Skip to main content
NCBI home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1989 Aug;84(2):665–671. doi: 10.1172/JCI114213

The erythrocyte as instigator of inflammation. Generation of amidated C3 by erythrocyte adenosine deaminase.

M K Hostetter 1, G M Johnson 1
PMCID: PMC548930  PMID: 2788175

Abstract

Myocardial ischemia is characterized by the liberation of adenosine and by complement-mediated inflammation. We have reported that amidated C3, formed when ammonia (NH3) disrupts the thiolester bond of C3, serves as an alternative pathway convertase, generates C5b-9, and stimulates phagocytic oxidative metabolism. We investigated whether the deamination of adenosine by adenosine deaminase in hematopoietic cells might liberate sufficient ammonia to form amidated C3 and thereby trigger complement-mediated inflammation at ischemic sites. In the presence of 4 mM adenosine, NH3 production per erythrocyte (RBC) was equal to that per neutrophil (PMN) (3.3 X 10(-15) mol/cell per h). Because RBC outnumber PMN in normal blood by a thousandfold, RBC are the major source of NH3 production in the presence of adenosine. NH3 production derived only from the deamination of adenosine by the enzyme adenosine deaminase and was abolished by 0.4 microM 2'-deoxycoformycin, a specific inhibitor of adenosine deaminase. When purified human C3 was incubated with 5 X 10(8) human RBC in the presence of adenosine, disruption of the C3 thiolester increased more than twofold over that measured in C3 incubated with buffer, or in C3 incubated with RBC (P less than 0.05). The formation of amidated C3 was abolished by the preincubation of RBC with 2'-deoxycoformycin (P less than 0.001). Amidated C3 elicited statistically significant release of superoxide, myeloperoxidase, and lactoferrin from PMN. Thus, the formation of amidated C3 by RBC deamination of adenosine triggers a cascade of complement-mediated inflammatory reactions.

