Involvement of superoxide and myeloperoxidase in oxygen-dependent killing of Staphylococcus aureus by neutrophils
Involvement of superoxide and myeloperoxidase in oxygen-dependent killing of Staphylococcus aureus by neutrophils
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DOI：
10.1128/iai.64.9.3512-3517.1996
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发表时间：
1996-09-01
影响因子：
3.1
通讯作者：
Winterbourn, CC
中科院分区：
文献类型：
--
作者：
Hampton, MB;Kettle, AJ;Winterbourn, CC
为了抑制超氧化物依赖的过程，我们将超氧化物歧化酶与免疫球蛋白G交联，并将其偶联物通过细胞壁中的蛋白a附着在金黄色葡萄球菌的表面。叠氮化物抑制髓过氧化物酶，并使用髓过氧化物酶缺陷中性粒细胞。加入NADPH氧化酶抑制剂二苯二酮，以防止超氧化物的产生，将杀伤率降低到25%，表明氧化杀伤机制在该系统中占主导地位。附超氧化物歧化酶的金黄色葡萄球菌的杀灭率常数为附超氧化物歧化酶的对照菌的70%。超氧化物歧化酶对二苯基偶氮肼没有作用，对叠氮化物的杀伤率降至33%，对髓过氧化物酶缺乏的中性粒细胞的杀伤率降至40%，对叠氮化物没有作用。假设杀伤的氧化和非氧化成分可以分开考虑，超氧化物歧化酶使氧化速率降低了近一半，当髓过氧化物酶失活时，氧化速率降低了约六倍。我们得出结论，髓过氧化物酶依赖过程是人类中性粒细胞强烈支持的，是它们氧化杀死金黄色葡萄球菌的主要机制，超氧化物对杀伤有直接的贡献。我们的研究结果还表明，超氧化物与髓过氧化物酶依赖途径一起起作用。
We have used a quantitative assay that measures independent rate constants for phagocytosis and killing of Staphylococcus aureus to investigate the involvement of superoxide and myeloperoxidase in bacterial killing by human neutrophils, To inhibit superoxide-dependent processes, superoxide dismutase was cross-linked to immunoglobulin G and the conjugate was attached to the surface of S. aureus via protein A in its cell wall. Myeloperoxidase was inhibited with azide, and myeloperoxidase-deficient neutrophils were used. Adding the NADPH oxidase inhibitor diphenyleneiodonium, to prevent superoxide production, decreased the killing rate to 25%, indicating that oxidative killing mechanisms predominate in this system. The rate constant for killing of S. aureus with superoxide dismutase attached was 70% of that for control bacteria linked to inactivated enzyme. Superoxide dismutase had no effect in the presence of diphenyleneiodonium, The rate of killing was decreased to 33% in the presence of azide and to 40% with myeloperoxidase-deficient neutrophils, Superoxide dismutase had no effect in the presence of azide. On the assumption that the oxidative and nonoxidative components of killing can be considered separately, the oxidative rate was decreased by almost half by superoxide dismutase and was about six times lower when myeloperoxidase was inactive, We conclude that myeloperoxidase-dependent processes are strongly favored by human neutrophils as their prime mechanism of oxidative killing of S. aureus and that superoxide makes a direct contribution to killing. Our results also suggest that superoxide acts in conjunction with a myeloperoxidase-dependent pathway.