Abstract
Formyl-methionyl-leucyl-phenylalanine (FMLP), platelet activating factor (PAF) and leukotriene B4 (LTB4) are potent activators of human neutrophils. Using human neutrophils prelabelled with the fluorescent indicator dye, Quin 2, or with [32P]-orthophosphate, we examined the effects of these stimuli on intracellular free calcium concentration, [Ca2+]i, and, on various indices of phosphoinositide metabolism, including [32P]-phosphatidic acid (PtdA) formation. The concentration-dependence of the observed changes in [Ca2+]i or [32P]-PtdA were then compared to stimulus-induced aggregation and enzyme release (β-N-acetylglucosaminidase (NAG) and lysozyme).
FMLP, PAF and LTB4 caused a concentration-dependent elevation of [Ca2+]i, aggregation and enzyme release. However, unlike FMLP and PAF, LTB4 (≦2.5 μM) did not cause significant formation of [32P]-PtdA. The concentration response curves for agonist-induced elevation of [Ca2+]i lie to the left of those for aggregation and enzyme release. FMLP and PAF also caused an elevation of [Ca2+]i at concentrations lower than those required to elicit [32P]-PtdA formation.
These observations suggest that [Ca2+]i elevationper se cannot mediate human neutrophil functional responses to FMLP, PAF and LTB4. Consequently there may exist other mediator(s) that act in concert with [Ca2+]i or are triggered by [Ca2+]i elevation to promote human neutrophil activation. Both the elevation of [Ca2+]i and the formation of these putative mediator(s) in response to LTB4 apparently occur independently of inositol phospholipid hydrolysis.
We’re sorry, something doesn't seem to be working properly.
Please try refreshing the page. If that doesn't work, please contact support so we can address the problem.
References
S. B. Kramer, B. N. Cronstein and G. Weissmann,Polymorphonuclear leukocytes. InArthritis and Allied Conditions. 10th Ed. (Ed. D. J. McCarthy) pp. 324–341, Philadelphia, USA 1985.
M. Baggiolini and B. Dewald.,The neutrophil, Int. Archs. Allergy Appl. Immun.76, 13–20 (1985).
I. M. Goldstein, C. L. Malmsten, H. B. Kaplan, O. Radmark, B. Samuelsson and G. Weissmann,Thromboxane generation by human peripheral blood polymorphonuclear leukocytes. J. Exp. Med.148, 787–792 (1978).
A. W. Ford-Hutchinson, M. A. Bray, M. V. Doig, M. E. Shipley and M. J. H. Smith,Leukotriene B, a potent chemokinetic and aggregating substance released from polymorphonuclear leukocytes, Nature286, 264–265 (1980).
W. Jubiz, O. Radmark, C. Malmsten, G. Hansson, J. A. Lindgren, J. Palmblad, A. M. Uden and B. Samuelsson,A novel leukotriene produced by stimulation of leukocytes with formylmethionyl-leucylphenylalanine. J. Biol. Chem.257, 6106–6110 (1982).
H. W. Mueller, J. T. O'Flaherty and R. L. Wykle,Biosynthesis of platelet activating factor in rabbit polymorphonuclear neutrophils. J. Biol. Chem.258, 6213–6218 (1983).
D. E. Chenoweth and T. E. Hugli,Demonstration of specific C5a receptor on intact human polymorphonuclear leukocytes. Proc. Natl. Acad. Sci. USA75, 3943–3947 (1978).
S. Aswanikumar, B. Corcoran, E. Schiffmann, A. R. Day, R. J. Freer, H. J. Showell, E. L. Becker and C. Pert,Demonstration of a receptor on rabbit neutrophils for chemotactic peptides, Biochem. Biophys. Res. Commun.74, 810–817 (1977).
L. T. Williams, R. Snyderman, M. C. Pike and R. J. Lefkowitz,Specific receptor sites for chemotactic peptides on human polymorphonuclear leukocytes, Proc. Natl. Acad. Sci. USA74, 1204–1208 (1977).
D. W. Goldman and E. J. Goetzl,Specific binding of leukotriene B 4 to receptors on human polymorphonuclear leukocytes, J. Immunol.129, 1600–1604 (1982).
F. H. Valone and E. J. Goetzl,Specific binding by human polymorphonuclear leukocytes of the immunological mediator I-O-hexadecyl/octadecyl-2-acetyl-sn-glycero-3-phosphorylcholine. Immunology48, 141–149 (1983).
