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Updated: Aug 8, 2026
Generation of Induced Regulatory T Cells from Primary Human Naïve and Memory T Cells
Published on: April 16, 2012
Spontaneous generation of human CD8+ TCR alpha beta+ cells derived from precursors within the double negative
Y Fujimiya1, M Nakayama, T Shibata
1Division of Immunology, National Children's Medical Research Center, Tokyo, Japan.
Insights
Human thymocytes, including double-negative (DN) cells, can develop T-cell receptor (TCR) expression outside the thymus. Double-positive (DP) cells influence DN cell differentiation, promoting TCR alpha beta development.
Area of Science:
- Immunology
- Cell Biology
- Developmental Biology
Background:
- Human thymocytes are crucial for T-cell development, with distinct populations like double-negative (DN) and double-positive (DP) cells.
- Understanding T-cell differentiation pathways, particularly T-cell receptor (TCR) expression, is vital for immune system function.
Purpose of the Study:
- To investigate the in vitro expansion and differentiation of human thymocytes, focusing on TCR expression.
- To determine the influence of double-positive (DP) cells on the development of double-negative (DN) thymocytes.
Main Methods:
- Flow cytometry was used to analyze thymocyte populations (DN, DP, mature T cells).
- In vitro expansion of unfractionated and purified thymocyte subsets (DN, DP) using specific stimuli (TPA, PHA, IL-2).
- Long-term culture (18 days) to assess cell proliferation and surface marker expression (CD3, TCR alpha beta, TCR gamma delta, CD8).
Main Results:
- Unfractionated thymocytes expanded in vitro predominantly expressed CD3, TCR alpha beta, and CD8.
- Purified DN thymocytes expanded with DP cells acquired CD3, TCR alpha beta, and CD8 expression.
- Purified DN thymocytes cultured alone expressed TCR gamma delta but not CD8 or TCR alpha beta.
- Purified DP cells showed limited proliferation despite expressing IL-2 receptors and TCR alpha beta.
Conclusions:
- Human precursor T cells in the DN compartment can differentiate and express TCRs outside the thymus.
- DP thymocytes play a role in directing the TCR expression profile of developing DN thymocytes.
- This study provides evidence for extrathymic T-cell differentiation pathways in humans.
Abstract:
Flow cytometric analysis demonstrated that fresh human thymocytes contain only a low level of mature CD8+ TCR alpha beta + or CD8+ TCR gamma delta + cells and they consist consist of approximately 70% double positive (DP) and approximately 10% double negative (DN) cells. These unfractionated thymocytes could be selectively expanded in vitro by stimulation with 12-O-tetradecanoylphorbol 13-acetate (TPA) and PHA in the presence of IL-2. The majority of the cells expanded from unfractionated thymocytes expressed CD3, TCR alpha beta and CD8 molecules after long-term culture (18 days). When highly purified DN thymocytes were expanded over a period of 18 days in the presence of DP cells, they also co-expressed CD3, TCR alpha beta and CD8 molecules on their surface. However, when purified DN thymocytes were expanded alone, that is, in the absence of DP cells for 18 days, they expressed CD3-associated TCR gamma delta, but not CD8 or TCR alpha beta. Despite the expression of measurable levels of IL-2 alpha and beta receptors, as well as a significant level of TCR alpha beta, purified DP cells failed to proliferate. These findings provide the first evidence, in humans, that the progression of precursor cells in the DN compartment to a later stage of differentiation can be induced outside the thymus and that DP cells can affect the development of TCR expression in proliferating DN thymocytes.
