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Effect of iron and phagocytosis on murine macrophage activation in vitro
1Unité de Microbiologie, Centre de Recherche du Service de Santé des Armées, La Tronche, France.
Abstract:
Iron-exposed murine macrophages have a modified bactericidal activity as shown by previous observations. In order to assess the role of iron in macrophage activation, as measured by free radical production and by intracellular bacterial killing, murine peritoneal macrophages were cultivated in the presence of various sources of iron, human iron-saturated transferrin and ammonium ferric citrate, or iron chelators, Desferal, and human Apo-transferrin, and were infected with an enteropathogenic strain of E. coli. The release of nitrite (NO2-), and the production of superoxide anion (O2-) and hydrogen peroxide (H2O2) by the phagocytes were measured and compared to the production by uninfected macrophages. The synergistic action with murine r.IFN-gamma was also studied in the radical production reaction and for the bactericidal activity of macrophages. Our results show that in vitro phagocytosis of E. coli induced elevated production of NO2- and H2O2 by macrophages, and that oxygen derivatives were released independently of the presence of added iron or chelator. Despite a phagocytosis-related enhancement of NO2- release, reactive nitrogen intermediates (RNI) are not directly involved in the bactericidal mechanism, as revealed by increased intracellular killing owing to RNI inhibitors. Moreover, bacterial killing may depend on oxygen derivatives, as suggested by the effect of the antioxidant sodium ascorbate leading to both a diminished H2O2 production and a decreased bactericidal activity of macrophages.
Insights
Iron influences macrophage function, impacting bacterial killing. Reactive oxygen and nitrogen species are involved, but iron
Area of Science:
- Immunology
- Cell Biology
- Microbiology
Background:
- Macrophages play a crucial role in host defense against bacterial infections.
- Iron availability is known to modulate macrophage functions, including bactericidal activity.
Purpose of the Study:
- To investigate the role of iron in macrophage activation and its impact on intracellular bacterial killing.
- To determine the involvement of reactive oxygen species (ROS) and reactive nitrogen intermediates (RNI) in iron-modulated macrophage bactericidal activity.
Main Methods:
- Murine peritoneal macrophages were cultured with various iron sources (transferrin, ammonium ferric citrate) or iron chelators (Desferal, Apo-transferrin).
- Macrophages were infected with an enteropathogenic E. coli strain.
- Production of nitrite (NO2-), superoxide anion (O2-), and hydrogen peroxide (H2O2) was measured.
- Synergistic effects with murine r.IFN-gamma were assessed.
- The impact of RNI inhibitors and the antioxidant sodium ascorbate on bacterial killing was evaluated.
Main Results:
- Phagocytosis of E. coli induced increased production of NO2- and H2O2 by macrophages, independent of added iron or chelators.
- Reactive nitrogen intermediates (RNI) were not directly involved in the bactericidal mechanism, as indicated by enhanced killing with RNI inhibitors.
- Bacterial killing appears to depend on reactive oxygen species (ROS), as sodium ascorbate diminished H2O2 production and reduced macrophage bactericidal activity.
Conclusions:
- Iron availability does not directly influence the production of reactive oxygen and nitrogen species in macrophages during E. coli infection.
- While RNI are not the primary bactericidal agents, ROS play a significant role in the intracellular killing of E. coli by macrophages.