Skip to main content
PMC home page
Infection and Immunity logoLink to Infection and Immunity
. 1997 Dec;65(12):5262–5271. doi: 10.1128/iai.65.12.5262-5271.1997

Apoptosis of human monocytes and macrophages by Mycobacterium avium sonicate.

T Hayashi 1, A Catanzaro 1, S P Rao 1
PMCID: PMC175758  PMID: 9393825

Abstract

Mycobacterium avium is an intracellular organism which multiplies predominantly within human macrophages. This organism has previously been shown to induce apoptosis in human macrophages. With a view to identifying M. avium components that induce cell death in infected host cells, sonicated extracts of M. avium as well as individual components isolated from the M. avium sonicate were tested in various assays with a human monocytic cell line (THP-1). THP-1 cells incubated with M. avium sonicate showed significantly reduced viability after a 2-day exposure compared to control cells incubated with media alone. This effect was dose dependent, with only 6.6% +/- 5.2% and 48.8% +/- 10.3% of the cells being viable by trypan blue exclusion at 600 and 300 microg/ml, respectively. Control cells, on the other hand, exhibited a viability of 98.8% +/- 1.0%. In addition, an 80% ammonium sulfate fraction of the M. avium sonicate and the previously characterized 68-kDa protein were found to have similar effects on THP-1 cells. In both cases, the reduction in viability was due to apoptosis characterized by chromatin condensation, DNA fragmentation by agarose gel electrophoresis, or terminal deoxynucleotidyl transferase-mediated d-UTP nick end labeling (TUNEL) and release of nuclear matrix protein (NMP) into the culture medium. M. avium sonicate-induced apoptosis of THP-1 cells was completely inhibited by the commonly used antioxidants pyrrolidinedithiocarbamate (PDTC) and butylated hydroxyanisole (BHA), indicating that the generation of free oxygen radicals may be responsible for inducing cell death. M. avium sonicate was found to induce apoptosis of monocyte-derived macrophages (MDMs) as well. This effect was not reversed in the presence of PDTC and was not accompanied with DNA fragmentation when determined by agarose gel electrophoresis, as seen in the case of THP-1 cells. However, these MDMs were found to contain fragmented DNA by TUNEL. These findings suggest that the mechanism of cell death in MDMs may be different from that observed with THP-1 cells. Furthermore, these results provide new insight into the effect of M. avium components on host cell responses during M. avium infection.

Full Text

The Full Text of this article is available as a PDF (1.1 MB).

