SimplySmart_v1, a new tool for the analysis of DNA damage optimized in primary neuronal cultures

• Published in: BMC bioinformatics Vol. 25; no. 1; pp. 318 - 18
• Main Authors: Koirala, Sushma, Sharma, Harman, Chew, Yee Lian, Konopka, Anna
• Format: Journal Article
• Language: English
• Published: England BioMed Central Ltd 01.10.2024; BioMed Central; BMC
• Subjects: Amyotrophic lateral sclerosis; Analysis; Animals; Bioinformatics; Carbon dioxide; Cell cycle; Cells, Cultured; Cloning; Comparative analysis; Computational biology; Damage detection; Dementia; Deoxyribonucleic acid; DNA; DNA Breaks, Double-Stranded; DNA Damage; DNA damage foci; DNA damage in neurons; Etoposide; Methods; Mice; Neurodegeneration; Neuronal culture; Neurons; Neurons - cytology; Neurons - metabolism; Open source software; Pathophysiology; Physiological aspects; Physiology; Python application; RNA-binding protein; Software; Source programs; Australia; Python application; Neuronal culture; DNA damage foci; Neurodegeneration; Analysis; DNA damage in neurons
• DOI: 10.1186/s12859-024-05947-8
• ISSN: 1471-2105

The increased interest in research on DNA damage in neurodegeneration has created a need for the development of tools dedicated to the analysis of DNA damage in neurons. Double-stranded breaks (DSBs) are among the most detrimental types of DNA damage and have become a subject of intensive research. DSBs result in DNA damage foci, which are detectable with the marker γH2AX. Manual counting of DNA damage foci is challenging and biased, and there is a lack of open-source programs optimized specifically in neurons. Thus, we developed a new, fully automated application, SimplySmart_v1, for DNA damage quantification and optimized its performance specifically in primary neurons cultured in vitro. Compared with control neurons, SimplySmart_v1 accurately identifies the induction of DNA damage with etoposide in primary neurons. It also accurately quantifies DNA damage in the desired fraction of cells and processes a batch of images within a few seconds. SimplySmart_v1 was also capable of quantifying DNA damage effectively regardless of the cell type (neuron or NSC-34). The comparative analysis of SimplySmart_v1 with other open-source tools, such as Fiji, CellProfiler and a focinator, revealed that SimplySmart_v1 is the most 'user-friendly' and the quickest tool among others and provides highly accurate results free of variability between measurements. In the context of neurodegenerative research, SimplySmart_v1 revealed an increase in DNA damage in primary neurons expressing abnormal TAR DNA/RNA binding protein (TDP-43). These findings showed that SimplySmart_v1 is a new and effective tool for research on DNA damage and can successfully replace other available software.