Skip to main content
NCBI home page
Clinical and Experimental Immunology logoLink to Clinical and Experimental Immunology
. 1994 Aug;97(2):266–272. doi: 10.1111/j.1365-2249.1994.tb06079.x

Flow cytometric analysis of the stimulatory response of T cell subsets from normal and HIV-1+ individuals to various mitogenic stimuli in vitro.

E Medina 1, N Borthwick 1, M A Johnson 1, S Miller 1, M Bofill 1
PMCID: PMC1534693  PMID: 7914156

Abstract

A novel technique is described which allows the study of the responses of T cell subpopulations stimulated in bulk cultures without interfering with cell-cell interactions. The number and phenotype of lymphoblasts developing following stimulation with phytohaemagglutinin (PHA), anti-CD3, staphylococcal protein A (SPA) and pokeweed mitogen (PWM) was determined in HIV-1- and HIV-1+ patients using a new five-parameter flow cytometric method. We found that normal T cells responded faster to PHA than to any of the other mitogens tested. The peak of the PHA response occurred on day 3, followed by anti-CD3 and SPA on day 4 and PWM mitogen on day 5. Although PHA and anti-CD3 stimulated up to 95% and 80% of lymphocytes, respectively, SPA and PWM stimulated only 40% and 30% of cells, respectively. A defective T cell response was observed in lymphocytes cultured from asymptomatic HIV-1+ patients compared with negative controls. This loss of response was related to a selective mortality of T cells following mitogenic stimulation, referred to as activation-associated lymphocyte death (AALD). The results showed that stronger mitogens (PHA and anti-CD3) induced AALD in a larger proportion (50-60%) of T cells than weaker mitogens such as SPA and PWM (30-40%), and that AALD affected different lymphocyte subsets to different extents. AALD occurred more frequently in total CD8+ and CD45RO+ T cells compared with CD4+ and CD45RA+ T cells, but memory CD4+ T cells were the population most severely affected in samples from HIV-1+ donors.

