Interleukin (IL)-21-independent pathogen-specific CD8+ T-cell expansion, and IL-21-dependent suppression of CD4+ T-cell IL-17 production

Abstract

Although interleukin-21 (IL-21) potently activates and controls the differentiation of immune cells after stimulation in vitro, the role for this pleiotropic cytokine during in vivo infection remains poorly defined. Herein, the requirement for IL-21 in innate and adaptive host defence after Listeria monocytogenes infection was examined. In the innate phase, IL-21 deficiency did not cause significant defects in infection susceptibility, or in the early activation of natural killer and T cells. In the adaptive phase, L. monocytogenes-specific CD8(+) T cells expand to a similar magnitude in IL-21-deficient mice compared with control mice. Interestingly, the IL-21-independent expansion of L. monocytogenes-specific CD8(+) T cells was maintained even in the combined absence of IL-12 and type I interferon (IFN) receptor. Similarly, L. monocytogenes-specific CD4(+) T cells expanded and produced similar levels of IFN-γ regardless of IL-21 deficiency. Unexpectedly however, IL-21 deficiency caused significantly increased CD4(+) T-cell IL-17 production, and this effect became even more pronounced after L. monocytogenes infection in mice with combined defects in both IL-12 and type I IFN receptor that develop a T helper type 17-dominated CD4(+) T-cell response. Despite increased CD4(+) T-cell IL-17 production, L. monocytogenes-specific T cells re-expanded and conferred protection against secondary challenge with virulent L. monocytogenes regardless of IL-21 deficiency, or combined defects in IL-21, IL-12, and type I IFN receptor. Together, these results demonstrate non-essential individual and combined roles for IL-21, IL-12 and type I IFNs in priming pathogen-specific CD8(+) T cells, and reveal IL-21-dependent suppression of IL-17 production by CD4(+) T cells during in vivo infection.

Figure 1

Interleukin-21 (IL-21) plays a non-essential role in Listeria monocytogenes innate host defence. Number of recoverable L. monocytogenes colony-forming units (CFUs) in the spleen and liver at the indicated time-point after infection with either (a) 10⁵ CFUs, 1 LD₅₀ or (b) 10³ CFUs wild-type L. monocytogenes strain 10403s in B6 control and IL-21-deficient mice. These results are from six to eight mice per group, and are representative of two independent experiments each with similar results. Bar represents one standard error.

Figure 2

Interferon-γ (IFN-γ) production by innate immune cells in the absence of interleukin-21 (IL-21). (a) IFN-γ serum concentration at the indicated time-points after infection with 10⁵ colony-forming units (CFUs) of Listeria monocytogenes 10403s in B6 control and IL-21-deficient mice. (b) Percent (top), and total numbers (bottom) of IFN-γ-producing natural killer (NK) cells (NKp46⁺) in B6 control and IL-21-deficient mice 24 hr post-infection. (c) Percentage (top), and total numbers (bottom) of IFN-γ-producing T cells (CD3⁺) in B6 control and IL-21-deficient mice 24 hr post-infection. These results are from six to nine mice per group, and are representative of three independent experiments each with similar results. Bar represents one standard error; ns, not significant.

Figure 3

Listeria monocytogenes-specific CD8⁺ T-cell expansion in the absence of interleukin-21 (IL-21), or combined absence of IL-21, IL-12, and type I interferons (IFNs). (a) Percentage ovalbumin (OVA_257–264) dimer-positive CD8⁺ T cells in control B6, IL-21-deficient, IL-12 and type I IFN receptor-deficient (DKO), and IL-21, IL-12 and type I IFN receptor-deficient (TKO) mice day 7 post-infection with 10⁶L. monocytogenes-OVA ΔactA. (b) Percentage and total numbers of OVA_257–264 dimer-positive CD8⁺ T cells for each group of mice at the indicated time-points after infection. (c) Percentage and total numbers of IFN-γ-producing CD8⁺ T cells after stimulation with OVA_257–264 peptide (1 μm) for each group of mice at the indicated time-points after infection. These data represent 10–12 mice per experimental group, and are representative of four independent experiments each with similar results. Bar represents standard error.

Figure 4

Listeria monocytogenes-specific CD4⁺ T-cell differentiation in the absence of interleukin-21 (IL-21), or combined absence of IL-21, IL-12 and type I interferons (IFNs). (a) Percentage IL-17- and IFN-γ-producing CD4⁺ T cells after stimulation with listeriolysin O (LLO_189–201) peptide or no peptide for control B6, IL-21-deficient, IL-12 and type I IFN receptor-deficient (DKO), and IL-21, IL-12 and type I IFN receptor-deficient (TKO) mice on day 7 post-infection with 10⁶L. monocytogenes-OVA ΔactA. (b) Percentage and total numbers of IL-17-producing CD4⁺ T cells after stimulation with LLO_189–201 peptide for each group of mice at the indicated time-points after infection. (c) Percentage and total numbers of IFN-γ-producing CD4⁺ T cells after stimulation with LLO_189–201 peptide for each group of mice at the indicated time-points after infection. These data represent 10–12 mice per experimental group, and are representative of four independent experiments, each with similar results. Bar represents standard error. *P < 0·05, **P < 0·01.

Figure 5

Protective immunity to secondary Listeria monocytogenes infection and re-expansion of L. monocytogenes-specific CD8⁺ and CD4⁺ T cells in the absence of interleukin-21 (IL-21), or combined absence of IL-21, IL-12 and type I interferons (IFNs). (a) Number of recoverable L. monocytogenes colony-forming units (CFUs) at the indicated time-points post-infection with 10⁵L. monocytogenes–ovalbumin (OVA) for groups of either naive mice or mice inoculated with L. monocytogenes-OVA ΔactA 30 days before infection. (b) Total numbers of OVA_257–264 dimer-positive (left) and IFN-γ-producing right CD8⁺ T cells after stimulation with OVA_257–264 peptide before (D30), and on day 3 (30 + 3) and day 5 (30 + 5) after secondary infection with L. monocytogenes-OVA. (c) Total numbers of IL-17-producing (left) and IFN-γ-producing (right) CD4⁺ T cells after stimulation with listeriolysin O (LLO_189–201) peptide before (D30), and on day 3 (30 + 3) and day 5 (30 + 5) after secondary infection with L. monocytogenes-OVA. These data represent 12 mice per experimental group, and are representative of three independent experiments each with similar results. Bar represents standard error.