J. T. O'Flaherty, J. R. Sutles, J. Redman, D. Jacobson, C. Piantadosi and R. Wykle,Binding and metabolism of platelet-activating factor by human neutrophils, J. Clin. Invest.78, 381–388 (1986).
F. Divirgilio, D. P. Lew and T. Pozzan,Protein kinase C activation of physiological processes in human neutrophils at vanishingly small cytosolic Ca 2+ levels. Nature301, 691–693 (1984).
D. P. Lew, C. B. Wollheim, F. A. Waldvogel and T. Pozzan,Modulation of cytosolic-free calcium transients by changes in intracellular calcium-buffering capacity: correlation with exocytosis and O 2 production in human neutrophils, J. Cell. Biol.99, 1212–1220 (1984).
J. Westwick and C. Poll,Mechanism of calcium homeestasis in the polymorphonuclear leucocyte, Agents and Actions19, 80–86 (1986).
T. Pozzan, D. P. Lew, C. B. Wollheim and R. Y. Tsien,Is cytosolic free calcium regulating neutrophil activation? Science221, 1413–1415 (1983).
M. Volpi, R. Yassin, W. Tao, T. F. P. Molski, P. H. Naccache and R. I. Sha'afi,Leukotriene B 4 mobilises calcium without the breakdown of polyphosphoinositides and the production of phosphatidic acid in rabbit neutrophils; Proc. Natl. Acad. Sci. USA81, 5966–5969 (1984).
P. G. Bradford and R. P. Rubin,Characterization of formylmethionyl-leucyl-phenylalanine stimulation of inositol triphosphate accumulation in rabbit neutrophils, Mol. Pharmacol.27, 74–78 (1985).
Y. Nishizuka,The role of protein kinase C in cell surface signal transduction and tumour promotion, Nature308, 693–698 (1984).
H. Streb, R. F. Irvine, M. J. Berridge and I. Schulz,Release of Ca 2+ from a nonmitochondrial intracellular store in pancreatic acinar cells by inositol-1,4,5,-triphosphate, Nature306, 67–69 (1983).
M. Prentki, C. B. Wollheim and P. D. Lew,Characterization of Ca 2+ sequestering pools and the action of inositol-1,4,5-trisphosphate, J. Biol. Chem.259, 13777–13782 (1984).
C. A. Tyson, H. V. Zande and D. E. Green,Phospholipids as ionophores, J. Bio. Chem.251, 1326–1332 (1976).
R. P. Holmes and N. L. Yoss,Failure of phosphatidic acid to translocate Ca 2+ across phosphatidylcholine membranes, Nature305, 637–638 (1983).
J. M. Robinson, J. A. Badwey, M. L. Karnovski and M. J. Karnovski,Superoxide release by neutrophils: synergistic effects of a phorbol ester and a calcium ionophore, Biochem. Biophys. Res. Commun.122, 734–739 (1984).
A. Penfield and M. M. Dale,Synergism between A23187 and I-oleoyl-2-acetyl-glycerol in superoxide production by human neutrophils, Biochem. Biophys. Res. Commun.125, 332–336 (1984).
C. D. Smith, C. Cox and R. Snyderman,Receptor-coupled activation of phosphoinositide-specific-phospholipase C by an N protein, Science 232, 97–100 (1986).
H. Ohta, F. Okajima and M. Ui,Inhibition by islet-activating protein of a chemotactic peptide-induced early breakdown of inositol phospholipids and Ca 2+ mobilisation in guinea pig neutrophils. J. Biol. Chem.260, 15771–15780 (1985).
P. M. Lad, C. V. Olson, I. S. Grewal and S. J. Scott,A pertussis toxin-sensitive GTP-binding protein in the human neutrophil regulates multiple receptors, calcium mobilisation and lectin-induced capping, Proc. Natl. Acad. Sci. USA82, 8643–8647 (1985).
M. W. Verghese, C. D. Smith and R. Snyderman,Potential role of guanine nucleotide regulator protein in chemoattractant receptor mediated phosphoinositide metabolism, Ca 2+ mobilisation and cellular responses by leukocytes, Biochem. Biophys. Res. Commun.127, 450–457 (1985).
K. G. Lazzari, P. J. Proto and E. R. Simons,Simultaneous measurements of stimulus-induced changes in cytoplasmic Ca 2+ and in membrane potential of human neutrophils, J. Biol. Chem.261, 9710–9713 (1986).
