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Role of macrophage oxidative burst in the action of anthrax lethal toxin
P C Hanna1, B A Kruskal, R A Ezekowitz
1Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, Massachusetts 02115, USA.
Background:
Major symptoms and death from systemic Bacillus anthracis infections are mediated by the action of the pathogen's lethal toxin on host macrophages. High levels of the toxin are cytolytic to macrophages, whereas low levels stimulate these cells to produce cytokines (interleukin-1 beta and tumor necrosis factor-alpha), which induce systemic shock and death.
Materials And Methods:
Experiments were performed to assess the possibility that the oxidative burst may be involved in one or both of lethal toxin's effects on macrophages. Toximediated cell lysis, superoxide anion and cytokine production were measured. Effects of antioxidants and macrophage mutations were examined.
Results:
RAW264.7 murine macrophages treated with high levels of toxin released large amounts of superoxide anion, beginning at about 1 hr, which correlates with the onset of cytolysis. Cytolysis could be blocked with various exogenous antioxidants or with N-acetyl-L-cysteine and methionine, which promote production of the endogenous antioxidant, glutathione. Mutant murine macrophage lines deficient in production of reactive oxygen intermediates (ROIs) were relatively insensitive to the lytic effects of the toxin, whereas a line with increased oxidative burst potential showed elevated sensitivity. Also, cultured blood monocyte-derived macrophages from a patient with Chronic Granulomatous Disease, a disorder in which the phagocyte's oxidative burst is disabled, were totally resistant to toxin, in contrast to control monocytes.
Conclusions:
These results imply that the cytolytic effect of the toxin is mediated by ROIs. Additionally, cytokine production and consequent pathologies showed partial dependence on macrophage ROIs. Antioxidants moderately inhibited toxin-induced cytokine production in vitro, and BALB/c mice pretreated with N-acetyl-L-cysteine or mepacrine showed partial protection against lethal toxin. Thus ROIs are involved in both the cytolytic action of anthrax lethal toxin and the overall pathologic process in vivo.
Insights
Anthrax lethal toxin causes macrophage death via reactive oxygen intermediates (ROIs). Antioxidants and blocked oxidative bursts protect against toxin effects, indicating ROIs
Area of Science:
- Immunology
- Pathogen-host interactions
- Cellular toxicology
Background:
- Bacillus anthracis lethal toxin induces major symptoms and death by targeting host macrophages.
- High toxin levels cause macrophage lysis; low levels stimulate cytokine production, leading to shock.
Purpose of the Study:
- To investigate the role of the oxidative burst in Bacillus anthracis lethal toxin's effects on macrophages.
- To determine if reactive oxygen intermediates (ROIs) mediate toxin-induced cytolysis and cytokine production.
Main Methods:
- Assessed toxin-mediated cell lysis, superoxide anion, and cytokine production in macrophages.
- Examined the effects of antioxidants and macrophage mutations on toxin response.
- Utilized RAW264.7 murine macrophages and patient-derived monocytes with impaired oxidative bursts.
Main Results:
- High toxin levels induced superoxide anion release and cytolysis in macrophages, which was blocked by antioxidants.
- Macrophage lines with deficient or enhanced oxidative burst potential showed reduced or increased sensitivity to toxin, respectively.
- Monocytes from Chronic Granulomatous Disease patients were resistant to toxin-induced lysis.
Conclusions:
- The cytolytic effect of Bacillus anthracis lethal toxin on macrophages is mediated by reactive oxygen intermediates (ROIs).
- Cytokine production and associated pathologies demonstrate partial dependence on macrophage ROIs.
- ROIs are implicated in both the direct cytolytic action of anthrax lethal toxin and the broader in vivo pathological process.