Investigating human KIR+CD8+ and NKG2A+CD8+ T cells biology in peripheral blood and in pediatric thymuses

Virtual memory T cells(TVMs) are a recently described population of TCRαβ+CD8+T cells described in mice that, despite their antigen inexperience, express markers of conventional memory cells1. Recently, two populations of circulating TVMs expressing KIRs or NKG2A have been described also in humans and, as their murine counterpart, human TVMs are characterized by a high expression of the transcription-factor Eomes2. Contrary to mice, little is known about human TVM thymic development. Few studies suggest a possible role of IL4, IL15 and type-I-interferons(IFNs-I) in TVM development in humans3-4. To investigate the role of IL4, IL15 and IFNs-I and their combination during thymic development of human TVMs but also in the peripheral counterpart, we stimulated human pediatric thymocytes and adult peripheral-blood-mononuclear-cells (PBMCs) in-vitro, in the presence or absence of these cytokines, and we analyzed the cellular phenotype by flow-cytometry. After 9 days of thymocyte culture, we observed the development of a population of TCRαβ+CD8+Eomes+cells only in cells cultured with IL4 and IL4+IL15. Unlike their circulating counterpart, these cells were characterized by a naïve phenotype, lacking the expression of NKG2A/KIRs that may possibly be acquired later during their development. Performing similar experiments on PBMCs, we observed that, as in mice, IL4+IL15 stimulation induced the expansion of TCRαβ+CD8+Eomes+cells either expressing KIRs or NKG2A. Moreover, PBMCs stimulated with IL15+IFNs-I showed increased cytotoxic-potential characterized by an increased production of GranzymeB and Perforin. Altogether, our results demonstrate that, similar to mice, human TVM development is dependent on IL4+IL15 but, TVMs functional properties are sustained by IL15+IFN stimulation.