Skip to main content
PMC home page
The Journal of Experimental Medicine logoLink to The Journal of Experimental Medicine
. 1988 Mar 1;167(3):1154–1170. doi: 10.1084/jem.167.3.1154

Analysis of the nonfunctional respiratory burst in murine Kupffer cells

PMCID: PMC2188895  PMID: 3127523

Abstract

Murine Kupffer cells (KCs), which constitute one of the largest populations of tissue macrophages, differ from most other cells of the myelomonocytic lineage in lacking the capacity for a respiratory burst. A collagenase perfusion technique followed by adherence to plastic at low temperature yielded pure cultures of KCs uniformly expressing receptors for Fc and C3bi, and containing virtually no morphologically detectable intracytoplasmic debris. Such KCs took up and oxidized glucose via the hexose monophosphate shunt about the same as peritoneal macrophages (PCs). Respiratory burst stimuli failed to enhance the hexose monophosphate shunt in KCs, probably because no H2O2 was produced. Detergent-permeabilized KCs generated no O2- in the presence of 1 mM NADPH, in striking contrast to all PC populations studied. Yet, KCs contained at least one component of the O2(-)-producing oxidase, cytochrome b559, in the same quantities as PCs and neutrophils. Cytochrome b559 was demonstrated by a novel double-reduction spectral technique that eliminated interference from hemoglobin and mitochondrial cytochromes. Consistent with the presence of the oxidase, KCs acquired normal respiratory burst capacity after prolonged incubation in vitro. The defect in triggering the respiratory burst in KCs was selective for the reduction of O2 by NADPH, in that reduction of O2 by endogenous arachidonate was readily demonstrate in response to zymosan. The percent of arachidonate released, the percent oxygenated, and the suppression of prostacyclin and leukotriene C production, as well as the pattern of LFA-1 expression, all resembled the pattern reported with PCs several days after exposure to bacteria. Indeed, exposure of PCs to low numbers of zymosan particles led gradually to complete suppression of respiratory burst capacity and refractoriness to its enhancement by rIFN-gamma, as evident in KCs both before and after their explanation. Thus, the modulation of oxidative metabolism that characterizes KCs probably arises from frequent endocytic encounters. This phenomenon may permit macrophages to act as scavengers without oxidative damage to bystander cells.

Full Text

The Full Text of this article is available as a PDF (1.1 MB).

