Borisovs Vitālijs, Ļeonova Elīna, Baumane Larisa, Kalniņa Jolanta, Mjagkova Natalja, Sjakste Nikolajs
Genomics and Bioinformatics, Institute of Biology of the University of Latvia, Miera Str. 3, LV2169, Salaspils, Latvia.
Latvian Institute of Organic Synthesis, Aizkraukles Str. 3, Riga LV1006, Latvia.
Mutat Res Genet Toxicol Environ Mutagen. 2019 Jul;843:90-94. doi: 10.1016/j.mrgentox.2018.11.008. Epub 2018 Nov 23.
Oxidative stress, especially overproduction of nitric oxide (NO), is considered to be one of the crucial factors in the pathogenesis of multifactorial multiple sclerosis (MS). DNA breaks could be one of the consequences of oxidative stress; however, data on DNA breakage in MS are very few and contradictory. There are no data on direct measurements of NO production in the blood of MS patients. The goal of this study was to determine the level of single-stranded DNA breaks in whole blood or isolated peripheral blood mononuclear cells (PBMNCs) by means of alkaline single cell gel electrophoresis (comet assay) and to evaluate production of NO in the human blood by applying electron paramagnetic resonance (EPR) spectroscopy. Groups of healthy subjects and MS patients were enrolled in the study. Blood samples were obtained by vein puncture and divided in aliquots for the analysis of the whole blood and isolated PBMNC with comet assay. Alkaline single cell gel electrophoresis was performed on whole blood and isolated PBMNC samples of 28 patients and 15 controls. A separate blood sample was mixed with a spin-trap, frozen in liquid nitrogen and used for NO detection by EPR; 22 MS patients and 22 controls were tested. A statistically significant increase in the level of DNA breakage was observed in specimens taken from MS patients compared to healthy persons. The level of DNA damage in whole blood and PBMNCs of the same group was similar. NO production was significantly higher in the blood of MS patients.
氧化应激，尤其是一氧化氮（NO）的过量产生，被认为是多因素多发性硬化症（MS）发病机制中的关键因素之一。DNA断裂可能是氧化应激的后果之一；然而，关于MS中DNA断裂的数据非常少且相互矛盾。目前尚无关于直接测量MS患者血液中NO产生量的数据。本研究的目的是通过碱性单细胞凝胶电泳（彗星试验）确定全血或分离的外周血单核细胞（PBMNC）中单链DNA断裂的水平，并应用电子顺磁共振（EPR）光谱评估人体血液中NO的产生。健康受试者组和MS患者组参与了该研究。通过静脉穿刺采集血样，并分成小份用于全血分析和用彗星试验分离PBMNC。对28例患者和15例对照的全血和分离的PBMNC样本进行了碱性单细胞凝胶电泳。将一份单独的血样与自旋捕获剂混合，在液氮中冷冻，用于通过EPR检测NO；对22例MS患者和22例对照进行了检测。与健康人相比，在取自MS患者的样本中观察到DNA断裂水平有统计学意义的增加。同一组全血和PBMNC中的DNA损伤水平相似。MS患者血液中的NO产生量明显更高。