Selected References

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

  1. Arends M. J., Wyllie A. H. Apoptosis: mechanisms and roles in pathology. Int Rev Exp Pathol. 1991;32:223–254. doi: 10.1016/b978-0-12-364932-4.50010-1. [DOI] [PubMed] [Google Scholar]
  2. Bingisser R., Stey C., Weller M., Groscurth P., Russi E., Frei K. Apoptosis in human alveolar macrophages is induced by endotoxin and is modulated by cytokines. Am J Respir Cell Mol Biol. 1996 Jul;15(1):64–70. doi: 10.1165/ajrcmb.15.1.8679223. [DOI] [PubMed] [Google Scholar]
  3. Buttke T. M., Sandstrom P. A. Oxidative stress as a mediator of apoptosis. Immunol Today. 1994 Jan;15(1):7–10. doi: 10.1016/0167-5699(94)90018-3. [DOI] [PubMed] [Google Scholar]
  4. Chen Y., Smith M. R., Thirumalai K., Zychlinsky A. A bacterial invasin induces macrophage apoptosis by binding directly to ICE. EMBO J. 1996 Aug 1;15(15):3853–3860. [PMC free article] [PubMed] [Google Scholar]
  5. Chen Y., Zychlinsky A. Apoptosis induced by bacterial pathogens. Microb Pathog. 1994 Oct;17(4):203–212. doi: 10.1006/mpat.1994.1066. [DOI] [PubMed] [Google Scholar]
  6. Cohen G. M., Sun X. M., Snowden R. T., Dinsdale D., Skilleter D. N. Key morphological features of apoptosis may occur in the absence of internucleosomal DNA fragmentation. Biochem J. 1992 Sep 1;286(Pt 2):331–334. doi: 10.1042/bj2860331. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Edwards D., Kirkpatrick C. H. The immunology of mycobacterial diseases. Am Rev Respir Dis. 1986 Nov;134(5):1062–1071. doi: 10.1164/arrd.1986.134.5.1062. [DOI] [PubMed] [Google Scholar]
  8. Fadok V. A., Savill J. S., Haslett C., Bratton D. L., Doherty D. E., Campbell P. A., Henson P. M. Different populations of macrophages use either the vitronectin receptor or the phosphatidylserine receptor to recognize and remove apoptotic cells. J Immunol. 1992 Dec 15;149(12):4029–4035. [PubMed] [Google Scholar]
  9. Fattorini L., Xiao Y., Li B., Santoro C., Ippoliti F., Orefici G. Induction of IL-1 beta, IL-6, TNF-alpha, GM-CSF and G-CSF in human macrophages by smooth transparent and smooth opaque colonial variants of Mycobacterium avium. J Med Microbiol. 1994 Feb;40(2):129–133. doi: 10.1099/00222615-40-2-129. [DOI] [PubMed] [Google Scholar]
  10. Gan H., Newman G. W., Remold H. G. Plasminogen activator inhibitor type 2 prevents programmed cell death of human macrophages infected with Mycobacterium avium, serovar 4. J Immunol. 1995 Aug 1;155(3):1304–1315. [PubMed] [Google Scholar]
  11. Hayashi T., Rao S. P., Catanzaro A. Binding of the 68-kilodalton protein of Mycobacterium avium to alpha(v)beta3 on human monocyte-derived macrophages enhances complement receptor type 3 expression. Infect Immun. 1997 Apr;65(4):1211–1216. doi: 10.1128/iai.65.4.1211-1216.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Johnson J. L., Shiratsuchi H., Toba H., Ellner J. J. Preservation of monocyte effector functions against Mycobacterium avium-M. intracellulare in patients with AIDS. Infect Immun. 1991 Oct;59(10):3639–3645. doi: 10.1128/iai.59.10.3639-3645.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Jonas D., Walev I., Berger T., Liebetrau M., Palmer M., Bhakdi S. Novel path to apoptosis: small transmembrane pores created by staphylococcal alpha-toxin in T lymphocytes evoke internucleosomal DNA degradation. Infect Immun. 1994 Apr;62(4):1304–1312. doi: 10.1128/iai.62.4.1304-1312.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Keane J., Balcewicz-Sablinska M. K., Remold H. G., Chupp G. L., Meek B. B., Fenton M. J., Kornfeld H. Infection by Mycobacterium tuberculosis promotes human alveolar macrophage apoptosis. Infect Immun. 1997 Jan;65(1):298–304. doi: 10.1128/iai.65.1.298-304.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Khelef N., Zychlinsky A., Guiso N. Bordetella pertussis induces apoptosis in macrophages: role of adenylate cyclase-hemolysin. Infect Immun. 1993 Oct;61(10):4064–4071. doi: 10.1128/iai.61.10.4064-4071.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kochi S. K., Collier R. J. DNA fragmentation and cytolysis in U937 cells treated with diphtheria toxin or other inhibitors of protein synthesis. Exp Cell Res. 1993 Sep;208(1):296–302. doi: 10.1006/excr.1993.1249. [DOI] [PubMed] [Google Scholar]
  17. Mahida Y. R., Makh S., Hyde S., Gray T., Borriello S. P. Effect of Clostridium difficile toxin A on human intestinal epithelial cells: induction of interleukin 8 production and apoptosis after cell detachment. Gut. 1996 Mar;38(3):337–347. doi: 10.1136/gut.38.3.337. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Mangan D. F., Taichman N. S., Lally E. T., Wahl S. M. Lethal effects of Actinobacillus actinomycetemcomitans leukotoxin on human T lymphocytes. Infect Immun. 1991 Sep;59(9):3267–3272. doi: 10.1128/iai.59.9.3267-3272.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Mangan D. F., Wahl S. M. Differential regulation of human monocyte programmed cell death (apoptosis) by chemotactic factors and pro-inflammatory cytokines. J Immunol. 1991 Nov 15;147(10):3408–3412. [PubMed] [Google Scholar]
