Interleukin-2 signals during priming are required for secondary expansion of CD8+ memory T cells

Abstract

Although interleukin-2 (IL-2) was initially characterized as the primary T-cell growth factor following in vitro activation, less is known about its role in shaping T-cell responses to acute infections in vivo. The use of IL-2- or IL-2-receptor-deficient mice is problematic owing to their early development of autoimmunity, attributable to the central role of IL-2 in the generation, maintenance and function of CD4+CD25+ regulatory T cells. To bypass these inherent difficulties, we have studied the effect of IL-2 on T-cell responses to acute infections by adopting a mixed chimaera strategy in which T cells lacking the high-affinity IL-2 receptor could be studied in an otherwise healthy mouse containing a full complement of regulatory T cells. Here we show that although IL-2 signalling to pathogen-specific CD8+ T cells affects the number of developing effector and memory cells very little, it is required for the generation of robust secondary responses. This is not due to an altered T-cell-receptor repertoire development or selection, and does not reflect an acute requirement for IL-2 during secondary activation and expansion. Rather, we demonstrate a previously unappreciated role for IL-2 during primary infection in programming the development of CD8+ memory T cells capable of full secondary expansion. These results have important implications for the development of vaccination or immunotherapeutic strategies aimed at boosting memory T-cell function.

Figure 1. IL-2Rα-deficient CD8⁺ T cells generate robust primary but defective secondary responses

a, WT/IL-2Rα^−/− mixed chimaeras were infected with 2 × 10⁵ plaque-forming units (PFU) LCMV and assessed for the frequency of interferon-γ (IFNγ)-producing cells specific for the GP_33–41 epitope among either WTor IL-2Rα-deficient CD8⁺ T cells in the spleen. In all experiments, CD45.1⁺ host cells were excluded. Mice were re-challenged 150 days post-infection with 1 × 10⁵ colony-forming units (CFU) LM-GP, and GP_33–41-specific responses in the spleen were analysed 5 days later (day 150 + 5). b, The ratio of GP_33–41-specific WT to IL-2Rα-deficient responders is shown at each time point. c, The fold expansion by 5 days after re-challenge is shown for WT and IL-2Rα-deficient T cells. Error bars display s.e.m. (n = 3–4) and results are representative of five separate time courses.

Figure 2. IL-2Rα-deficient memory cells are maintained at normal levels

a, 1 × 10⁴ naive WT and IL-2Rα-deficient P14 cells were co-transferred into B6 hosts, infected with LCMV, and frequencies of P14 cells were measured in the spleen. b, The total number of WT or IL-2Rα-deficient P14 cells in the spleen is shown over a 90 day time course. c, At day 90 post-infection, mice were fed BrdU in their drinking water for 7 days and stained for BrdU incorporation by WT or IL-2Rα-deficient P14 cells. d, Cell surface expression of the indicated molecules by WT or IL-2Rα-deficient P14 cells at days 8, 15, 42 and 90 post-infection. Results are representative of 3 separate time courses and error bars display s.e.m. (n = 3–4).

Figure 3. IL-2Rα-deficient memory cells proliferate but do not accumulate following rechallenge

a, At 42 days post-infection, WT and IL-2Rα-deficient memory P14 cells were sorted, transferred into new B6 hosts (1 × 10⁴ of each) and rechallenged with LCMV. Eight days after secondary challenge, we measured their frequency in the spleen. Results were consistent for four separate experiments. b, The ratio of WT to IL-2Rα-deficient P14 cells was assessed at days 8 and 42 post-infection, as well as day 8 after re-challenge. Error bars display s.e.m. (n = 3–5) and are representative of four separate experiments. c, At day 90 post-infection, WT and IL-2Rα-deficient memory P14 cells were labelled with CFSE, transferred to a new host and re-challenged with LCMV. Flow plots indicate CFSE divisions after three days, with dashed lines representing uninfected controls, and the graph displays the fold expansion of each group. Error bars display s.e.m. (n = 3).

Figure 4. IL-2 signalling during the primary response promotes secondary CD8⁺ T cell responsiveness

a, Following co-transfer of 1 × 10⁴ naive WT and IL-2Rα-deficient P14 cells, B6 hosts were infected with LCMV and received daily intraperitoneal injections of 50μg anti-IL-2 (clone S4B6) alone, daily co-injections of anti-IL-2 and 1.5μg recombinant mouse IL-2 on days 0–6 of the primary infection, or no treatment (No Rx). Plots display the frequency and CD62L expression of WT and IL-2Rα-deficient P14 cells at day 19 post-infection. b, WT/IL-2Rα^−/− mixed chimaeras were infected with LCMV and re-challenged with LM-GP 6 weeks later. Mice were treated on either days 0–6 of the primary infection or days 0–4 of the secondary infection. Plots display the frequency of GP_33–41-specific WT or IL-2Rα-deficient CD8⁺ T cells at day 5 post-rechallenge. c, The graph displays the fold expansion by 5 days post-rechallenge. Error bars display s.e.m. (n = 3).