T cell activation induced by anti-CD3 antibodies requires prolonged stimulation of protein kinase C.
Accessory cell-depleted T cells required the presence of a protein kinase C (PKC) stimulating phorbol ester, such as phorbol 12,13-dibutyrate (PDB), to be activated by soluble antibodies to the CD3 molecular complex. To determine the duration of PDB costimulation necessary to induce a proliferative response, highly purified T cells were pulsed with anti-CD3, incubated with PDB for limited periods of time, and then washed and recultured in the absence of PDB. T cells stimulated with anti-CD3 and PDB for 2 hr were unable to proliferate unless IL-2 or PDB was added to the second culture. With more prolonged exposure to PDB (4-18 hr), anti-CD3-pulsed cells exhibited an increased capacity to proliferate in the absence of additional PDB. Proliferation could be augmented by exogenous IL-2, but remained submaximal. Optimal DNA synthetic responses required the presence of PDB throughout the entire culture. Despite this, costimulation with anti-CD3 and PDB induced a significant number of cells to express IL-2 receptors and enter the cell cycle after 18 hr of costimulation with PDB. Moreover, T cells costimulated by anti-CD3 and PDB produced IL-2 within 4 hr. However, T cells that were stimulated with anti-CD3 and PDB for 4 hr, washed, and recultured rapidly lost the ability to continue to produce IL-2, which reflected a decrease in the content of mRNA encoding IL-2. This loss of IL-2 production was prevented by reculturing the cells with PDB. These studies therefore indicate that after initial T cell activation by anti-CD3, continued stimulation of PKC is necessary for ongoing IL-2 production. These results suggest a model of T cell activation in which sustained stimulation of PKC after cell cycle entry is required to maintain growth factor production and continued proliferation.
Davis
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LS
Harold C. Simmons Arthritis Research Center, Dallas, Texas 75235.
Lipsky
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PE
D000328:Adult; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000945:Antigens, Differentiation, T-Lymphocyte / Q000276:immunology*; D017252:CD3 Complex; D002455:Cell Division / Q000187:drug effects; D004261:DNA Replication / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D007376:Interleukin-2 / Q000494:pharmacology; D008213:Lymphocyte Activation / Q000187:drug effects*; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism / Q000502:physiology*; D012333:RNA, Messenger / Q000032:analysis; D011948:Receptors, Antigen, T-Cell / Q000276:immunology*; D015398:Signal Transduction / Q000187:drug effects; D013601:T-Lymphocytes / Q000201:enzymology / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000945:Antigens, Differentiation, T-Lymphocyte; D017252:CD3 Complex; D007376:Interleukin-2; D012333:RNA, Messenger; D011948:Receptors, Antigen, T-Cell; D015240:Phorbol 12,13-Dibutyrate; D011493:Protein Kinase C