Role of Macrophage Oxidative Burst in the Action of Anthrax Lethal Toxin

Abstract

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(3 and tumor necrosis factor-a), 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.

Hypothetical Integrative Medical Strategies for the Prevention and Treatment of Bio-terrorism Incidents

“…These include interleukins and interferons. The main cytokines involved in this discussion are interleukin-1-beta and tumor necrosis factor-alpha [6,7]. This sophisticated defense system, however, teeters on a precarious balance because the same molecules (reactive oxygen and nitrogen intermediates and cytokines) produced by the macrophages in defense against invading organisms also have the potential to cause serious damage and destruction of the macrophages themselves.…”

“…This sophisticated defense system, however, teeters on a precarious balance because the same molecules (reactive oxygen and nitrogen intermediates and cytokines) produced by the macrophages in defense against invading organisms also have the potential to cause serious damage and destruction of the macrophages themselves. Furthermore, if excessive amounts of these molecules escape into the extracellular space and general circulation, they can also cause severe damage and in ammation in distant tissues and ultimately cause systemic shock and death [6,7]. Normally, this potential for self-injury/destruction is well compensated for by an elegant built-in self-protection system, whereby the macrophage (a) simultaneously produces its own free radical scavenger/antioxidant molecules and associated enzyme catalysts, such as glutathione, glutathione peroxidase, and superoxide dismutase; (b) tightly regulates the synthesis/inhibition signaling governing the production and release of these 'double-edged sword' molecules; (c) restricts them to subcellular locales where their effects can be tolerated [6,7].…”

“…Neutrophils are disarmed by inhibition of the oxidative burst pathway due to interference of signal transduction by the action of the anthrax toxins. The clinical and symptomatic manifestations of anthrax infection, on the other hand, are mediated by the effects of the toxins on the host macrophages [6,7].…”

“…There, the lethal factor is cleaved from the protective antigen by a protease and begins to exert its pathogenic effects (the Bacillus anthracis version of the Trojan Horse story). The third toxic protein of signi cance, the edema factor, produces swelling and in ammation and contributes to the pulmonary pathology of inhalation anthrax (pneumonitis) [6,7,60,61].…”

Anthrax toxins

“…Hanna et al [129] found that cultured macrophages treated with LeTx release superoxide anion at about the time that cell lysis begins, and that macrophage lysis could be prevented by the addition of anti-oxidants. When LeTx resistant cell lines were examined, they were found to be deficient in the production of reactive oxygen intermediates (ROIs).…”