Full text

PDF
665

Images in this article

669Image
on p.669

Selected References

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

  1. Agarwal R. P. In vivo inhibition of adenosine deaminase by 2'-deoxycoformycin in mouse blood and leukemia L1210 cells. Biochem Pharmacol. 1980 Feb;29(2):187–193. doi: 10.1016/0006-2952(80)90327-5. [DOI] [PubMed] [Google Scholar]
  2. Agarwal R. P. Inhibitors of adenosine deaminase. Pharmacol Ther. 1982;17(3):399–429. doi: 10.1016/0163-7258(82)90023-7. [DOI] [PubMed] [Google Scholar]
  3. Agarwal R. P., Spector T., Parks R. E., Jr Tight-binding inhibitors--IV. Inhibition of adenosine deaminases by various inhibitors. Biochem Pharmacol. 1977 Mar 1;26(5):359–367. doi: 10.1016/0006-2952(77)90192-7. [DOI] [PubMed] [Google Scholar]
  4. Bolli R., Patel B. S., Jeroudi M. O., Lai E. K., McCay P. B. Demonstration of free radical generation in "stunned" myocardium of intact dogs with the use of the spin trap alpha-phenyl N-tert-butyl nitrone. J Clin Invest. 1988 Aug;82(2):476–485. doi: 10.1172/JCI113621. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Giclas P. C., Pinckard R. N., Olson M. S. In vitro activation of complement by isolated human heart subcellular membranes. J Immunol. 1979 Jan;122(1):146–151. [PubMed] [Google Scholar]
  6. Gordon D. L., Johnson G. M., Hostetter M. K. Ligand-receptor interactions in the phagocytosis of virulent Streptococcus pneumoniae by polymorphonuclear leukocytes. J Infect Dis. 1986 Oct;154(4):619–626. doi: 10.1093/infdis/154.4.619. [DOI] [PubMed] [Google Scholar]
  7. Gordon D. L., Krueger R. A., Quie P. G., Hostetter M. K. Amidation of C3 at the thiolester site: stimulation of chemiluminescence and phagocytosis by a new inflammatory mediator. J Immunol. 1985 May;134(5):3339–3345. [PubMed] [Google Scholar]
  8. Hartmann J. R., Robinson J. A., Gunnar R. M. Chemotactic activity in the coronary sinus after experimental myocardial infarction: effects of pharmacologic interventions on ischemic injury. Am J Cardiol. 1977 Oct;40(4):550–555. doi: 10.1016/0002-9149(77)90070-4. [DOI] [PubMed] [Google Scholar]
  9. Henderson J. F., Brox L., Zombor G., Hunting D., Lomax C. A. Specificity of adenosine deaminase inhibitors. Biochem Pharmacol. 1977 Nov 1;26(21):1967–1972. doi: 10.1016/0006-2952(77)90003-x. [DOI] [PubMed] [Google Scholar]
  10. Hetherington S. V., Spitznagel J. K., Quie P. G. An enzyme-linked immunoassay (ELISA) for measurement of lactoferrin. J Immunol Methods. 1983 Dec 16;65(1-2):183–190. doi: 10.1016/0022-1759(83)90314-9. [DOI] [PubMed] [Google Scholar]
  11. Hill J. H., Ward P. A. The phlogistic role of C3 leukotactic fragments in myocardial infarcts of rats. J Exp Med. 1971 Apr 1;133(4):885–900. doi: 10.1084/jem.133.4.885. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hostetter M. K., Gordon D. L. Biochemistry of C3 and related thiolester proteins in infection and inflammation. Rev Infect Dis. 1987 Jan-Feb;9(1):97–109. doi: 10.1093/clinids/9.1.97. [DOI] [PubMed] [Google Scholar]
  13. Hostetter M. K., Krueger R. A., Schmeling D. J. The biochemistry of opsonization: central role of the reactive thiolester of the third component of complement. J Infect Dis. 1984 Nov;150(5):653–661. doi: 10.1093/infdis/150.5.653. [DOI] [PubMed] [Google Scholar]
  14. Isenman D. E., Kells D. I., Cooper N. R., Müller-Eberhard H. J., Pangburn M. K. Nucleophilic modification of human complement protein C3: correlation of conformational changes with acquisition of C3b-like functional properties. Biochemistry. 1981 Jul 21;20(15):4458–4467. doi: 10.1021/bi00518a034. [DOI] [PubMed] [Google Scholar]
  15. Janatova J., Lorenz P. E., Schechter A. N., Prahl J. W., Tack B. F. Third component of human complement: appearance of a sulfhydryl group following chemical or enzymatic inactivation. Biochemistry. 1980 Sep 16;19(19):4471–4478. [PubMed] [Google Scholar]
  16. Kovacsovics T., Tschopp J., Kress A., Isliker H. Antibody-independent activation of C1, the first component of complement, by cardiolipin. J Immunol. 1985 Oct;135(4):2695–2700. [PubMed] [Google Scholar]
  17. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  18. Maclean D., Fishbein M. C., Braunwald E., Maroko P. R. Long-term preservation of ischemic myocardium after experimental coronary artery occlusion. J Clin Invest. 1978 Mar;61(3):541–551. doi: 10.1172/JCI108965. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Maroko P. R., Carpenter C. B., Chiariello M., Fishbein M. C., Radvany P., Knostman J. D., Hale S. L. Reduction by cobra venom factor of myocardial necrosis after coronary artery occlusion. J Clin Invest. 1978 Mar;61(3):661–670. doi: 10.1172/JCI108978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Nath K. A., Hostetter M. K., Hostetter T. H. Pathophysiology of chronic tubulo-interstitial disease in rats. Interactions of dietary acid load, ammonia, and complement component C3. J Clin Invest. 1985 Aug;76(2):667–675. doi: 10.1172/JCI112020. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Parks R. E., Jr, Crabtree G. W., Kong C. M., Agarwal R. P., Agarwal K. C., Scholar E. M. Incorporation of analog purine nucleosides into the formed elements of human blood: erythrocytes, platelets, and lymphocytes. Ann N Y Acad Sci. 1975 Aug 8;255:412–434. doi: 10.1111/j.1749-6632.1975.tb29249.x. [DOI] [PubMed] [Google Scholar]
