Decreased pulmonary inflammation after ethanol exposure and burn injury in intercellular adhesion molecule-1 knockout mice

Abstract

Clinical and laboratory evidence suggests that alcohol consumption dysregulates immune function. Burn patients who consume alcohol before their injuries demonstrate higher rates of morbidity and mortality, including acute respiratory distress syndrome, than patients without alcohol at the time of injury. Our laboratory observed higher levels of proinflammatory cytokines and leukocyte infiltration in the lungs of mice after ethanol exposure and burn injury than with either insult alone. To understand the mechanism of the increased pulmonary inflammatory response in mice treated with ethanol and burn injury, we investigated the role of intercellular adhesion molecule (ICAM)-1. Wild-type and ICAM-1 knockout (KO) mice were treated with vehicle or ethanol and subsequently given a sham or burn injury. Twenty-four hours postinjury, lungs were harvested and analyzed for indices of inflammation. Higher numbers of neutrophils were observed in the lungs of wild-type mice after burn and burn with ethanol treatment. This increase in pulmonary inflammatory cell accumulation was significantly lower in the KO mice. In addition, levels of KC, interleukin-1beta, and interleukin-6 in the lung were decreased in the ICAM-1 KO mice after ethanol exposure and burn injury. Interestingly, no differences were observed in serum or lung tissue content of soluble ICAM-1 24 hours postinjury. These data suggest that upregulation of adhesion molecules such as ICAM-1 on the vascular endothelium may play a critical role in the excessive inflammation seen after ethanol exposure and burn injury.

Figure 1

Inflammation after ethanol exposure and burn injury in the lung. A, Lung sections from wild-type (WT) and intercellular adhesion molecule-1 knockout (ICAM-1 KO) mice were examined for degree of inflammation 24 hours after ethanol exposure and burn injury. Representative micrographs of hematoxylin and eosin lung sections are shown. All images are at magnification, 400×. B, Enlarged image of lung sections in A from WT and ICAM-1 KO mice after ethanol exposure and burn injury. Arrowheads indicate areas of neutrophils and thickened alveolar walls.

Figure 2

Neutrophil infiltration in the lung. Neutrophils were counted by light microscopy in hemotoxylin and eosin-stained lung sections 24 hours after ethanol exposure and burn injury. Data are shown as the total number of neutrophils in 10 high-power fields (magnification, 400×) ± SEM. *P < .001 compared with all groups. N = 7–12 animals per group.

Figure 3

Neutrophil infiltration after ethanol exposure and burn injury in the lung. Lung sections from wild-type (WT) were stained by flurochrome-conjugated antibodies against Gr-1 (green) and MOMA-2 (red) to detect the presence of neutrophils and macrophages, respectively. Representative micrographs are shown. All images are at magnification, 400×.

Figure 4

Decreased pulmonary levels of proinflammatory mediators after ethanol exposure and burn injury in intercellular adhesion molecule-1 knockout (ICAM-1 KO) mice. Levels of KC (A) and interleukin (IL)-6 (B) in the lung were quantified by multiplex assay in total lung homogenates. Cytokine concentrations were normalized to total protein in the sample as determined by BioRad protein assay and presented as concentration ± SEM. *P < .01 compared with all other groups, #P < .05 compared with all groups but sham ethanol knockout. N = 7–12 animals per group. No significant differences were seen in monocyte inflammatory protein-2 production in the lung.

Figure 5

No differences in soluble ICAM-1 in serum or lung after ethanol exposure and burn injury. Levels of soluble ICAM (sICAM)-1 in serum (A) and lung homogenates (B) were quantified by ELISA. Soluble ICAM-1 levels in the lung were normalized to total protein in the sample as determined by BioRad protein assay and presented as concentration ± SEM. Dashed line represents the level of sICAM-1 in the serum and lungs of ICAM-1 KO mice. N = 7–12 animals per group.