Selected References

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

  1. Aderem A. A., Scott W. A., Cohn Z. A. A selective defect in arachidonic acid release from macrophage membranes in high potassium media. J Cell Biol. 1984 Oct;99(4 Pt 1):1235–1241. doi: 10.1083/jcb.99.4.1235. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Assoian R. K., Fleurdelys B. E., Stevenson H. C., Miller P. J., Madtes D. K., Raines E. W., Ross R., Sporn M. B. Expression and secretion of type beta transforming growth factor by activated human macrophages. Proc Natl Acad Sci U S A. 1987 Sep;84(17):6020–6024. doi: 10.1073/pnas.84.17.6020. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Assoian R. K., Komoriya A., Meyers C. A., Miller D. M., Sporn M. B. Transforming growth factor-beta in human platelets. Identification of a major storage site, purification, and characterization. J Biol Chem. 1983 Jun 10;258(11):7155–7160. [PubMed] [Google Scholar]
  4. Berton G., Gordon S. Desensitization of macrophages to stimuli which induce secretion of superoxide anion. Down-regulation of receptors for phorbol myristate acetate. Eur J Immunol. 1983 Aug;13(8):620–627. doi: 10.1002/eji.1830130804. [DOI] [PubMed] [Google Scholar]
  5. Black C. M., Catterall J. R., Remington J. S. In vivo and in vitro activation of alveolar macrophages by recombinant interferon-gamma. J Immunol. 1987 Jan 15;138(2):491–495. [PubMed] [Google Scholar]
  6. Bonney R. J., Wightman P. D., Davies P., Sadowski S. J., Kuehl F. A., Jr, Humes J. L. Regulation of prostaglandin synthesis and of the selective release of lysosomal hydrolases by mouse peritoneal macrophages. Biochem J. 1978 Nov 15;176(2):433–442. doi: 10.1042/bj1760433. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cohn Z. A. Activation of mononuclear phagocytes: fact, fancy, and future. J Immunol. 1978 Sep;121(3):813–816. [PubMed] [Google Scholar]
  8. Crocker P. R., Gordon S. Isolation and characterization of resident stromal macrophages and hematopoietic cell clusters from mouse bone marrow. J Exp Med. 1985 Sep 1;162(3):993–1014. doi: 10.1084/jem.162.3.993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Cross A. R., Higson F. K., Jones O. T., Harper A. M., Segal A. W. The enzymic reduction and kinetics of oxidation of cytochrome b-245 of neutrophils. Biochem J. 1982 May 15;204(2):479–485. doi: 10.1042/bj2040479. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. De la Harpe J., Nathan C. F. A semi-automated micro-assay for H2O2 release by human blood monocytes and mouse peritoneal macrophages. J Immunol Methods. 1985 Apr 22;78(2):323–336. doi: 10.1016/0022-1759(85)90089-4. [DOI] [PubMed] [Google Scholar]
  11. Dinauer M. C., Orkin S. H., Brown R., Jesaitis A. J., Parkos C. A. The glycoprotein encoded by the X-linked chronic granulomatous disease locus is a component of the neutrophil cytochrome b complex. 1987 Jun 25-Jul 1Nature. 327(6124):717–720. doi: 10.1038/327717a0. [DOI] [PubMed] [Google Scholar]
  12. Ding A. H., Nathan C. F. Trace levels of bacterial lipopolysaccharide prevent interferon-gamma or tumor necrosis factor-alpha from enhancing mouse peritoneal macrophage respiratory burst capacity. J Immunol. 1987 Sep 15;139(6):1971–1977. [PubMed] [Google Scholar]
  13. Ding A., Wright S. D., Nathan C. Activation of mouse peritoneal macrophages by monoclonal antibodies to Mac-1 (complement receptor type 3). J Exp Med. 1987 Mar 1;165(3):733–749. doi: 10.1084/jem.165.3.733. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Esparza I., Green R., Schreiber R. D. Inhibition of macrophage tumoricidal activity by immune complexes and altered erythrocytes. J Immunol. 1983 Nov;131(5):2117–2121. [PubMed] [Google Scholar]
  15. Ezekowitz R. A., Sim R. B., MacPherson G. G., Gordon S. Interaction of human monocytes, macrophages, and polymorphonuclear leukocytes with zymosan in vitro. Role of type 3 complement receptors and macrophage-derived complement. J Clin Invest. 1985 Dec;76(6):2368–2376. doi: 10.1172/JCI112249. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Fels A. O., Nathan C. F., Cohn Z. A. Hydrogen peroxide release by alveolar macrophages from sarcoid patients and by alveolar macrophages from normals after exposure to recombinant interferons alpha A, beta, and gamma and 1,25-dihydroxyvitamin D3. J Clin Invest. 1987 Aug;80(2):381–386. doi: 10.1172/JCI113083. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Jaffe C. J., Vierling J. M., Jones E. A., Lawley T. J., Frank M. M. Receptor specific clearance by the reticuloendothelial system in chronic liver diseases. Demonstration of defective C3b-specific clearance in primary biliary cirrhosis. J Clin Invest. 1978 Nov;62(5):1069–1077. doi: 10.1172/JCI109212. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kakinuma K., Kaneda M., Chiba T., Ohnishi T. Electron spin resonance studies on a flavoprotein in neutrophil plasma membranes. Redox potentials of the flavin and its participation in NADPH oxidase. J Biol Chem. 1986 Jul 15;261(20):9426–9432. [PubMed] [Google Scholar]
  19. Karnovsky M. L., Lazdins J. K. Biochemical criteria for activated macrophages. J Immunol. 1978 Sep;121(3):809–813. [PubMed] [Google Scholar]
  20. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  21. Lepay D. A., Nathan C. F., Steinman R. M., Murray H. W., Cohn Z. A. Murine Kupffer cells. Mononuclear phagocytes deficient in the generation of reactive oxygen intermediates. J Exp Med. 1985 May 1;161(5):1079–1096. doi: 10.1084/jem.161.5.1079. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Lepay D. A., Steinman R. M., Nathan C. F., Murray H. W., Cohn Z. A. Liver macrophages in murine listeriosis. Cell-mediated immunity is correlated with an influx of macrophages capable of generating reactive oxygen intermediates. J Exp Med. 1985 Jun 1;161(6):1503–1512. doi: 10.1084/jem.161.6.1503. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Locksley R. M., Nelson C. S., Fankhauser J. E., Klebanoff S. J. Loss of granule myeloperoxidase during in vitro culture of human monocytes correlates with decay in antiprotozoa activity. Am J Trop Med Hyg. 1987 May;36(3):541–548. doi: 10.4269/ajtmh.1987.36.541. [DOI] [PubMed] [Google Scholar]
