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T cell receptor-induced activation and apoptosis in cycling human T cells occur throughout the cell cycle
1Diabetes Branch, National Institute of Diabetes and Digestive and Kidney Diseases, Bethesda, Maryland 20892-1770, USA. karas@box-k.nih.gov
Abstract:
Previous studies have found conflicting associations between susceptibility to activation-induced cell death and the cell cycle in T cells. However, most of the studies used potentially toxic pharmacological agents for cell cycle synchronization. A panel of human melanoma tumor-reactive T cell lines, a CD8+ HER-2/neu-reactive T cell clone, and the leukemic T cell line Jurkat were separated by centrifugal elutriation. Fractions enriched for the G0-G1, S, and G2-M phases of the cell cycle were assayed for T cell receptor-mediated activation as measured by intracellular Ca(2+) flux, cytolytic recognition of tumor targets, and induction of Fas ligand mRNA. Susceptibility to apoptosis induced by recombinant Fas ligand and activation-induced cell death were also studied. None of the parameters studied was specific to a certain phase of the cell cycle, leading us to conclude that in nontransformed human T cells, both activation and apoptosis through T cell receptor activation can occur in all phases of the cell cycle.
Insights
T cell activation and apoptosis can occur in any cell cycle phase. This study used centrifugal elutriation to avoid toxic agents, finding no cell cycle specificity for T cell receptor activation or cell death.
Area of Science:
- Immunology
- Cell Biology
Background:
- Conflicting data exists on T cell susceptibility to activation-induced cell death (AICD) across the cell cycle.
- Previous synchronization methods used toxic pharmacological agents, potentially confounding results.
Purpose of the Study:
- To investigate the relationship between T cell cycle phases and activation-induced cell death.
- To determine if T cell activation and apoptosis are restricted to specific cell cycle phases using non-toxic methods.
Main Methods:
- Human T cell lines (melanoma-reactive, CD8+ HER-2/neu-reactive, Jurkat) were synchronized using centrifugal elutriation.
- Cell cycle fractions (G0-G1, S, G2-M) were analyzed for T cell receptor (TCR)-mediated activation (intracellular Ca2+ flux, cytolytic activity, Fas ligand mRNA).
- Susceptibility to Fas ligand-induced apoptosis and AICD was assessed across cell cycle phases.
Main Results:
- TCR-mediated activation markers were detected in all cell cycle phases (G0-G1, S, G2-M).
- Cytolytic recognition and Fas ligand mRNA induction occurred irrespective of the cell cycle phase.
- Susceptibility to both Fas ligand-induced apoptosis and AICD was not confined to any specific cell cycle phase.
Conclusions:
- In non-transformed human T cells, T cell receptor (TCR) activation and subsequent apoptosis can occur throughout all phases of the cell cycle.
- Centrifugal elutriation provides a non-toxic method for cell cycle synchronization, yielding reliable data on T cell activation and death pathways.
- The findings resolve previous discrepancies by demonstrating cell cycle-independent T cell activation and apoptosis.