enhances selectivity of chemotherapy through changes in glutathione metabolism. Ann. Surg. 221:420–426 doi: 10.1097/00000658-199504000-00014.PubMedCrossRefGoogle Scholar
  24. 24.
    Goeters, C., A. Wenn, N. Mertes, C. Wempe, H. Van Aken, P. Stehle et al. 2004. Parenteral L-alanyl-L-glutamine improves 6-month outcome in critically ill patients. Crit. Care Med. 30:2032–2037 doi: 10.1097/00003246-200209000-00013.CrossRefGoogle Scholar
  25. 25.
    Lai, Y. N., S. L. Yeh, M. T. Lin, H. F. Shang, C. L. Yeh, and W. J. Chen. 2004. Glutamine supplementation enhances mucosal immunity in rats with gut-derived sepsis. Nutrition 20:286–291 doi: 10.1016/j.nut.2003.11.015.PubMedCrossRefGoogle Scholar
  26. 26.
    Chang, W. K., K. D. Yang, H. Chuang, J. T. Jan, and M. F. Shaio. 2002. Glutamine protects activated human T cells from apoptosis by up-regulating glutathione and Bcl-2 levels. Clin. Immunol. 104:151–160 doi: 10.1006/clim.2002.5257.PubMedCrossRefGoogle Scholar
  27. 27.
    Glauser, M. P., G. Zanett, J. D. Baumgartner, and J. Cohen. 1991. Septic shock: pathogenesis. Lancet 338:732–736 doi: 10.1016/0140-6736(91)91452-Z.PubMedCrossRefGoogle Scholar
  28. 28.
    Pena, L. R., D. B. Hill, and C. J. McClain. 1999. Treatment with glutathione precursor decreases cytokine activity. J. Parenter. Enter. Nutr. 23:1–6 doi: 10.1177/014860719902300101.CrossRefGoogle Scholar
  29. 29.
    Victor, V. M., M. Rocha, and M. D. L. Fuente. 2003. N-acetylcysteine protects mice from lethal endotoxemia by regulating the redox state of immune cells. Free Radic. Res. 37:919–929 doi: 10.1080/1071576031000148727.PubMedCrossRefGoogle Scholar
  30. 30.
    Wischmeyer, P. E., M. Kahana, R. Wolfson, H. Ren, M. M. Musch, and E. B. Chang. 2001. Glutamine reduces cytokine release, organ damage, and mortality in a rat model of endotoxemia. Shock 16:398–402.PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2008

Authors and Affiliations

  • Feng Zhang
    • 1
  • Xinying Wang
    • 1
  • Weiya Wang
    • 1
  • Ning Li
    • 1
  • Jieshou Li
    • 1
  1. 1.Research Institute of General Surgery, Department of General Surgery, Jinling Hospital, Medical SchoolNanjing UniversityNanjingPeople’s Republic of China

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