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. 1997 Dec 15;100(12):3173–3183. doi: 10.1172/JCI119873

Following the fate of individual T cells throughout activation and clonal expansion. Signals from T cell receptor and CD28 differentially regulate the induction and duration of a proliferative response.

A D Wells 1, H Gudmundsdottir 1, L A Turka 1
PMCID: PMC508531  PMID: 9399965

Abstract

A detailed understanding of the effects of costimulatory signals on primary T cell expansion has been limited by experimental approaches that measure the bulk response of a cell population, without distinguishing responses of individual cells. Here, we have labeled live T cells in vitro with a stable, fluorescent dye that segregates equally between daughter cells upon cell division, allowing the proliferative history of any T cell present or generated during a response to be monitored over time. This system permits simultaneous evaluation of T cell surface markers, allowing concomitant assessment of cellular activation and quantitative determination of T cell receptor (TCR) occupancy on individual cells. Through this approach, we find that TCR engagement primarily regulates the frequency of T cells that enter the proliferative pool, but has relatively little effect on the number of times these cells will ultimately divide. In contrast, CD28-costimulation regulates both the frequency of responding cells (particularly at sub-maximal levels of TCR engagement), and more prominently, the number of mitotic events that responding cells undergo. When CD28-stimulation is blocked, provision of IL-2 restores the frequency of responding cells and the normal pattern of mitotic progression, indicating that the other CD28-induced genes are not required for this effect. An unexpected finding was that even at maximal levels of TCR engagement and CD28-mediated costimulation, only 50-60% of the original T cells in culture can be induced to divide. The nondividing cells are heterogeneous for naive versus memory markers, suggesting a more complex relationship between expression of memory markers and the ability to be recruited into the dividing pool. From these studies, we conclude that a stringent checkpoint regulates the participation of activated T cells in clonal expansion, with TCR and CD28 signals having both overlapping and differential effects on the induction and maintenance of T cell responses.

