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. 1997 Nov;65(11):4747–4753. doi: 10.1128/iai.65.11.4747-4753.1997

Azurophilic granules of human neutrophils contain CD14.

D A Rodeberg 1, R E Morris 1, G F Babcock 1
PMCID: PMC175681  PMID: 9353060

Abstract

CD14, the leukocyte receptor for lipopolysaccharide (LPS), is important in the response of human polymorphonuclear leukocytes (PMNs) to infection with gram-negative bacteria. The level of CD14 on the PMN surface increases after exposure to some inflammatory stimuli such as N-formyl-methionyl-leucyl-phenylalanine (fMLP). These newly expressed CD14 molecules probably come from an intracellular pool of preformed receptors. We sought to further characterize PMN CD14 expression, upregulation, and shedding and to define the intracellular location of CD14 molecules. Our results demonstrate that both LPS and fMLP significantly increased CD14 cell surface expression; however, neither phorbol myristate acetate (PMA) or A23187 increased receptor levels on the PMN surface. Neither fMLP, PMA, or A23187 stimulated the release of soluble CD14 from PMNs. Intracellular CD14 was observed in >90% of PMNs examined by flow cytometry and confocal microscopy. Additional analyses using CD14 enzyme-linked immunosorbent assays and electron microscopy studies, examining PMN granules separated by discontinuous sucrose or Percoll gradients, showed that CD14 was present in both the plasma membrane-secretory vesicle fractions and azurophilic granules.

