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. 1979 Oct 1;150(4):938–949. doi: 10.1084/jem.150.4.938

Macrophage oxygen-dependent antimicrobial activity. I. Susceptibility of Toxoplasma gondii to oxygen intermediates

PMCID: PMC2185675  PMID: 92521

Abstract

A sensitive method for evaluating extracellular parasite viability was used to determine the in vitro susceptibility of virulent Toxoplasma gondii to selected oxygen intermediates. By acridine orange fluorescent staining criteria, toxoplasmas were resistant to up to either 10(-3) M reagent H2O2 or H2O2 generated by glucose-glucose oxidase. In keeping with a lack of sensitivity to H2O2, toxoplasmas contained endogenous catalase (5.7 x 10(-4) Baudhuin units/10(6) organisms). The addition of a peroxidase and halide, however, markedly accelerated killing and lowered the H2O2 requirement by 1,000-fold. In contrast, toxoplasmas were promptly killed after exposure to products generated by xanthine (1.5 x 10(-4) M) and xanthine oxidase (50 micrograms). The inhibition of this system's microbicidal activity by scavengers of O2- (superoxide dismutase) and H2O2 (catalase) indicated that although neither O2- nor H2O2 were toxoplasmacidal, their interaction was required for parasite killing. Quenching OH. and 1O2, presumed products of O2--H2O2 interaction, by mannitol, benzoate, diazabicyclooctane, and histidine, also inhibited toxoplasma killing by xanthine-xanthine oxidase. These findings suggested that O2- and H2O2 functioned in precursor roles and that OH. and 1O2 were toxoplasmacidal. The capacity of normal peritoneal macrophages to pinocytose an oxygen intermediate scavenger, soluble catalase, was also demonstrated. Appreciable extraphagosomal concentrations of catalase were achieved by exposing macrophages to 1 mg/ml of the enzyme for 3 h. Maintenance of high intracellular levels required constant exposure because interiorized catalase was rapidly degraded.

