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Journal of General Virology header logo Journal of General Virology

Volume 85, Issue 4

Proliferation and differentiation in isogenic populations of peripheral B cells activated by Epstein–Barr virus or T cell-derived mitogens No Access

Abstract

Human B cells isolated from peripheral blood were activated and induced to proliferate by either Epstein–Barr virus (EBV) or the T cell-derived mitogens CD40 ligand (CD40L) plus interleukin (IL)-4. Although both populations initially proliferated as B-blasts, significant differences were revealed over a longer period. EBV infection resulted in continuously proliferating lymphoblastoid cell lines (LCLs), whereas most of the CD40L/IL-4-stimulated B cells had a finite proliferative lifespan of 3–4 weeks. Cell cycle analysis, trypan blue staining and Western blot analysis for cleavage of poly(ADP-ribose) polymerase (PARP) all demonstrated that the decrease in proliferation in CD40L/IL-4-stimulated B cells is not due to cell death. Instead, these cells arrest, accumulate in G/G and undergo alterations in cell surface marker expression, cellular morphology and immunoglobulin production, all consistent with plasmacytoid differentiation. In contrast, B cells infected with EBV continued to proliferate and retained a blast-like phenotype. Differences in both cytokine production and the expression of cell cycle regulators were identified between the two B-cell populations, which might contribute to the differentiation of the CD40L/IL-4-stimulated B cells and suggest potential mechanisms by which EBV may overcome this. The study has also identified a window of opportunity during which a comparison of isogenic populations of EBV- and mitogen-driven B blasts can be made.

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2004-04-01
2026-02-09

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References

  1. Agematsu K. 2000; Memory B cells and CD27. Histol Histopathol 15:573–576
     [Google Scholar]
  2. Allday M. J., Sinclair A., Parker G., Crawford D. H., Farrell P. J. 1995; Epstein–Barr virus efficiently immortalizes human B cells without neutralizing the function of p53. EMBO J 14:1382–1391
     [Google Scholar]
  3. Altmeyer A., Simmons R. C., Krajewski S., Reed J. C., Bornkamm G. W., Chen-Kiang S. 1997; Reversal of EBV immortalization precedes apoptosis in IL-6-induced human B cell terminal differentiation. Immunity 7:667–677
     [Google Scholar]
  4. Aman P., Ehlin-Henriksson B., Klein G. 1984; Epstein–Barr virus susceptibility of normal human B lymphocyte populations. J Exp Med 159:208–220
     [Google Scholar]
  5. Armitage R. J., Fanslow W. C., Strockbine L. other authors 1992; Molecular and biological characterization of a murine ligand for CD40. Nature 357:80–82
     [Google Scholar]
  6. Arpin C., Dechanet J., Van Kooten C., Merville P., Grouard G., Briere F., Banchereau J., Liu Y. J. 1995; Generation of memory B cells and plasma cells in vitro. Science 268:720–722
     [Google Scholar]
  7. Banchereau J., Rousset F. 1991; Growing human B lymphocytes in the CD40 system. Nature 353:678–679
     [Google Scholar]
  8. Banchereau J., de Paoli P., Valle A., Garcia E., Rousset F. 1991; Long-term human B cell lines dependent on interleukin-4 and antibody to CD40. Science 251:70–72
     [Google Scholar]
  9. Burdin N., Van Kooten C., Galibert L., Abrams J. S., Wijdenes J., Banchereau J., Rousset F. 1995; Endogenous IL-6 and IL-10 contribute to the differentiation of CD40-activated human B lymphocytes. J Immunol 154:2533–2544
     [Google Scholar]
  10. Burdin N., Rousset F., Banchereau J. 1997; B-cell-derived IL-10: production and function. Methods Enzymol 11:98–111
     [Google Scholar]
