Hanna P C, Kruskal B A, Ezekowitz R A, Bloom B R, Collier R J
Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, Massachusetts 02115, USA.
Mol Med. 1994 Nov;1(1):7-18.
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.
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.
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.
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.
全身性炭疽芽孢杆菌感染的主要症状和死亡是由病原体的致死毒素作用于宿主巨噬细胞介导的。高浓度的毒素对巨噬细胞具有细胞溶解作用，而低浓度则刺激这些细胞产生细胞因子（白细胞介素-1β和肿瘤坏死因子-α），从而诱发全身性休克和死亡。
进行实验以评估氧化爆发可能参与致死毒素对巨噬细胞的一种或两种作用的可能性。测量了毒素介导的细胞溶解、超氧阴离子和细胞因子的产生。研究了抗氧化剂和巨噬细胞突变的影响。
用高浓度毒素处理的RAW264.7小鼠巨噬细胞从约1小时开始释放大量超氧阴离子，这与细胞溶解的开始相关。细胞溶解可被各种外源性抗氧化剂或N-乙酰-L-半胱氨酸和蛋氨酸阻断，这两种物质可促进内源性抗氧化剂谷胱甘肽的产生。缺乏活性氧中间体（ROIs）产生的突变小鼠巨噬细胞系对毒素的溶解作用相对不敏感，而具有增强氧化爆发潜力的细胞系则表现出更高的敏感性。此外，与对照单核细胞相比，慢性肉芽肿病患者（一种吞噬细胞氧化爆发功能丧失的疾病）的培养血单核细胞衍生巨噬细胞对毒素完全耐药。
这些结果表明毒素的细胞溶解作用是由ROIs介导的。此外，细胞因子的产生及随之而来的病理变化部分依赖于巨噬细胞ROIs。抗氧化剂在体外适度抑制毒素诱导的细胞因子产生，用N-乙酰-L-半胱氨酸或米帕林预处理的BALB/c小鼠对致死毒素表现出部分保护作用。因此，ROIs参与了炭疽致死毒素的细胞溶解作用和体内的整体病理过程。