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Selected References

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  1. Abuchowski A., McCoy J. R., Palczuk N. C., van Es T., Davis F. F. Effect of covalent attachment of polyethylene glycol on immunogenicity and circulating life of bovine liver catalase. J Biol Chem. 1977 Jun 10;252(11):3582–3586. [PubMed] [Google Scholar]
  2. Anderson S. E., Jr, Remington J. S. Effect of normal and activated human macrophages on Toxoplasma gondii. J Exp Med. 1974 May 1;139(5):1154–1174. doi: 10.1084/jem.139.5.1154. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Babior B. M., Curnutte J. T., Kipnes R. S. Biological defense mechanisms. Evidence for the participation of superoxide in bacterial killing by xanthine oxidase. J Lab Clin Med. 1975 Feb;85(2):235–244. [PubMed] [Google Scholar]
  4. Babior B. M. Oxygen-dependent microbial killing by phagocytes (second of two parts). N Engl J Med. 1978 Mar 30;298(13):721–725. doi: 10.1056/NEJM197803302981305. [DOI] [PubMed] [Google Scholar]
  5. Baudhuin P., Beaufay H., Rahman-Li Y., Sellinger O. Z., Wattiaux R., Jacques P., De Duve C. Tissue fractionation studies. 17. Intracellular distribution of monoamine oxidase, aspartate aminotransferase, alanine aminotransferase, D-amino acid oxidase and catalase in rat-liver tissue. Biochem J. 1964 Jul;92(1):179–184. doi: 10.1042/bj0920179. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Beauchamp C., Fridovich I. A mechanism for the production of ethylene from methional. The generation of the hydroxyl radical by xanthine oxidase. J Biol Chem. 1970 Sep 25;245(18):4641–4646. [PubMed] [Google Scholar]
  7. Borges J. S., Johnson W. D., Jr Inhibition of multiplication of Toxoplasma gondii by human monocytes exposed to T-lymphocyte products. J Exp Med. 1975 Feb 1;141(2):483–496. doi: 10.1084/jem.141.2.483. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Boveris A., Martino E., Stoppani A. O. Evaluation of the horseradish peroxidase-scopoletin method for the measurement of hydrogen peroxide formation in biological systems. Anal Biochem. 1977 May 15;80(1):145–158. doi: 10.1016/0003-2697(77)90634-0. [DOI] [PubMed] [Google Scholar]
  9. Boveris A., Stoppani A. O. Hydrogen peroxide generation in Trypanosoma cruzi. Experientia. 1977 Oct 15;33(10):1306–1308. doi: 10.1007/BF01920148. [DOI] [PubMed] [Google Scholar]
  10. COHEN G., HOCHSTEIN P. GENERATION OF HYDROGEN PEROXIDE IN ERYTHROCYTES BY HEMOLYTIC AGENTS. Biochemistry. 1964 Jul;3:895–900. doi: 10.1021/bi00895a006. [DOI] [PubMed] [Google Scholar]
  11. COHN Z. A., BENSON B. THE DIFFERENTIATION OF MONONUCLEAR PHAGOCYTES. MORPHOLOGY, CYTOCHEMISTRY, AND BIOCHEMISTRY. J Exp Med. 1965 Jan 1;121:153–170. doi: 10.1084/jem.121.1.153. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Docampo R., de Boiso J. F., Boveris A., Stoppani A. O. Localization of peroxidase activity in Trypanosoma cruzi microbodies. Experientia. 1976 Aug 15;32(8):972–975. doi: 10.1007/BF01933918. [DOI] [PubMed] [Google Scholar]
  13. Drath D. B., Karnovsky M. L. Superoxide production by phagocytic leukocytes. J Exp Med. 1975 Jan 1;141(1):257–262. doi: 10.1084/jem.141.1.257. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. FULTON J. D., SPOONER D. F. Metabolic studies on Toxoplasma gondii. Exp Parasitol. 1960 Jun;9:293–301. doi: 10.1016/0014-4894(60)90037-0. [DOI] [PubMed] [Google Scholar]
  15. FULTON J. D., SPOONER D. F. Terminal respiration in certain mammalian trypanosomes. Exp Parasitol. 1959 Apr;8(2):137–162. doi: 10.1016/0014-4894(59)90014-1. [DOI] [PubMed] [Google Scholar]
  16. Fridovich I. Superoxide dismutases. Adv Enzymol Relat Areas Mol Biol. 1974;41(0):35–97. doi: 10.1002/9780470122860.ch2. [DOI] [PubMed] [Google Scholar]
  17. Johnson W. D., Jr, Mei B., Cohn Z. A. The separation, long-term cultivation, and maturation of the human monocyte. J Exp Med. 1977 Dec 1;146(6):1613–1626. doi: 10.1084/jem.146.6.1613. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Johnston R. B., Jr, Godzik C. A., Cohn Z. A. Increased superoxide anion production by immunologically activated and chemically elicited macrophages. J Exp Med. 1978 Jul 1;148(1):115–127. doi: 10.1084/jem.148.1.115. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Johnston R. B., Jr, Keele B. B., Jr, Misra H. P., Lehmeyer J. E., Webb L. S., Baehner R. L., RaJagopalan K. V. The role of superoxide anion generation in phagocytic bactericidal activity. Studies with normal and chronic granulomatous disease leukocytes. J Clin Invest. 1975 Jun;55(6):1357–1372. doi: 10.1172/JCI108055. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Jones T. C., Hirsch J. G. The interaction between Toxoplasma gondii and mammalian cells. II. The absence of lysosomal fusion with phagocytic vacuoles containing living parasites. J Exp Med. 1972 Nov 1;136(5):1173–1194. doi: 10.1084/jem.136.5.1173. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Jones T. C., Len L., Hirsch J. G. Assessment in vitro of immunity against Toxoplasma gondii. J Exp Med. 1975 Feb 1;141(2):466–482. doi: 10.1084/jem.141.2.466. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jones T. C., Yeh S., Hirsch J. G. The interaction between Toxoplasma gondii and mammalian cells. I. Mechanism of entry and intracellular fate of the parasite. J Exp Med. 1972 Nov 1;136(5):1157–1172. doi: 10.1084/jem.136.5.1157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kellogg E. W., 3rd, Fridovich I. Superoxide, hydrogen peroxide, and singlet oxygen in lipid peroxidation by a xanthine oxidase system. J Biol Chem. 1975 Nov 25;250(22):8812–8817. [PubMed] [Google Scholar]