Full text

PDF
266

Selected References

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

  1. Adriaansen H. J., Osman C., van Dongen J. J., Wijdenes-de Bresser J. H., Kappetijn-van Tilborg C. M., Hooijkaas H. Immunological marker analysis of mitogen-induced proliferating lymphocytes using BrdU incorporation or screening of metaphases. Staphylococcal protein A is a potent mitogen for CD4+ lymphocytes. Scand J Immunol. 1990 Dec;32(6):687–694. doi: 10.1111/j.1365-3083.1990.tb03211.x. [DOI] [PubMed] [Google Scholar]
  2. Akbar A. N., Salmon M., Janossy G. The synergy between naive and memory T cells during activation. Immunol Today. 1991 Jun;12(6):184–188. doi: 10.1016/0167-5699(91)90050-4. [DOI] [PubMed] [Google Scholar]
  3. Ameisen J. C., Capron A. Cell dysfunction and depletion in AIDS: the programmed cell death hypothesis. Immunol Today. 1991 Apr;12(4):102–105. doi: 10.1016/0167-5699(91)90092-8. [DOI] [PubMed] [Google Scholar]
  4. Creemers P. C. Determination of co-expression of activation antigens on proliferating CD4+, CD4+ CD8+ and CD8+ lymphocyte subsets by dual parameter flow cytometry. J Immunol Methods. 1987 Mar 12;97(2):165–171. doi: 10.1016/0022-1759(87)90456-x. [DOI] [PubMed] [Google Scholar]
  5. Ferrer J. M., Plaza A., Kreisler M., Díaz-Espada F. Differential interleukin secretion by in vitro activated human CD45RA and CD45RO CD4+ T cell subsets. Cell Immunol. 1992 Apr 15;141(1):10–20. doi: 10.1016/0008-8749(92)90123-7. [DOI] [PubMed] [Google Scholar]
  6. Geppert T. D., Lipsky P. E. Immobilized anti-CD3-induced T cell growth: comparison of the frequency of responding cells within various T cell subsets. Cell Immunol. 1991 Mar;133(1):206–218. doi: 10.1016/0008-8749(91)90192-e. [DOI] [PubMed] [Google Scholar]
  7. Gruters R. A., Terpstra F. G., De Jong R., Van Noesel C. J., Van Lier R. A., Miedema F. Selective loss of T cell functions in different stages of HIV infection. Early loss of anti-CD3-induced T cell proliferation followed by decreased anti-CD3-induced cytotoxic T lymphocyte generation in AIDS-related complex and AIDS. Eur J Immunol. 1990 May;20(5):1039–1044. doi: 10.1002/eji.1830200514. [DOI] [PubMed] [Google Scholar]
  8. Gruters R. A., Terpstra F. G., Lange J. M., Roos M. T., Harkema T., Mulder J. W., De Wolf F., Schellekens P. T., Miedema F. Differences in clinical course in zidovudine-treated asymptomatic HIV-infected men associated with T-cell function at intake. AIDS. 1991 Jan;5(1):43–47. doi: 10.1097/00002030-199101000-00006. [DOI] [PubMed] [Google Scholar]
  9. Hofmann B., Jakobsen K. D., Odum N., Dickmeiss E., Platz P., Ryder L. P., Pedersen C., Mathiesen L., Bygbjerg I. B., Faber V. HIV-induced immunodeficiency. Relatively preserved phytohemagglutinin as opposed to decreased pokeweed mitogen responses may be due to possibly preserved responses via CD2/phytohemagglutinin pathway. J Immunol. 1989 Mar 15;142(6):1874–1880. [PubMed] [Google Scholar]
  10. Hofmann B., Lindhardt B. O., Gerstoft J., Petersen C. S., Platz P., Ryder L. P., Odum N., Dickmeiss E., Nielsen P. B., Ullman S. Lymphocyte transformation response to pokeweed mitogen as a predictive marker for development of AIDS and AIDS related symptoms in homosexual men with HIV antibodies. Br Med J (Clin Res Ed) 1987 Aug 1;295(6593):293–296. doi: 10.1136/bmj.295.6593.293. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Holter W., Majdic O., Liszka K., Stockinger H., Knapp W. Kinetics of activation antigen expression by in vitro-stimulated human T lymphocytes. Cell Immunol. 1985 Feb;90(2):322–330. doi: 10.1016/0008-8749(85)90197-2. [DOI] [PubMed] [Google Scholar]
  12. Jacob M. C., Rose M. L. Staphylococcal protein-A is a potent T-cell mitogen of tonsillar lymphocytes. Immunol Lett. 1983;7(2):77–80. doi: 10.1016/0165-2478(83)90037-8. [DOI] [PubMed] [Google Scholar]
  13. Janeway C. A., Jr, Yagi J., Conrad P. J., Katz M. E., Jones B., Vroegop S., Buxser S. T-cell responses to Mls and to bacterial proteins that mimic its behavior. Immunol Rev. 1989 Feb;107:61–88. doi: 10.1111/j.1600-065x.1989.tb00003.x. [DOI] [PubMed] [Google Scholar]
