Broad DNA methylation changes of spermatogenesis, inflammation and immune response‐related genes in a subgroup of sperm samples for assisted reproduction

Abstract

Aberrant sperm DNA methylation patterns, mainly in imprinted genes, have been associated with male subfertility and oligospermia. Here, we performed a genome-wide methylation analysis in sperm samples representing a wide range of semen parameters. Sperm DNA samples of 38 males attending a fertility centre were analysed with Illumina HumanMethylation27 BeadChips, which quantify methylation of >27 000 CpG sites in cis-regulatory regions of almost 15 000 genes. In an unsupervised analysis of methylation of all analysed sites, the patient samples clustered into a major and a minor group. The major group clustered with samples from normozoospermic healthy volunteers and, thus, may more closely resemble the normal situation. When correlating the clusters with semen and clinical parameters, the sperm counts were significantly different between groups with the minor group exhibiting sperm counts in the low normal range. A linear model identified almost 3000 CpGs with significant methylation differences between groups. Functional analysis revealed a broad gain of methylation in spermatogenesis-related genes and a loss of methylation in inflammation- and immune response-related genes. Quantitative bisulfite pyrosequencing validated differential methylation in three of five significant candidate genes on the array. Collectively, we identified a subgroup of sperm samples for assisted reproduction with sperm counts in the low normal range and broad methylation changes (affecting approximately 10% of analysed CpG sites) in specific pathways, most importantly spermatogenesis-related genes. We propose that epigenetic analysis can supplement traditional semen parameters and has the potential to provide new insights into the aetiology of male subfertility.

DNA methylation in spermatogenesis and male infertility

“…The methylation of constitutive heterochromatic and promoter regions is generally associated with reduced gene transcription (Fig. 1A) (15)(16)(17)(18). Therefore, DNA methylation is a type of epigenetic modification that can effectively promote gene silencing (Fig.…”

“…1B). The methyl group for this chemical modification of the DNA is donated by S-adenosyl-L-methionine (SAM) (17)(18)(19)(20)(21)(22), and the methylation reaction is catalyzed by members of the DNA methyltransferases (DNMT) family, which are classified into the following three types in mammals: DNMT1, DNMT2 and DNMT3. It was demonstrated that DNMT1, DNMT3a and DNMT3b are robustly expressed in the early embryonic stage; however, the biological functions of DNMT2 remain elusive (23-25).…”

The role of epigenetics in idiopathic male infertility

“…DAZL promoter methylated only in infertile patents [47] Global Infertile males (38) • Hypermethylation in spermatogenesis-related genes • Loss of methylation in inflammation and immune response-related genes [18] Global NOA (65) and oligoastenozoospermic (29)…”

“…These epigenetic modifications can be inherited through both mitotic and meiotic divisions. Recent studies demonstrate that aberrant DNA methylation of imprinted genes and reproduction-related genes in particular might be helpful to explain unknown infertility cases [17][18][19][20][21][22]. Therefore, epigenetics appears to be a promising research area for studying idiopathic male infertility.…”

Sperm DNA Methylation, Infertility and Transgenerational Epigenetics

“…However, methylation at non-imprinted sites may also be important for expression of the paternal genome in the developing embryo. High throughput array technology has allowed the exploration of methylation across the whole genome, and has resulted in the finding that many non-imprinted genes are also linked to poor semen parameters [59,65].…”