  24. Klebanoff S. J. Antimicrobial activity of catalase at acid pH. Proc Soc Exp Biol Med. 1969 Nov;132(2):571–574. doi: 10.3181/00379727-132-34263. [DOI] [PubMed] [Google Scholar]
  25. Klebanoff S. J. Antimicrobial mechanisms in neutrophilic polymorphonuclear leukocytes. Semin Hematol. 1975 Apr;12(2):117–142. [PubMed] [Google Scholar]
  26. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  27. Lynch R. E., Fridovich I. Effects of superoxide on the erythrocyte membrane. J Biol Chem. 1978 Mar 25;253(6):1838–1845. [PubMed] [Google Scholar]
  28. MACKANESS G. B. Cellular resistance to infection. J Exp Med. 1962 Sep 1;116:381–406. doi: 10.1084/jem.116.3.381. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Mandell G. L. Catalase, superoxide dismutase, and virulence of Staphylococcus aureus. In vitro and in vivo studies with emphasis on staphylococcal--leukocyte interaction. J Clin Invest. 1975 Mar;55(3):561–566. doi: 10.1172/JCI107963. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Marklund S. Tryptic digestion and alkaline denaturation of catalase. The influence on catalatic activity, peroxidatic activity towards phenolic compounds, and the reactivity with methyl- and ethyl-hydroperoxide. Biochim Biophys Acta. 1973 Sep 15;321(1):90–97. doi: 10.1016/0005-2744(73)90062-4. [DOI] [PubMed] [Google Scholar]
  31. Mauel J., Buchmüller Y., Behin R. Studies on the mechanisms of macrophage activation. I. Destruction of intracellular Leishmania enriettii in macrophages activated by cocultivation with stimulated lymphocytes. J Exp Med. 1978 Aug 1;148(2):393–407. doi: 10.1084/jem.148.2.393. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. McLeod R., Remington J. S. Studies on the specificity of killing of intracellular pathogens by macrophages. Cell Immunol. 1977 Nov;34(1):156–174. doi: 10.1016/0008-8749(77)90238-6. [DOI] [PubMed] [Google Scholar]
  33. Nathan C. F., Root R. K. Hydrogen peroxide release from mouse peritoneal macrophages: dependence on sequential activation and triggering. J Exp Med. 1977 Dec 1;146(6):1648–1662. doi: 10.1084/jem.146.6.1648. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Nathan C., Nogueira N., Juangbhanich C., Ellis J., Cohn Z. Activation of macrophages in vivo and in vitro. Correlation between hydrogen peroxide release and killing of Trypanosoma cruzi. J Exp Med. 1979 May 1;149(5):1056–1068. doi: 10.1084/jem.149.5.1056. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Nogueira N., Cohn Z. A. Trypanosoma cruzi: in vitro induction of macrophage microbicidal activity. J Exp Med. 1978 Jul 1;148(1):288–300. doi: 10.1084/jem.148.1.288. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Norrby R., Lindholm L., Lycke E. Lysosomes of Toxoplasma gondii and their possible relation to the host-cell penetration of toxoplasma parasites. J Bacteriol. 1968 Oct;96(4):916–919. doi: 10.1128/jb.96.4.916-919.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. North R. J. Suppression of cell-mediated immunity to infection by an antimitotic drug. Further evidence that migrant macrophages express immunity. J Exp Med. 1970 Sep 1;132(3):535–545. doi: 10.1084/jem.132.3.535. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. North R. J. The concept of the activated macrophage. J Immunol. 1978 Sep;121(3):806–809. [PMC free article] [PubMed] [Google Scholar]
  39. Novikoff A. B., Goldfischer S. Visualization of peroxisomes (microbodies) and mitochondria with diaminobenzidine. J Histochem Cytochem. 1969 Oct;17(10):675–680. doi: 10.1177/17.10.675. [DOI] [PubMed] [Google Scholar]
  40. Reiss M., Roos D. Differences in oxygen metabolism of phagocytosing monocytes and neutrophils. J Clin Invest. 1978 Feb;61(2):480–488. doi: 10.1172/JCI108959. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Rosen H., Klebanoff S. J. Bactericidal activity of a superoxide anion-generating system. A model for the polymorphonuclear leukocyte. J Exp Med. 1979 Jan 1;149(1):27–39. doi: 10.1084/jem.149.1.27. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Simmons S. R., Karnovsky M. L. Iodinating ability of various leukocytes and their bactericidal activity. J Exp Med. 1973 Jul 1;138(1):44–63. doi: 10.1084/jem.138.1.44. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Steinman R. M., Cohn Z. A. The interaction of soluble horseradish peroxidase with mouse peritoneal macrophages in vitro. J Cell Biol. 1972 Oct;55(1):186–204. doi: 10.1083/jcb.55.1.186. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Weiss S. J., King G. W., LoBuglio A. F. Evidence for hydroxyl radical generation by human Monocytes. J Clin Invest. 1977 Aug;60(2):370–373. doi: 10.1172/JCI108785. [DOI] [PMC free article] [PubMed] [Google Scholar]

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