Defense and inflammation by alveolar macrophages in response to inhaled fungi

Abstract

Background: Serious fungal infections have increased notably in patients receiving modern treatment like organ transplantation, parenteral nutrition, long term steroid therapy and broad spectrum antibiotics. Patients with HIV infections and drug addicts also contract fungal infections. Especially, infections with species of Candida, Cryptococcus and Aspergillus are increasing. The normal portal of entry for Aspergillus species and C. neoformans is the respiratory tract. Aspergillus conidia are found in the atmosphere throughout the world. These conidia might give toxic and allergic reactions and may contribute to the development of asthma, allergic bronchopulmonary aspergillosis and hypersensitivity pneumonitis. As desiccated fungal yeasts and conidia have a diameter well below 10 um, they can easily be aerosolized and when inhaled, most of them are deposited in the alveolar region. In this region alveolar macrophages (AM) constitute a first defense. After phagocytosis, the fungi will be localized in phagolysosomes. The phagolysosomal pH in AM is around 5 and low pH is important for the killing of microorganisms. An increase of oxidative metabolism by AM is also important for the killing, but may lead to adverse toxic effects.

Purpose: To study the interaction between AM and various fungal yeasts and conidia. Reactions of rat AM to highly pathogenic and low pathogenic yeasts as Candida species, C. neoformans and S. cerevisiae, as well as pathogenic and non-pathogenic Aspergillus conidia were studied in vitro. The reactions of rat and human AM on exposure to A. fumigatus were compared. For C. neoformans the reactions of rabbit AM in vivo were also studied.

Material and methods: Fungal yeasts and conidia were cultured and labelled with fluorescein. Intratracheal instillation of C. neoformans into rabbit lung was performed. AM were obtained by lung lavage from rat, rabbit and humans. Assays of phagocytosis, oxidative metabolism and phagolysosomal pH were used. Release of cytokines (IL-6, IL-8 and TNF-a) from human AM was measured. Electron microscopy of AM was performed.

Results: Candida and Aspergillus species were phagocytized faster than control particles of inert silica mainly due to a faster ingestion process and they induced a 34 fold increase in oxidative metabolism during phagocytosis as well as 24 hrs after the onset of phagocytosis. Phagocytosis of C. neoformans in vitro by AM was negligible during the first hour in comparison with the silica particles, but was remarkable after 24 h at which time they induced a significantly increased oxidative metabolism. When rabbits were exposed in vivo to C. neoformans for 24 h, the phagocytosis of C. neoformans was similar to that of the silica particles and C. neoformans induced a 2 fold increase in oxidative metabolism. After 3 h, in the in vitro experiments around 10% of phagolysosomes had neutral pH and after 24 h, in the in vitro and in vivo experiments, 1-3 % of phagolysosomes had neutral pH for all the yeasts and conidia. Phagolysosomes with neutral pH was not observed for the control particles. In spite of the difference in pathogenicity among yeasts and conidia, there were no significant differences in phagocytosis, oxidative burst and phagolysosomal pH. In spite of the markedly increased oxidative metabolism there was no increase in cytokine production by human AM after exposure to A. fumigatus conidia. Human AM exposed to silica particles had significantly decreased IL-6 and IL-8 production both after 8 and 24 h and had a tendency to decreased TNF-a production. Electron microscopic examination of AM with the fungal yeasts and conidia showed narrow passages between phagolysosomes and the cell surface.

Conclusion: The increased phagocytic activity of AM, the markedly increased oxidative metabolism and the low phagolysosomal pH after 24 h indicate an adequate defense by AM against the fungal yeasts and conidia, including C. neoformans. But the increased oxidative metabolism even after 24 h in combination with unsealed phagolysosomes is of interest. The narrow passages might facilitate the extracellular escape of reactive oxidative metabolites which can cause damage to surrounding tissues and in the long run might cause inflammation and emphysema.