Skip to main content
NCBI home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1986 Aug;78(2):381–388. doi: 10.1172/JCI112588

Binding and metabolism of platelet-activating factor by human neutrophils.

J T O'Flaherty, J R Surles, J Redman, D Jacobson, C Piantadosi, R L Wykle
PMCID: PMC423558  PMID: 3016027

Abstract

Human polymorphonuclear neutrophils rapidly incorporated radiolabeled platelet-activating factor, 1-O-[hexadecyl-9, 10-3H2]-2-acetyl-sn-glycero-3-phosphocholine ([3H]PAF), and then metabolized it into its sn-2-fatty acyl derivative. Fractionation of radiolabel-pretreated cells over Percoll gradients revealed that virtually all of the intact [3H]PAF was located in nongranule membranes that were enriched with alkaline phosphatase and cell surface glycoproteins. While still membrane associated, the ligand was rapidly converted to its acyl derivative and then more slowly transferred to specific granules and, to a lesser extent, azurophilic granules. In contrast, neutrophils did not metabolize [3H]PAF at 4 degrees C but rather gradually accumulated it in their alkaline phosphatase-enriched membrane subfractions. These same subfractions contained receptors for the ligand, as determined by their capacity to bind [3H]PAF specifically. Binding was readily saturated, partially reversible, and fit a two receptor model; dissociation constant (Kd) values for high and low affinity sites were 0.2 and 500 nM, respectively. Receptors with similar affinities were detected in whole cells. Furthermore, the potencies of several structural analogues in inhibiting binding of [3H]PAF to membranes correlated closely with their respective potencies in stimulating degranulation responses. Finally, quantitative studies suggested all or most of the cell's receptors were membrane associated. We conclude that PAF rapidly enters cellular membranes to bind with specific receptors that trigger function. The intramembranous ligand is also deacetylated, acylated, and then transferred to granules. This metabolism may be sufficiently rapid to limit ligand-receptor binding and distort quantitative analyses of receptors.

