Daan Pieren

168 Chapter 6 as bystander cells [38]. Moreover, virtual memory cells with characteristics of cellular senescence accumulate with age in unprimed aged mice [39]. Together, these observations imply that virtual memory cells are capable of contributing to protective immunity, which is subject to age-related changes that may diminish their effectivity despite their accumulation with age. Virtual memory cells have also been identified in humans, characterized by expression of CD45RA + CD8 + T cells that express several Killer-cell Immunoglobulin like Receptors (KIR) and/or the inhibitory natural-killer cell (NK cell) receptor NKG2A [40]. Similar to findings in mice, human virtual memory cells accumulate with age and show reduced capacity to proliferate [41]. However, further phenotypical and functional characterization of human virtual memory cells and their clinical relevance in the context of infectious disease in older adults remained unexplored. Identification of KIR + RA + and NKG2A + RA + CD8 + T cells within the Human Virtual Memory Subset In chapter 5 we discovered that the accumulation of virtual memory cells with age was caused by an increase in the proportion of KIR + NKG2A - cells, whereas we found a decline in KIR - NKG2A + cells with age ( Figure 2A ). These findings indicated that the human virtual memory subset may actually consist of two different cell subsets. We show that these cells indeed express differing transcriptional phenotypes and that the transcriptional phenotype of CD45RA + KIR + NKG2A - cells (which we named KIR + RA + T cells based on their distinctive co-expression of KIR and CD45RA) was enriched for regulatory markers ( Figure 2B ). These findings encouraged us to further explore the phenotype and functionality of KIR + RA + T cells, as these cells accumulated with age and appeared to be a relatively large subset amongst the CD8 + T-cell population. At the protein level, we found that KIR + RA + T cells express a phenotype that could be linked to regulatory T cells, as they expressed TIGIT, Helios, and CD122 [42-46]. Furthermore, we found that KIR + RA + T cells have a low proliferative capacity and low IL-2 production. Interestingly, we found that KIR + RA + T cells suppressed proliferation of other CD8 + T cells and therefore can be considered to be a regulatory cell subset. Moreover, we found KIR + RA + T cells to be highly activated in the blood of older adults in response to acute respiratory viral infection. Importantly, activation of KIR + RA + T cells was associated with prolonged presence of symptoms in older adults infected with Influenza A virus, indicating that these regulatory KIR + RA + T cells may aggravate respiratory disease following viral infection, possibly by suppressing anti-viral CD8 + T cells.