The preTCR‐dependent DN3 to DP transition requires Notch signaling, is improved by CXCL12 signaling and is inhibited by IL‐7 signaling

• Published in: European journal of immunology Vol. 41; no. 11; pp. 3371 - 3380
• Main Authors: Tussiwand, Roxane, Engdahl, Corinne, Gehre, Nadine, Bosco, Nabil, Ceredig, Rod, Rolink, Antonius G.
• Format: Journal Article
• Language: English
• Published: Weinheim WILEY‐VCH Verlag 01-11-2011; Wiley Subscription Services, Inc
• Subjects: Animals; Cell Differentiation - immunology; Cell Separation; Chemokine CXCL12 - immunology; Chemokine CXCL12 - metabolism; Delta‐like 4 and preTCR; DN3 cells; Double‐positive cells; Female; Flow Cytometry; Interleukin-7 - immunology; Interleukin-7 - metabolism; Intracellular Signaling Peptides and Proteins - immunology; Intracellular Signaling Peptides and Proteins - metabolism; Membrane Proteins - immunology; Membrane Proteins - metabolism; Mice; Mice, Inbred C57BL; Mice, Transgenic; Notch; Real-Time Polymerase Chain Reaction; Recombinant Proteins - immunology; Recombinant Proteins - metabolism; Signal Transduction - immunology; Thymocytes - cytology; Thymocytes - immunology; Thymocytes - metabolism
• ISSN: 0014-2980; 1521-4141
• DOI: 10.1002/eji.201141824

The requirement for Notch signaling during T‐cell development has been extensively studied. Nevertheless, the developmental stage at which it is required and whether additional signaling pathways are needed are still poorly understood. By using a stromal‐cell‐free culture system, we show that sorted double‐negative 3 (DN3) thymocytes only require a Delta‐like‐4‐induced Notch signal to differentiate into double‐positive (DP) cells. This differentiation process is preTCR‐α dependent. DN3 cells undergo 4–5 proliferation cycles, and the addition of the chemokine CXCL12 improves proliferation. IL‐7 blocks the differentiation of DN3 cells to DP cells but not the Notch‐induced proliferation of cultured DN3 cells. The impaired differentiation correlates with an inhibition of Rag‐2 up‐regulation. Overall, the in vitro stromal‐cell‐free culture system presented here also provides a powerful and unique tool for studying the mechanisms involved in the positive and negative selection of T cells.