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. 1996 Aug 1;184(2):473–483. doi: 10.1084/jem.184.2.473

Differentiation and stability of T helper 1 and 2 cells derived from naive human neonatal CD4+ T cells, analyzed at the single-cell level

PMCID: PMC2192741  PMID: 8760801

Abstract

The development of CD4+ T helper (Th) type 1 and 2 cells is essential for the eradication of pathogens, but can also be responsible for various pathological disorders. Therefore, modulation of Th cell differentiation may have clinical utility in the treatment of human disease. Here, we show that interleukin (IL) 12 and IL-4 directly induce human neonatal CD4- T cells, activated via CD3 and CD28, to differentiate into Th1 and Th2 subsets. In contrast, IL-13, which shares many biological activities with IL-4, failed to induce T cell differentiation, consistent with the observation that human T cells do not express IL-13 receptors. Both the IL-12-induced Th1 subset and the IL-4-induced Th2 subset produce large quantities of IL-10, confirming that human IL-10 is not a typical human Th2 cytokine. Interestingly, IL- 4-driven Th2 cell differentiation was completely prevented by an IL-4 mutant protein (IL-4.Y124D), indicating that this molecule acts as a strong IL-4 receptor antagonist. Analysis of single T cells producing interferon gamma or IL-4 revealed that induction of Th1 cell differentiation occurred rapidly and required only 4 d of priming of the neonatal CD4+ T cells in the presence of IL-12. The IL-12-induced Th1 cell phenotype was stable and was not significantly affected when repeatedly stimulated in the presence of recombinant IL-4. In contrast, the differentiation of Th2 cells occurred slowly and required not only 6 d of priming, but also additional restimulation of the primed CD4+ T cells in the presence of IL-4. Moreover, IL-4-induced Th2 cell phenotypes were not stable and could rapidly be reverted into a population predominantly containing Th0 and Th1 cells, after a single restimulation in the presence of IL-12. The observed differences in stability of IL-12- and IL-4-induced human Th1 and Th2 subsets, respectively, may have implications for cytokine-based therapies of chronic disease.

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Selected References

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  1. Andersson U., Andersson J., Lindfors A., Wagner K., Möller G., Heusser C. H. Simultaneous production of interleukin 2, interleukin 4 and interferon-gamma by activated human blood lymphocytes. Eur J Immunol. 1990 Jul;20(7):1591–1596. doi: 10.1002/eji.1830200727. [DOI] [PubMed] [Google Scholar]
  2. Assenmacher M., Schmitz J., Radbruch A. Flow cytometric determination of cytokines in activated murine T helper lymphocytes: expression of interleukin-10 in interferon-gamma and in interleukin-4-expressing cells. Eur J Immunol. 1994 May;24(5):1097–1101. doi: 10.1002/eji.1830240513. [DOI] [PubMed] [Google Scholar]
  3. Aversa G., Punnonen J., Cocks B. G., de Waal Malefyt R., Vega F., Jr, Zurawski S. M., Zurawski G., de Vries J. E. An interleukin 4 (IL-4) mutant protein inhibits both IL-4 or IL-13-induced human immunoglobulin G4 (IgG4) and IgE synthesis and B cell proliferation: support for a common component shared by IL-4 and IL-13 receptors. J Exp Med. 1993 Dec 1;178(6):2213–2218. doi: 10.1084/jem.178.6.2213. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Azuma M., Cayabyab M., Phillips J. H., Lanier L. L. Requirements for CD28-dependent T cell-mediated cytotoxicity. J Immunol. 1993 Mar 15;150(6):2091–2101. [PubMed] [Google Scholar]
