The Journal of Immunology, Vol 155, Issue 4 2085-2095, Copyright © 1995 by American Association of Immunologists

ARTICLES

Endotoxin receptors (CD14) are found with CD16 (Fc gamma RIII) in an intracellular compartment of neutrophils that contains alkaline phosphatase

PA Detmers, D Zhou, D Powell, H Lichenstein, M Kelley and R Pironkova
Laboratory of Cellular Physiology and Immunology, Rockefeller University, New York, NY 10021, USA.

CD14 is a glycosylphosphatidylinositol (GPI)-anchored protein on the surfaces of monocytes and polymorphonuclear leukocytes (PMN) that binds and initiates cellular responses to bacterial LPS. PMN also contain an intracellular pool of CD14 that can be deployed rapidly to the cell surface in response to stimulation with a variety of agonists. To determine which of the well-characterized subcellular compartments of PMN contains CD14, cells were cavitated and fractionated on Percoll gradients. The gradient fractions were assayed for CD14 by ELISA and Western blot and for the marker proteins beta-glucuronidase (azurophil granules), vitamin B12 binding protein (specific granules), alkaline phosphatase (secretory vesicles and plasma membrane), and HLA (plasma membrane). Approximately one-half of the CD14 ran with plasma membrane fractions and one-half with intracellular membranes of light density. Both intracellular and cell surface CD14 were associated tightly with membrane, and both forms showed identical electrophoretic mobility. The intracellular CD14 was clearly not present in azurophil granules or specific granules, but ran precisely with alkaline phosphatase, a marker for secretory vesicles. Parallel studies showed that an additional GPI-linked protein, Fc gamma RIII (CD16), also fractionated precisely with CD14 and alkaline phosphatase. Association of CD14 with secretory vesicles were confirmed by studies on cells stimulated with the formyl peptide fNLLP for 20 min at 37 degrees C before fractionation. This treatment caused translocation of CD14 from intracellular fractions to plasma membrane fractions. No release of the specific granule marker vitamin B12 binding protein was observed under these conditions, whereas two other GPI-anchored proteins, alkaline phosphatase and CD16, moved coincidentally with CD14 to comigrate with the plasma membrane. Time course studies of CD14 and CD16 surface expression confirmed the rapid and coordinate up-regulation of these proteins. Thus, the intracellular compartment containing CD14 and CD16 had the properties of secretory vesicles. These vesicles may represent a specialized membrane domain of PMN enriched in GPI-anchored proteins.

This article has been cited by other articles:

				J. X. Wei, A. Verity, M. Garle, R. Mahajan, and V. Wilson Examination of the effect of procalcitonin on human leucocytes and the porcine isolated coronary artery Br. J. Anaesth., May 1, 2008; 100(5): 612 - 621. [Abstract] [Full Text] [PDF]

				S. W. Waldo, Y. Li, C. Buono, B. Zhao, E. M. Billings, J. Chang, and H. S. Kruth Heterogeneity of Human Macrophages in Culture and in Atherosclerotic Plaques Am. J. Pathol., April 1, 2008; 172(4): 1112 - 1126. [Abstract] [Full Text] [PDF]

				O. Popa-Nita, E. Rollet-Labelle, N. Thibault, C. Gilbert, S. G. Bourgoin, and P. H. Naccache Crystal-induced neutrophil activation. IX. Syk-dependent activation of class Ia phosphatidylinositol 3-kinase J. Leukoc. Biol., September 1, 2007; 82(3): 763 - 773. [Abstract] [Full Text] [PDF]

				N. Yin, M. Peng, Y. Xing, and W. Zhang Intracellular pools of Fc{alpha}R (CD89) in human neutrophils are localized in tertiary granules and secretory vesicles, and two Fc{alpha}R isoforms are found in tertiary granules J. Leukoc. Biol., September 1, 2007; 82(3): 551 - 558. [Abstract] [Full Text] [PDF]

				P. Scott, H. Ma, S. Viriyakosol, R. Terkeltaub, and R. Liu-Bryan Engagement of CD14 Mediates the Inflammatory Potential of Monosodium Urate Crystals J. Immunol., November 1, 2006; 177(9): 6370 - 6378. [Abstract] [Full Text] [PDF]

