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Abstract
BackgroundCD117 is expressed on double-negative (DN; CD4–CD8–) cells (Nat Rev Immunol 14:529–545; 2014), but whether it is expressed in other stages and its subsequent functions are unclear. We used an improved method of flow cytometry to analyze different populations of thymocytes (Sci Rep 4:5781; 2014). The expression of CD117 and CTLA-4 were directly assayed in the early stage of thymocytes.MethodsFlow cytometry was used to analyze different populations of thymocytes, and T-cell proliferation assays, RT-PCR, and real-time RT-PCR were used to characterize the stem cells and examine the function of CD44+CD117+ cells.ResultsIn DN cells, CD117 expression was greatest on CD44+CD25+ cells (DN2), followed by CD44+CD25– (DN1), CD44–CD25+ (DN3), and CD44–CD25– (DN4) cells. In thymocytes, CD117 expression was highest in DN cells, followed by single-positive (SP; CD4 or CD8) and double-positive (DP; CD4+CD8+) cells. Especially, CD117 expression was positively associated with CD44 and CTLA-4 expression. CTLA-4 expression was highest in DN cells, followed by SP and DP cells. CTLA-4 expression was positively associated with CD25, CD44, and Foxp3 expression. CD44+CD117+ T cells expressed more CTLA-4, which suppressed T-cell proliferation and blocked CTLA-4 to cause antibody-induced T-cell proliferation.ConclusionThese results suggest that CD44+CD117+ T cells are stem cells and a specific T-cell phenotype that initially develops in the thymus, but they do not progress through DN3 and DN4 stages, lack a DP stage, and potently suppress T-cell proliferation and modulate the CTLA-4 pathway.
Highlights
CD117 is expressed on double-negative (DN; CD4–CD8–) cells (Nat Rev Immunol 14:529–545; 2014), but whether it is expressed in other stages and its subsequent functions are unclear
To confirm CD117 expression in the thymus during T-cell development, thymocytes were harvested from C57BL/6 mice, stained with CD4 (FITC), CD8 (PercP5.5), CD44 (APC-CY7), CD25 (PE-CY7), and CD117 (PE), and analyzed by flow cytometry
DN cells had more CD117-expressing cells in C57BL/6 mice followed by CD8+ SP cells, CD4+ SP cells, and DP cells (Fig. 1b); the pooled data from three independent experiments (Fig. 1d) indicate that CD117 expression was lowest in DP cells
Summary
CD117 is expressed on double-negative (DN; CD4–CD8–) cells (Nat Rev Immunol 14:529–545; 2014), but whether it is expressed in other stages and its subsequent functions are unclear. The earliest precursors in the T-cell lineage are found within thymocyte populations and lack CD4 and CD8 expression, so they are referred to as CD4−CD8− or double-negative (DN) thymocytes [3]. DN cells that express CD4 and CD8 are CD4+CD8+ or double-positive (DP) thymocytes before. Mammalian CD117 (c-kit) gene protein is a stem cell marker and a type III tyrosine kinase receptor [9, 10], widely expressed in hematopoietic stem cells (HSC), myeloid progenitor cells, as well as pro-B cells and proT cells [11,12,13].
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