  5. Carballido J. M., Schols D., Namikawa R., Zurawski S., Zurawski G., Roncarolo M. G., de Vries J. E. IL-4 induces human B cell maturation and IgE synthesis in SCID-hu mice. Inhibition of ongoing IgE production by in vivo treatment with an IL-4/IL-13 receptor antagonist. J Immunol. 1995 Nov 1;155(9):4162–4170. [PubMed] [Google Scholar]
  6. Chatelain R., Varkila K., Coffman R. L. IL-4 induces a Th2 response in Leishmania major-infected mice. J Immunol. 1992 Feb 15;148(4):1182–1187. [PubMed] [Google Scholar]
  7. Chrétien I., Van Kimmenade A., Pearce M. K., Banchereau J., Abrams J. S. Development of polyclonal and monoclonal antibodies for immunoassay and neutralization of human interleukin-4. J Immunol Methods. 1989 Feb 8;117(1):67–81. doi: 10.1016/0022-1759(89)90120-8. [DOI] [PubMed] [Google Scholar]
  8. Cocks B. G., Chang C. C., Carballido J. M., Yssel H., de Vries J. E., Aversa G. A novel receptor involved in T-cell activation. Nature. 1995 Jul 20;376(6537):260–263. doi: 10.1038/376260a0. [DOI] [PubMed] [Google Scholar]
  9. Del Prete G. F., De Carli M., Mastromauro C., Biagiotti R., Macchia D., Falagiani P., Ricci M., Romagnani S. Purified protein derivative of Mycobacterium tuberculosis and excretory-secretory antigen(s) of Toxocara canis expand in vitro human T cells with stable and opposite (type 1 T helper or type 2 T helper) profile of cytokine production. J Clin Invest. 1991 Jul;88(1):346–350. doi: 10.1172/JCI115300. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Demeure C. E., Wu C. Y., Shu U., Schneider P. V., Heusser C., Yssel H., Delespesse G. In vitro maturation of human neonatal CD4 T lymphocytes. II. Cytokines present at priming modulate the development of lymphokine production. J Immunol. 1994 May 15;152(10):4775–4782. [PubMed] [Google Scholar]
  11. Ehlers S., Smith K. A. Differentiation of T cell lymphokine gene expression: the in vitro acquisition of T cell memory. J Exp Med. 1991 Jan 1;173(1):25–36. doi: 10.1084/jem.173.1.25. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Favre C., Wijdenes J., Cabrillat H., Djossou O., Banchereau J., de Vries J. E. Epitope mapping of recombinant human gamma interferon using monoclonal antibodies. Mol Immunol. 1989 Jan;26(1):17–25. doi: 10.1016/0161-5890(89)90015-1. [DOI] [PubMed] [Google Scholar]
  13. Firestein G. S., Roeder W. D., Laxer J. A., Townsend K. S., Weaver C. T., Hom J. T., Linton J., Torbett B. E., Glasebrook A. L. A new murine CD4+ T cell subset with an unrestricted cytokine profile. J Immunol. 1989 Jul 15;143(2):518–525. [PubMed] [Google Scholar]
  14. Freeman G. J., Boussiotis V. A., Anumanthan A., Bernstein G. M., Ke X. Y., Rennert P. D., Gray G. S., Gribben J. G., Nadler L. M. B7-1 and B7-2 do not deliver identical costimulatory signals, since B7-2 but not B7-1 preferentially costimulates the initial production of IL-4. Immunity. 1995 May;2(5):523–532. doi: 10.1016/1074-7613(95)90032-2. [DOI] [PubMed] [Google Scholar]
  15. Haanen J. B., de Waal Malefijt R., Res P. C., Kraakman E. M., Ottenhoff T. H., de Vries R. R., Spits H. Selection of a human T helper type 1-like T cell subset by mycobacteria. J Exp Med. 1991 Sep 1;174(3):583–592. doi: 10.1084/jem.174.3.583. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Heinzel F. P., Sadick M. D., Holaday B. J., Coffman R. L., Locksley R. M. Reciprocal expression of interferon gamma or interleukin 4 during the resolution or progression of murine leishmaniasis. Evidence for expansion of distinct helper T cell subsets. J Exp Med. 1989 Jan 1;169(1):59–72. doi: 10.1084/jem.169.1.59. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hodes R. J. Molecular alterations in the aging immune system. J Exp Med. 1995 Jul 1;182(1):1–3. doi: 10.1084/jem.182.