Roles of interleukin-17 in an experimental Legionella pneumophila pneumonia model

Abstract

Interleukin-17 (IL-17) is a key factor in T helper type 17 (Th17) lineage host responses and plays critical roles in immunological control of a variety of infectious diseases. Although Legionella pneumophila, an intracellular bacterium found widely in the environment, often causes a serious and life-threatening pneumonia in humans, the contribution of IL-17 to immune function during Legionella pneumonia is unknown. In the present study, we used an experimental Legionella pneumonia infection to clarify the role of IL-17 in the resulting immune response. We observed robust production of pulmonary IL-17A and IL-17F (IL-17A/F), peaking on day 1 and declining thereafter. Upregulated production of tumor necrosis factor alpha (TNF-α), IL-6, and IL-1β, but not monocyte chemotactic protein 1 (MCP-1), was observed in Legionella-infected bone marrow-derived macrophages from BALB/c mice that had been stimulated with IL-17A or IL-17F. A significant decrease in the production of proinflammatory cytokines IL-6 and TNF-α was observed in IL-17A/F-deficient mice (BALB/c background) infected with L. pneumophila. Moreover, we found impaired neutrophil migration and lower numbers of chemokines (KC, LIX, and MIP-2) in IL-17A/F-deficient mice. IL-17A/F-deficient mice also eliminated L. pneumophila more slowly and were less likely to survive a lethal challenge. These results demonstrate that IL-17A/F plays a critical role in L. pneumophila pneumonia, probably through induction of proinflammatory cytokines and accumulation of neutrophils at the infection site.

Fig 1

Production of IL-17A and IL-17F in the lung during L. pneumophila pneumonia in a mouse model. IL-17A (A) and IL-17F (B) levels are shown as a function of time after the inoculation of approximately 10⁵ CFU of L. pneumophila. All bars indicate mean ± SE (n = 6/group). *, P < 0.05 in comparison with a negative control. Similar results were obtained across three repeat experiments.

Fig 2

Stimulation of macrophage (BALB/c) proinflammatory cytokines, as indicated on the figure, by IL-17A and IL-17F at 24 h after infection with L. pneumophila (10⁶ CFU/ml; MOI of 1.0). All bars indicate mean ± SE (n = 5/group). *, P < 0.01 in a comparison with the infection-only group; **, P < 0.05 in a comparison between IL-17A and IL-17F. The results are representative of three independent experiments.

Fig 3

Proinflammatory cytokine production in the lungs of wild-type and IL-17A/F-deficient mice (BALB/c background) during the week following infection with L. pneumophila. The production of IL-6, TNF-α, IL-1β, and MCP-1 proteins was examined at 1, 2, 4, and 7 days after infection with L. pneumophila (10⁵ CFU). All bars indicate mean ± SE (n = 6/group). *, P < 0.05 in comparisons between Il17a^−/− Il17f^−/− (A/F*) and wild-type (WT) mice. Results are representative of two independent experiments.

Fig 4

Leukocyte accumulation and gene expression of CXC chemokines in lungs of BALB/c mice infected with Legionella. (A) Total cell numbers and proportions of different inflammatory cells in the BALF of wild-type and IL-17A/F-deficient (A/F*) mice (BALB/c background) at 1, 2, 4, and 7 days after infection with 10⁵ CFU of L. pneumophila. *, P < 0.05 for a comparison between the neutrophil numbers of Il17a^−/− Il17f^−/− (A/F*) and wild-type (WT) mice. (B) Expression of the chemokines CXCL-1 (KC), CXCL-5 (LIX), CXCL-2 (MIP-2), IL-12p35, IFN-γ, and IL-4 in whole-lung tissue of mice at 48 h after infection with L. pneumophila. Data are expressed as fold increase. *, P < 0.05 for a comparison between Il17a^−/− Il17f^−/− (A/F*) and wild-type (WT) mice. Bars indicate mean ± SE (n = 5/group). Results are representative of three independent experiments.

Fig 5

Numbers of bacteria in the lungs of wild-type and IL-17A/F-deficient mice (BALB/c background) infected with L. pneumophila. Bacteria were counted at 1, 2, 4, and 7 days after inoculation of 10⁵ CFU of L. pneumophila. *, P < 0.01 for a comparison between Il17a^−/− Il17f^−/− (A/F*) and wild-type (WT) mice. Bars indicate mean ± SE (n = 6/group). Results are representative of two independent experiments.

Fig 6

Survival curves for wild-type (WT; n = 26) and IL-17A/F-deficient (A/F*; n = 16) mice (BALB/c background) infected with L. pneumophila. Results are representative of two independent experiments.