Sperm double-strand DNA breaks in patients with normozoospermia and pathozoospermia.

The levels of double-strand DNA breaks as a severe disruption of genome integrity were studied using the neutral version of the comet assay in the sperm samples of the men of the Belarusian population with normozoospermia and pathozoospermia, including asthenozoospermia, oligozoospermia, oligoasthenozoospermia, and other combined forms of pathozoospermia. It was demonstrated that double-strand DNA breaks have a large proportion (about 44–50 % on average) of the total number of DNA damage (singleand double-strand breaks, alkaline-labile sites, etc.) analyzed with the alkaline version of the comet assay recommended by the World Health Organization. Higher levels of sperm double-strand DNA breaks were established in the pathozoospermia group and in asthenozoospermia subgroup compared to the normozoospermia group. It was also shown that the levels of double-strand breaks observed at pathozoospermia correspond to the mutagenic effect in vitro of high concentrations (10–30 μg/ml) of bleomycin sulfate that acts as a strong radiomimetic, which points to a significant disruption of the DNA integrity at pathozoospermia. In general, the data obtained demonstrate the usefulness of the sperm double-strand DNA break analysis for male infertility diagnostics.

1. O’Neill C. L., Palermo G. D. A worldwide profile of the utilization of sperm DNA fragmentation testing in relation to reproductive outcome. Translational Andrology and Urology, 2017, vol. 6, no. 4, pp. 320−321. https://doi.org/10.21037/tau.2017.09.14

2. Zini A. The benefits and limitations of sperm DNA testing in clinical practice. Translational Andrology and Urology, 2017, vol. 6, no. 4, pp. 326−327. https://doi.org/10.21037/tau.2017.09.09

3. World Health Organization. WHO laboratory manual for the examination and processing of human semen. 6th ed. Geneva, Switzerland, 2021. 276 p.

4. Agarwal A., Barbăroșie C., Ambar R., Finelli R. The impact of single- and double-strand DNA breaks in human spermatozoa on assisted reproduction. International Journal of Molecular Sciences, 2020, vol. 21, no. 11, art. 3882. https://doi.org/10.3390/ijms21113882

5. Esteves S. C., Zini A., Coward R. M., Evenson D. P., Gosálvez J., Lewis S. E. M., Sharma R., Humaidan P. Sperm DNA fragmentation testing: Summary evidence and clinical practice recommendations. Andrologia, 2021, vol. 53, no. 2, art. e13874. https://doi.org/10.1111/and.13874

6. Ribas-Maynou J., Benet J. Single and double strand sperm DNA damage: different reproductive effects on male fertility. Genes, 2019, vol. 10, no. 2, art. 105. https://doi.org/10.3390/genes10020105

7. van den Berg J., Manjón A. G., Kielbassa K., Feringa F. M., Freire R., Medema R. H. A limited number of doublestrand DNA breaks is sufficient to delay cell cycle progression. Nucleic Acids Research, 2018, vol. 46, no. 19, pp. 10132–10144. https://doi.org/10.1093/nar/gky786

8. Casanovas A., Ribas-Maynou J., Lara-Cerrillo S., Jimenez-Macedo A. R., Hortal O., Benet J., Carrera J., García-Peiró A. Double-stranded sperm DNA damage is a cause of delay in embryo development and can impair implantation rates. Fertility and Sterility, 2019, vol. 111, no. 4, pp. 699–707. https://doi.org/10.1016/j.fertnstert.2018.11.035

9. Garolla A., Cosci I., Bertoldo A., Sartini B., Boudjema E., Foresta C. DNA double strand breaks in human spermatozoa can be predictive for assisted reproductive outcome. Reproductive BioMedicine Online, 2015, vol. 31, no. 1, pp. 100–107. https://doi.org/10.1016/j.rbmo.2015.03.009

10. Coban O., Serdarogullari M., Yarkiner Z., Serakinci N. Investigating the level of DNA double-strand break in human spermatozoa and its relation to semen characteristics and IVF outcome using phospho-histone H2AX antibody as a biomarker. Andrology, 2020, vol. 8, no. 2, pp. 421–426. https://doi.org/10.1111/andr.12689

11. Ribas-Maynou J., García-Peiró A., Fernandez-Encinas A., Amengual M. J., Prada E., Cortés P., Navarro J., Benet J. Double stranded sperm DNA breaks, measured by Comet assay, are associated with unexplained recurrent miscarriage in couples without a female factor. PLoS One, 2012, vol. 7, no. 9, art. e44679. https://doi.org/10.1371/journal.pone.0044679

12. Agarwal A., Majzoub A., Esteves S. C., Ko E., Ramasamy R., Zini A. Clinical utility of sperm DNA fragmentation testing: practice recommendations based on clinical scenarios. Translational Andrology and Urology, 2016, vol. 5, no. 6, pp. 935−950. https://doi.org/10.21037/tau.2016.10.03

13. Ribas-Maynou J., Gawecka J. E., Benet J., Ward W. S. Double-stranded DNA breaks hidden in the neutral Comet assay suggest a role of the sperm nuclear matrix in DNA integrity maintenance. Molecular Human Reproduction, 2014, vol. 20, no. 4, pp. 330−340. https://doi.org/10.1093/molehr/gat090

14. Enciso M., Iglesias M., Galán I., Sarasa J., Gosálvez A., Gosálvez J. The ability of sperm selection techniques to remove single- or double-strand DNA damage. Asian Journal of Andrology, 2011, vol. 13, no. 5, pp. 764−768. https://doi.org/10.1038/aja.2011.46