P. H. Naccache, M. M. Molski, M. Volpi, E. L. Becker and R. I. Sha'Afi,Unique inhibitory profile of platelet activating factor induced calcium mobilisation, polyphosphoinositide turnover and granule enzyme secretion in rabbit neutrophils towards pertussis toxin and phorbol ester, Biochem. Biophys. Res. Commun. 130, 677–684 (1985).
J. L. Gordon,Blood platelet lysosomes and their contribution to the pathophysiological role of platelets, InLysosomes in Biology and Pathology, Vol. 4 (Eds. J. T. Dingle and R. T. Dean) pp. 3–31, Amsterdam, Holland 1975.
J. R. White, C.-K. Huang, J. M. Hill, P. H. Naccache, E. L. Becker and R. I. Sha'Afi,Effect of phorbol 12-myristate 13-acetate and its anologue 4 a phorbol 12,13-didecanoate on protein phosphorylation and lysosomal enzyme release in rabbit neutrophils, J. Biol. Chem.259, 8605–8611 (19884).
J. R. White, P. H. Naccache, T. F. P. Molski, P. Borgeat and R. I. Sha'Afi,Direct demonstration of increased intracellular concentration of free calcium in rabbit and human neutrophils following stimulation by chemotactic factor, Biochem. Biophys. Res. Commun.113, 44–50 (1983).
R. W. Dougherty, P. P. Godfrey, P. C. Hoyle, J. W. Putney, and R. J. Freer,Secretagogue-induced phosphoinositide metabolism in human leucocytes, Biochem. J.222, 307–314 (1984).
J. V. Lloyd, E. E. Nishizawa, J. Halder and J. F. Mustard,Changes in 32 p-labelling of platelet phospholipids in response to ADP, Br. J. Haematol.23, 571–585 (1972).
E. Yavin and A. Zutra,Separation and analysis of 32 p-labelled phospholipids by a simple and rapid thin-layer chromatographic procedure and its application to cultured neuroblastoma cells, Anal. Biochem.80, 430–437 (1977).
J. Schacht,Extraction and purification of polyphosphoinositides, Meth. Enzymol.72, 616–631 (1981).
J. Jolles, H. Zwiers, A. Dekker, K. W. A. Wirtz and W. H. Gispen,Corticotrophin-(1-24) tetracosapeptide affects protein phosphorylation and polyphosphoinositide metabolism in rat brain, Biochem. J.194, 283–291 (1981).
K. Yano, J. Hattori, A. Imai and Y. Nozawa,Modification of positional distribution of fatty acids in phosphatidylinositol of rabbit neutrophils stimulated with forylmethionyl-leucylphenyl-alanine. Biochem. Biophys. Acta.752, 137–144 (1983).
C. D. Smith, B. C. Lane, I. Kusaka, M.W. Verghese and R. Snyderman,Chemiattractant receptor-induced hydrolysis of phosphatidyl-inositol 4,5-bisphosphate in human polymorphonuclear leukocyte membranes, J. Biol. Chem.260, 5875–5878 (1985).
L. A. Kater, E. J. Goetzl and K. F. Austen,Isolation of human eosinophil phospholipase D, J. Clin. Invest.57, 1173–1180 (1976).
D. Allan and R. H. Michell,A comparison of the effects of phytohaemagglutinin and of calcium ionophore A23187 on the metabolism of glycerolipids in small lymphocytes, Biochem. J.164, 389–397 (1977).
A. G. Rossi and D. E. Macintyre,Pertussis toxin inhibits the transduction processes activated by LTB 4 and PAF in human neutrophils, Adv. PG. Tx. LTs. Research (1987) in press.
A. G. Rossi and D. E. Macintyre,The effect of pertussis toxin on agonist-induced phosphoinositide hydrolysis and changes in cytosolic free Ca 2+ in human neutrophils, Br. J. Pharmacol.88, 654P (1986).
F. Okajima and M. Ui,ADP-ribosylation of the specific membrane protein by islet-activating protein, pertussis toxin, associated with inhibition of a chemotactic peptide-induced arachidonate release in neutrophils: a possible role of the toxin substrate in Ca 2+ mobilising biosignaling, J. Biol. Chem.259, 13863–13871 (1984).
P. Gierschik, J. Fallon, G. Milligan, M. Pines, I. Gallin and A. Spiegel,Immunochemical evidence for a novel pertussis toxin substrate in human neutrophils, J. Biol. Chem.261, 8058–8062 (1986).
S. K. Joseph,Receptor-stimulated phosphoinositide metabolism: a role for GTP-binding proteins? Trends. Biochem. Sci.10, 297–298 (1985).