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hsieh C. S., Heimberger A. B., Gold J. S., O'Garra A., Murphy K. M. Differential regulation of T helper phenotype development by interleukins 4 and 10 in an alpha beta T-cell-receptor transgenic system. Proc Natl Acad Sci U S A. 1992 Jul 1;89(13):6065–6069. doi: 10.1073/pnas.89.13.6065. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hsieh C. S., Macatonia S. E., Tripp C. S., Wolf S. F., O'Garra A., Murphy K. M. Development of TH1 CD4+ T cells through IL-12 produced by Listeria-induced macrophages. Science. 1993 Apr 23;260(5107):547–549. doi: 10.1126/science.8097338. [DOI] [PubMed] [Google Scholar]
  20. Jeannin P., Delneste Y., Seveso M., Life P., Bonnefoy J. Y. IL-12 synergizes with IL-2 and other stimuli in inducing IL-10 production by human T cells. J Immunol. 1996 May 1;156(9):3159–3165. [PubMed] [Google Scholar]
  21. Jung T., Schauer U., Heusser C., Neumann C., Rieger C. Detection of intracellular cytokines by flow cytometry. J Immunol Methods. 1993 Feb 26;159(1-2):197–207. doi: 10.1016/0022-1759(93)90158-4. [DOI] [PubMed] [Google Scholar]
  22. Kamogawa Y., Minasi L. A., Carding S. R., Bottomly K., Flavell R. A. The relationship of IL-4- and IFN gamma-producing T cells studied by lineage ablation of IL-4-producing cells. Cell. 1993 Dec 3;75(5):985–995. doi: 10.1016/0092-8674(93)90542-x. [DOI] [PubMed] [Google Scholar]
  23. Kelso A., Groves P., Troutt A. B., Francis K. Evidence for the stochastic acquisition of cytokine profile by CD4+ T cells activated in a T helper type 2-like response in vivo. Eur J Immunol. 1995 May;25(5):1168–1175. doi: 10.1002/eji.1830250506. [DOI] [PubMed] [Google Scholar]
  24. Kuchroo V. K., Das M. P., Brown J. A., Ranger A. M., Zamvil S. S., Sobel R. A., Weiner H. L., Nabavi N., Glimcher L. H. B7-1 and B7-2 costimulatory molecules activate differentially the Th1/Th2 developmental pathways: application to autoimmune disease therapy. Cell. 1995 Mar 10;80(5):707–718. doi: 10.1016/0092-8674(95)90349-6. [DOI] [PubMed] [Google Scholar]
  25. Lanier L. L., O'Fallon S., Somoza C., Phillips J. H., Linsley P. S., Okumura K., Ito D., Azuma M. CD80 (B7) and CD86 (B70) provide similar costimulatory signals for T cell proliferation, cytokine production, and generation of CTL. J Immunol. 1995 Jan 1;154(1):97–105. [PubMed] [Google Scholar]
  26. Le Gros G., Ben-Sasson S. Z., Seder R., Finkelman F. D., Paul W. E. Generation of interleukin 4 (IL-4)-producing cells in vivo and in vitro: IL-2 and IL-4 are required for in vitro generation of IL-4-producing cells. J Exp Med. 1990 Sep 1;172(3):921–929. doi: 10.1084/jem.172.3.921. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Levine B. L., Ueda Y., Craighead N., Huang M. L., June C. H. CD28 ligands CD80 (B7-1) and CD86 (B7-2) induce long-term autocrine growth of CD4+ T cells and induce similar patterns of cytokine secretion in vitro. Int Immunol. 1995 Jun;7(6):891–904. doi: 10.1093/intimm/7.6.891. [DOI] [PubMed] [Google Scholar]
  28. Maggi E., Del Prete G., Macchia D., Parronchi P., Tiri A., Chrétien I., Ricci M., Romagnani S. Profiles of lymphokine activities and helper function for IgE in human T cell clones. Eur J Immunol. 1988 Jul;18(7):1045–1050. doi: 10.1002/eji.1830180712. [DOI] [PubMed] [Google Scholar]
  29. Maggi E., Parronchi P., Manetti R., Simonelli C., Piccinni M. P., Rugiu F. S., De Carli M., Ricci M., Romagnani S. Reciprocal regulatory effects of IFN-gamma and IL-4 on the in vitro development of human Th1 and Th2 clones. J Immunol. 1992 Apr 1;148(7):2142–2147. [PubMed] [Google Scholar]