  11. Caldwell R. G., Wilson J. B., Anderson S. J., Longnecker R. 1998; Epstein–Barr virus LMP2A drives B cell development and survival in the absence of normal B cell receptor signals. Immunity 9:405–411
     [Google Scholar]
  12. Caldwell R. G., Brown R. C., Longnecker R. 2000; Epstein–Barr virus LMP2A-induced B-cell survival in two unique classes of EmuLMP2A transgenic mice. J Virol 74:1101–1113
     [Google Scholar]
  13. Callard R. E., Herbert J., Smith S. H., Armitage R. J., Costelloe K. E. 1995; CD40 cross-linking inhibits specific antibody production by human B cells. Int Immunol 11:1809–1815
     [Google Scholar]
  14. Campisi J. 2003; Cancer and ageing: rival demons?. Nat Rev Cancer 3:339–349
     [Google Scholar]
  15. Cannell E. J., Farrell P. J., Sinclair A. J. 1996; Epstein–Barr virus exploits the normal cell pathway to regulate Rb activity during the immortalisation of primary B-cells. Oncogene 13:1413–1421
     [Google Scholar]
  16. Casamayor-Palleja M., Khan M., MacLennan I. C. 1995; A subset of CD4+ memory T cells contains preformed CD40 ligand that is rapidly but transiently expressed on their surface after activation through the T cell receptor complex. J Exp Med 181:1293–1301
     [Google Scholar]
  17. Chen-Kiang S. 1995; Regulation of terminal differentiation of human B-cells by IL-6. Curr Top Microbiol Immunol 194:189–198
     [Google Scholar]
  18. Coleman M. L., Marshall C. J., Olson M. F. 2003; Ras promotes p21Waf1/Cip1 protein stability via a cyclin D1-imposed block in proteasome-mediated degradation. EMBO J 22:2036–2046
     [Google Scholar]
  19. Crawford D. H. 1986; Use of the virus to prepare human-derived monoclonal antibodies. In The Epstein–Barr Virus: Recent Advances pp  249–271 Edited by Epstein M. A., Achong B. G. London: William Heinemann Medical Books;
     [Google Scholar]
  20. Croft M., Swain S. L. 1991; B cell response to fresh and effector T helper cells. Role of cognate T-B interaction and the cytokines IL-2, IL-4, and IL-6. J Immunol 146:4055–4064
     [Google Scholar]
  21. Ehlin-Henriksson B., Gordon J., Klein G. 2003; B-lymphocyte subpopulations are equally susceptible to Epstein–Barr virus infection, irrespective of immunoglobulin isotype expression. Immunology 108:427–430
     [Google Scholar]
  22. Finke J., Rowe M., Kallin B., Ernberg I., Rosen A., Dillner J., Klein G. 1987; Monoclonal and polyclonal antibodies against Epstein–Barr virus nuclear antigen 5 (EBNA-5) detect multiple protein species in Burkitt's lymphoma and lymphoblastoid cell lines. J Virol 61:3870–3878
     [Google Scholar]
  23. Franklin D. S., Godfrey V. L., Lee H., Kovalev G. I., Schoonhoven R., Chen-Kiang S., Su L., Xiong Y. 1998; CDK inhibitors p18INK4c and p27Kip1 mediate two separate pathways to collaboratively suppress pituitary tumorigenesis. Genes Dev 12:2899–2911
     [Google Scholar]
  24. Fukuda M., Satoh T. A., Takanashi M., Hirai K., Ohnishi E., Sairenji T. 2000; Inhibition of cell growth and Epstein–Barr virus reactivation by CD40 stimulation in Epstein–Barr virus-transformed B cells. Viral Immunol 13:215–229
     [Google Scholar]
  25. Garrone P., Neidhardt E. M., Garcia E., Galibert L., van Kooten C., Banchereau J. 1995; Fas ligation induces apoptosis of CD40-activated human B lymphocytes. J Exp Med 182:1265–1273
     [Google Scholar]
  26. Gires O., Zimber-Strobl U., Gonnella R., Ueffing M., Marschall G., Zeidler R., Pich D., Hammerschmidt W. 1997; Latent membrane protein 1 of Epstein–Barr virus mimics a constitutively active receptor molecule. EMBO J 16:6131–6140
     [Google Scholar]
  27. Gordon J., Pound J. D. 2000; Fortifying B cells with CD154: an engaging tale of many hues. Immunology 100:269–280
     [Google Scholar]