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Selected References

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  1. Allen R. C., Pruitt B. A., Jr Humoral-phagocyte axis of immune defense in burn patients. Chemoluminigenic probing. Arch Surg. 1982 Feb;117(2):133–140. doi: 10.1001/archsurg.1982.01380260019004. [DOI] [PubMed] [Google Scholar]
  2. Babcock G. F., Taylor A. F., Hynd B. A., Sramkoski R. M., Alexander J. W. Flow cytometric analysis of lymphocyte subset phenotypes comparing normal children and adults. Diagn Clin Immunol. 1987;5(4):175–179. [PubMed] [Google Scholar]
  3. Ball E. D., Graziano R. F., Shen L., Fanger M. W. Monoclonal antibodies to novel myeloid antigens reveal human neutrophil heterogeneity. Proc Natl Acad Sci U S A. 1982 Sep;79(17):5374–5378. doi: 10.1073/pnas.79.17.5374. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bazil V., Strominger J. L. Shedding as a mechanism of down-modulation of CD14 on stimulated human monocytes. J Immunol. 1991 Sep 1;147(5):1567–1574. [PubMed] [Google Scholar]
  5. Borregaard N., Heiple J. M., Simons E. R., Clark R. A. Subcellular localization of the b-cytochrome component of the human neutrophil microbicidal oxidase: translocation during activation. J Cell Biol. 1983 Jul;97(1):52–61. doi: 10.1083/jcb.97.1.52. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Detmers P. A., Zhou D., Powell D., Lichenstein H., Kelley M., Pironkova R. Endotoxin receptors (CD14) are found with CD16 (Fc gamma RIII) in an intracellular compartment of neutrophils that contains alkaline phosphatase. J Immunol. 1995 Aug 15;155(4):2085–2095. [PubMed] [Google Scholar]
  7. Ferrero E., Hsieh C. L., Francke U., Goyert S. M. CD14 is a member of the family of leucine-rich proteins and is encoded by a gene syntenic with multiple receptor genes. J Immunol. 1990 Jul 1;145(1):331–336. [PubMed] [Google Scholar]
  8. Frey E. A., Miller D. S., Jahr T. G., Sundan A., Bazil V., Espevik T., Finlay B. B., Wright S. D. Soluble CD14 participates in the response of cells to lipopolysaccharide. J Exp Med. 1992 Dec 1;176(6):1665–1671. doi: 10.1084/jem.176.6.1665. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Freyer D. R., Morganroth M. L., Rogers C. E., Arnaout M. A., Todd R. F., 3rd Modulation of surface CD11/CD18 glycoproteins (Mo1, LFA-1, p150,95) by human mononuclear phagocytes. Clin Immunol Immunopathol. 1988 Feb;46(2):272–283. doi: 10.1016/0090-1229(88)90189-4. [DOI] [PubMed] [Google Scholar]
  10. GOTTLIEBLAU K. S., WASSERMAN L. R., HERBERT V. RAPID CHARCOAL ASSAY FOR INTRINSIC FACTOR (IF), GASTRIC JUICE UNSATURATED B12 BINDING CAPACITY, ANTIBODY TO IF, AND SERUM UNSATURATED B12 BINDING CAPACITY. Blood. 1965 Jun;25:875–884. [PubMed] [Google Scholar]
  11. Gallay P., Jongeneel C. V., Barras C., Burnier M., Baumgartner J. D., Glauser M. P., Heumann D. Short time exposure to lipopolysaccharide is sufficient to activate human monocytes. J Immunol. 1993 Jun 1;150(11):5086–5093. [PubMed] [Google Scholar]
  12. Gallin J. I., Fletcher M. P., Seligmann B. E., Hoffstein S., Cehrs K., Mounessa N. Human neutrophil-specific granule deficiency: a model to assess the role of neutrophil-specific granules in the evolution of the inflammatory response. Blood. 1982 Jun;59(6):1317–1329. [PubMed] [Google Scholar]
  13. Gegner J. A., Ulevitch R. J., Tobias P. S. Lipopolysaccharide (LPS) signal transduction and clearance. Dual roles for LPS binding protein and membrane CD14. J Biol Chem. 1995 Mar 10;270(10):5320–5325. doi: 10.1074/jbc.270.10.5320. [DOI] [PubMed] [Google Scholar]
  14. Golenbock D. T., Bach R. R., Lichenstein H., Juan T. S., Tadavarthy A., Moldow C. F. Soluble CD14 promotes LPS activation of CD14-deficient PNH monocytes and endothelial cells. J Lab Clin Med. 1995 May;125(5):662–671. [PubMed] [Google Scholar]
  15. Goyert S. M., Ferrero E., Rettig W. J., Yenamandra A. K., Obata F., Le Beau M. M. The CD14 monocyte differentiation antigen maps to a region encoding growth factors and receptors. Science. 1988 Jan 29;239(4839):497–500. doi: 10.1126/science.2448876. [DOI] [PubMed] [Google Scholar]
  16. Grunwald U., Krüger C., Westermann J., Lukowsky A., Ehlers M., Schütt C. An enzyme-linked immunosorbent assay for the quantification of solubilized CD14 in biological fluids. J Immunol Methods. 1992 Nov 5;155(2):225–232. doi: 10.1016/0022-1759(92)90289-6. [DOI] [PubMed] [Google Scholar]
  17. Hailman E., Lichenstein H. S., Wurfel M. M., Miller D. S., Johnson D. A., Kelley M., Busse L. A., Zukowski M. M., Wright S. D. Lipopolysaccharide (LPS)-binding protein accelerates the binding of LPS to CD14. J Exp Med. 1994 Jan 1;179(1):269–277. doi: 10.1084/jem.179.1.269. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Haziot A., Chen S., Ferrero E., Low M. G., Silber R., Goyert S. M. The monocyte differentiation antigen, CD14, is anchored to the cell membrane by a phosphatidylinositol linkage. J Immunol. 1988 Jul 15;141(2):547–552. [PubMed] [Google Scholar]
  19. Haziot A., Rong G. W., Bazil V., Silver J., Goyert S. M. Recombinant soluble CD14 inhibits LPS-induced tumor necrosis factor-alpha production by cells in whole blood. J Immunol. 1994 Jun 15;152(12):5868–5876. [PubMed] [Google Scholar]
  20. Haziot A., Rong G. W., Lin X. Y., Silver J., Goyert S. M. Recombinant soluble CD14 prevents mortality in mice treated with endotoxin (lipopolysaccharide). J Immunol. 1995 Jun 15;154(12):6529–6532. [PubMed] [Google Scholar]
  21. Henson P. M., Johnston R. B., Jr Tissue injury in inflammation. Oxidants, proteinases, and cationic proteins. J Clin Invest. 1987 Mar;79(3):669–674. doi: 10.1172/JCI112869. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jack R. S., Grunwald U., Stelter F., Workalemahu G., Schütt C. Both membrane-bound and soluble forms of CD14 bind to gram-negative bacteria. Eur J Immunol. 1995 May;25(5):1436–1441. doi: 10.1002/eji.1830250545. [DOI] [PubMed] [Google Scholar]
