Daan Pieren

109 Co-expression of TIGIT and Helios by CD8+ T cells and indicate that TIGIT + Helios + T cells accumulate in older adults not only as late-differentiated cells but also as intermediate-differentiated cells. Figure 6, aang past TIGIT + Helios + A B C Helios + TIGIT + 0 20 40 60 80 100 0 5 10 15 20 40 60 80 Age (Years) % of subset in CD8 + T cells Early TIGIT + Helios + Intermediate TIGIT + Helios + Late TIGIT + Helios + r p 0.367 ** 0.290 * 0.212 ns 0 20 40 60 80 100 0 20 40 60 80 Age (Years) % of subset in CD8 + T cells 0.454 *** 0.246 0.0268 ns 0.08 Early TIGIT + Intermediate TIGIT + Late TIGIT + r p 0 20 40 60 80 100 0 20 40 60 80 Age (Years) % of subset in CD8 + T cells 0.340 * 0.206 ns -0.297 * Early Helios + Intermediate Helios + Late Helios + r p Figure 6. TIGIT + Helios + intermediate- and TIGIT + Helios + late-differentiated cells accumulate by aging within the CD8 + T-cell population. Frequency of ( A ) early-, intermediate-, or late-differentiated TIGIT + cells, ( B ) early-, intermediate-, or late-differentiated Helios + cells, ( C ) early-, intermediate-, or late-differentiated TIGIT + Helios + cells within the CD8 + T-cell population and their relationship with age (n=50). Correlations ( r and p values) were assessed by Spearman test with p <0.05 considered as statistically significant. (* p <0.05, ** p <0.01, *** p <0.001, ns=not significant). DISCUSSION The accumulation of late-differentiated CD8 + T cells at older age is a widely used marker of immunological aging due to features of immunosenescence [10,12]. We here identified co-expression of TIGIT and Helios as an accurate hallmark of immunosenescent CD8 + T cells. We show that these immunosenescent cells accumulate with age not only by the increase of the late-differentiated population, but also by aging-related development of immunosenescence already within earlier stages of differentiation. Therefore, accumulation of functionally immunosenescent cells at old age is partly independent of the final progression into late stage of differentiation. These findings indicate that aging- related development of immunosenescence should be analyzed by hallmarks of immunosenescence that complement classic analyses of late differentiation state using markers such as CD27 and CD28. TIGIT has recently been identified as a marker for immunosenescent CD8 + T cells that accumulate at older age [13-15]. By including Helios in our analyses, we found that the TIGIT + T-cell population is heterogeneous and that only a part of the TIGIT + T cells co-expressed Helios. Particularly the TIGIT + Helios + T-cell subset 4