Daan Pieren

36 Chapter 2 older age ( Figure 2A ). Stronger stimulation induced higher rates of proliferation in Th and Tc cells of young mice, which was most pronounced after four days ( Figure 2B ). In contrast, the Th and Tc cells of mice of 22 months and older did not show any proliferative response to these stimuli over time. Proliferation of Treg cells of young mice did not increase in response to higher strength of stimulation, but Treg cells of young mice proliferated significantly more compared to aged mice under any condition tested. These data indicate a loss of capacity to proliferate at old age across all T cell subsets that cannot be overcome by increasing the stimulatory strength. Young (2 months) Aged (17-18 months) Aged (22-24 months) % of max CellTrace A Proliferation B Low Intermediate High Time (Days) Proliferation (Fold change Cellterace MFI) - Low Int High - Low Int High - Low Int High - Low Int High - Low Int High - Low Int High - Low Int High - Low Int High - Low Int High Th cells Tc cells Treg cells Young (2 months) Aged (22-24 months) 1 2 3 4 0 1 2 3 4 0 1 2 3 4 0 2 4 6 8 10 *** *** *** *** 1 2 3 4 0 1 2 3 4 0 1 2 3 4 0 2 4 6 8 10 20 25 30 * ** *** *** 1 2 3 4 0 1 2 3 4 0 1 2 3 4 0 2 4 6 8 10 *** *** * *** Figure 2. Gradual loss of proliferative capacity in Th, Tc, and Treg cells with ageing. ( A ) The proliferative capacity of Th, Tc, and Treg cells of young (2 months old) and aged (17-18 and 22-24 months old) mice was measured four days after exposure to a low, intermediate, or high stimulatory strength. For each condition graphs show data of one mouse that is representative of n=6 per age group. ( B ) Graphs show proliferation of Th, Tc, and Treg cells by fold change of proliferation marker Celltrace in young (n=6, 2 months old) and aged (n=6, 22-24 months old) mice after four days cultured in the presence of a low, intermediate, or high stimulatory strength. Mean ± SEM; * p < 0.05, ** p < 0.01, *** p < 0.001 for difference between young and aged mice using Two-way ANOVA.