S. Cockcroft and B. D. GompertsRole of guanine nucleotide binding protein in the activation of polyphosphoinositide phosphodiesterase, Nature314, 534–536 (1985).
A. Holian,Leukotriene B 4 stimulation of phosphatidylinositol turnover in macrophages and inhibition by pertussis toxin, Febs Letters201, 15–19 (1986).
T. Andersson, W. Schlegel, A. Monod, K.-H. Krause, O. Stendahl and D. P. Lew,Leukotriene B 4 stimulation of phagocytes results in the formation of inositol 1,4,5-trisphosphate, Biochem. J.240, 333–340 (1986).
M. J. Berridge,The molecular basis of cummunication within the cell, Scientific American, 253, 142–152 (1985).
D. Chandler, G. Meusel, E. Schumaker and C. Stapleton,FMLP-induced enzyme release from neutrophils: a role for intracellular calcium, Am. J. Physiol.245, C196-C202 (1983).
P. H. Naccache, T. F. P. Molski, P. Borgeat and R. I. Sha'Afi,Intracellular calcium redistribution and its relationship to f-Met-Leu-Phe, leukotriene B 4 and phorbol ester induced rabbit neutrophil degranulation, J. Cell. Physiol.122, 273–280 (1985).
F. Divirgilio, L. M. Vicentini, S. Treves, G. Riz and T. Pozzan,Inositol phosphate formation in f-Met-Leu-Phe-stimulated human neutrophils does not require an increase in cytoplasmic free Ca 2+ concentration, Biochem. J.229, 361–367 (1985).
G. M. Burgess, J. S. McKinney, R. F. Irvine, M. J. Berridge, P. C. Hoyle and J. W. Putney,Inositol 1,4,5-trisphosphate may be a signal for f-Met-Leu-Phe-induced intracellular Ca 2+ mobilisation in human leucocytes (HL-60 cells) Febs Letters176, 193–196 (1984).
C. N. Serhan, J. Fridovich, E. G. Goetzl, P. B. Dunham and G. WeissmannLeukotriene B 4 and phosphatidic acid are calcium ionophores, J. Biol. Chem.257, 4746–4752 (1984).
M. W. Verghese, L. Charles, L. Jakoi, S. B. Dillon and R. Snyderman,Role of a guanine nucleotide regulatory protein in the activation of phospholipase C by different chemoattractants, J. Immunol.138, 4374–4380 (1987).
I. R. Batty, S. R. Nahorski and R. F. Irvine,Rapid formation of inositol 1,3,4,5-tetracisphosphate following muscarinic receptor stimulation of rat cerebral cortical slices, Biochem. J.232, 211–215 (1985).
R. F. Irvine, A. J. Letcher, J. P. Heslop and M. J. Berridge,The inositol tris/tetracisphosphate pathway — demonstration of Ins (1,4,5)p 3 3-kinase activity in animal tissues, Nature320, 631–634 (1986).
C. A. Hansen, S. Mah and J. R. Williamson,Formation and metabolism of inositol 1,3,4,5-tetrakisphosphate in liver, J. Biol. Chem.261, 8100–8103 (1986).
R. F. Irvine and R. M. Moor,Micro-injection of inositol 1,3,4,5-tetracisphosphate activates sea urchin eggs by a mechanism dependent on external Ca 2+, Biochem. J.240, 917–920 (1986).
C. W. Taylor,Receptor regulation of calcium entry, Trends. Pharmacol. Sci.8, 79–80 (1987).
F. Rossi, V. Della Bianca, M. Grezeskowiak, P. de Togni and G. Cabrini,Relationships between phosphoinositide metabolism, Ca 2+ changes and respiratory burst in formylmethionyl-leucyl-phenylalanine-stimulated human neutrophils, Febs Letters.181, 253–258 (1985).
P. H. Naccache, M. M. Molski, M. Volpi, J. Shefcyk, T. F. P. Molski, L. Loew, E. L. Becker and R. I. Sha'Afi,Biochemical events associated with the stimulation of rabbit neutrophils by platelet activating factor, J. Leukocyte Biology40, 533–548 (1986).
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Rossi, A.G., McMillan, R.M. & MacIntyre, D.E. Agonist-induced calcium flux, phosphoinositide metabolism, aggregation and enzyme secretion in human neutrophils. Agents and Actions 24, 272–282 (1988). https://doi.org/10.1007/BF02028283
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF02028283