  20. Merien F., Baranton G., Perolat P. Invasion of Vero cells and induction of apoptosis in macrophages by pathogenic Leptospira interrogans are correlated with virulence. Infect Immun. 1997 Feb;65(2):729–738. doi: 10.1128/iai.65.2.729-738.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Miller T., Beausang L. A., Meneghini M., Lidgard G. Death-induced changes to the nuclear matrix: the use of anti-nuclear matrix antibodies to study agents of apoptosis. Biotechniques. 1993 Dec;15(6):1042–1047. [PubMed] [Google Scholar]
  22. Moore K. J., Matlashewski G. Intracellular infection by Leishmania donovani inhibits macrophage apoptosis. J Immunol. 1994 Mar 15;152(6):2930–2937. [PubMed] [Google Scholar]
  23. Müller A., Hacker J., Brand B. C. Evidence for apoptosis of human macrophage-like HL-60 cells by Legionella pneumophila infection. Infect Immun. 1996 Dec;64(12):4900–4906. doi: 10.1128/iai.64.12.4900-4906.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Newman G. W., Gan H. X., McCarthy P. L., Jr, Remold H. G. Survival of human macrophages infected with Mycobacterium avium intracellulare correlates with increased production of tumor necrosis factor-alpha and IL-6. J Immunol. 1991 Dec 1;147(11):3942–3948. [PubMed] [Google Scholar]
  25. Oberhammer F., Wilson J. W., Dive C., Morris I. D., Hickman J. A., Wakeling A. E., Walker P. R., Sikorska M. Apoptotic death in epithelial cells: cleavage of DNA to 300 and/or 50 kb fragments prior to or in the absence of internucleosomal fragmentation. EMBO J. 1993 Sep;12(9):3679–3684. doi: 10.1002/j.1460-2075.1993.tb06042.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Rao S. P., Ogata K., Catanzaro A. Mycobacterium avium-M. intracellulare binds to the integrin receptor alpha v beta 3 on human monocytes and monocyte-derived macrophages. Infect Immun. 1993 Feb;61(2):663–670. doi: 10.1128/iai.61.2.663-670.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Rao S. P., Ogata K., Morris S. L., Catanzaro A. Identification of a 68 kd surface antigen of Mycobacterium avium that binds to human macrophages. J Lab Clin Med. 1994 Apr;123(4):526–535. [PubMed] [Google Scholar]
  28. Rao S. P., Ratnakar P., Catanzaro A. Macrophage release of tumor necrosis factor-alpha by Mycobacterium avium antigens. FEMS Immunol Med Microbiol. 1996 Aug;15(1):27–34. doi: 10.1111/j.1574-695X.1996.tb00355.x. [DOI] [PubMed] [Google Scholar]
  29. Rawadi G., Roman-Roman S., Castedo M., Dutilleul V., Susin S., Marchetti P., Geuskens M., Kroemer G. Effects of Mycoplasma fermentans on the myelomonocytic lineage. Different molecular entities with cytokine-inducing and cytocidal potential. J Immunol. 1996 Jan 15;156(2):670–678. [PubMed] [Google Scholar]
  30. Togashi H., Sasaki M., Frohman E., Taira E., Ratan R. R., Dawson T. M., Dawson V. L. Neuronal (type I) nitric oxide synthase regulates nuclear factor kappaB activity and immunologic (type II) nitric oxide synthase expression. Proc Natl Acad Sci U S A. 1997 Mar 18;94(6):2676–2680. doi: 10.1073/pnas.94.6.2676. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Verhaegen S., McGowan A. J., Brophy A. R., Fernandes R. S., Cotter T. G. Inhibition of apoptosis by antioxidants in the human HL-60 leukemia cell line. Biochem Pharmacol. 1995 Sep 28;50(7):1021–1029. doi: 10.1016/0006-2952(95)00233-p. [DOI] [PubMed] [Google Scholar]
  32. Wyllie A. H., Kerr J. F., Currie A. R. Cell death: the significance of apoptosis. Int Rev Cytol. 1980;68:251–306. doi: 10.1016/s0074-7696(08)62312-8. [DOI] [PubMed] [Google Scholar]
  33. Yanagisawa-Shiota F., Sakagami H., Kuribayashi N., Iida M., Sakagami T., Takeda M. Endonuclease activity and induction of DNA fragmentation in human myelogenous leukemic cell lines. Anticancer Res. 1995 Mar-Apr;15(2):259–265. [PubMed] [Google Scholar]
  34. Zakowski P., Fligiel S., Berlin G. W., Johnson L., Jr Disseminated Mycobacterium avium-intracellulare infection in homosexual men dying of acquired immunodeficiency. JAMA. 1982 Dec 10;248(22):2980–2982. doi: 10.1001/jama.1982.03330220024029. [DOI] [PubMed] [Google Scholar]
  35. Zhang Y., Doerfler M., Lee T. C., Guillemin B., Rom W. N. Mechanisms of stimulation of interleukin-1 beta and tumor necrosis factor-alpha by Mycobacterium tuberculosis components. J Clin Invest. 1993 May;91(5):2076–2083. doi: 10.1172/JCI116430. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Zychlinsky A., Prevost M. C., Sansonetti P. J. Shigella flexneri induces apoptosis in infected macrophages. Nature. 1992 Jul 9;358(6382):167–169. doi: 10.1038/358167a0. [DOI] [PubMed] [Google Scholar]
  37. Zychlinsky A. Programmed cell death in infectious diseases. Trends Microbiol. 1993 Jun;1(3):114–117. doi: 10.1016/0966-842x(93)90118-b. [DOI] [PubMed] [Google Scholar]

Articles from Infection and Immunity are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES