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L-arginine-dependent killing of intracellular Ehrlichia risticii by macrophages treated with gamma interferon
1Department of Veterinary Pathobiology, College of Veterinary Medicine, Ohio State University, Columbus 43210-1092.
Abstract:
Thioglycolate-induced murine peritoneal macrophages infected with Ehrlichia risticii and treated in vitro with gamma interferon (IFN-gamma) developed antiehrlichial activity that eliminated the intracellular bacteria. This antiehrlichial activity was suppressed by NG-monomethyl-L-arginine, a competitive inhibitor of nitric oxide synthesis from L-arginine, but not by L-tryptophan. Increased levels of nitrite, an oxidative product of nitric oxide, were measured in cultures of infected macrophages treated with IFN-gamma. Sodium nitroprusside, which spontaneously releases nitric oxide, also showed the antiehrlichial activity. The antiehrlichial activity by reactive nitrogen intermediates was not mediated by elevation of the cellular concentration of cyclic GMP since the addition of 8-bromo-cyclic GMP itself had no influence on ehrlichial infection of macrophages. Addition of the intracellular iron chelator deferoxamine also inhibited E. risticii infection in vitro. These results suggest that intracellular E. risticii survival is iron dependent and that production of reactive nitrogen intermediates triggers iron loss from critical target enzymes of E. risticii, leading to lethal metabolic inhibition. However, addition of excess FeSO4, ferric citrate, or iron-saturated transferrin did not counteract the antiehrlichial effect induced by IFN-gamma.
Insights
Gamma interferon (IFN-gamma) activates macrophages to kill Ehrlichia risticii. This killing mechanism involves nitric oxide intermediates and iron depletion, suggesting a novel therapeutic strategy against ehrlichial infections.
Area of Science:
- Immunology
- Microbiology
- Infectious Diseases
Background:
- Ehrlichia risticii is an intracellular bacterium that infects macrophages.
- Gamma interferon (IFN-gamma) is a cytokine known to activate immune cells.
- The mechanisms by which macrophages combat intracellular pathogens are complex and involve various immune mediators.
Purpose of the Study:
- To investigate the antiehrlichial activity of gamma interferon (IFN-gamma) in murine peritoneal macrophages infected with Ehrlichia risticii.
- To elucidate the role of nitric oxide and iron metabolism in the IFN-gamma-mediated killing of intracellular E. risticii.
Main Methods:
- Murine peritoneal macrophages were infected with E. risticii and treated with IFN-gamma in vitro.
- Nitric oxide synthesis was inhibited using NG-monomethyl-L-arginine.
- Nitrite levels were measured as an indicator of nitric oxide production.
- Iron metabolism was assessed using the chelator deferoxamine and iron supplements.
Main Results:
- IFN-gamma treatment induced antiehrlichial activity, eliminating intracellular E. risticii.
- This activity was dependent on nitric oxide synthesis, as shown by suppression with NG-monomethyl-L-arginine and increased nitrite levels.
- Deferoxamine inhibited E. risticii infection, indicating iron dependency for bacterial survival.
- Reactive nitrogen intermediates likely trigger iron loss from essential E. risticii enzymes, leading to metabolic inhibition.
Conclusions:
- IFN-gamma-induced antiehrlichial activity against E. risticii is mediated by reactive nitrogen intermediates.
- Intracellular survival of E. risticii is iron-dependent, and nitric oxide contributes to iron deprivation.
- These findings suggest a mechanism involving iron sequestration as a key component of macrophage defense against E. risticii.