  14. Janossy G., Borthwick N., Lomnitzer R., Medina E., Squire S. B., Phillips A. N., Lipman M., Johnson M. A., Lee C., Bofill M. Lymphocyte activation in HIV-1 infection. I. Predominant proliferative defects among CD45R0+ cells of the CD4 and CD8 lineages. AIDS. 1993 May;7(5):613–624. doi: 10.1097/00002030-199305000-00002. [DOI] [PubMed] [Google Scholar]
  15. Janossy G., Gomez de la Concha E., Luquetti A., Snajdr M. J., Waxdal M. J., Platts-Mills T. A. T-cell regulation of immunoglobulin synthesis and proliferation in pokeweed (Pa-1)-stimulated human lymphocyte cultures. Scand J Immunol. 1977;6(1-2):109–123. doi: 10.1111/j.1365-3083.1977.tb00326.x. [DOI] [PubMed] [Google Scholar]
  16. Leca G., Boumsell L., Fabbi M., Reinherz E. L., Kanellopoulos J. M. The sheep erythrocyte receptor and both alpha and beta chains of the human T-lymphocyte antigen receptor bind the mitogenic lectin (phytohaemagglutinin) from Phaseolus vulgaris. Scand J Immunol. 1986 May;23(5):535–544. doi: 10.1111/j.1365-3083.1986.tb01985.x. [DOI] [PubMed] [Google Scholar]
  17. Li Y., Hui H., Burgess C. J., Price R. W., Sharp P. M., Hahn B. H., Shaw G. M. Complete nucleotide sequence, genome organization, and biological properties of human immunodeficiency virus type 1 in vivo: evidence for limited defectiveness and complementation. J Virol. 1992 Nov;66(11):6587–6600. doi: 10.1128/jvi.66.11.6587-6600.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. O'Flynn K., Krensky A. M., Beverley P. C., Burakoff S. J., Linch D. C. Phytohaemagglutinin activation of T cells through the sheep red blood cell receptor. Nature. 1985 Feb 21;313(6004):686–687. doi: 10.1038/313686a0. [DOI] [PubMed] [Google Scholar]
  19. Pantaleo G., Koenig S., Baseler M., Lane H. C., Fauci A. S. Defective clonogenic potential of CD8+ T lymphocytes in patients with AIDS. Expansion in vivo of a nonclonogenic CD3+CD8+DR+CD25- T cell population. J Immunol. 1990 Mar 1;144(5):1696–1704. [PubMed] [Google Scholar]
  20. Prince H. E., Jensen E. R. HIV-related alterations in CD8 cell subsets defined by in vitro survival characteristics. Cell Immunol. 1991 May;134(2):276–286. doi: 10.1016/0008-8749(91)90302-r. [DOI] [PubMed] [Google Scholar]
  21. Prince H. E., John J. K. Early activation marker expression to detect impaired proliferative responses to pokeweed mitogen and tetanus toxoid: studies in patients with AIDS and related disorders. Diagn Immunol. 1986;4(6):306–311. [PubMed] [Google Scholar]
  22. Schellekens P. T., Roos M. T., De Wolf F., Lange J. M., Miedema F. Low T-cell responsiveness to activation via CD3/TCR is a prognostic marker for acquired immunodeficiency syndrome (AIDS) in human immunodeficiency virus-1 (HIV-1)-infected men. J Clin Immunol. 1990 Mar;10(2):121–127. doi: 10.1007/BF00918194. [DOI] [PubMed] [Google Scholar]
  23. Schnittman S. M., Lane H. C., Greenhouse J., Justement J. S., Baseler M., Fauci A. S. Preferential infection of CD4+ memory T cells by human immunodeficiency virus type 1: evidence for a role in the selective T-cell functional defects observed in infected individuals. Proc Natl Acad Sci U S A. 1990 Aug;87(16):6058–6062. doi: 10.1073/pnas.87.16.6058. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Valentine M. A., Tsoukas C. D., Rhodes G., Vaughan J. H., Carson D. A. Phytohemagglutinin binds to the 20-kDa molecule of the T3 complex. Eur J Immunol. 1985 Aug;15(8):851–854. doi: 10.1002/eji.1830150821. [DOI] [PubMed] [Google Scholar]
  25. Van Wauwe J. P., De Mey J. R., Goossens J. G. OKT3: a monoclonal anti-human T lymphocyte antibody with potent mitogenic properties. J Immunol. 1980 Jun;124(6):2708–2713. [PubMed] [Google Scholar]
  26. White J., Herman A., Pullen A. M., Kubo R., Kappler J. W., Marrack P. The V beta-specific superantigen staphylococcal enterotoxin B: stimulation of mature T cells and clonal deletion in neonatal mice. Cell. 1989 Jan 13;56(1):27–35. doi: 10.1016/0092-8674(89)90980-x. [DOI] [PubMed] [Google Scholar]

Articles from Clinical and Experimental Immunology are provided here courtesy of British Society for Immunology

RESOURCES