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Receptor modulation and early signal transduction events in cytotoxic T lymphocytes inactivated by sensitive target
G A Hommel-Berrey1, A M Shenoy, Z Brahmi
1Department of Medicine, Indiana University School of Medicine, Indianapolis 46202.
Abstract:
In previous studies, we demonstrated that NK cells and lymphokine-activated killer cells were inactivated early in the lytic process by susceptible but not by resistant target cells (TC). We examined the functional status of human MHC-restricted CTL, after interaction with sensitive TC. Two CTL lines were generated in vitro by stimulation with irradiated PAMO, an EBV-transformed cell line. CTL were incubated for up to 4 h with an equal number of PAMO, then separated by a SRBC rosette assay. CTL lost greater than 60% of their lytic activity during the first 30 min of incubation, and greater than 90% by 4 h as assessed by their inability to lyse fresh TC. Inactivated CTL had 35% less serine esterase activity than did control CTL. IL-2 restored the lytic potential and serine esterase activity to normal values within 72 h. Exposure of CTL to PAMO for 4 h induced the modulation of 22 to 44% of TCR/CD3, CD4/CD8, and class I Ag from the cell surface. In contrast, the expression of CD69, and class II Ag increased and there was no change in the expression of CD2, CD28, or LFA-1 Ag. Furthermore, early metabolic events that usually follow CTL-ligand interaction such as phosphatidylinositol metabolism and transient increase in intracellular calcium, did not occur in inactivated CTL upon challenge with PAMO. PMA and the calcium ionophore A23187, restored cytolytic activity, indicating that protein kinase C can be activated and translocated in inactivated CTL. Our data suggest that TC-induced inactivation of CTL may be due to the modulation of key membrane molecules and the lack of certain secondary messengers involved in signal transduction.
Insights
Cytotoxic T-lymphocytes (CTL) lose lytic activity when interacting with target cells (TC). This inactivation involves altered cell surface molecules and impaired signal transduction, but can be restored by IL-2.
Area of Science:
- Immunology
- Cellular Biology
Background:
- Natural killer (NK) cells and lymphokine-activated killer (LAK) cells are inactivated by susceptible target cells (TC).
- The functional status of human MHC-restricted cytotoxic T-lymphocytes (CTL) after interaction with sensitive TC was investigated.
Purpose of the Study:
- To examine the functional status and molecular changes in human CTL after interaction with sensitive target cells.
- To understand the mechanisms behind TC-induced CTL inactivation.
Main Methods:
- Generated human CTL lines in vitro.
- Incubated CTL with sensitive TC (PAMO) for up to 4 hours.
- Assessed CTL lytic activity, serine esterase activity, and cell surface molecule expression (TCR/CD3, CD4/CD8, MHC class I/II, CD69, CD2, CD28, LFA-1).
- Investigated early metabolic events (phosphatidylinositol metabolism, intracellular calcium) and the effect of IL-2, PMA, and A23187.
Main Results:
- CTL lost >60% lytic activity within 30 min and >90% by 4 hours.
- Inactivated CTL showed reduced serine esterase activity, which was restored by IL-2.
- Exposure to TC modulated TCR/CD3, CD4/CD8, and MHC class I Ag, while increasing CD69 and MHC class II Ag expression.
- Early metabolic events and signal transduction pathways were impaired in inactivated CTL.
- PMA and A23187 restored cytolytic activity, suggesting protein kinase C involvement.
Conclusions:
- TC-induced inactivation of CTL involves modulation of key membrane molecules.
- Impaired secondary messengers in signal transduction contribute to CTL inactivation.
- Interleukin-2 (IL-2) can restore CTL function and serine esterase activity.