Full text

PDF
381

Selected References

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

  1. Albert D. H., Snyder F. Biosynthesis of 1-alkyl-2-acetyl-sn-glycero-3-phosphocholine (platelet-activating factor) from 1-alkyl-2-acyl-sn-glycero-3-phosphocholine by rat alveolar macrophages. Phospholipase A2 and acetyltransferase activities during phagocytosis and ionophore stimulation. J Biol Chem. 1983 Jan 10;258(1):97–102. [PubMed] [Google Scholar]
  2. Benveniste J., Chignard M., Le Couedic J. P., Vargaftig B. B. Biosynthesis of platelet-activating factor (PAF-ACETHER). II. Involvement of phospholipase A2 in the formation of PAF-ACETHER and lyso-PAF-ACETHER from rabbit platelets. Thromb Res. 1982 Mar 1;25(5):375–385. doi: 10.1016/0049-3848(82)90128-1. [DOI] [PubMed] [Google Scholar]
  3. Betz S. J., Henson P. M. Production and release of platelet-activating factor (PAF); dissociation from degranulation and superoxide production in the human neutrophil. J Immunol. 1980 Dec;125(6):2756–2763. [PubMed] [Google Scholar]
  4. Blank M. L., Lee T., Fitzgerald V., Snyder F. A specific acetylhydrolase for 1-alkyl-2-acetyl-sn-glycero-3-phosphocholine (a hypotensive and platelet-activating lipid). J Biol Chem. 1981 Jan 10;256(1):175–178. [PubMed] [Google Scholar]
  5. Blank M. L., Snyder F., Byers L. W., Brooks B., Muirhead E. E. Antihypertensive activity of an alkyl ether analog of phosphatidylcholine. Biochem Biophys Res Commun. 1979 Oct 29;90(4):1194–1200. doi: 10.1016/0006-291x(79)91163-x. [DOI] [PubMed] [Google Scholar]
  6. 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]
  7. Camussi G., Aglietta M., Coda R., Bussolino F., Piacibello W., Tetta C. Release of platelet-activating factor (PAF) and histamine. II. The cellular origin of human PAF: monocytes, polymorphonuclear neutrophils and basophils. Immunology. 1981 Feb;42(2):191–199. [PMC free article] [PubMed] [Google Scholar]
  8. Camussi G., Aglietta M., Malavasi F., Tetta C., Piacibello W., Sanavio F., Bussolino F. The release of platelet-activating factor from human endothelial cells in culture. J Immunol. 1983 Nov;131(5):2397–2403. [PubMed] [Google Scholar]
  9. Chap H., Mauco G., Simon M. F., Benveniste J., Douste-Blazy L. Biosynthetic labelling of platelet activating factor from radioactive acetate by stimulated platelets. Nature. 1981 Jan 22;289(5795):312–314. doi: 10.1038/289312a0. [DOI] [PubMed] [Google Scholar]
  10. Chesney C. M., Pifer D. D., Huch K. M. Desensitization of human platelets by platelet activating factor. Biochem Biophys Res Commun. 1985 Feb 28;127(1):24–30. doi: 10.1016/s0006-291x(85)80120-0. [DOI] [PubMed] [Google Scholar]
  11. Chilton F. H., O'Flaherty J. T., Ellis J. M., Swendsen C. L., Wykle R. L. Metabolic fate of platelet-activating factor in neutrophils. J Biol Chem. 1983 May 25;258(10):6357–6361. [PubMed] [Google Scholar]
  12. Chilton F. H., O'Flaherty J. T., Ellis J. M., Swendsen C. L., Wykle R. L. Selective acylation of lyso platelet activating factor by arachidonate in human neutrophils. J Biol Chem. 1983 Jun 25;258(12):7268–7271. [PubMed] [Google Scholar]
  13. Hanahan D. J., Demopoulos C. A., Liehr J., Pinckard R. N. Identification of platelet activating factor isolated from rabbit basophils as acetyl glyceryl ether phosphorylcholine. J Biol Chem. 1980 Jun 25;255(12):5514–5516. [PubMed] [Google Scholar]
  14. Handley D. A., Arbeeny C. M., Lee M. L., Van Valen R. G., Saunders R. N. Effect of platelet activating factor on endothelial permeability to plasma macromolecules. Immunopharmacology. 1984 Dec;8(3-4):137–142. doi: 10.1016/0162-3109(84)90017-1. [DOI] [PubMed] [Google Scholar]
  15. Hartung H. P. Acetyl glyceryl ether phosphorylcholine (platelet-activating factor) mediates heightened metabolic activity in macrophages. Studies on PGE, TXB2 and O2- production, spreading, and the influence of calmodulin-inhibitor W-7. FEBS Lett. 1983 Aug 22;160(1-2):209–212. doi: 10.1016/0014-5793(83)80968-5. [DOI] [PubMed] [Google Scholar]
  16. Humphrey D. M., McManus L. M., Hanahan D. J., Pinckard R. N. Morphologic basis of increased vascular permeability induced by acetyl glyceryl ether phosphorylcholine. Lab Invest. 1984 Jan;50(1):16–25. [PubMed] [Google Scholar]
