Fig 1.
Diagram from C.L. Pavaux [26]. The black circles correspond to areas where samples were taken on each pig (cranial left lung lobe, caudal left lung lobe, cranial right lung lobe, middle right lung lobe, accessory right lung lobe and caudal right lung lobe), trachea (d: bronchial crossroads) and tracheal lymph nodes.
Table 1.
Clinical follow-up, blood biochemistry and C-reactive protein in pigs after P. aeruginosa infection.
Fig 2.
Cytokine concentrations in the BAL and serum of pigs after P. aeruginosa infection.
A. IL-6, IL-8 and TNF-α in BAL fluid. B. IL-6, IL-8 and TNF-α in serum. Data were analysed by two-way ANOVA followed by Bonferroni's post hoc test. Data are means ± S.E.M. * indicates p<0.05. *** indicates p<0.001.
Fig 3.
Effect of P. aeruginosa infection on the reactivity of porcine bronchial smooth muscle cells.
A. Influence of P. aeruginosa infection on the constriction of porcine bronchial rings induced by 1μM carbachol 6 h after infection. B. Concentration dependence of the dilatation of carbachol-contracted bronchial rings induced by salbutamol in control and 6 h after P. aeruginosa infection. Control and infected groups were compared using the Mann-Whitney U test. Data are means ± S.E.M. * indicates p<0.05. A sigmoidal dose-response curve was used to generate the IC50.
Fig 4.
Histology of pig lung tissue sections and trachea sections taken 6 h and 24 h after P. aeruginosa infection.
Sections of pig lung tissue were stained with haematoxylin & eosin. Degrees of inflammation are given in Table 2. A. Section of lung (right middle lobe; x 20) from a control PBS-instilled pig. B. P. aeruginosa-induced level 1 inflammatory response 6 h after infection showing blood cells in the dilated interlobular septa (*) and in lung tissue (left cranial lobe; x 20). C. Blood cells in the dilated interlobular septa (left cranial lobe) (x 100). D. Leukocytes, mainly neutrophils, in the alveolar spaces (left cranial lobe) (x 400). E. Inflammatory response (level 3) appearing as early as 6 h after infection with small areas of healthy tissue and occlusion of alveolar spaces by neutrophils and red blood cells (right cranial lobe; x 20). F. Foci of inflamed areas around a bronchus and blood vessels 24 h p.i., indicating a level 4 inflammatory response (right middle lobe; x 20). G. Section of trachea sampled 6 h p.i. (x 200). H. Inflammatory response 24 h p.i., with a great increase of leukocytes and red blood cells leading to intense dilation of the interlobular septa, occlusion of most alveolar spaces, and hepatisation of the lung tissue (left cranial lobe; x 20). I. Neutrophils in the tracheal lymph nodes 24 h p.i. (x 200).
Table 2.
Histological evaluation of the P. aeruginosa-induced inflammatory response in the lungs of control and infected pigs.
Fig 5.
Bacterial loads in the BAL fluid, lung lobes, trachea, thoracic lymph nodes and spleen of pigs infected with P. aeruginosa.
Pigs were inoculated with 70 mL of an 8 x 106 cfu/mL suspension of P. aeruginosa PAK strain. Controls were obtained by inoculating 70 mL of 1X sterile PBS. Data were analysed by two-way ANOVA followed by Bonferroni’s post hoc test. Data are means ± S.E.M. * indicates p<0.05. ** indicates p<0.01. *** indicates p<0.001.
Fig 6.
Total white blood cell counts and cell profiles in the peripheral blood and BAL fluid of pigs at different times after P. aeruginosa infection.
A. Total white blood cells (WBC) in peripheral blood. B. Profile of the WBC populations in peripheral blood. C. Total WBC in BAL fluid. D. Profile of the WBC populations in BAL fluid. Total WBC and cell profiles of the control and infected groups were compared using the Mann-Whitney U test for each time point. Data are means ± S.E.M. * indicates p<0.05. ** indicates p<0.01. *** indicates p<0.001.
Fig 7.
Neutrophil serine proteases in BAL fluids of pigs infected with P. aeruginosa and purified blood neutrophils.
Confocal microscopy showing DNA (blue) and elastase (NE; green). Arrows show NET filaments.
Fig 8.
Peptidase activity of neutrophil serine proteases from the BAL fluid of pigs infected with P. aeruginosa.
A. Peptidase activities of NE, Pr3 and cat G (means ± SEM; n = 6). Increased protease activity were analysed using the non-parametric Mann-Whitney U test. *** indicates a significant increase in enzyme activity (p<0.001). B. Western blotting of neutrophil elastase showing free protease (25 kDa) and an irreversible complex with serpins (63 kDa).