Developmentally regulated expression of the PD-1 protein on the surface of double-negative(CD4–CD8–) thymocytes

PD-1, a member of the Ig superfamily, was previously isolated from an apoptosis-induced T cell hybridoma 2B4.11 by subtractive hybridization. Expression of the PD-1 mRNA is restricted to thymus in adult mice. Using an anti-PD-1 mAb (J43), we examined expression of the PD-1 protein during differentiation of thymocytes in normal adult, fetal and RAG-2-/- mice with or without anti-CD3 mAb stimulation. While PD-1 was expressed only on 3–5% of total normal thymocytes, –34% of the CD4-CD8- double-negative (DN) fraction are PD-1+ cells with two distinct expression levels (low and high). PD-1high thymocytes belonged to TCR γδ lineage cells. In the DN compartment of the TCR αβ lineage, PD-1 expression started at the low level from the CD44+CD25+ stage and the majority of thymocytes expressed PD-1 at the CD44-CD25- stage in which thymocytes express TCR β chains. The anti-CD3ε antibody administration augmented the PD-1 expression as well as the differentiation of the CD44-CD25+ DN cells into the CD44-CD25- DN stage, not only in normal mice but also in RAG-2-deficient mice. The fraction of the PD-1low cells in the CD4+CD8+ double-positive (DP) compartment was very small (>5%) but increased by stimulation with the anti-CD3 antibody, although the total number of DP cells was drastically reduced. The results show that PD-1 expression is specifically induced at the stages preceding clonal selection.