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Capacity of recombinant gamma interferon to activate macrophages for Salmonella-killing activity
K Kagaya1, K Watanabe, Y Fukazawa
1Department of Microbiology, Yamanashi Medical College, Japan.
Abstract:
The ability of recombinant gamma interferon (rIFN-gamma) to activate macrophages for Salmonella-killing activity was kinetically examined in relation to phagosome-lysosome fusion and H2O2 generation. Resident peritoneal macrophages of BALB/c mice incubated with 10(2) to 10(3) U of rIFN-gamma per ml for 12 h exhibited enhanced bactericidal activity against Salmonella typhimurium, although H2O2 generation was unaltered. In contrast, macrophages incubated with equal doses of rIFN-gamma for 48 h showed both an enhanced Salmonella-killing activity and an increased generation of H2O2. To evaluate Salmonella-killing activities of macrophages, intracellular bacteria were assayed at 0, 2, and 8 h after infection. During the initial 2 h of infection, 12-h-activated macrophages, as well as the unstimulated control macrophages, showed a decline in bacterial population at the same rate. Over the next 6 h of infection, however, the number of viable bacteria in activated macrophages remained unchanged, whereas the number of bacteria in control macrophages significantly (P less than 0.05) increased. Similar results were obtained in 48-h-activated macrophages. On the other hand, macrophages incubated with 10 to 10(3) U of rIFN-gamma exhibited enhanced fusion of lysosomes to Salmonella-containing phagosomes in both the 12-h- and 48-h-stimulated stages. Moreover, when 48-h-activated macrophages were incubated concomitantly with superoxide dismutase and catalase, Salmonella-killing activity was not affected. These results indicate that rIFN-gamma per se is able to activate peritoneal macrophages to induce Salmonella-killing activity and suggest that increased phagosome-lysosome fusion followed by an oxygen-independent killing mechanism is primarily responsible for the enhanced Salmonella-killing activity in rIFN-gamma-activated macrophages.
Insights
Recombinant gamma interferon (rIFN-gamma) activates macrophages to kill Salmonella. This involves enhanced phagosome-lysosome fusion and an oxygen-independent mechanism, particularly after 48 hours of activation.
Area of Science:
- Immunology
- Cell Biology
- Microbiology
Background:
- Macrophages are key immune cells involved in pathogen clearance.
- Salmonella typhimurium is a bacterial pathogen that infects macrophages.
- Interferon-gamma (IFN-gamma) is a cytokine known to modulate immune responses.
Purpose of the Study:
- To investigate the kinetic effects of recombinant gamma interferon (rIFN-gamma) on macrophage activation for Salmonella killing.
- To examine the relationship between rIFN-gamma-induced Salmonella-killing activity, phagosome-lysosome fusion, and hydrogen peroxide (H2O2) generation.
- To elucidate the mechanisms underlying enhanced macrophage bactericidal activity.
Main Methods:
- Peritoneal macrophages from BALB/c mice were incubated with varying doses of rIFN-gamma for 12 and 48 hours.
- Macrophage bactericidal activity against Salmonella typhimurium was assessed by quantifying intracellular viable bacteria over time.
- Phagosome-lysosome fusion was evaluated using microscopy.
- Hydrogen peroxide (H2O2) generation was measured.
- The role of oxygen-dependent mechanisms was tested using superoxide dismutase and catalase.
Main Results:
- rIFN-gamma (10^2-10^3 U/ml) enhanced Salmonella-killing activity in macrophages after 12 and 48 hours of incubation.
- While H2O2 generation was not altered after 12 hours, it increased significantly after 48 hours of rIFN-gamma treatment.
- Enhanced phagosome-lysosome fusion was observed in macrophages stimulated with rIFN-gamma for both 12 and 48 hours.
- Inhibition of H2O2 production did not affect the Salmonella-killing activity in 48-hour activated macrophages.
Conclusions:
- rIFN-gamma effectively activates peritoneal macrophages to exhibit enhanced Salmonella-killing capacity.
- Increased phagosome-lysosome fusion is a key event in rIFN-gamma-mediated macrophage activation.
- The enhanced Salmonella-killing activity appears to be primarily mediated by an oxygen-independent mechanism.