A14-09 Intercellular Adhesion Molecule-1 (ICAM-1) Regulates Bacterial Clearance and Lung Injury in the Naïve and Pneumonic Lung

Rationale ICAM-1 is a type 1 transmembrane glycoprotein. Its primary ligands are the β2 leukocyte integrins. Interactions between ICAM-1 and β2 integrins contribute to leukocyte adhesion, migration and motility. Our studies showed that Icam1-/- mice generated by homologous recombination have impaired clearance of S. pneumoniae, the leading cause of community-acquired pneumonia. The impaired clearance was not due to defective leukocyte recruitment (PMC11905799). Curiously, they also appeared to have no NK cells, as assessed by NK1.1 expression. Further investigation revealed that this absence was due to the presence of a divergent Nkrp1c allele, which was retained from the 129P3/J background used to generate this strain. Because of uncertainty regarding the rest of their genome, we generated mice through CRISPR/Cas9 mediated deletion of Icam1 and studied host defense against S. pneumoniae. Methods Pneumonias were induced by intratracheal instillation of S. pneumoniae suspension into the left lung. Bacterial clearance was determined by plating serial dilutions of lung homogenates. Immunophenotyping was performed by flow cytometry. Data represent mean±SEM. Results In naïve mice, there was no difference in the numbers of lavageable alveolar macrophages and neutrophils, alveolar, interstitial or monocyte-derived macrophages in lung homogenates between genotypes. Twenty-four hours post-inoculation, the number of viable bacteria in the lung of Icam1-/- mice (1.62±0.46x106 CFU) was significantly greater than in WT mice (0.04±0.02x106 CFU). In lung homogenates, the number of alveolar macrophages decreased by 83-85% and neutrophils increased 9–13-fold. By day 7, alveolar macrophage numbers began to recover in WT mice but remained low in Icam1-/- animals, which also exhibited persistently elevated neutrophil and monocyte-derived macrophage numbers. NK1.1+ NK cells were identified and were significantly higher in Icam1-/- compared to WT mice. Mechanistic studies in vivo showed that a similar percentage of unchallenged Icam1-/- and WT alveolar macrophages phagocytosed and acidified S. aureus pHRhodo bioparticles, but Icam1-/- macrophages internalized a greater number of particles. Mutant macrophages were more granular, as indicated by increased side-scatter properties, and contained a greater number of lysosomes, as determined by fluorescent markers and by tEM. Conclusions CRISPR/Cas9-generated Icam1-/- mice retain defective bacterial clearance and the delayed resolution observed in the homologous recombinant mice, but have identifiable NK cells. The defect in bacterial clearance is not due to defects in either myeloid recruitment or alveolar macrophage phagocytosis. However, unchallenged alveolar macrophages show evidence of enhanced phagocytic activity and lysosome formation, suggesting that an altered microenvironment may contribute to defective bacterial clearance. This abstract is funded by: R01HL171598