  28. Greipp P. R., Witzig T. E., Gonchoroff N. J. 1985; Immunofluorescent plasma cell labeling indices (LI) using a monoclonal antibody (BU-1). Am J Hematol 20:289–292
     [Google Scholar]
  29. Harada H., Kawano M. M., Huang N. 7 other authors 1993; Phenotypic difference of normal plasma cells from mature myeloma cells. Blood 81:2658–2663
     [Google Scholar]
  30. Henle W., Diehl V., Kohn G., Zur Hausen H., Henle G. 1967; Herpes-type virus and chromosome marker in normal leukocytes after growth with irradiated Burkitt cells. Science 157:1064–1065
     [Google Scholar]
  31. Janeway C. A., Travers P., Walport M., Shlomchik M. J. 2001 Immunobiology: the immune system in health and disease, 5th edn. New York: Garland Publishing;
     [Google Scholar]
  32. Jego G., Bataille R., Pellat-Deceunynck C. 2001; Interleukin-6 is a growth factor for nonmalignant human plasmablasts. Blood 97:1817–1822
     [Google Scholar]
  33. Jung D., Neron S., Lemieux R., Roy A., Richard M. 2001; Telomere-independent reduction of human B lymphocyte: proliferation during long-term culture. Immunol Invest 30:157–168
     [Google Scholar]
  34. Kamijo T., Weber J. D., Zambetti G., Zindy F., Roussel M. F., Sherr C. J. 1998; Functional and physical interactions of the ARF tumor suppressor with p53 and Mdm2. Proc Natl Acad Sci U S A 95:8292–8297
     [Google Scholar]
  35. Kataoka H., Tahara H., Watanabe T., Sugawara M., Ide T., Goto M., Furuichi Y., Sugimoto M. 1997; Immortalization of immunologically committed Epstein–Barr virus-transformed human B-lymphoblastoid cell lines accompanied by a strong telomerase activity. Differentiation 62:203–211
     [Google Scholar]
  36. Kieff E., Rickinson A. B. 2001; Epstein Barr Virus and its replication. In Fields Virology , 4th dn pp  2511–2574 Edited by Knipe D. M., Howley P. M. Philadelphia: Lippincott Williams & Wilkins;
     [Google Scholar]
  37. Klein U., Küppers R., Rajewsky K. 1997; Evidence for a large compartment of IgM-expressing memory B cells in humans. Blood 89:1288–1298
     [Google Scholar]
  38. Klein U., Rajewsky K., Küppers R. 1998; Human immunoglobulin (Ig)M+IgD+ peripheral blood B cells expressing the CD27 cell surface antigen carry somatically mutated variable region genes: CD27 as a general marker for somatically mutated (memory) B cells. J Exp Med 188:1679–1689
     [Google Scholar]
  39. Lam E. W., Glassford J., van der Sman J., Banerji L., Pizzey A. R., Shaun N., Thomas B., Klaus G. G. 1999; Modulation of E2F activity in primary mouse B cells following stimulation via surface IgM and CD40 receptors. Eur J Immunol 29:3380–3389
     [Google Scholar]
  40. Lam E. W., Glassford J., Banerji L., Thomas N. S., Sicinski P., Klaus G. G. 2000; Cyclin D3 compensates for loss of cyclin D2 in mouse B-lymphocytes activated via the antigen receptor and CD40. J Biol Chem 275:3479–3484
     [Google Scholar]
  41. Lane P., Traunecker A., Hubele S., Inui S., Lanzavecchia A., Gray D. 1992; Activated human T cells express a ligand for the human B cell-associated antigen CD40 which participates in T cell-dependent activation of B lymphocytes. Eur J Immunol 22:2573–2578
     [Google Scholar]
  42. Lohoff M., Koch A., Rollinghoff M. 1992; Two signals are involved in polyclonal B cell stimulation by T helper type 2 cells: a role for LFA-1 molecules and interleukin 4. Eur J Immunol 22:599–602
     [Google Scholar]
  43. Mathon N. F., Lloyd A. C. 2001; Cell senescence and cancer. Nat Rev Cancer 1:203–213
     [Google Scholar]
  44. McCloskey N., Pound J. D., Holder M. J., Williams J. M., Roberts L. M., Lord J. M., Gordon J. 1999; The extrafollicular-to-follicular transition of human B lymphocytes: induction of functional globotriaosylceramide (CD77) on high threshold occupancy of CD40. Eur J Immunol 29:3236–3244