  23. Kitchens R. L., Ulevitch R. J., Munford R. S. Lipopolysaccharide (LPS) partial structures inhibit responses to LPS in a human macrophage cell line without inhibiting LPS uptake by a CD14-mediated pathway. J Exp Med. 1992 Aug 1;176(2):485–494. doi: 10.1084/jem.176.2.485. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Labeta M. O., Landmann R., Obrecht J. P., Obrist R. Human B cells express membrane-bound and soluble forms of the CD14 myeloid antigen. Mol Immunol. 1991 Jan-Feb;28(1-2):115–122. doi: 10.1016/0161-5890(91)90094-z. [DOI] [PubMed] [Google Scholar]
  25. Lanier L. L., Le A. M., Phillips J. H., Warner N. L., Babcock G. F. Subpopulations of human natural killer cells defined by expression of the Leu-7 (HNK-1) and Leu-11 (NK-15) antigens. J Immunol. 1983 Oct;131(4):1789–1796. [PubMed] [Google Scholar]
  26. Luchi M., Munford R. S. Binding, internalization, and deacylation of bacterial lipopolysaccharide by human neutrophils. J Immunol. 1993 Jul 15;151(2):959–969. [PubMed] [Google Scholar]
  27. Lynn W. A., Raetz C. R., Qureshi N., Golenbock D. T. Lipopolysaccharide-induced stimulation of CD11b/CD18 expression on neutrophils. Evidence of specific receptor-based response and inhibition by lipid A-based antagonists. J Immunol. 1991 Nov 1;147(9):3072–3079. [PubMed] [Google Scholar]
  28. Maliszewski C. R. CD14 and immune response to lipopolysaccharide. Science. 1991 May 31;252(5010):1321–1322. doi: 10.1126/science.1718034. [DOI] [PubMed] [Google Scholar]
  29. Morris R. E., Gerstein A. S., Bonventre P. F., Saelinger C. B. Receptor-mediated entry of diphtheria toxin into monkey kidney (Vero) cells: electron microscopic evaluation. Infect Immun. 1985 Dec;50(3):721–727. doi: 10.1128/iai.50.3.721-727.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Pugin J., Ulevitch R. J., Tobias P. S. A critical role for monocytes and CD14 in endotoxin-induced endothelial cell activation. J Exp Med. 1993 Dec 1;178(6):2193–2200. doi: 10.1084/jem.178.6.2193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Schumann R. R. Function of lipopolysaccharide (LPS)-binding protein (LBP) and CD14, the receptor for LPS/LBP complexes: a short review. Res Immunol. 1992 Jan;143(1):11–15. doi: 10.1016/0923-2494(92)80074-u. [DOI] [PubMed] [Google Scholar]
  32. Schütt C., Schilling T., Grunwald U., Schönfeld W., Krüger C. Endotoxin-neutralizing capacity of soluble CD14. Res Immunol. 1992 Jan;143(1):71–78. doi: 10.1016/0923-2494(92)80082-v. [DOI] [PubMed] [Google Scholar]
  33. Sengeløv H., Kjeldsen L., Diamond M. S., Springer T. A., Borregaard N. Subcellular localization and dynamics of Mac-1 (alpha m beta 2) in human neutrophils. J Clin Invest. 1993 Sep;92(3):1467–1476. doi: 10.1172/JCI116724. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Setoguchi M., Nasu N., Yoshida S., Higuchi Y., Akizuki S., Yamamoto S. Mouse and human CD14 (myeloid cell-specific leucine-rich glycoprotein) primary structure deduced from cDNA clones. Biochim Biophys Acta. 1989 Jul 7;1008(2):213–222. doi: 10.1016/0167-4781(80)90012-3. [DOI] [PubMed] [Google Scholar]
  35. Terstappen L. W., Hollander Z., Meiners H., Loken M. R. Quantitative comparison of myeloid antigens on five lineages of mature peripheral blood cells. J Leukoc Biol. 1990 Aug;48(2):138–148. doi: 10.1002/jlb.48.2.138. [DOI] [PubMed] [Google Scholar]
  36. Ulevitch R. J., Mathison J. C., Schumann R. R., Tobias P. S. A new model of macrophage stimulation by bacterial lipopolysaccharide. J Trauma. 1990 Dec;30(12 Suppl):S189–S192. [PubMed] [Google Scholar]
  37. Ulevitch R. J., Tobias P. S. Receptor-dependent mechanisms of cell stimulation by bacterial endotoxin. Annu Rev Immunol. 1995;13:437–457. doi: 10.1146/annurev.iy.13.040195.002253. [DOI] [PubMed] [Google Scholar]
  38. Watson F., Robinson J. J., Edwards S. W. Neutrophil function in whole blood and after purification: changes in receptor expression, oxidase activity and responsiveness to cytokines. Biosci Rep. 1992 Apr;12(2):123–133. doi: 10.1007/BF02351217. [DOI] [PubMed] [Google Scholar]
  39. Weingarten R., Sklar L. A., Mathison J. C., Omidi S., Ainsworth T., Simon S., Ulevitch R. J., Tobias P. S. Interactions of lipopolysaccharide with neutrophils in blood via CD14. J Leukoc Biol. 1993 May;53(5):518–524. doi: 10.1002/jlb.53.5.518. [DOI] [PubMed] [Google Scholar]
  40. Wright S. D., Ramos R. A., Hermanowski-Vosatka A., Rockwell P., Detmers P. A. Activation of the adhesive capacity of CR3 on neutrophils by endotoxin: dependence on lipopolysaccharide binding protein and CD14. J Exp Med. 1991 May 1;173(5):1281–1286. doi: 10.1084/jem.173.5.1281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Wright S. D., Ramos R. A., Tobias P. S., Ulevitch R. J., Mathison J. C. CD14, a receptor for complexes of lipopolysaccharide (LPS) and LPS binding protein. Science. 1990 Sep 21;249(4975):1431–1433. doi: 10.1126/science.1698311. [DOI] [PubMed] [Google Scholar]
  42. Wright S. D., Tobias P. S., Ulevitch R. J., Ramos R. A. Lipopolysaccharide (LPS) binding protein opsonizes LPS-bearing particles for recognition by a novel receptor on macrophages. J Exp Med. 1989 Oct 1;170(4):1231–1241. doi: 10.1084/jem.170.4.1231. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Ziegler-Heitbrock H. W., Ulevitch R. J. CD14: cell surface receptor and differentiation marker. Immunol Today. 1993 Mar;14(3):121–125. doi: 10.1016/0167-5699(93)90212-4. [DOI] [PubMed] [Google Scholar]

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