Sperm Ribosomal DNA Promoter Methylation Levels Are Correlated With Paternal Aging and May Relate With in vitro Fertilization Outcomes

Abstract

The impact of aging on reproductive outcomes has received considerable critical attention; however, there is much less information available on the effects of paternal age compared to the effects of maternal age. In this study, methylation levels of sperm rDNA promoter regions and Long Interspersed Nucleotide Element 1 (LINE-1) were measured using pyrosequencing and fertilization, day 3 good-quality embryo, pregnancies, and implantation results were assessed. We observed significantly increasing levels of DNA methylation in the sperm rDNA promoter regions with age based on stratifying the samples by age alone (P = 0.0001) and performing linear regression analysis (P < 0.0001). Meanwhile, no statistically significant correlations were observed between global LINE-1 methylation with age. No statistically significant correlations were observed between sperm rDNA promoter methylation levels and either the day 3 goodquality embryo rate or clinical pregnancy rate. In contrast, the correlation between sperm rDNA promoter methylation levels and fertilization (2 pronuclei) rate was nearly significant (P = 0.0707), especially the methylation levels of some individual CpG units (CpG_10, P = 0.0176; CpG_11, P = 0.0438; CpG_14, P = 0.0232) and rDNA promoter methylation levels measured using primerS2 (P = 0.0513). No significant correlation was found between sperm rDNA promoter methylation levels and fertilization rates (2 pronuclei, 1 pronuclei, and 1 polypronuclei). Our results demonstrate that sperm are susceptible to age-associated alterations in methylation levels of rDNA promoter regions, suggesting that sperm rDNA promoter methylation levels can be applied to DNA methylation-based age prediction, and that the aberrant methylation of rDNA promoters may be partially responsible for enhanced disease susceptibility of offspring sired by older fathers. Methylation levels of sperm rDNA promoter regions may correlate with polypronuclei rates of IVF programs.

Mitochondrial Reactive Oxygen Species (ROS) Production Alters Sperm Quality

“…The father’s advanced age also correlates with abnormal sperm DNA methylation that is not only confined to differentially methylated regions, but widespread in the genome, especially at the level of regions associated with the control of schizophrenia and/or bipolar disorders [ 166 ]. Interestingly, the methylation of ribosomal DNA increases with age in both somatic and sperm cells, thus influencing nucleolar formation and embryo development [ 167 ].…”

Age-associated epigenetic changes in mammalian sperm: implications for offspring health and development

“…A model involving 15 of the most robust CpGs showed a correlation between the predicted epigenetic age and chronological age ( r = 0.67, median absolute difference between predicted and chronological age 2.91). In a similar study using 60 couples who underwent IVF, a positive correlation was found between the methylation of 21 CpG sites in sperm rDNA promoter and donor age, but no statistically significant correlation was observed between rDNA methylation and IVF outcomes (successful fertilization, Day 3 good-quality embryos, clinical pregnancies) ( Li et al , 2020 ). It should be noted that the rDNA methylation clock gives a lower correlation compared with the epigenetic clock of Jenkins et al described above ( Jenkins et al , 2018 ).…”

Role and Mechanism of Epigenetic Regulation in the Aging of Germ Cells: Prospects for Targeted Interventions

“…Moreover, evidence showed that sperm from older men had more hypermethylated than hypomethylated CpG sites [50]. Additionally, the level of DNA hypermethylation in the rDNA promoter region of sperm increased significantly with age [51].…”