Skip to main content
PMC home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1994 Mar;93(3):1042–1050. doi: 10.1172/JCI117053

Leukotriene A4 hydrolase in human bronchoalveolar lavage fluid.

D A Munafo 1, K Shindo 1, J R Baker 1, T D Bigby 1
PMCID: PMC294032  PMID: 8132742

Abstract

We examined cell-free human bronchoalveolar lavage fluid (BALF) for enzymes of the 5-lipoxygenase pathway. BALF was obtained from six patients who were active smokers and six nonsmokers. Enzymatic activity in cell-free BALF was assessed by specific assays for leukotriene (LT) A4 hydrolase, 5-lipoxygenase, and LTC4 synthase using HPLC. Only LTA4 hydrolase enzymatic activity was found. This activity ranged from 101 to 667 when expressed as picomoles of LTB4 produced per milliliter BALF. Enzymatic activity in smokers vs nonsmokers was 484 +/- 120 vs 129 +/- 32 pmol LTB4/ml BALF (mean +/- SD, P < 0.0001). There were no leukotrienes found in BALF before assay. Immunoblot analysis revealed an immunoreactive band at a relative molecular mass of 69,000 D in all samples, consistent with LTA4 hydrolase, but no evidence of 5-lipoxygenase. BALF had greater LTA4 hydrolase activity per milligram of protein than neutrophil cytosol, epithelial cell cytosol, plasma, or serum. The synthesis of LTB4 was significantly increased when neutrophils were stimulated in BALF. These data indicate the selective presence of LTA4 hydrolase in BALF which is significantly increased in smokers. This enzyme in BALF may contribute to the inflammatory response in tobacco-related lung disease.

