Starkebaum G, Harlan J M
J Clin Invest. 1986 Apr;77(4):1370-6. doi: 10.1172/JCI112442.
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 oxygen-dependent oxidation of homocysteine. The molar ratio of homocysteine oxidized to oxygen consumed was approximately 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. Homocystine 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.
我们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由过氧化氢的形成和作用所致。在最初的无细胞系统实验中，发现微摩尔量的铜可催化同型半胱氨酸的氧依赖性氧化。被氧化的同型半胱氨酸与消耗的氧的摩尔比约为4.0，这表明氧被还原为水。然而，加入过氧化氢酶后，氧消耗减少了近一半，这表明反应过程中形成了过氧化氢。荧光 scopoletin 的辣根过氧化物酶依赖性氧化检测证实了这一假设，即检测到了过氧化氢的形成。还发现铜蓝蛋白可催化同型半胱氨酸的氧化并产生与硫酸铜等量的过氧化氢。最后，正常人血清以浓度依赖性方式催化同型半胱氨酸的氧化。使用培养的人及牛内皮细胞，我们发现同型半胱氨酸加铜可呈剂量依赖性地裂解细胞，而过氧化氢酶可完全阻止这种作用。同型胱氨酸加铜对细胞无毒。仅在同型半胱氨酸加铜孵育4小时后才观察到对内皮细胞的特异性损伤。与生化研究结果一致，还发现铜蓝蛋白在同型半胱氨酸存在下对内皮细胞造成损伤的能力与铜离子相当。由于同型半胱氨酸水平升高与动脉粥样硬化的过早发生有关，这些发现可能与某些类型的慢性血管损伤机制有关。
