DNA methylation in spermatozoa as a prospective marker in andrology

Abstract

Clinics of M€ unster, M€ unster, Germany SUMMARYRecent studies have shown associations of aberrant DNA methylation in spermatozoa with idiopathic infertility. The analysis of DNA methylation of specific genes could therefore serve as a valuable diagnostic marker in clinical andrology. For this purpose, rapid and reliable detection methods, reference values and the temporal stability of spermatozoal DNA methylation need to be established and demonstrated. In this prospective study, swim-up purified semen samples from 212 consecutive patients (single samples), 31 normozoospermic volunteers (single samples) and 10 normozoospermic volunteers (four samples at days 1, 3, 42 and 45 plus a fifth sample after 180-951 days) were collected. Spermatozoal DNA was isolated, bisulphite converted and DNA methylation was analysed by pyrosequencing. DNA methylation of the maternally imprinted gene MEST was measured in samples of 212 patients and 31 normozoospermic volunteers and the temporal stability of eight different genes and two repetitive elements was examined in consecutive samples of 10 normozoospermic volunteers. MEST DNA methylation was significantly associated with oligozoospermia, decreased bi-testicular volume and increased FSH levels. A reference range for spermatozoal MEST DNA methylation (0-15%) was established using the 95th percentile of DNA methylation in normozoospermic volunteers. Using this reference range, around 23% of our patient cohort displayed an aberrant MEST DNA methylation. This epigenetic aberration was found to be significantly associated with bi-testicular volume, sperm concentration and total sperm count. DNA methylation in normozoospermic volunteers was stable over a time period of up to 951 days in contrast to classical semen parameters. Our data show that MEST DNA methylation fulfils the prerequisites to be used as routine parameter and support its use during andrological workup if a prognostic value can be shown in future.

DNA methylation in spermatogenesis and male infertility

“…However, DNMT1 heterozygous mice had a normal reproductive ability, thus suggesting that DNMT1 in the heterozygous state is sufficient to maintain the required DNA methylation pattern (30,75,76). DNMT3a expression in the testis is upregulated prior to birth and during early postnatal life, whereas DNMT3b expression is downregulated during embryonic development and upregulated postnatally (15)(16)(17)(18)(19)(20). Notably, mice lacking DNMT3L have smaller testes, and a negligible number of spermatozoa by adulthood, resulting in sterile animals.…”

“…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).…”

Use of FTA ® classic cards for epigenetic analysis of sperm DNA

“…This approach can be implemented in geographically distant laboratories, eliminating the problems associated with biological sample transportation. Our procedure allows repeated testing over an extended time-span of many years following sample collection, especially because it has been demonstrated that methylation of sperm DNA is not subject to change over time (14). Since assisted fertilization techniques have increasingly shown problems arising after a significant time-span (15), it becomes increasingly important to monitor the methylation status of germ cell genes.…”

Sperm DNA Methylation, Infertility and Transgenerational Epigenetics

“…MEST hypermethylation has repeatedly been linked to low sperm count, low progressive sperm motility and poor sperm morphology [8,61]. MEST is a candidate gene for the developmental disorder Silver-Russell Syndrome (SRS).…”