  24. Mathison J. C., Ulevitch R. J. The clearance, tissue distribution, and cellular localization of intravenously injected lipopolysaccharide in rabbits. J Immunol. 1979 Nov;123(5):2133–2143. [PubMed] [Google Scholar]
  25. Murray H. W. Pretreatment with phorbol myristate acetate inhibits macrophage activity against intracellular protozoa. J Reticuloendothel Soc. 1982 Jun;31(6):479–487. [PubMed] [Google Scholar]
  26. Murray H. W., Spitalny G. L., Nathan C. F. Activation of mouse peritoneal macrophages in vitro and in vivo by interferon-gamma. J Immunol. 1985 Mar;134(3):1619–1622. [PubMed] [Google Scholar]
  27. Murray H. W., Stern J. J., Welte K., Rubin B. Y., Carriero S. M., Nathan C. F. Experimental visceral leishmaniasis: production of interleukin 2 and interferon-gamma, tissue immune reaction, and response to treatment with interleukin 2 and interferon-gamma. J Immunol. 1987 Apr 1;138(7):2290–2297. [PubMed] [Google Scholar]
  28. Nakagawara A., Nathan C. F., Cohn Z. A. Hydrogen peroxide metabolism in human monocytes during differentiation in vitro. J Clin Invest. 1981 Nov;68(5):1243–1252. doi: 10.1172/JCI110370. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Nathan C. F., Karnovsky M. L., David J. R. Alterations of macrophage functions by mediators from lymphocytes. J Exp Med. 1971 Jun 1;133(6):1356–1376. doi: 10.1084/jem.133.6.1356. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Nathan C. F., Silverstein S. C., Brukner L. H., Cohn Z. A. Extracellular cytolysis by activated macrophages and granulocytes. II. Hydrogen peroxide as a mediator of cytotoxicity. J Exp Med. 1979 Jan 1;149(1):100–113. doi: 10.1084/jem.149.1.100. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Nathan C. F., Tsunawaki S. Secretion of toxic oxygen products by macrophages: regulatory cytokines and their effects on the oxidase. Ciba Found Symp. 1986;118:211–230. doi: 10.1002/9780470720998.ch14. [DOI] [PubMed] [Google Scholar]
  32. North R. J. The concept of the activated macrophage. J Immunol. 1978 Sep;121(3):806–809. [PMC free article] [PubMed] [Google Scholar]
  33. Pasquier C., Marquetty C., Chollet-Martin S., Hakim J. Spectroscopic interference of hemoglobin with neutrophil cytochrome b-245 and its elimination by carbon monoxide. J Immunol Methods. 1985 Feb 28;77(1):147–153. doi: 10.1016/0022-1759(85)90192-9. [DOI] [PubMed] [Google Scholar]
  34. Reed P. W. Glutathione and the hexose monophosphate shunt in phagocytizing and hydrogen peroxide-treated rat leukocytes. J Biol Chem. 1969 May 10;244(9):2459–2464. [PubMed] [Google Scholar]
  35. Scott W. A., Zrike J. M., Hamill A. L., Kempe J., Cohn Z. A. Regulation of arachidonic acid metabolites in macrophages. J Exp Med. 1980 Aug 1;152(2):324–335. doi: 10.1084/jem.152.2.324. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Segal A. W., Cross A. R., Garcia R. C., Borregaard N., Valerius N. H., Soothill J. F., Jones O. T. Absence of cytochrome b-245 in chronic granulomatous disease. A multicenter European evaluation of its incidence and relevance. N Engl J Med. 1983 Feb 3;308(5):245–251. doi: 10.1056/NEJM198302033080503. [DOI] [PubMed] [Google Scholar]
  37. Strassmann G., Springer T. A., Haskill S. J., Miraglia C. C., Lanier L. L., Adams D. O. Antigens associated with the activation of murine mononuclear phagocytes in vivo: differential expression of lymphocyte function-associated antigen in the several stages of development. Cell Immunol. 1985 Aug;94(1):265–275. doi: 10.1016/0008-8749(85)90103-0. [DOI] [PubMed] [Google Scholar]
  38. Sung S. S., Nelson R. S., Silverstein S. C. Yeast mannans inhibit binding and phagocytosis of zymosan by mouse peritoneal macrophages. J Cell Biol. 1983 Jan;96(1):160–166. doi: 10.1083/jcb.96.1.160. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Tsunawaki S., Nathan C. F. Enzymatic basis of macrophage activation. Kinetic analysis of superoxide production in lysates of resident and activated mouse peritoneal macrophages and granulocytes. J Biol Chem. 1984 Apr 10;259(7):4305–4312. [PubMed] [Google Scholar]
  40. Tsunawaki S., Nathan C. F. Macrophage deactivation. Altered kinetic properties of superoxide-producing enzyme after exposure to tumor cell-conditioned medium. J Exp Med. 1986 Oct 1;164(4):1319–1331. doi: 10.1084/jem.164.4.1319. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Tsunawaki S., Nathan C. F. Release of arachidonate and reduction of oxygen. Independent metabolic bursts of the mouse peritoneal macrophage. J Biol Chem. 1986 Sep 5;261(25):11563–11570. [PubMed] [Google Scholar]
  42. Unger W. G., Stamford I. F., Bennett A. Extraction of prostaglandins from human blood. Nature. 1971 Oct 1;233(5318):336–337. doi: 10.1038/233336b0. [DOI] [PubMed] [Google Scholar]
  43. Wright S. D., Silverstein S. C. Phagocytosing macrophages exclude proteins from the zones of contact with opsonized targets. Nature. 1984 May 24;309(5966):359–361. doi: 10.1038/309359a0. [DOI] [PubMed] [Google Scholar]
  44. van Furth R. Macrophage activity and clinical immunology. Origin and kinetics of mononuclear phagocytes. Ann N Y Acad Sci. 1976;278:161–175. doi: 10.1111/j.1749-6632.1976.tb47027.x. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Experimental Medicine are provided here courtesy of The Rockefeller University Press

RESOURCES