CD4⁺ and CD8⁺ regulatory T cell characterization in the rat using a unique transgenic Foxp3-EGFP model

Regulatory T cells (Treg) in diverse species include CD4⁺ and CD8⁺ T cells. In all species, CD8⁺ Treg have been only partially characterized and there is no rat model in which CD4⁺ and CD8⁺ FOXP3⁺ Treg are genetically tagged.

We generated a Foxp3-EGFP rat transgenic line in which FOXP3 gene was expressed and controlled EGFP. CD4⁺ and CD8⁺ T cells were the only cells that expressed EGFP, in similar proportion as observed with anti-FOXP3 antibodies and co-labeled in the same cells. CD4⁺EGFP⁺ Treg were 5-10 times more frequent than CD8⁺EGFP⁺ Treg. The suppressive activity of CD4⁺ and CD8⁺ Treg was largely confined to EGFP⁺ cells. RNAseq analyses showed similarities but also differences among CD4⁺ and CD8⁺ EGFP⁺ cells and provided the first description of the natural FOXP3⁺CD8⁺ Treg transcriptome. In vitro culture of CD4⁺ and CD8⁺ EGFP^- cells with TGFbeta and IL-2 generated induced EGFP⁺ Treg. CD4⁺ and CD8⁺ EGFP⁺ Treg were expanded upon in vivo administration of a low dose of IL-2.

This new and unique rat line constitutes a useful model to identify and isolate viable CD4⁺ and CD8⁺ FOXP3⁺ Treg. Additionally, it allows to identify molecules expressed in CD8⁺ Treg that may allow to better define their phenotype and function not only in rats but also in other species.