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Updated: Aug 9, 2026
Examination of Thymic Positive and Negative Selection by Flow Cytometry
Published on: October 8, 2012
Blocked negative selection of developing T cells in mice expressing the baculovirus p35 caspase inhibitor
M Izquierdo1, A Grandien, L M Criado
1Department of Immunology and Oncology, Centro Nacional de Biotecnología, UAM Campus de Cantoblanco, E-28049, Madrid, Spain.
Abstract:
Clonal deletion in the thymus by apoptosis is involved in purging the immune system of self-reactive T lymphocytes (negative selection). Cysteine proteases (caspases) belonging to the CPP32 family are activated during this process. We have produced transgenic mice expressing baculovirus p35, a broad-range caspase inhibitor. Thymocytes from p35 transgenic mice were resistant in vitro to several apoptosis-inducing agents; this resistance correlated with the inhibition of CPP32-like activity. Negative selection in vivo of thymocytes triggered by two exogenous antigens, staphylococcal enterotoxin B superantigen and an antigenic peptide in the F5 T-cell receptor transgenic model, was specifically inhibited in p35 transgenic mice. Our results provide direct evidence for caspase involvement in negative selection during thymocyte development.
Insights
This study shows that caspases (cysteine proteases) are essential for negative selection, a process that eliminates self-reactive T cells in the thymus. Inhibiting caspases prevented this crucial immune system purging.
Area of Science:
- Immunology
- Molecular Biology
- Cell Biology
Background:
- Negative selection in the thymus eliminates self-reactive T lymphocytes via apoptosis.
- Caspases, specifically CPP32 family proteases, are activated during thymocyte apoptosis.
- Understanding the role of caspases in negative selection is crucial for immune system development.
Purpose of the Study:
- To investigate the direct involvement of caspases in thymocyte negative selection.
- To determine if caspase inhibition affects the in vivo purging of self-reactive T cells.
Main Methods:
- Generated transgenic mice expressing baculovirus p35, a caspase inhibitor.
- Assessed thymocyte resistance to apoptosis in vitro.
- Analyzed in vivo negative selection using exogenous antigens (staphylococcal enterotoxin B and an antigenic peptide) in a T-cell receptor transgenic model.
Main Results:
- Thymocytes from p35 transgenic mice exhibited resistance to apoptosis-inducing agents in vitro.
- This resistance correlated with the inhibition of CPP32-like caspase activity.
- In vivo negative selection of thymocytes was specifically inhibited in p35 transgenic mice.
Conclusions:
- Caspases play a direct and essential role in thymocyte negative selection.
- Inhibiting caspases effectively blocks the elimination of self-reactive T lymphocytes during thymic development.
- These findings provide direct evidence for caspase involvement in immune tolerance.
