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Comparison of 6-hydroxylmelatonin or melatonin in protecting neurons against ischemia/reperfusion-mediated injury

Authors


Address reprint requests to Ximing Wang, Department of Biochemistry and Molecular biology, Tongji Medical College, Huazhong University of Science and Technology, Hangkong Road No. 13, Wuhan 430030, China.
E-mail: biocwx@mails.tjmu.edu.cn

Abstract

Abstract:  The protective effect of exogenous melatonin or 6-hydroxylmelatonin on neurons was examined in N2a cells following exposure to oxygen–glucose–serum deprivation insults. After N2a cells cultured in vitro were deprived of glucose, serum and oxygen for 90 min, the different concentrations of melatonin or 6-hydroxylmelatonin were added to the medium. Then, treated cells were cultured for different intervals. At the end of the treatment, the collected culture solution was used for the analysis of the activity of lactate dehydrogenase (LDH) and the cells were used for the examination of the following parameters: cell viability (MTT), DNA fragmentation, reactive oxygen species (ROS) production, mitochondrial transmembrane potential, cytochrome C and caspase 3 activity. The results show that melatonin and 6-hydroxylmelatonin both reduced oxygen–glucose–serum deprivation-mediated N2a cell apoptosis, but they could not completely inhibit the apoptosis of the cells and the inhibitory effect of melatonin was stronger than that of 6-hydroxylmelatonin. Both of them could inhibit LDH and cytochrome C release and caspase 3 activity. Although 6-hydroxylmelatonin could no longer maintain mitochondrial transmembrane potential 6 h after reperfusion, its inhibitory effect on cytochrome C release from mitochondria and the scavenging role of ROS were stronger than those of melatonin. Moreover, melatonin promoted ROS production at the 15th min of the reperfusion, and then it began to remove ROS from cells. Our study showed that melatonin and 6-hydroxylmelatonin can be used as supplements in the treatment of neurological disorders involving oxidative stress. Melatonin serves as more than a ROS scavenger and its other roles await further study.

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