Simultaneous determination of 8-oxo-2'-deoxyguanosine and 8-oxo-2'-deoxyadenosine in human retinal DNA by liquid chromatography nanoelectrospray-tandem mass spectrometry.
Simultaneous determination of 8-oxo-2'-deoxyguanosine and 8-oxo-2'-deoxyadenosine in human retinal DNA by liquid chromatography nanoelectrospray-tandem mass spectrometry.
复制标题
DOI：
10.1038/srep22375
复制
发表时间：
2016-03-16
影响因子：
4.6
通讯作者：
Stepanov I
中科院分区：
文献类型：
--
作者：
Ma B;Jing M;Villalta PW;Kapphahn RJ;Montezuma SR;Ferrington DA;Stepanov I
视网膜相关性黄斑变性（AMD）是发达国家老年人失明的主要原因。视网膜色素上皮（RPE）中线粒体DNA（mtDNA）的氧化损伤可能在AMD中起关键作用。检测RPE中8-氧代-2 '-脱氧鸟苷（8-oxo-dG）和8-氧代-2'-脱氧腺苷（8-oxo-dA）等氧化性DNA损伤可为AMD的发病机制提供重要的信息。我们建立了一种液相色谱-纳米电喷雾电离-串联质谱法同时分析人视网膜DNA中8-oxo-dG和8-oxo-dA的方法。将所建立的方法应用于5名AMD供体和5名对照供体的视网膜DNA分析。在线粒体DNA中，对照组和AMD供体的8-oxo-dG水平平均为170和188，8-oxo-dA平均为11和17个加合物/106个碱基。在核DNA中，对照组和AMD供体中的8-oxo-dG水平平均为0.54和0.96，8-oxo-dA平均为0.04和0.05加合物/106个碱基。这种高灵敏度的方法允许在非常少量的DNA中测量两种加合物，并且可以用于未来的研究中，研究8-oxo-dG和8-oxo-dA在AMD和人类其他氧化损伤相关疾病中的病理生理作用。
Age-related macular degeneration (AMD) is the leading cause of blindness among older adults in the developed world. Oxidative damage to mitochondrial DNA (mtDNA) in the retinal pigment epithelium (RPE) may play a key role in AMD. Measurement of oxidative DNA lesions such as 8-oxo-2’-deoxyguanosine (8-oxo-dG) and 8-oxo-2’-deoxyadenosine (8-oxo-dA) in diseased RPE could provide important insights into the mechanism of AMD development. We have developed a liquid chromatography-nanoelectrospray ionization-tandem mass spectrometry method for simultaneous analysis of 8-oxo-dG and 8-oxo-dA in human retinal DNA. The developed method was applied to the analysis of retinal DNA from 5 donors with AMD and 5 control donors without AMD. In mtDNA, the levels of 8-oxo-dG in controls and AMD donors averaged 170 and 188, and 8-oxo-dA averaged 11 and 17 adducts per 106 bases, respectively. In nuclear DNA, the levels of 8-oxo-dG in controls and AMD donors averaged 0.54 and 0.96, and 8-oxo-dA averaged 0.04 and 0.05 adducts per 106 bases, respectively. This highly sensitive method allows for the measurement of both adducts in very small amounts of DNA and can be used in future studies investigating the pathophysiological role of 8-oxo-dG and 8-oxo-dA in AMD and other oxidative damage-related diseases in humans.