Evaluation of DNA Damage in Brain Tissue Using the Comet Assay in a Cuprizone-Induced Mouse Model of Demyelination

Abstract

Oxidative stress and DNA damage play a critical role in the pathogenesis of multiple sclerosis (MS). This study aimed to evaluate DNA damage in brain cells using the alkaline Comet assay in a cuprizone-induced mouse model of demyelination.
Brain tissue from the left hemisphere was collected from control and cuprizone-treated mice. Single-cell gel electrophoresis was performed, and DNA fragmentation was quantified using fluorescence microscopy and CometScore 2.0 software. Key parameters analyzed included Tail DNA percentage, Tail Length (TL), and Olive Tail Moment (OTM). Cells exhibiting catastrophic fragmentation were excluded from statistical analysis.
The cuprizone model demonstrated a profound genotoxic effect compared to controls. The proportion of DNA in the tail significantly increased from 4.7% to 15.6%. Furthermore, Tail Length expanded from 7 µm to 22 µm, and the Olive Tail Moment rose from 2.2 to 12.3 arbitrary units. Additionally, approximately 8% of cells in the treated group exhibited catastrophic DNA fragmentation, indicative of severe cellular distress.
These findings confirm that cuprizone-induced demyelination is accompanied by severe oxidative stress and extensive DNA strand breaks in brain tissue. The Comet assay proved to be a highly sensitive tool for quantifying neurodegeneration, holding significant potential for screening neuroprotective compounds and identifying DNA damage biomarkers in complex neurodegenerative diseases like MS.

Speaker

Ulyana Apukhtina
Federal Research Centre of Biotechnology of the Russian Academy of Sciences, Moscow, Russia
Russia

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