GM-CSF-deficient mice are susceptible to pulmonary group B streptococcal infection

Abstract

Granulocyte-macrophage colony-stimulating factor (GM-CSF) gene-targeted mice (GM-/-) cleared group B streptococcus (GBS) from the lungs more slowly than wild-type mice. Expression of GM-CSF in the respiratory epithelium of GM-/- mice improved bacterial clearance to levels greater than that in wild-type GM+/+ mice. Acute aerosolization of GM-CSF to GM+/+ mice significantly enhanced clearance of GBS at 24 hours. GBS infection was associated with increased neutrophilic infiltration in lungs of GM-/- mice, while macrophage infiltrates predominated in wild-type mice, suggesting an abnormality in macrophage clearance of bacteria in the absence of GM-CSF. While phagocytosis of GBS was unaltered, production of superoxide radicals and hydrogen peroxide was markedly deficient in macrophages from GM-/- mice. Lipid peroxidation, assessed by measuring the isoprostane 8-iso-PGF2alpha, was decreased in the lungs of GM-/- mice. GM-CSF plays an important role in GBS clearance in vivo, mediated in part by its role in enhancing superoxide and hydrogen peroxide production and bacterial killing by alveolar macrophages.

Figure 1

Increased GBS in lung homogenates from GM^–/– mice. Concentration of GBS was determined by quantitative cultures of lung homogenates. Colony counts were significantly greater 6, 24, and 48 h after administration of 10⁴ CFU GBS in GM^–/– (solid bars) compared with GM^+/+ (hatched bars) mice. Six hours after GBS infection, clearance of GBS from the lung was enhanced in SP-C-GM (open bars) mice compared with GM^+/+ mice. Data were not obtained for SP-C-GM mice at 48 h. Data are mean ± SEM with n = 10 mice per group. *P < 0.05 compared with GM^+/+ mice. GBS, group B streptococcus; GM^–/–, GM-CSF gene-targeted mice; SP-C-GM, human surfactant protein C (SP-C) gene promoter expressing GM-CSF.

Figure 2

Decreased 8-iso-PGF_2α in lung homogenates and BAL fluid from GM^–/– mice. Concentrations of 8-iso-prostaglandin F_2α (8-iso-PGF_2α) were assessed in lung homogenates (a) and BAL fluid (b) from GM^–/– (solid bars), and GM^+/+ (hatched bars) mice. Prostaglandin F₂–like compounds are formed by free radical–catalyzed peroxidation of arachidonic acid, independent of cyclooxygenase enzyme. Decreased concentrations of 8-iso-PGF_2α were found in lung homogenates and BAL fluid from the GM ^–/– mice 18 h after GBS infection. Data are expressed as ng/ml for the lung homogenates and pg/ml for BAL fluid and represent mean ± SEM with n = 8 mice per group. *P < 0.05 compared with GM^+/+ mice. BAL, bronchoalveolar lavage.

Figure 3

Increased TNF-α, IL-6, and MIP-2 in lung homogenates from GM^–/– mice after GBS infection. Concentrations of TNF-α, IL-6, and MIP-2 were assessed in lung homogenates from GM^–/– (solid bars), GM^+/+ (hatched bar), and SP-C-GM (open bars) mice. Increased concentrations of the proinflammatory cytokines TNF-α, IL-6, and MIP-2 were found in lung homogenates from the GM^–/– mice at 24 h. TNF-α levels were increased in lung homogenates from SP-C-GM compared with GM^+/+ mice at 24 h. Data are expressed as pg/ml and represent mean ± SEM with n = 10 mice per group. *P < 0.05 compared with GM^+/+ mice. IL-6, interleukin-6; MIP-2, macrophage inflammatory protein-2; TNF-α, tumor necrosis factor-α.

Figure 4

Increased nitrite concentrations in BAL fluid from GM^–/– mice after GBS infection. Nitrite in the BAL fluid was measured by the Griess reaction as described in Methods. Six and 24 h after intratracheal inoculation of 10⁴ CFU GBS, nitrite (a) and nitrite/nitrate (b) levels in BAL fluid from GM^–/– mice (solid bars) were increased compared with GM^+/+ (hatched bars) and SP-C-GM (open bars) mice. Data represent mean ± SEM with n = 5 mice per group. *P < 0.05 compared with GM^+/+ mice.

Figure 5

Exogenous GM-CSF enhances GBS clearance from the lung. Concentration of GBS was determined by quantitative cultures of lung homogenates. Colony counts were significantly decreased 24 h after administration of 10⁴ CFU GBS to wild-type mice (solid) treated with three doses of aerosolized recombinant mouse GM-CSF (4 ng/mouse) compared with untreated controls (hatched bars). Data are mean ± SEM with n = 15 mice per group. *P < 0.05 compared with untreated wild-type mice.