Single human T cells stimulated in the absence of feeder cells transcribe interleukin-2 and undergo long-term clonal growth in response to defined monoclonal antibodies and cytokine stimulation.
The two-signal model of T-cell activation postulates that T lymphocytes require at least two distinct signals for activation. This model has been established with bulk cultures of T cells in which T-cell-T-cell interaction can occur, possibly delivering further unrecognized costimulatory signals. The signal requirements of single T cells for the induction of clonal cell growth or the transcription of cytokines would best be studied in a cell cloning system in the absence of feeder cells; however, such an experimental system has not been reported so far. In this study, we report the long-term cloning of human resting peripheral blood CD4+CD45RO- T cells under feeder cell-free conditions in response to CD3 and CD28 stimulation in the presence of exogenous interleukin-2 (IL-2). Cloning efficiency ranged from 40% to 60% depending on the presence of additional cytokines IL-1 and IL-6. Single-call polymerase chain reaction showed that transcription of IL-2 occurred in cells stimulated through CD3 plus CD28 alone. T cells grown in response to CD3 plus CD28 plus IL-2 stimulation produced both IL-4 and interferon-gamma (IFN-gamma) on restimulation (Th0 cells) and could be functionally differentiated into Th1- or Th2-type cells by the addition of IFN-gamma or IL-4, respectively, during cell cloning. These data show on the single-cell level a two-signal model of T- cell activation for the transcription of IL-2. In addition, these experiments show that IFN-gamma and IL-4 exert their T-cell-differentiating effects directly on the T cell without any further need for antigen-presenting cells. Together, our experiments show the feasability of a defined long-term clonal cell culture system to study the growth and differentiation of human T lymphocytes.
Sunder-Plassmann
R
R
Institute of Immunology, University of Vienna, Austria.
Breiteneder
H
H
Zimmermann
K
K
Strunk
D
D
Majdic
O
O
Knapp
W
W
Holter
W
W
D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology*; D001483:Base Sequence; D018106:CD28 Antigens / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000187:drug effects / Q000276:immunology*; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D002999:Clone Cells; D016207:Cytokines / Q000276:immunology / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects; D015333:Gene Rearrangement, beta-Chain T-Cell Antigen Receptor; D006801:Humans; D007371:Interferon-gamma / Q000096:biosynthesis / Q000494:pharmacology; D007375:Interleukin-1 / Q000494:pharmacology; D007376:Interleukin-2 / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology; D015847:Interleukin-4 / Q000096:biosynthesis / Q000494:pharmacology; D015850:Interleukin-6 / Q000494:pharmacology; D008213:Lymphocyte Activation / Q000187:drug effects*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D017260:Receptor-CD3 Complex, Antigen, T-Cell / Q000276:immunology; D018417:Th1 Cells / Q000166:cytology; D018418:Th2 Cells / Q000166:cytology
D000911:Antibodies, Monoclonal; D018106:CD28 Antigens; D016207:Cytokines; D007375:Interleukin-1; D007376:Interleukin-2; D015850:Interleukin-6; D017260:Receptor-CD3 Complex, Antigen, T-Cell; D015847:Interleukin-4; D007371:Interferon-gamma