     [Google Scholar]
  45. Morse L., Chen D., Franklin D., Xiong Y., Chen-Kiang S. 1997; Induction of cell cycle arrest and B cell terminal differentiation by CDK inhibitor p18INK4c and IL-6. Immunity 6:47–56
     [Google Scholar]
  46. Nakagomi H., Dolcetti R., Bejarano M. T., Pisa P., Kiessling R., Masucci M. G. 1994; The Epstein–Barr virus latent membrane protein-1 (LMP1) induces interleukin-10 production in Burkitt lymphoma lines. Int J Cancer 57:240–244
     [Google Scholar]
  47. Nicoletti I., Migliorati G., Pagliacci M. C., Grignani F., Riccardi C. 1991; A rapid and simple method for measuring thymocyte apoptosis by propidium iodide staining and flow cytometry. J Immunol Methods 139:271–279
     [Google Scholar]
  48. Nilsson K. 1979; The nature of lymphoid cell lines and their relationship to the virus. In The Epstein–Barr Virus pp  225–281 Edited by Epstein M. A., Achong B. G. Berlin: Springer;
     [Google Scholar]
  49. Nilsson K., Ponten J. 1975; Classification and biological nature of established human hematopoietic cell lines. Int J Cancer 15:321–341
     [Google Scholar]
  50. Nilsson K., Klein G., Henle W., Henle G. 1971; The establishment of lymphoblastoid lines from adult and fetal human lymphoid tissue and its dependence on EBV. Int J Cancer 8:443–450
     [Google Scholar]
  51. Noelle R. J., Roy M., Shepherd D. M., Stamenkovic I., Ledbetter J. A., Aruffo A. 1992; A 39-kDa protein on activated helper T cells binds CD40 and transduces the signal for cognate activation of B cells. Proc Natl Acad Sci U S A 89:6550–6554
     [Google Scholar]
  52. O'Nions J., Allday M. J. 2003; Epstein–Barr virus can inhibit genotoxin-induced G1 arrest downstream of p53 by preventing the inactivation of CDK2. Oncogene 22:7181–7191
     [Google Scholar]
  53. Ormerod M. G., Sun X. M., Snowden R. T., Davies R., Fearnhead H., Cohen G. M. 1993; Increased membrane permeability of apoptotic thymocytes: a flow cytometric study. Cytometry 14:595–602
     [Google Scholar]
  54. Parker S. B., Eichele G., Zhang P., Rawls A., Sands A. T., Bradley A., Olson E. N., Harper J. W., Elledge S. J. 1995; p53-independent expression of p21Cip1 in muscle and other terminally differentiating cells. Science 267:1024–1027
     [Google Scholar]
  55. Parker G. A., Crook T., Bain M., Sara E. A., Farrell P. J., Allday M. J. 1996; Epstein–Barr virus nuclear antigen (EBNA)3C is an immortalizing oncoprotein with similar properties to adenovirus E1A and papillomavirus E7. Oncogene 13:2541–2549
     [Google Scholar]
  56. Pokrovskaja K., Ehlin-Henriksson B., Kiss C., Challa A., Gordon J., Gogolak P., Klein G., Szekely L. 2002; CD40 ligation downregulates EBNA-2 and LMP-1 expression in EBV-transformed lymphoblastoid cell lines. Int J Cancer 99:705–712
     [Google Scholar]
  57. Polack A., Hortnagel K., Pajic A. 7 other authors 1996; c-myc activation renders proliferation of Epstein–Barr virus (EBV)-transformed cells independent of EBV nuclear antigen 2 and latent membrane protein 1. Proc Natl Acad Sci U S A 93:10411–10416
     [Google Scholar]
  58. Pope J. H. 1967; Establishment of cell lines from peripheral leucocytes in infectious mononucleosis. Nature 216:810–811
     [Google Scholar]
  59. Quelle D. E., Zindy F., Ashmun R. A., Sherr C. J. 1995; Alternative reading frames of the INK4a tumor suppressor gene encode two unrelated proteins capable of inducing cell cycle arrest. Cell 83:993–1000
     [Google Scholar]
  60. Randall T. D., Heath A. W., Santos-Argumedo L., Howard M. C., Weissman I. L., Lund F. E. 1998; Arrest of B lymphocyte terminal differentiation by CD40 signalling: mechanism for lack of antibody-secreting cells in germinal centres. Immunity 8:733–742
     [Google Scholar]