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Selected References

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  1. Boise L. H., González-García M., Postema C. E., Ding L., Lindsten T., Turka L. A., Mao X., Nuñez G., Thompson C. B. bcl-x, a bcl-2-related gene that functions as a dominant regulator of apoptotic cell death. Cell. 1993 Aug 27;74(4):597–608. doi: 10.1016/0092-8674(93)90508-n. [DOI] [PubMed] [Google Scholar]
  2. Boise L. H., Minn A. J., Noel P. J., June C. H., Accavitti M. A., Lindsten T., Thompson C. B. CD28 costimulation can promote T cell survival by enhancing the expression of Bcl-XL. Immunity. 1995 Jul;3(1):87–98. doi: 10.1016/1074-7613(95)90161-2. [DOI] [PubMed] [Google Scholar]
  3. Cai Z., Kishimoto H., Brunmark A., Jackson M. R., Peterson P. A., Sprent J. Requirements for peptide-induced T cell receptor downregulation on naive CD8+ T cells. J Exp Med. 1997 Feb 17;185(4):641–651. doi: 10.1084/jem.185.4.641. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chen L., Linsley P. S., Hellström K. E. Costimulation of T cells for tumor immunity. Immunol Today. 1993 Oct;14(10):483–486. doi: 10.1016/0167-5699(93)90262-J. [DOI] [PubMed] [Google Scholar]
  5. Croft M., Bradley L. M., Swain S. L. Naive versus memory CD4 T cell response to antigen. Memory cells are less dependent on accessory cell costimulation and can respond to many antigen-presenting cell types including resting B cells. J Immunol. 1994 Mar 15;152(6):2675–2685. [PubMed] [Google Scholar]
  6. Cross A. H., Girard T. J., Giacoletto K. S., Evans R. J., Keeling R. M., Lin R. F., Trotter J. L., Karr R. W. Long-term inhibition of murine experimental autoimmune encephalomyelitis using CTLA-4-Fc supports a key role for CD28 costimulation. J Clin Invest. 1995 Jun;95(6):2783–2789. doi: 10.1172/JCI117982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Green J. M., Noel P. J., Sperling A. I., Walunas T. L., Gray G. S., Bluestone J. A., Thompson C. B. Absence of B7-dependent responses in CD28-deficient mice. Immunity. 1994 Sep;1(6):501–508. doi: 10.1016/1074-7613(94)90092-2. [DOI] [PubMed] [Google Scholar]
  8. Harding F. A., McArthur J. G., Gross J. A., Raulet D. H., Allison J. P. CD28-mediated signalling co-stimulates murine T cells and prevents induction of anergy in T-cell clones. Nature. 1992 Apr 16;356(6370):607–609. doi: 10.1038/356607a0. [DOI] [PubMed] [Google Scholar]
  9. Janeway C. A., Jr, Bottomly K. Signals and signs for lymphocyte responses. Cell. 1994 Jan 28;76(2):275–285. doi: 10.1016/0092-8674(94)90335-2. [DOI] [PubMed] [Google Scholar]
  10. Jenkins M. K., Schwartz R. H. Antigen presentation by chemically modified splenocytes induces antigen-specific T cell unresponsiveness in vitro and in vivo. J Exp Med. 1987 Feb 1;165(2):302–319. doi: 10.1084/jem.165.2.302. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. June C. H., Bluestone J. A., Nadler L. M., Thompson C. B. The B7 and CD28 receptor families. Immunol Today. 1994 Jul;15(7):321–331. doi: 10.1016/0167-5699(94)90080-9. [DOI] [PubMed] [Google Scholar]
  12. Krangel M. S. Endocytosis and recycling of the T3-T cell receptor complex. The role of T3 phosphorylation. J Exp Med. 1987 Apr 1;165(4):1141–1159. doi: 10.1084/jem.165.4.1141. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kündig T. M., Shahinian A., Kawai K., Mittrücker H. W., Sebzda E., Bachmann M. F., Mak T. W., Ohashi P. S. Duration of TCR stimulation determines costimulatory requirement of T cells. Immunity. 1996 Jul;5(1):41–52. doi: 10.1016/s1074-7613(00)80308-8. [DOI] [PubMed] [Google Scholar]
  14. Lenschow D. J., Ho S. C., Sattar H., Rhee L., Gray G., Nabavi N., Herold K. C., Bluestone J. A. Differential effects of anti-B7-1 and anti-B7-2 monoclonal antibody treatment on the development of diabetes in the nonobese diabetic mouse. J Exp Med. 1995 Mar 1;181(3):1145–1155. doi: 10.1084/jem.181.3.1145. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Lenschow D. J., Walunas T. L., Bluestone J. A. CD28/B7 system of T cell costimulation. Annu Rev Immunol. 1996;14:233–258. doi: 10.1146/annurev.immunol.14.1.233. [DOI] [PubMed] [Google Scholar]