Full text

PDF

Images in this article

1046Image
on p.1046

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Antonelli M., Bufi M., De Blasi R. A., Crimi G., Conti G., Mattia C., Vivino G., Lenti L., Lombardi D., Dotta A. Detection of leukotrienes B4, C4 and of their isomers in arterial, mixed venous blood and bronchoalveolar lavage fluid from ARDS patients. Intensive Care Med. 1989;15(5):296–301. doi: 10.1007/BF00263864. [DOI] [PubMed] [Google Scholar]
  2. Bigby T. D., Lee D. M., Meslier N., Gruenert D. C. Leukotriene A4 hydrolase activity of human airway epithelial cells. Biochem Biophys Res Commun. 1989 Oct 16;164(1):1–7. doi: 10.1016/0006-291x(89)91674-4. [DOI] [PubMed] [Google Scholar]
  3. Bigby T. D., Meslier N. Transcellular lipoxygenase metabolism between monocytes and platelets. J Immunol. 1989 Sep 15;143(6):1948–1954. [PubMed] [Google Scholar]
  4. Borgeat P., Samuelsson B. Metabolism of arachidonic acid in polymorphonuclear leukocytes. Structural analysis of novel hydroxylated compounds. J Biol Chem. 1979 Aug 25;254(16):7865–7869. [PubMed] [Google Scholar]
  5. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  6. Böyum A. Isolation of mononuclear cells and granulocytes from human blood. Isolation of monuclear cells by one centrifugation, and of granulocytes by combining centrifugation and sedimentation at 1 g. Scand J Clin Lab Invest Suppl. 1968;97:77–89. [PubMed] [Google Scholar]
  7. Drazen J. M., Austen K. F. Leukotrienes and airway responses. Am Rev Respir Dis. 1987 Oct;136(4):985–998. doi: 10.1164/ajrccm/136.4.985. [DOI] [PubMed] [Google Scholar]
  8. Feinmark S. J., Cannon P. J. Endothelial cell leukotriene C4 synthesis results from intercellular transfer of leukotriene A4 synthesized by polymorphonuclear leukocytes. J Biol Chem. 1986 Dec 15;261(35):16466–16472. [PubMed] [Google Scholar]
  9. Fiore S., Serhan C. N. Phospholipid bilayers enhance the stability of leukotriene A4 and epoxytetraenes: stabilization of eicosanoids by liposomes. Biochem Biophys Res Commun. 1989 Mar 15;159(2):477–481. doi: 10.1016/0006-291x(89)90017-x. [DOI] [PubMed] [Google Scholar]
  10. Fitzpatrick F., Haeggström J., Granström E., Samuelsson B. Metabolism of leukotriene A4 by an enzyme in blood plasma: a possible leukotactic mechanism. Proc Natl Acad Sci U S A. 1983 Sep;80(17):5425–5429. doi: 10.1073/pnas.80.17.5425. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Fitzpatrick F., Liggett W., McGee J., Bunting S., Morton D., Samuelsson B. Metabolism of leukotriene A4 by human erythrocytes. A novel cellular source of leukotriene B4. J Biol Chem. 1984 Sep 25;259(18):11403–11407. [PubMed] [Google Scholar]
  12. Ford-Hutchinson A. W. Leukotriene B4 in inflammation. Crit Rev Immunol. 1990;10(1):1–12. [PubMed] [Google Scholar]
  13. Fu J. Y., Haeggström J., Collins P., Meijer J., Rådmark O. Leukotriene A4 hydrolase: analysis of some human tissues by radioimmunoassay. Biochim Biophys Acta. 1989 Nov 6;1006(1):121–126. doi: 10.1016/0005-2760(89)90332-9. [DOI] [PubMed] [Google Scholar]
  14. Funk C. D., FitzGerald G. A. Eicosanoid forming enzyme mRNA in human tissues. Analysis by quantitative polymerase chain reaction. J Biol Chem. 1991 Jul 5;266(19):12508–12513. [PubMed] [Google Scholar]
  15. Haeggström J. Z., Wetterholm A., Shapiro R., Vallee B. L., Samuelsson B. Leukotriene A4 hydrolase: a zinc metalloenzyme. Biochem Biophys Res Commun. 1990 Nov 15;172(3):965–970. doi: 10.1016/0006-291x(90)91540-9. [DOI] [PubMed] [Google Scholar]
  16. Haeggström J. Z., Wetterholm A., Vallee B. L., Samuelsson B. Leukotriene A4 hydrolase: an epoxide hydrolase with peptidase activity. Biochem Biophys Res Commun. 1990 Nov 30;173(1):431–437. doi: 10.1016/s0006-291x(05)81076-9. [DOI] [PubMed] [Google Scholar]
  17. Haeggström J., Rådmark O., Fitzpatrick F. A. Leukotriene A4-hydrolase activity in guinea pig and human liver. Biochim Biophys Acta. 1985 Jul 9;835(2):378–384. doi: 10.1016/0005-2760(85)90294-2. [DOI] [PubMed] [Google Scholar]
  18. Hastings R. H., Grady M., Sakuma T., Matthay M. A. Clearance of different-sized proteins from the alveolar space in humans and rabbits. J Appl Physiol (1985) 1992 Oct;73(4):1310–1316. doi: 10.1152/jappl.1992.73.4.1310. [DOI] [PubMed] [Google Scholar]
  19. Hsueh W. A., Carlson E. J., Luetscher J. A., Grislis G. Activation and characterization of inactive big renin in plasma of patients with diabetic nephropathy and unusual active renin. J Clin Endocrinol Metab. 1980 Sep;51(3):535–543. doi: 10.1210/jcem-51-3-535. [DOI] [PubMed] [Google Scholar]
  20. Hunter J. A., Finkbeiner W. E., Nadel J. A., Goetzl E. J., Holtzman M. J. Predominant generation of 15-lipoxygenase metabolites of arachidonic acid by epithelial cells from human trachea. Proc Natl Acad Sci U S A. 1985 Jul;82(14):4633–4637. doi: 10.1073/pnas.82.14.4633. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Jakschik B. A., Kuo C. G. Characterization of leukotriene A4 and B4 biosynthesis. Prostaglandins. 1983 Jun;25(6):767–782. doi: 10.1016/0090-6980(83)90002-3. [DOI] [PubMed] [Google Scholar]
  22. Kargman S., Rouzer C. A. Studies on the regulation, biosynthesis, and activation of 5-lipoxygenase in differentiated HL60 cells. J Biol Chem. 1989 Aug 5;264(22):13313–13320. [PubMed] [Google Scholar]
