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. 1971 Sep 1;134(3):713–740. doi: 10.1084/jem.134.3.713

RESPONSE OF CULTURED MACROPHAGES TO MYCOBACTERIUM TUBERCULOSIS, WITH OBSERVATIONS ON FUSION OF LYSOSOMES WITH PHAGOSOMES

J A Armstrong 1, P D'Arcy Hart 1
PMCID: PMC2139093  PMID: 15776571

Abstract

The cytological response to the ingestion of tubercle bacilli by cultured mouse peritoneal macrophages has been studied by electron microscopy. Methods included a quantitative assessment based on systematic surveying of cell profiles, and of phagosomes and their contained bacteria, encountered in thin sections; classification of the sectioned bacteria into visibly damaged and apparently intact categories; prelabeling of dense granules (secondary lysosomes) with ferritin as an aid to identifying the occurrence and frequency of phagosome-lysosome fusion; and monitoring of bacterial growth and viability by light microscopy and cultural counts. The situations studied were as follows: progressive infection with the multiplying virulent strain H37Rv; ingestion of the same strain previously inactivated by gamma radiation; infection with an attenuated strain (BCG); and a stabilized virulent infection induced by the surfactant Macrocyclon. In the bacterial suspensions used routinely for inoculation, about half the bacilli were viable, matching closely the proportions of intact and damaged organisms identified with the electron microscope. In the inoculated macrophages, some phagosomes containing intact bacilli and others containing damaged bacilli were always to be found; but the proportion of organisms scored as damaged increased, and that of intact organisms decreased, in situations where the population as a whole had been rendered nonviable before inoculation, or where they became so intracellularly as in the late stages of a BCG infection. Evidence of fusion of ferritin-marked lysosomes with some bacterium-containing phagosomes was obtained in all experiments, but a significant difference was regularly observed according to whether the bacilli were damaged or intact. Virtually all phagosomes containing damaged bacilli showed signs of fusion; but when many phagosomes were present containing apparently intact organisms (as with actively multiplying strain H37Rv or with this strain held at a steady level of viability by Macrocyclon, and also with strain BCG at an early stage of that infection), signs of fusion of lysosomes with these phagosomes were infrequent. From these findings it is inferred that intracellular survival of M. tuberculosis in cultured macrophages is associated with a tendency to nonfusion of dense granules with the phagosome, thus avoiding direct exposure of the bacilli to the contents of these organelles. It is suggested, further, that fusion of dense granules with the phagosome, leading to digestion, is determined by recognition of the bacillus as nonviable. The possibility is discussed that the cytological response to different mycobacterial infections may reflect differences of a basic nature between facultative and obligate intracellular parasitism.

