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Updated: Aug 8, 2026
Examination of Thymic Positive and Negative Selection by Flow Cytometry
Published on: October 8, 2012
CD8 requirements for negative selection events are directly related to the TCR-antigen interaction
S J Curnow1, A Guimezanes, A M Schmitt-Verhulst
1Centre d'Immunologie INSERM-CNRS de Marseille-Luminy, France.
Insights
CD8 molecule expression is essential for positive selection of T cells. However, negative selection can occur without CD8, depending on the T cell receptor-antigen interaction, revealing differential CD8 dependencies.
Area of Science:
- Immunology
- T cell biology
- Thymocyte development
Background:
- Positive selection of T cells typically requires CD8 expression on thymocytes.
- Negative selection's dependence on CD8 varies with the antigen encountered by the T cell receptor (TCR).
Purpose of the Study:
- To investigate the role of CD8 in positive and negative selection of T cells.
- To analyze CD8-dependency using TCR-transgenic (Tg) mice deficient in CD8 expression.
Main Methods:
- Utilized two TCR-transgenic (Tg) mouse models reacting to the H-2kb allo-antigen.
- Generated CD8-deficient mice by backcrossing Tg mice with CD8-null strains.
- Analyzed thymocyte populations based on CD4, CD8, and Tg-TCR expression.
Main Results:
- CD8 expression was consistently required for positive selection of high-Tg-TCR expressing cells.
- Negative selection showed differential CD8 dependency, mirroring the original T cell clone's CD8 dependence.
- One model showed partial CD8-dependent negative selection, while the other was unaffected by CD8 absence.
- CD4-8- Tg-TCR positive populations remained unchanged in CD8-deficient mice.
Conclusions:
- CD8 expression appears necessary for positive selection of T cells.
- Negative selection can proceed independently of CD8, driven by specific TCR-antigen interactions.
- These findings highlight distinct CD8 roles in thymocyte selection pathways.
Abstract:
Positive selection of class I-restricted T cells has been suggested to always require the surface expression of CD8 molecules on CD4+8+ thymocytes, whilst negative selection was found to be differentially dependent, relating to the antigen being engaged by the T cell receptor (TCR). We have studied the CD8-dependency of positive and negative selection using two TCR-transgenic (Tg) mice models, which both react against the same allo-antigen, H-2kb, back crossed with mice which are deficient for the expression of CD8. Whilst CD8 expression was always required for positive selection of cells expressing high levels of the Tg-TCR, events of negative selection were differentially dependent upon CD8, reflecting the CD8-dependency of the original CTL clones. For one TCR-Tg model (derived from a CD8-dependent CTL clone), deletion of CD4+8+ thymocytes was partially dependent upon the expression of CD8, though there was still selection against Tg-TCR positive CD4+ cells, which normally exit to the periphery expressing low levels of Tg-TCR. For the other model (derived from a CD8-independent CTL clone) negative selection was unaffected by the absence of CD8. For both models the CD4-8- Tg-TCR positive population was unaffected by the absence of CD8, including, for the CD8-independent model, reduced expression of the Tg-TCR/CD3 complex. These results suggest that CD8 expression may be a prerequisite for positive selection, whilst negative selection events can occur in the absence of CD8, depending directly upon the TCR-antigen interaction.