  17. Hwang S. B., Lee C. S., Cheah M. J., Shen T. Y. Specific receptor sites for 1-O-alkyl-2-O-acetyl-sn-glycero-3-phosphocholine (platelet activating factor) on rabbit platelet and guinea pig smooth muscle membranes. Biochemistry. 1983 Sep 27;22(20):4756–4763. doi: 10.1021/bi00289a022. [DOI] [PubMed] [Google Scholar]
  18. Kornecki E., Ehrlich Y. H., Lenox R. H. Platelet-activating factor-induced aggregation of human platelets specifically inhibited by triazolobenzodiazepines. Science. 1984 Dec 21;226(4681):1454–1456. doi: 10.1126/science.6150550. [DOI] [PubMed] [Google Scholar]
  19. Kramer R. M., Patton G. M., Pritzker C. R., Deykin D. Metabolism of platelet-activating factor in human platelets. Transacylase-mediated synthesis of 1-O-alkyl-2-arachidonoyl-sn-glycero-3-phosphocholine. J Biol Chem. 1984 Nov 10;259(21):13316–13320. [PubMed] [Google Scholar]
  20. Lee T., Lenihan D. J., Malone B., Roddy L. L., Wasserman S. I. Increased biosynthesis of platelet-activating factor in activated human eosinophils. J Biol Chem. 1984 May 10;259(9):5526–5530. [PubMed] [Google Scholar]
  21. Lumb R. H., Pool G. L., Bubacz D. G., Blank M. L., Snyder F. Spontaneous and protein-catalyzed transfer of 1-alkyl-2-acetyl-sn-glycero-3-phosphocholine (platelet-activating factor) between phospholipid bilayers. Biochim Biophys Acta. 1983 Feb 7;750(2):217–222. doi: 10.1016/0005-2760(83)90021-8. [DOI] [PubMed] [Google Scholar]
  22. McManus L. M., Hanahan D. J., Demopoulos C. A., Pinckard R. N. Pathobiology of the intravenous infusion of acetyl glyceryl ether phosphorylcholine (AGEPC), a synthetic platelet-activating factor (PAF), in the rabbit. J Immunol. 1980 Jun;124(6):2919–2924. [PubMed] [Google Scholar]
  23. Mencia-Huerta J. M., Roubin R., Morgat J. L., Benveniste J. Biosynthesis of platelet-activating factor (PAF)acether). III. Formation of PAF-acether from synthetic substrates by stimulated murine macrophages. J Immunol. 1982 Aug;129(2):804–808. [PubMed] [Google Scholar]
  24. Mueller H. W., O'Flaherty J. T., Wykle R. L. Biosynthesis of platelet activating factor in rabbit polymorphonuclear neutrophils. J Biol Chem. 1983 May 25;258(10):6213–6218. [PubMed] [Google Scholar]
  25. Mueller H. W., O'Flaherty J. T., Wykle R. L. The molecular species distribution of platelet-activating factor synthesized by rabbit and human neutrophils. J Biol Chem. 1984 Dec 10;259(23):14554–14559. [PubMed] [Google Scholar]
  26. Nishihira J., O'Flaherty J. T. Phorbol myristate acetate receptors in human polymorphonuclear neutrophils. J Immunol. 1985 Nov;135(5):3439–3447. [PubMed] [Google Scholar]
  27. O'Flaherty J. T., Lees C. J., Miller C. H., McCall C. E., Lewis J. C., Love S. H., Wykle R. L. Selective desensitization of neutrophils: further studies with 1-O-alkyl-sn-glycero-3-phosphocholine analogues. J Immunol. 1981 Aug;127(2):731–737. [PubMed] [Google Scholar]
  28. O'Flaherty J. T., Miller C. H., Lewis J. C., Wykle R. L., Bass D. A., McCall C. E., Waite M., DeChatelet L. R. Neutrophil responses to platelet-activating factor. Inflammation. 1981 Sep;5(3):193–201. doi: 10.1007/BF00914443. [DOI] [PubMed] [Google Scholar]
  29. O'Flaherty J., Kosfeld S., Nishihira J. Binding and metabolism of leukotriene B4 by neutrophils and their subcellular organelles. J Cell Physiol. 1986 Mar;126(3):359–370. doi: 10.1002/jcp.1041260306. [DOI] [PubMed] [Google Scholar]
  30. Pieroni G., Hanahan D. J. Metabolic behavior of acetyl glyceryl ether phosphorylcholine on interaction with rabbit platelets. Arch Biochem Biophys. 1983 Jul 15;224(2):485–493. doi: 10.1016/0003-9861(83)90236-9. [DOI] [PubMed] [Google Scholar]
  31. Polonsky J., Tencé M., Varenne P., Das B. C., Lunel J., Benveniste J. Release of 1-O-alkylglyceryl 3-phosphorylcholine, O-deacetyl platelet-activating factor, from leukocytes: chemical ionization mass spectrometry of phospholipids. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7019–7023. doi: 10.1073/pnas.77.12.7019. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Prescott S. M., Zimmerman G. A., McIntyre T. M. Human endothelial cells in culture produce platelet-activating factor (1-alkyl-2-acetyl-sn-glycero-3-phosphocholine) when stimulated with thrombin. Proc Natl Acad Sci U S A. 1984 Jun;81(11):3534–3538. doi: 10.1073/pnas.81.11.3534. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Purdon A. D., Smith J. B. Turnover of arachidonic acid in the major diacyl and ether phospholipids of human platelets. J Biol Chem. 1985 Oct 15;260(23):12700–12704. [PubMed] [Google Scholar]