  16. Lenschow D. J., Zeng Y., Thistlethwaite J. R., Montag A., Brady W., Gibson M. G., Linsley P. S., Bluestone J. A. Long-term survival of xenogeneic pancreatic islet grafts induced by CTLA4lg. Science. 1992 Aug 7;257(5071):789–792. doi: 10.1126/science.1323143. [DOI] [PubMed] [Google Scholar]
  17. Lin H., Bolling S. F., Linsley P. S., Wei R. Q., Gordon D., Thompson C. B., Turka L. A. Long-term acceptance of major histocompatibility complex mismatched cardiac allografts induced by CTLA4Ig plus donor-specific transfusion. J Exp Med. 1993 Nov 1;178(5):1801–1806. doi: 10.1084/jem.178.5.1801. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Linsley P. S., Brady W., Urnes M., Grosmaire L. S., Damle N. K., Ledbetter J. A. CTLA-4 is a second receptor for the B cell activation antigen B7. J Exp Med. 1991 Sep 1;174(3):561–569. doi: 10.1084/jem.174.3.561. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Linsley P. S., Ledbetter J. A. The role of the CD28 receptor during T cell responses to antigen. Annu Rev Immunol. 1993;11:191–212. doi: 10.1146/annurev.iy.11.040193.001203. [DOI] [PubMed] [Google Scholar]
  20. Liu Y., Linsley P. S. Costimulation of T-cell growth. Curr Opin Immunol. 1992 Jun;4(3):265–270. doi: 10.1016/0952-7915(92)90075-p. [DOI] [PubMed] [Google Scholar]
  21. Lucas P. J., Negishi I., Nakayama K., Fields L. E., Loh D. Y. Naive CD28-deficient T cells can initiate but not sustain an in vitro antigen-specific immune response. J Immunol. 1995 Jun 1;154(11):5757–5768. [PubMed] [Google Scholar]
  22. Lyons A. B., Parish C. R. Determination of lymphocyte division by flow cytometry. J Immunol Methods. 1994 May 2;171(1):131–137. doi: 10.1016/0022-1759(94)90236-4. [DOI] [PubMed] [Google Scholar]
  23. Murphy K. M., Heimberger A. B., Loh D. Y. Induction by antigen of intrathymic apoptosis of CD4+CD8+TCRlo thymocytes in vivo. Science. 1990 Dec 21;250(4988):1720–1723. doi: 10.1126/science.2125367. [DOI] [PubMed] [Google Scholar]
  24. Perrin P. J., Scott D., Quigley L., Albert P. S., Feder O., Gray G. S., Abe R., June C. H., Racke M. K. Role of B7:CD28/CTLA-4 in the induction of chronic relapsing experimental allergic encephalomyelitis. J Immunol. 1995 Feb 1;154(3):1481–1490. [PubMed] [Google Scholar]
  25. Sloan-Lancaster J., Allen P. M. Altered peptide ligand-induced partial T cell activation: molecular mechanisms and role in T cell biology. Annu Rev Immunol. 1996;14:1–27. doi: 10.1146/annurev.immunol.14.1.1. [DOI] [PubMed] [Google Scholar]
  26. Sperling A. I., Auger J. A., Ehst B. D., Rulifson I. C., Thompson C. B., Bluestone J. A. CD28/B7 interactions deliver a unique signal to naive T cells that regulates cell survival but not early proliferation. J Immunol. 1996 Nov 1;157(9):3909–3917. [PubMed] [Google Scholar]
  27. Thompson C. B., Lindsten T., Ledbetter J. A., Kunkel S. L., Young H. A., Emerson S. G., Leiden J. M., June C. H. CD28 activation pathway regulates the production of multiple T-cell-derived lymphokines/cytokines. Proc Natl Acad Sci U S A. 1989 Feb;86(4):1333–1337. doi: 10.1073/pnas.86.4.1333. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Tough D. F., Sprent J. Turnover of naive- and memory-phenotype T cells. J Exp Med. 1994 Apr 1;179(4):1127–1135. doi: 10.1084/jem.179.4.1127. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Turka L. A., Linsley P. S., Lin H., Brady W., Leiden J. M., Wei R. Q., Gibson M. L., Zheng X. G., Myrdal S., Gordon D. T-cell activation by the CD28 ligand B7 is required for cardiac allograft rejection in vivo. Proc Natl Acad Sci U S A. 1992 Nov 15;89(22):11102–11105. doi: 10.1073/pnas.89.22.11102. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Viola A., Lanzavecchia A. T cell activation determined by T cell receptor number and tunable thresholds. Science. 1996 Jul 5;273(5271):104–106. doi: 10.1126/science.273.5271.104. [DOI] [PubMed] [Google Scholar]
  31. Weston S. A., Parish C. R. New fluorescent dyes for lymphocyte migration studies. Analysis by flow cytometry and fluorescence microscopy. J Immunol Methods. 1990 Oct 4;133(1):87–97. doi: 10.1016/0022-1759(90)90322-m. [DOI] [PubMed] [Google Scholar]

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