  30. Manetti R., Gerosa F., Giudizi M. G., Biagiotti R., Parronchi P., Piccinni M. P., Sampognaro S., Maggi E., Romagnani S., Trinchieri G. Interleukin 12 induces stable priming for interferon gamma (IFN-gamma) production during differentiation of human T helper (Th) cells and transient IFN-gamma production in established Th2 cell clones. J Exp Med. 1994 Apr 1;179(4):1273–1283. doi: 10.1084/jem.179.4.1273. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Mocci S., Coffman R. L. Induction of a Th2 population from a polarized Leishmania-specific Th1 population by in vitro culture with IL-4. J Immunol. 1995 Apr 15;154(8):3779–3787. [PubMed] [Google Scholar]
  32. Mosmann T. R., Cherwinski H., Bond M. W., Giedlin M. A., Coffman R. L. Two types of murine helper T cell clone. I. Definition according to profiles of lymphokine activities and secreted proteins. J Immunol. 1986 Apr 1;136(7):2348–2357. [PubMed] [Google Scholar]
  33. Murphy E., Shibuya K., Hosken N., Openshaw P., Maino V., Davis K., Murphy K., O'Garra A. Reversibility of T helper 1 and 2 populations is lost after long-term stimulation. J Exp Med. 1996 Mar 1;183(3):901–913. doi: 10.1084/jem.183.3.901. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Nabors G. S., Afonso L. C., Farrell J. P., Scott P. Switch from a type 2 to a type 1 T helper cell response and cure of established Leishmania major infection in mice is induced by combined therapy with interleukin 12 and Pentostam. Proc Natl Acad Sci U S A. 1995 Apr 11;92(8):3142–3146. doi: 10.1073/pnas.92.8.3142. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Paliard X., de Waal Malefijt R., Yssel H., Blanchard D., Chrétien I., Abrams J., de Vries J., Spits H. Simultaneous production of IL-2, IL-4, and IFN-gamma by activated human CD4+ and CD8+ T cell clones. J Immunol. 1988 Aug 1;141(3):849–855. [PubMed] [Google Scholar]
  36. Peltz G. A., Trounstine M. L., Moore K. W. Cloned and expressed human Fc receptor for IgG mediates anti-CD3-dependent lymphoproliferation. J Immunol. 1988 Sep 15;141(6):1891–1896. [PubMed] [Google Scholar]
  37. Perez V. L., Lederer J. A., Lichtman A. H., Abbas A. K. Stability of Th1 and Th2 populations. Int Immunol. 1995 May;7(5):869–875. doi: 10.1093/intimm/7.5.869. [DOI] [PubMed] [Google Scholar]
  38. Picker L. J., Singh M. K., Zdraveski Z., Treer J. R., Waldrop S. L., Bergstresser P. R., Maino V. C. Direct demonstration of cytokine synthesis heterogeneity among human memory/effector T cells by flow cytometry. Blood. 1995 Aug 15;86(4):1408–1419. [PubMed] [Google Scholar]
  39. Seder R. A., Gazzinelli R., Sher A., Paul W. E. Interleukin 12 acts directly on CD4+ T cells to enhance priming for interferon gamma production and diminishes interleukin 4 inhibition of such priming. Proc Natl Acad Sci U S A. 1993 Nov 1;90(21):10188–10192. doi: 10.1073/pnas.90.21.10188. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Seder R. A., Paul W. E., Davis M. M., Fazekas de St Groth B. The presence of interleukin 4 during in vitro priming determines the lymphokine-producing potential of CD4+ T cells from T cell receptor transgenic mice. J Exp Med. 1992 Oct 1;176(4):1091–1098. doi: 10.1084/jem.176.4.1091. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Shu U., Demeure C. E., Byun D. G., Podlaski F., Stern A. S., Delespesse G. Interleukin 12 exerts a differential effect on the maturation of neonatal and adult human CD45R0- CD4 T cells. J Clin Invest. 1994 Oct;94(4):1352–1358. doi: 10.1172/JCI117469. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Spits H., Keizer G., Borst J., Terhorst C., Hekman A., de Vries J. E. Characterization of monoclonal antibodies against cell surface molecules associated with cytotoxic activity of natural and activated killer cells and cloned CTL lines. Hybridoma. 1983;2(4):423–437. doi: 10.1089/hyb.1983.2.423. [DOI] [PubMed] [Google Scholar]