				N. L. Webster, K. Kedzierska, R. Azzam, G. Paukovics, J. Wilson, S. M. Crowe, and A. Jaworowski Phagocytosis stimulates mobilization and shedding of intracellular CD16A in human monocytes and macrophages: inhibition by HIV-1 infection J. Leukoc. Biol., February 1, 2006; 79(2): 294 - 302. [Abstract] [Full Text] [PDF]

				A. Tuin, A. Huizinga-Van der Vlag, A.-M. M. A. van Loenen-Weemaes, D. K. F. Meijer, and K. Poelstra On the role and fate of LPS-dephosphorylating activity in the rat liver Am J Physiol Gastrointest Liver Physiol, February 1, 2006; 290(2): G377 - G385. [Abstract] [Full Text] [PDF]

				K. Hattar, S. van Burck, A. Bickenbach, U. Grandel, U. Maus, J. Lohmeyer, E. Csernok, T. Hartung, W. Seeger, F. Grimminger, et al. Anti-proteinase 3 antibodies (c-ANCA) prime CD14-dependent leukocyte activation J. Leukoc. Biol., October 1, 2005; 78(4): 992 - 1000. [Abstract] [Full Text] [PDF]

				A. A. Rarok, P. C. Limburg, and C. G. M. Kallenberg Neutrophil-activating potential of antineutrophil cytoplasm autoantibodies J. Leukoc. Biol., July 1, 2003; 74(1): 3 - 15. [Abstract] [Full Text] [PDF]

				J.-W. Lee, M. J. Paape, T. H. Elsasser, and X. Zhao Elevated Milk Soluble CD14 in Bovine Mammary Glands Challenged with Escherichia coli Lipopolysaccharide J Dairy Sci, July 1, 2003; 86(7): 2382 - 2389. [Abstract] [Full Text] [PDF]

				V. S. Bansal, S. Vaidya, E. P. Somers, M. Kanuga, D. Shevell, R. Weikel, and P. A. Detmers Small Molecule Antagonists of Complement Receptor Type 3 Block Adhesion and Adhesion-Dependent Oxidative Burst in Human Polymorphonuclear Leukocytes J. Pharmacol. Exp. Ther., March 1, 2003; 304(3): 1016 - 1024. [Abstract] [Full Text] [PDF]

				V. L. Vega and A. De Maio Geldanamycin Treatment Ameliorates the Response to LPS in Murine Macrophages by Decreasing CD14 Surface Expression Mol. Biol. Cell, February 1, 2003; 14(2): 764 - 773. [Abstract] [Full Text] [PDF]

				P. Peyron, C. Bordier, E.-N. N'Diaye, and I. Maridonneau-Parini Nonopsonic Phagocytosis of Mycobacterium kansasii by Human Neutrophils Depends on Cholesterol and Is Mediated by CR3 Associated with Glycosylphosphatidylinositol-Anchored Proteins J. Immunol., November 1, 2000; 165(9): 5186 - 5191. [Abstract] [Full Text] [PDF]

				C. Hess, S. Sadallah, A. Hefti, R. Landmann, and J.-A. Schifferli Ectosomes Released by Human Neutrophils Are Specialized Functional Units J. Immunol., October 15, 1999; 163(8): 4564 - 4573. [Abstract] [Full Text] [PDF]

				T. Vasselon, E. Hailman, R. Thieringer, and P. A. Detmers Internalization of Monomeric Lipopolysaccharide Occurs after Transfer out of Cell Surface CD14 J. Exp. Med., August 16, 1999; 190(4): 509 - 522. [Abstract] [Full Text] [PDF]

				N. Thieblemont and S. D. Wright Mice Genetically Hyporesponsive to Lipopolysaccharide (LPS) Exhibit a Defect in Endocytic Uptake of LPS and Ceramide J. Exp. Med., June 16, 1997; 185(12): 2095 - 2100. [Abstract] [Full Text] [PDF]

				N. Borregaard and J. B. Cowland Granules of the Human Neutrophilic Polymorphonuclear Leukocyte Blood, May 15, 1997; 89(10): 3503 - 3521. [Full Text] [PDF]