ENDOTHELIAL-CELL INJURY DUE TO COPPER-CATALYZED HYDROGEN-PEROXIDE GENERATION FROM HOMOCYSTEINE
ENDOTHELIAL-CELL INJURY DUE TO COPPER-CATALYZED HYDROGEN-PEROXIDE GENERATION FROM HOMOCYSTEINE
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DOI：
10.1172/jci112442
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发表时间：
1986-04-01
影响因子：
15.9
通讯作者：
HARLAN, JM
中科院分区：
文献类型：
--
作者：
STARKEBAUM, G;HARLAN, JM
我们已经研究了同型半胱氨酸对培养的内皮细胞的毒性作用是否可能是由于过氧化氢的形成和作用。在无细胞系统的初始实验中，发现微摩尔量的铜催化同型半胱氨酸的氧依赖性氧化。氧化的高半胱氨酸与消耗的氧的摩尔比为约。4.0，这表明氧气被还原为水。然而，添加过氧化氢酶，减少了近一半的耗氧量，这表明在反应过程中形成了H2O2。重申这一假设，H2O2的形成检测使用辣根过氧化物酶依赖的氧化的荧光东莨菪内酯。铜蓝蛋白还被发现催化同型半胱氨酸的氧化和产生H2O2的摩尔量相当于硫酸铜。最后，同型半胱氨酸的氧化被催化的正常人血清中的浓度依赖性的方式。使用培养的人和牛内皮细胞，我们发现，同型半胱氨酸加铜可以溶解细胞的剂量依赖性的方式，这种作用是完全防止过氧化氢酶。同型半胱氨酸加铜对细胞没有毒性。内皮细胞的特异性损伤后，只看到4小时的孵育同型半胱氨酸加铜。在生化研究中，还发现铜蓝蛋白在高半胱氨酸存在下引起内皮细胞损伤的能力与Cu++相当。由于同型半胱氨酸水平升高与动脉粥样硬化的过早发展有关，这些发现可能与某些类型的慢性血管损伤的机制有关。
We have examined whether the toxic effects of homocysteine on cultured endothelial cells could result from the formation and action of hydrogen peroxide. In initial experiments with a cell-free system, micromolar amounts of copper were found to catalyze an oxyge-dependent oxidation of homocysteine. The molar ratio of homocysteine oxidized to oxygen consumed was .apprx. 4.0, which suggests that oxygen was reduced to water. The addition of catalase, however, decreased oxygen consumption by nearly one-half, which suggests that H2O2 was formed during the reaction. Confirming this hypothesis, H2O2 formation was detected using the horseradish peroxidase-dependent oxidation of fluorescent scopoletin. Ceruloplasmin was also found to catalyze oxidation of homocysteine and generation of H2O2 in molar amounts equivalent to copper sulfate. Finally, homocysteine oxidation was catalyzed by normal human serum in a concentration-dependent manner. Using cultured human and bovine endothelial cells, we found that homocysteine plus copper could lyse the cells in a dose-dependent manner, an effect that was completely prevented by catalase. Homocysteine plus copper was not toxic to the cells. Specific injury to endothelial cells was seen only after 4 h of incubation with homocysteine plus copper. Confirming the biochemical studies, ceruloplasmin was also found to be equivalent to Cu++ in its ability to cause injury to endothelial cells in the presence of homocysteine. Since elevated levels of homocysteine have been implicated in premature development of atherosclerosis, these findings may be relevant to the mechanism of some types of chronic vascular injury.