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Selected References

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  1. ALLEN J. M., BRIEGER E. M., REES R. J. ELECTRON MICROSCOPY OF THE HOST-CELL PARASITE RELATION IN MURINE LEPROSY. J Pathol Bacteriol. 1965 Jan;89:301–306. doi: 10.1002/path.1700890131. [DOI] [PubMed] [Google Scholar]
  2. Allison A. C., Hart P. D. Potentiation by silica of the growth of Mycobacterium tuberculosis in macrophage cultures. Br J Exp Pathol. 1968 Oct;49(5):465–476. [PMC free article] [PubMed] [Google Scholar]
  3. Aquino T. I., Skinsnes O. K. Pathobiologic significance of the subcellular organelles of lepra cells. Int J Lepr Other Mycobact Dis. 1970 Apr-Jun;38(2):134–148. [PubMed] [Google Scholar]
  4. Axline S. G., Cohn Z. A. In vitro induction of lysosomal enzymes by phagocytosis. J Exp Med. 1970 Jun 1;131(6):1239–1260. doi: 10.1084/jem.131.6.1239. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. BRUMFITT W., GLYNN A. A. Intracellular killing of Micrococcus lysodeikticus by macrophages and polymorphonuclear leucocytes: a comparative study. Br J Exp Pathol. 1961 Oct;42:408–423. [PMC free article] [PubMed] [Google Scholar]
  6. Blanden R. V. Modification of macrophage function. J Reticuloendothel Soc. 1968 Jun;5(3):179–202. [PubMed] [Google Scholar]
  7. Brown C. A., Draper P. An electron-microscope study of rat fibroblasts infected with Mycobacterium lepraemurium. J Pathol. 1970 Sep;102(1):21–26. doi: 10.1002/path.1711020105. [DOI] [PubMed] [Google Scholar]
  8. Brown C. A., Draper P., Hart P. D. Mycobacteria and lysosomes: a paradox. Nature. 1969 Feb 15;221(5181):658–660. doi: 10.1038/221658a0. [DOI] [PubMed] [Google Scholar]
  9. CHANG Y. T. LONG-TERM CULTIVATION OF MOUSE PERITONEAL MACROPHAGES. J Natl Cancer Inst. 1964 Jan;32:19–35. [PubMed] [Google Scholar]
  10. COHN Z. A. The fate of bacteria within phagocytic cells. I. The degradation of isotopically labeled bacteria by polymorphonuclear leucocytes and macrophages. J Exp Med. 1963 Jan 1;117:27–42. doi: 10.1084/jem.117.1.27. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Carr I. The fine structure of the mammalian lymphoreticular system. Int Rev Cytol. 1970;27:283–348. doi: 10.1016/s0074-7696(08)61249-8. [DOI] [PubMed] [Google Scholar]
  12. Cohn Z. A., Ehrenreich B. A. The uptake, storage, and intracellular hydrolysis of carbohydrates by macrophages. J Exp Med. 1969 Jan 1;129(1):201–225. doi: 10.1084/jem.129.1.201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Cohn Z. A., Fedorko M. E., Hirsch J. G. The in vitro differentiation of mononuclear phagocytes. V. The formation of macrophage lysosomes. J Exp Med. 1966 Apr 1;123(4):757–766. doi: 10.1084/jem.123.4.757. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Cohn Z. A., Hirsch J. G., Fedorko M. E. The in vitro differentiation of mononuclear phagocytes. IV. The ultrastructure of macrophage differentiation in the peritoneal cavity and in culture. J Exp Med. 1966 Apr 1;123(4):747–756. doi: 10.1084/jem.123.4.747. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Draper P., Rees R. J. Electron-transparent zone of mycobacteria may be a defence mechanism. Nature. 1970 Nov 28;228(5274):860–861. doi: 10.1038/228860a0. [DOI] [PubMed] [Google Scholar]
  16. Dumont A., Robert A. Electron microscopic study of phagocytosis of Histoplasma capsulatum by hamster peritoneal macrophages. Lab Invest. 1970 Sep;23(3):278–286. [PubMed] [Google Scholar]
  17. Dumont A., Sheldon H. Changes in the fine structure of macrophages in experimentally produced tuberculous granulomas in hamsters. Lab Invest. 1965 Nov;14(11):2034–2055. [PubMed] [Google Scholar]
  18. Ericsson J. L. Studies on induced cellular autophagy. I. Electron microscopy of cells with in vivo labelled lysosomes. Exp Cell Res. 1969 Apr;55(1):95–106. doi: 10.1016/0014-4827(69)90462-5. [DOI] [PubMed] [Google Scholar]
  19. Fildes C., Hart P. D. Use of a detergent-acid mixture to rupture tissue culture cells. Nature. 1967 May 6;214(5088):624–625. doi: 10.1038/214624a0. [DOI] [PubMed] [Google Scholar]
  20. HART P. D., REES R. J. Effect of macrocyclon in acute and chronic pulmonary tuberculous infection in mice as shown by viable and total bacterial counts. Br J Exp Pathol. 1960 Aug;41:414–421. [PMC free article] [PubMed] [Google Scholar]
  21. Hart P. D. Mycobacterium tuberculosis in macrophages: effect of certain surfactants and other membrane-active compounds. Science. 1968 Nov 8;162(3854):686–689. doi: 10.1126/science.162.3854.686. [DOI] [PubMed] [Google Scholar]
  22. Hirsch J. G., Fedorko M. E. Ultrastructure of human leukocytes after simultaneous fixation with glutaraldehyde and osmium tetroxide and "postfixation" in uranyl acetate. J Cell Biol. 1968 Sep;38(3):615–627. doi: 10.1083/jcb.38.3.615. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Horn R. G., Koenig M. G., Goodman J. S., Collins R. D. Phagocytosis of Staphylococcus aureus by hepatic reticuloendothelial cells. An ultrastructural study. Lab Invest. 1969 Nov;21(5):406–414. [PubMed] [Google Scholar]
  24. Imaeda T. Electron microscopy. Approach to leprosy research. Int J Lepr. 1965 Jul-Sep;33(3 Suppl):669–688. [PubMed] [Google Scholar]
  25. MERCKX J. J., BROWN A. L., Jr, KARLSON A. G. AN ELECTRON-MICROSCOPIC STUDY OF EXPERIMENTAL INFECTIONS WITH ACID-FAST BACILLI. Am Rev Respir Dis. 1964 Apr;89:485–496. doi: 10.1164/arrd.1964.89.4.485. [DOI] [PubMed] [Google Scholar]
  26. Trouet A. Lysosomes et infection. Brux Med. 1966 Nov 20;46(47):1139–1144. [PubMed] [Google Scholar]

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