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Secretion of TNF-alpha by alveolar macrophages in response to Candida albicans mannan
R E Garner1, K Rubanowice, R T Sawyer
1Division of Biomedical Science, Mercer University School of Medicine, Macon, Georgia 31207.
Abstract:
Resident alveolar macrophages (AM phi) were tested for their ability to respond to Candida albicans mannan. AM phi were found to produce tumor necrosis factor alpha (TNF-alpha) in vitro in response to mannan stimulation. TNF-alpha secretion was measured using ELISA and L929B cellular cytotoxicity assays. Cytotoxicity was neutralized in parallel L929B cell cultures by the addition of rabbit anti-TNF-alpha antibody. Mannan preparations were found to be free of contaminating LPS by Limulus assay. When AM phi were cultivated for 18 h at 37 degrees C, 67 micrograms of mannan stimulated the secretion of approximately 207 U/ml of TNF-alpha. By comparison, AM phi treated with 6.7 micrograms of LPS secreted approximately 257 U/ml of TNF-alpha. Optimal TNF-alpha production occurred between 9 and 18 h after mannan stimulation. Disparate mechanisms for stimulation of TNF-alpha secretion were suggested by differential sugar blockade of LPS- and mannan-induced TNF-alpha secretion. The addition of 2% D-mannose or 2% alpha-methyl-D-mannoside to AM phi cultures blocked mannan- but not LPS-stimulated TNF-alpha secretion. Furthermore, the addition of rabbit anti-mannan antibody to mannan-coated plastic culture dishes prevented TNF-alpha secretion by the mannan-sensitive RAW 264.7 cell line. Moreover, the data suggest that C. albicans mannan stimulated AM phi to secrete TNF-alpha by an LPS-independent receptor mechanism which may also function as a mannose receptor.
Insights
Resident alveolar macrophages (AM phi) produce tumor necrosis factor alpha (TNF-alpha) in response to Candida albicans mannan. This immune response is mediated by an LPS-independent mechanism, potentially involving a mannose receptor.
Area of Science:
- Immunology
- Microbiology
- Cell Biology
Background:
- Alveolar macrophages (AM phi) are key immune cells in the lung.
- Candida albicans mannan is a component of the fungal cell wall.
- Tumor necrosis factor alpha (TNF-alpha) is a critical pro-inflammatory cytokine.
Purpose of the Study:
- To investigate the capacity of resident alveolar macrophages to respond to Candida albicans mannan.
- To characterize the mechanism by which mannan stimulates TNF-alpha secretion by AM phi.
Main Methods:
- AM phi were stimulated with purified Candida albicans mannan.
- TNF-alpha production was quantified using ELISA and L929B cytotoxicity assays.
- LPS contamination was excluded using Limulus assay; specific sugar blockades and antibody neutralization were employed.
Main Results:
- AM phi secreted TNF-alpha in response to mannan stimulation, comparable to LPS stimulation.
- Optimal TNF-alpha secretion occurred between 9 and 18 hours post-stimulation.
- Mannan-induced TNF-alpha secretion was blocked by D-mannose and alpha-methyl-D-mannoside, unlike LPS-induced secretion.
Conclusions:
- Candida albicans mannan stimulates alveolar macrophages to secrete TNF-alpha.
- The data suggest an LPS-independent receptor mechanism for mannan recognition, possibly involving a mannose receptor.
- This finding elucidates a specific pathway for fungal-derived immune activation in the lung.