  22. Peitsch M. C., Kovacsovics T. J., Tschopp J., Isliker H. Antibody-independent activation of C1. II. Evidence for two classes of nonimmune activators of the classical pathway of complement. J Immunol. 1987 Mar 15;138(6):1871–1876. [PubMed] [Google Scholar]
  23. Pick E., Mizel D. Rapid microassays for the measurement of superoxide and hydrogen peroxide production by macrophages in culture using an automatic enzyme immunoassay reader. J Immunol Methods. 1981;46(2):211–226. doi: 10.1016/0022-1759(81)90138-1. [DOI] [PubMed] [Google Scholar]
  24. Pinckard R. N., O'Rourke R. A., Crawford M. H., Grover F. S., McManus L. M., Ghidoni J. J., Storrs S. B., Olson M. S. Complement localization and mediation of ischemic injury in baboon myocardium. J Clin Invest. 1980 Nov;66(5):1050–1056. doi: 10.1172/JCI109933. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Pinckard R. N., Olson M. S., Giclas P. C., Terry R., Boyer J. T., O'Rourke R. A. Consumption of classical complement components by heart subcellular membranes in vitro and in patients after acute myocardial infarction. J Clin Invest. 1975 Sep;56(3):740–750. doi: 10.1172/JCI108145. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Rogler-Brown T., Agarwal R. P., Parks R. E., Jr Tight binding inhibitors--VI. Interactions of deoxycoformycin and adenosine deaminase in intact human erythrocytes and sarcoma 180 cells. Biochem Pharmacol. 1978;27(19):2289–2296. doi: 10.1016/0006-2952(78)90133-8. [DOI] [PubMed] [Google Scholar]
  27. Rubio R., Berne R. M., Katori M. Release of adenosine in reactive hyperemia of the dog heart. Am J Physiol. 1969 Jan;216(1):56–62. doi: 10.1152/ajplegacy.1969.216.1.56. [DOI] [PubMed] [Google Scholar]
  28. Rubio R., Berne R. M. Release of adenosine by the normal myocardium in dogs and its relationship to the regulation of coronary resistance. Circ Res. 1969 Oct;25(4):407–415. doi: 10.1161/01.res.25.4.407. [DOI] [PubMed] [Google Scholar]
  29. Schreiber R. D., Pangburn M. K., Müller-Eberhard H. J. C3 modified at the thiolester site: acquisition of reactivity with cellular C3b receptors. Biosci Rep. 1981 Nov;1(11):873–880. doi: 10.1007/BF01114821. [DOI] [PubMed] [Google Scholar]
  30. Simpson P. J., Todd R. F., 3rd, Fantone J. C., Mickelson J. K., Griffin J. D., Lucchesi B. R. Reduction of experimental canine myocardial reperfusion injury by a monoclonal antibody (anti-Mo1, anti-CD11b) that inhibits leukocyte adhesion. J Clin Invest. 1988 Feb;81(2):624–629. doi: 10.1172/JCI113364. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Spiers A. S., Moore D., Cassileth P. A., Harrington D. P., Cummings F. J., Neiman R. S., Bennett J. M., O'Connell M. J. Remissions in hairy-cell leukemia with pentostatin (2'-deoxycoformycin). N Engl J Med. 1987 Apr 2;316(14):825–830. doi: 10.1056/NEJM198704023161401. [DOI] [PubMed] [Google Scholar]
  32. Steinbuch M., Audran R. The isolation of IgG from mammalian sera with the aid of caprylic acid. Arch Biochem Biophys. 1969 Nov;134(2):279–284. doi: 10.1016/0003-9861(69)90285-9. [DOI] [PubMed] [Google Scholar]
  33. Tack B. F., Harrison R. A., Janatova J., Thomas M. L., Prahl J. W. Evidence for presence of an internal thiolester bond in third component of human complement. Proc Natl Acad Sci U S A. 1980 Oct;77(10):5764–5768. doi: 10.1073/pnas.77.10.5764. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Tolins J. P., Hostetter M. K., Hostetter T. H. Hypokalemic nephropathy in the rat. Role of ammonia in chronic tubular injury. J Clin Invest. 1987 May;79(5):1447–1458. doi: 10.1172/JCI112973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Van Belle H. Uptake and deamination of adenosine by blood. Species differences, effect of pH, ions, temperature and metabolic inhibitors. Biochim Biophys Acta. 1969 Oct 7;192(1):124–132. doi: 10.1016/0304-4165(69)90017-8. [DOI] [PubMed] [Google Scholar]
  36. Webster R. O., Henson P. M. Rapid micromeasurement of neutrophil exocytosis. Inflammation. 1978 Jun;3(2):129–135. doi: 10.1007/BF00910734. [DOI] [PubMed] [Google Scholar]
  37. von Zabern I., Nolte R., Vogt W. Treatment of human complement components C4 and C3 with amines or chaotropic ions. Evidence of a functional and structural change that provides uncleaved C4 and C3 with properties of their soluble activated froms, C4b and C3b. Scand J Immunol. 1981;13(5):413–431. doi: 10.1111/j.1365-3083.1981.tb00152.x. [DOI] [PubMed] [Google Scholar]

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

RESOURCES