The interleukin-1β/CXCL1/2/neutrophil axis mediates host protection against group B streptococcal infection

Abstract

Previous studies have indicated that group B streptococcus (GBS), a frequent human pathogen, potently induces the release of interleukin-1β (IL-1β), an important mediator of inflammatory responses. Since little is known about the role of this cytokine in GBS disease, we analyzed the outcome of infection in IL-1β-deficient mice. These animals were markedly sensitive to GBS infection, with most of them dying under challenge conditions that caused no deaths in wild-type control mice. Lethality was due to the inability of the IL-1β-deficient mice to control local GBS replication and dissemination to target organs, such as the brain and the kidneys. Moreover, in a model of inflammation induced by the intraperitoneal injection of killed GBS, a lack of IL-1β was associated with selective impairment in the production of the neutrophil chemokines CXCL1 and CXCL2 and in neutrophil recruitment to the peritoneal cavity. Decreased blood neutrophil counts and impaired neutrophil recruitment to the brain and kidneys were also observed during GBS infection in IL-1β-deficient mice concomitantly with a reduction in CXCL1 and CXCL2 tissue levels. Notably, the hypersusceptibility to GBS infection observed in the immune-deficient animals was recapitulated by neutrophil depletion with anti-Gr1 antibodies. Collectively, our data identify a cytokine circuit that involves IL-1β-induced production of CXCL1 and CXCL2 and leads the recruitment of neutrophils to GBS infection sites. Moreover, our data point to an essential role of these cells in controlling the progression and outcome of GBS disease.

FIG 1

Mice lacking IL-1β are highly susceptible to GBS infection. (A) Survival of WT or IL-1β-deficient mice after i.p. challenge with 2 × 10⁵ CFU of GBS. Shown are cumulative data from two experiments, each involving 9 animals per group. *, P < 0.05 versus WT mice determined with Kaplan-Meier survival plots. (B and C) Numbers of bacterial CFU/ml in peritoneal lavage fluid (B) and blood (C) of WT or IL-1β-deficient mice at 24 h after i.p. challenge with 2 × 10⁵ CFU of GBS. Horizontal bars indicate mean values. Each determination was conducted on a different animal in the course of two experiments, each involving 7 animals per group. *, P < 0.05 versus WT mice determined by one-way analysis of variance and the Student's-Keuls-Newman test.

FIG 2

Effects of a lack of IL-1β on hematogenous colonization of target organs by GBS. (A to C) Colony counts in the blood, brain, and kidney at 24 and 48 h after i.v. infection with 1 × 10⁶ CFU of GBS. Horizontal bars indicate mean values. Each determination was conducted on a different animal in the course of two experiments, each involving 7 animals per group. *, P < 0.05 versus WT mice determined by one-way analysis of variance and the Student's-Keuls-Newman test. (D) Survival of WT C57BL/6 and IL-1β-deficient mice after i.v. challenge with 1 × 10⁶ CFU of GBS. Shown are the cumulative results of two experiments, each involving 8 animals per group. *, P < 0.05 versus WT mice determined with Kaplan-Meier survival plot.

FIG 3

Impaired neutrophil recruitment in IL-1β-deficient mice during GBS-induced peritonitis. (A to E) Cell counts in peritoneal lavage fluid samples after i.p. challenge with heat-killed GBS (0.5 mg/mouse). (A) Kinetics of total cell influx in WT and IL-1β-defective mice; (B to E) kinetics of recruitment of cells positive for F4/80 (macrophages; B), CD3 (T lymphocytes; C), CD19 (B lymphocytes; D), and GR1 (granulocytes; E) in WT and IL-1β-defective mice. (F) Blood neutrophil counts in WT and IL-1β-defective mice before and after i.p. challenge with 2 × 10⁵ GBS. Data are expressed as the means + SDs of three determinations, each conducted in a different animal, during the course of one of three experiments producing similar results. *, P < 0.05 relative to WT mice determined by one-way analysis of variance and the Student's-Keuls-Newman test.

FIG 4

Kinetics of cytokine production in peritoneal lavage fluid after challenge with killed GBS. (A to D) GM-CSF, CXCL1 (KC), CXCL2 (MIP-2), and TNF-α protein levels in peritoneal lavage fluid from WT and IL-1β-deficient mice were measured at the indicated times after i.p. injection of killed GBS (0.5 mg/mouse). Data are expressed as the means ± SDs of three observations, each conducted on a different animal, during the course of one of two experiments producing similar results. *, P < 0.05 versus WT mice determined by one-way analysis of variance and the Student's-Keuls-Newman test.

FIG 5

Effect of a lack of IL-1β on chemokine production and neutrophil influx during GBS infection. CXCL1 (KC), CXCL2 (MIP-2), IL-1β, and MPO protein levels (A to D) or numbers of CFU (E and F) in organ homogenates from WT and IL-1β-deficient mice were measured at the indicated times after i.v. infection with 1 × 10⁶ CFU of GBS. Data are expressed as the means ± SDs of three observations, each conducted on a different animal, during the course of one experiment. Horizontal bars indicate mean values. *, P < 0.05 versus WT mice determined by one-way analysis of variance and the Student's-Keuls-Newman test.

FIG 6

Effects of neutrophil depletion on anti-GBS defenses. (A and B) Number of peritoneal cells positive for F4/80 (macrophages), CD3 (T lymphocytes), CD19 (B lymphocytes), and GR1 (granulocytes) in WT mice pretreated with anti-Ly-6G antibody or the isotype control after challenge with heat-killed GBS (0.5 mg/mouse). Data are expressed as the means + SDs of three independent experiments. *, P < 0.05 relative to isotype control-pretreated mice by one-way analysis of variance and the Student's-Keuls-Newman test. (C) Survival of WT mice pretreated with anti-Ly-6G antibody or the isotype control after i.p. challenge with 2 × 10⁵ CFU of GBS. Shown are the cumulative results of two experiments, each involving 8 animals per group. *, P < 0.05 relative to isotype control-pretreated mice determined with Kaplan-Meier survival plots. (D to F) Blood, kidney, and peritoneal lavage fluid bacterial numbers in WT mice pretreated with anti-Ly-6G antibody or the isotype control at 24 h after i.p. challenge with 2 × 10⁵ CFU of GBS. Each determination was conducted on a different animal in the course of one experiment involving 8 animals per group. *, P < 0.05 relative to isotype control-pretreated mice by one-way analysis of variance and the Student's-Keuls-Newman test.

FIG 7

Administration of rIL-1β restores host defenses in IL-1β-deficient mice. (A) Survival of IL-1β-deficient mice treated with rIL-1β or vehicle after i.p. challenge with 2 × 10⁵ CFU of GBS. *, P < 0.05 versus IL-1β-deficient mice treated with rIL-1β determined with Kaplan-Meier survival plots. Shown are cumulative results from two experiments, each involving 8 animals per group. (B to D) Blood, peritoneal lavage fluid, and kidney colony counts in IL-1β-deficient mice treated with rIL-1β or vehicle at 24 h after i.p. challenge with 2 × 10⁵ CFU of GBS. *, P < 0.05 versus IL-1β-deficient mice treated with rIL-1β by one-way analysis of variance and the Student's-Keuls-Newman test. Each determination was conducted on a different animal in the course of one experiment involving 8 animals per group.