  34. Shak S., Goldstein I. M. Leukotriene B4 omega-hydroxylase in human polymorphonuclear leukocytes. Partial purification and identification as a cytochrome P-450. J Clin Invest. 1985 Sep;76(3):1218–1228. doi: 10.1172/JCI112077. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Shen T. Y., Hwang S. B., Chang M. N., Doebber T. W., Lam M. H., Wu M. S., Wang X., Han G. Q., Li R. Z. Characterization of a platelet-activating factor receptor antagonist isolated from haifenteng (Piper futokadsura): specific inhibition of in vitro and in vivo platelet-activating factor-induced effects. Proc Natl Acad Sci U S A. 1985 Feb;82(3):672–676. doi: 10.1073/pnas.82.3.672. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Siraganian R. P., Osler A. G. Destruction of rabbit platelets in the allergic response of sensitized leukocytes. I. Demonstration of a fluid phase intermediate. J Immunol. 1971 May;106(5):1244–1251. [PubMed] [Google Scholar]
  37. Stimler N. P., Bloor C. M., Hugli T. E., Wykle R. L., McCall C. E., O'Flaherty J. T. Anaphylactic actions of platelet-activating factor. Am J Pathol. 1981 Oct;105(1):64–69. [PMC free article] [PubMed] [Google Scholar]
  38. Stimler N. P., O'Flaherty J. T. Spasmogenic properties of platelet-activating factor: evidence for a direct mechanism in the contractile response of pulmonary tissues. Am J Pathol. 1983 Oct;113(1):75–84. [PMC free article] [PubMed] [Google Scholar]
  39. Surles J. R., Wykle R. L., O'Flaherty J. T., Salzer W. L., Thomas M. J., Snyder F., Piantadosi C. Facile synthesis of platelet-activating factor and racemic analogues containing unsaturation in the sn-1-alkyl chain. J Med Chem. 1985 Jan;28(1):73–78. doi: 10.1021/jm00379a015. [DOI] [PubMed] [Google Scholar]
  40. Swendsen C. L., Ellis J. M., Chilton F. H., 3rd, O'Flaherty J. T., Wykle R. L. 1-O-alkyl-2-acyl-sn-glycero-3-phosphocholine: a novel source of arachidonic acid in neutrophils stimulated by the calcium ionophore A23187. Biochem Biophys Res Commun. 1983 May 31;113(1):72–79. doi: 10.1016/0006-291x(83)90433-3. [DOI] [PubMed] [Google Scholar]
  41. Sánchez-Crespo M., Alonso F., Iñarrea P., Alvarez V., Egido J. Vascular actions of synthetic PAF-acether (a synthetic platelet-activating factor) in the rat: evidence for a platelet independent mechanism. Immunopharmacology. 1982 Apr;4(2):173–185. doi: 10.1016/0162-3109(82)90019-4. [DOI] [PubMed] [Google Scholar]
  42. Touqui L., Jacquemin C., Vargaftig B. B. Conversion of 3H-PAF acether by rabbit platelets is independent from aggregation: evidences for a novel metabolite. Biochem Biophys Res Commun. 1983 Feb 10;110(3):890–893. doi: 10.1016/0006-291x(83)91045-8. [DOI] [PubMed] [Google Scholar]
  43. Valone F. H., Coles E., Reinhold V. R., Goetzl E. J. Specific binding of phospholipid platelet-activating factor by human platelets. J Immunol. 1982 Oct;129(4):1637–1641. [PubMed] [Google Scholar]
  44. Valone F. H., Goetzl E. J. Specific binding by human polymorphonuclear leucocytes of the immunological mediator 1-O-hexadecyl/octadecyl-2-acetyl-sn-glycero-3-phosphorylcholine. Immunology. 1983 Jan;48(1):141–149. [PMC free article] [PubMed] [Google Scholar]
  45. Valone F. H. Inhibition of binding of the platelet-activating factor AGEPC to platelets by the AGEPC analog rac-3-(N-n-octadecylcarbamoyloxy)-2-methoxypropyl 2-thiazolioethyl phosphate (CV-3988). Biochem Biophys Res Commun. 1985 Jan 16;126(1):502–508. doi: 10.1016/0006-291x(85)90634-5. [DOI] [PubMed] [Google Scholar]
  46. Valone F. H. Isolation of a platelet membrane protein which binds the platelet-activating factor 1-0-hexadecyl-2-acetyl-SN-glycero-3-phosphorylcholine. Immunology. 1984 May;52(1):169–174. [PMC free article] [PubMed] [Google Scholar]
  47. Wykle R. L., Miller C. H., Lewis J. C., Schmitt J. D., Smith J. A., Surles J. R., Piantadosi C., O'Flaherty J. T. Stereospecific activity of 1-O-alkyl-2-O-acetyl-sn-glycero-3-phosphocholine and comparison of analogs in the degranulation of platelets and neutrophils. Biochem Biophys Res Commun. 1981 Jun;100(4):1651–1658. doi: 10.1016/0006-291x(81)90708-7. [DOI] [PubMed] [Google Scholar]

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

RESOURCES