Daan Pieren

82 Chapter 3 explanation might be that we used whole spleen cultures that allow interactions between different subsets of T cells, whereas Hurez et al . studied an isolated subfraction of CD4 + T cells. Our spleen cell cultures included eRapa-induced CD25 + and PD-1 + memory Tregs. CD25 + and PD-1 + memory Tregs of WT aged mice have been shown to comprise a Treg subset with enhanced suppressive capacity [8], which may suggest suppression of Th- and Tc-cell proliferation in our cultures by these eRapa-induced memory Tregs. Generation of CD25 + and PD-1 + memory Tregs by eRapa may be explained by upregulation of the memory marker CD44 on naive Tregs, since in vivo treatment with rapamycin can induce expression of CD44 and PD-1 FoxP3 + Tregs [48]. Alternatively, memory FoxP3 - CD4 + T cells may have upregulated FoxP3 following eRapa. However, this is less likely as Foxp3 - effector T cells and CD4 + CD25 - cells exposed in vitro to rapamycin do not develop into FoxP3 + Tregs [49,50]. Future studies on the effects of eRapa on proliferation of isolated T-cell fractions and whole-cell cultures will be important to provide full insight in the immune modulatory properties of eRapa. Our data show that compromised DNA repair contributes to increased frequencies of memory T cells, but not to the extent found in WT aged mice. Since antigens are a major driving force behind the formation of memory T cells [10], the higher level of inflation of memory T cells found in WT aged mice is likely explained by the higher antigenic exposure over their longer lifetime. These findings indicate that compromised DNA repair partly contributes to aging-related phenotypic T-cell alterations and that aging of the T-cell pool is a diverse process that is driven by multiple factors in addition to DNA-damage. Interestingly, eRapa decreased the frequency of memory T cells found in Ercc1 -/ Δ 7 mice at the benefit of a rise in the proportion of naive T cells, which is consistent with a previous report in WT mice [29]. Since higher frequencies of naive cells at young age are linked to better T-cell mediated protection, we predict that the eRapa-induced rise of the naive T-cell frequency may contribute to improved T-cell mediated protection. This is supported by studies showing rapamycin improved T-cell function against pathogens [51] and antigen-specific T-cell responses [52]. Although our data indicate that compromised DNA damage repair contributes to aging of the immune system, there are limitations regarding our study. First, although Ercc1 -deficient mice show compromised DNA repair in at least four repair pathways [12], it does not account for deficiencies in other DNA-repair pathways, such as nonhomologous end-joining or base-excision repair [53]. It is therefore possible that defects in DNA-repair mechanisms other than those mediated by Ercc1 -deficiency have their own characteristic impact