  23. Lam S., Chan H., LeRiche J. C., Chan-Yeung M., Salari H. Release of leukotrienes in patients with bronchial asthma. J Allergy Clin Immunol. 1988 Apr;81(4):711–717. doi: 10.1016/0091-6749(88)91043-3. [DOI] [PubMed] [Google Scholar]
  24. Lee T. H., Crea A. E., Gant V., Spur B. W., Marron B. E., Nicolaou K. C., Reardon E., Brezinski M., Serhan C. N. Identification of lipoxin A4 and its relationship to the sulfidopeptide leukotrienes C4, D4, and E4 in the bronchoalveolar lavage fluids obtained from patients with selected pulmonary diseases. Am Rev Respir Dis. 1990 Jun;141(6):1453–1458. doi: 10.1164/ajrccm/141.6.1453. [DOI] [PubMed] [Google Scholar]
  25. Maclouf J. A., Murphy R. C. Transcellular metabolism of neutrophil-derived leukotriene A4 by human platelets. A potential cellular source of leukotriene C4. J Biol Chem. 1988 Jan 5;263(1):174–181. [PubMed] [Google Scholar]
  26. Malfroy B., Kado-Fong H., Gros C., Giros B., Schwartz J. C., Hellmiss R. Molecular cloning and amino acid sequence of rat kidney aminopeptidase M: a member of a super family of zinc-metallohydrolases. Biochem Biophys Res Commun. 1989 May 30;161(1):236–241. doi: 10.1016/0006-291x(89)91586-6. [DOI] [PubMed] [Google Scholar]
  27. Maycock A. L., Anderson M. S., DeSousa D. M., Kuehl F. A., Jr Leukotriene A4: preparation and enzymatic conversion in a cell-free system to leukotriene B4. J Biol Chem. 1982 Dec 10;257(23):13911–13914. [PubMed] [Google Scholar]
  28. McGee J., Fitzpatrick F. Enzymatic hydration of leukotriene A4. Purification and characterization of a novel epoxide hydrolase from human erythrocytes. J Biol Chem. 1985 Oct 15;260(23):12832–12837. [PubMed] [Google Scholar]
  29. Meslier N., Aldrich A. J., Bigby T. D. Effect of interferon-gamma on the 5-lipoxygenase pathway of rat lung macrophages. Am J Respir Cell Mol Biol. 1992 Jan;6(1):93–99. doi: 10.1165/ajrcmb/6.1.93. [DOI] [PubMed] [Google Scholar]
  30. Murray J. J., Tonnel A. B., Brash A. R., Roberts L. J., 2nd, Gosset P., Workman R., Capron A., Oates J. A. Release of prostaglandin D2 into human airways during acute antigen challenge. N Engl J Med. 1986 Sep 25;315(13):800–804. doi: 10.1056/NEJM198609253151304. [DOI] [PubMed] [Google Scholar]
  31. Odlander B., Claesson H. E., Bergman T., Rådmark O., Jörnvall H., Haeggström J. Z. Leukotriene A4 hydrolase in the human B-lymphocytic cell line Raji: indications of catalytically divergent forms of the enzyme. Arch Biochem Biophys. 1991 May 15;287(1):167–174. doi: 10.1016/0003-9861(91)90402-5. [DOI] [PubMed] [Google Scholar]
  32. Ohishi N., Izumi T., Minami M., Kitamura S., Seyama Y., Ohkawa S., Terao S., Yotsumoto H., Takaku F., Shimizu T. Leukotriene A4 hydrolase in the human lung. Inactivation of the enzyme with leukotriene A4 isomers. J Biol Chem. 1987 Jul 25;262(21):10200–10205. [PubMed] [Google Scholar]
  33. Rouzer C. A., Kargman S. Translocation of 5-lipoxygenase to the membrane in human leukocytes challenged with ionophore A23187. J Biol Chem. 1988 Aug 5;263(22):10980–10988. [PubMed] [Google Scholar]
  34. Samuelsson B. Leukotrienes: mediators of immediate hypersensitivity reactions and inflammation. Science. 1983 May 6;220(4597):568–575. doi: 10.1126/science.6301011. [DOI] [PubMed] [Google Scholar]
  35. Seeger W., Grimminger F., Barden M., Becker G., Lohmeyer J., Heinrich D., Lasch H. G. Omega-oxidized leukotriene B4 detected in the broncho-alveolar lavage fluid of patients with non-cardiogenic pulmonary edema, but not in those with cardiogenic edema. Intensive Care Med. 1991;17(1):1–6. doi: 10.1007/BF01708400. [DOI] [PubMed] [Google Scholar]
  36. Serhan C. N., Radin A., Smolen J. E., Korchak H., Samuelsson B., Weissmann G. Leukotriene B4 is a complete secretagogue in human neutrophils: a kinetic analysis. Biochem Biophys Res Commun. 1982 Aug;107(3):1006–1012. doi: 10.1016/0006-291x(82)90622-2. [DOI] [PubMed] [Google Scholar]
  37. Stephenson A. H., Lonigro A. J., Hyers T. M., Webster R. O., Fowler A. A. Increased concentrations of leukotrienes in bronchoalveolar lavage fluid of patients with ARDS or at risk for ARDS. Am Rev Respir Dis. 1988 Sep;138(3):714–719. doi: 10.1164/ajrccm/138.3.714. [DOI] [PubMed] [Google Scholar]
  38. Weller P. F., Lee C. W., Foster D. W., Corey E. J., Austen K. F., Lewis R. A. Generation and metabolism of 5-lipoxygenase pathway leukotrienes by human eosinophils: predominant production of leukotriene C4. Proc Natl Acad Sci U S A. 1983 Dec;80(24):7626–7630. doi: 10.1073/pnas.80.24.7626. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Wenzel S. E., Larsen G. L., Johnston K., Voelkel N. F., Westcott J. Y. Elevated levels of leukotriene C4 in bronchoalveolar lavage fluid from atopic asthmatics after endobronchial allergen challenge. Am Rev Respir Dis. 1990 Jul;142(1):112–119. doi: 10.1164/ajrccm/142.1.112. [DOI] [PubMed] [Google Scholar]
  40. Yoshimoto T., Soberman R. J., Lewis R. A., Austen K. F. Isolation and characterization of leukotriene C4 synthetase of rat basophilic leukemia cells. Proc Natl Acad Sci U S A. 1985 Dec;82(24):8399–8403. doi: 10.1073/pnas.82.24.8399. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation

RESOURCES