  43. Swain S. L., Weinberg A. D., English M., Huston G. IL-4 directs the development of Th2-like helper effectors. J Immunol. 1990 Dec 1;145(11):3796–3806. [PubMed] [Google Scholar]
  44. Szabo S. J., Jacobson N. G., Dighe A. S., Gubler U., Murphy K. M. Developmental commitment to the Th2 lineage by extinction of IL-12 signaling. Immunity. 1995 Jun;2(6):665–675. doi: 10.1016/1074-7613(95)90011-x. [DOI] [PubMed] [Google Scholar]
  45. Taylor S., Bryson Y. J. Impaired production of gamma-interferon by newborn cells in vitro is due to a functionally immature macrophage. J Immunol. 1985 Mar;134(3):1493–1497. [PubMed] [Google Scholar]
  46. Wierenga E. A., Snoek M., de Groot C., Chrétien I., Bos J. D., Jansen H. M., Kapsenberg M. L. Evidence for compartmentalization of functional subsets of CD2+ T lymphocytes in atopic patients. J Immunol. 1990 Jun 15;144(12):4651–4656. [PubMed] [Google Scholar]
  47. Yssel H., De Waal Malefyt R., Roncarolo M. G., Abrams J. S., Lahesmaa R., Spits H., de Vries J. E. IL-10 is produced by subsets of human CD4+ T cell clones and peripheral blood T cells. J Immunol. 1992 Oct 1;149(7):2378–2384. [PubMed] [Google Scholar]
  48. Yssel H., Fasler S., de Vries J. E., de Waal Malefyt R. IL-12 transiently induces IFN-gamma transcription and protein synthesis in human CD4+ allergen-specific Th2 T cell clones. Int Immunol. 1994 Jul;6(7):1091–1096. doi: 10.1093/intimm/6.7.1091. [DOI] [PubMed] [Google Scholar]
  49. Yssel H., Johnson K. E., Schneider P. V., Wideman J., Terr A., Kastelein R., De Vries J. E. T cell activation-inducing epitopes of the house dust mite allergen Der p I. Proliferation and lymphokine production patterns by Der p I-specific CD4+ T cell clones. J Immunol. 1992 Feb 1;148(3):738–745. [PubMed] [Google Scholar]
  50. Yssel H., Shanafelt M. C., Soderberg C., Schneider P. V., Anzola J., Peltz G. Borrelia burgdorferi activates a T helper type 1-like T cell subset in Lyme arthritis. J Exp Med. 1991 Sep 1;174(3):593–601. doi: 10.1084/jem.174.3.593. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Zurawski S. M., Vega F., Jr, Huyghe B., Zurawski G. Receptors for interleukin-13 and interleukin-4 are complex and share a novel component that functions in signal transduction. EMBO J. 1993 Jul;12(7):2663–2670. doi: 10.1002/j.1460-2075.1993.tb05927.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. de Waal Malefyt R., Abrams J. S., Zurawski S. M., Lecron J. C., Mohan-Peterson S., Sanjanwala B., Bennett B., Silver J., de Vries J. E., Yssel H. Differential regulation of IL-13 and IL-4 production by human CD8+ and CD4+ Th0, Th1 and Th2 T cell clones and EBV-transformed B cells. Int Immunol. 1995 Sep;7(9):1405–1416. doi: 10.1093/intimm/7.9.1405. [DOI] [PubMed] [Google Scholar]
  53. de Waal Malefyt R., Figdor C. G., Huijbens R., Mohan-Peterson S., Bennett B., Culpepper J., Dang W., Zurawski G., de Vries J. E. Effects of IL-13 on phenotype, cytokine production, and cytotoxic function of human monocytes. Comparison with IL-4 and modulation by IFN-gamma or IL-10. J Immunol. 1993 Dec 1;151(11):6370–6381. [PubMed] [Google Scholar]
  54. de Waal Malefyt R., Verma S., Bejarano M. T., Ranes-Goldberg M., Hill M., Spits H. CD2/LFA-3 or LFA-1/ICAM-1 but not CD28/B7 interactions can augment cytotoxicity by virus-specific CD8+ cytotoxic T lymphocytes. Eur J Immunol. 1993 Feb;23(2):418–424. doi: 10.1002/eji.1830230218. [DOI] [PubMed] [Google Scholar]

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