  61. Richards M. L., Katz D. H. 1997; Analysis of the promoter elements necessary for IL-4 and anti-CD40 antibody induction of murine Fc epsilon RII (CD23): comparison with the germline epsilon promoter. J Immunol 158:263–272
     [Google Scholar]
  62. Rousset F., Peyrol S., Garcia E., Vezzio N., Andujar M., Grimaud J. A., Banchereau J. 1995; Long-term cultured CD40-activated B lymphocytes differentiate into plasma cells in response to IL-10 but not IL-4. Int Immunol 7:1243–1253
     [Google Scholar]
  63. Schonbeck U., Libby P. 2001; The CD40/CD154 receptor/ligand dyad. Cell Mol Life Sci 58:4–43
     [Google Scholar]
  64. Schrantz N., Beney G. E., Auffredou M. T., Bourgeade M. F., Leca G., Vazquez A. 2000; The expression of p18INK4 and p27kip1 cyclin-dependent kinase inhibitors is regulated differently during human B cell differentiation. J Immunol 165:4346–4352
     [Google Scholar]
  65. Sherr C. J., DePinho R. A. 2000; Cellular senescence: mitotic clock or culture shock?. Cell 102:407–410
     [Google Scholar]
  66. Shvarts A., Brummelkamp T. R., Scheeren F., Koh E., Daley G. Q., Spits H., Bernards R. 2002; A senescence rescue screen identifies BCL6 as an inhibitor of anti-proliferative p19ARF–p53 signaling. Genes Dev 16:681–686
     [Google Scholar]
  67. Smeland E. B., Blomhoff H. K., Funderud S., Shalaby M. R., Espevik T. 1989; Interleukin 4 induces selective production of interleukin 6 from normal human B lymphocytes. J Exp Med 170:1463–1468
     [Google Scholar]
  68. Spender L. C., Cannell E. J., Hollyoake M., Wensing B., Gawn J. M., Brimmell M., Packham G., Farrell P. J. 1999; Control of cell cycle entry and apoptosis in B lymphocytes infected by Epstein–Barr virus. J Virol 73:4678–4688
     [Google Scholar]
  69. Staudt L. M., Brown P. O. 2000; Genomic views of the immune system. Annu Rev Immunol 18:829–859
     [Google Scholar]
  70. Thorley-Lawson D. A. 2001; Epstein–Barr virus: exploiting the immune system. Nat Rev Immunol 1:75–82
     [Google Scholar]
  71. Tosato G., Tanner J., Jones K. D., Revel M., Pike S. E. 1990; Identification of interleukin-6 as an autocrine growth factor for Epstein–Barr virus-immortalized B cells. J Virol 64:3033–3041
     [Google Scholar]
  72. Tourigny M. R., Ursini-Siegel J., Lee H. 9 other authors 2002; CDK inhibitor p18INK4c is required for the generation of functional plasma cells. Immunity 17:179–189
     [Google Scholar]
  73. Urashima M., Chauhan D., Hatziyanni M., Ogata A., Hollenbaugh D., Aruffo A., Anderson K. C. 1996; CD40 ligand triggers interleukin-6 mediated B cell differentiation. Leuk Res 20:507–515
     [Google Scholar]
  74. van Kooten C., Banchereau J. 2000; CD40–CD40 ligand. J Leukoc Biol 67:2–17
     [Google Scholar]
  75. Vousden K. H., Lu X. 2002; Live or let die: the cell's response to p53. Nat Rev Cancer 2:594–604
     [Google Scholar]
  76. Wade M., Allday M. J. 2000; Epstein–Barr virus suppresses a G2/M checkpoint activated by genotoxins. Mol Cell Biol 20:1344–1360
     [Google Scholar]
  77. Walker P. R., Smith C., Youdale T., Leblanc J., Whitfield J. F., Sikorska M. 1991; Topoisomerase II-reactive chemotherapeutic drugs induce apoptosis in thymocytes. Cancer Res 51:1078–1085
     [Google Scholar]
  78. Worm M., Krah J. M., Manz R. A., Henz B. M. 1998; Retinoic acid inhibits CD40+interleukin-4-mediated IgE production in vitro. Blood 92:1713–1720
     [Google Scholar]
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Proliferation and differentiation in isogenic populations of peripheral B cells activated by Epstein–Barr virus or T cell-derived mitogens
J. Gen. Virol. 85, 881 (2004); https://doi.org/10.1099/vir.0.19704-0
/content/journal/jgv/10.1099/vir.0.19704-0
/content/journal/jgv/10.1099/vir.0.19704-0
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