Altered DNA methylation patterns in sperm from oligozoospermic men

Abstract

Investigate methylation changes in sperm DNA from infertile men. We characterized methylation changes in sperm DNA isolated from men with idiopathic oligozoospermic and normospermic controls. Human spermatozoa samples were collected in both Hopital Tenon and Laboratoire Eylau. Spermatozoa were separated from somatic cells and subjected to DNA isolation. Genomic DNA was bisulfite converted. We have investigated by PCR and cloning the methylation pattern H19 (Maternally expressed, methylated in sperm) in spermatozoa of oligozoospermic men. Genome-wide DNA methylation patterns at promoter elements from sperm DNA was analyzed on selected cases using the MeDIP (Methyl-DNA Immunoprecipitation) based technique. 3microg of genomic DNA were submitted to immunoprecipitation with an antibody directed against 5-methylCyosine.The immunoprecipitated and Input DNA samples were labeled and hybridized to 2.1 M deluxe human promoter array. Methylation status of H19 was analyzed in spermatozoa DNA from 294 men (119 normospermic [>20x106 spz/ ml], 116 mild oligozoospermic [5-20x106 spz /ml] and 59 severe oligozoospermic [<5x106 spz/ ml]). Hypomethylation of H19-DMR was found in 16 mild oligozoospermic (13.8%, P=2.7.10-5) men and 19 severe oligozoospermic (32%, P=5.7.10-11) men, respectively. MeDIP analysis in sperm of oligozoospermic men revealed global DNA methylation changes. These included the promoter elements of a series of imprinted genes as well as over a hundred genes involved in spermatogenesis. Mature sperm from 20% of oligozoospermic men have altered DNA methylation patterns. The degree of hypomethylation increased with reduced sperm counts and may be genome-wide. It remains to be determined if oligozoospermic men with sperm DNA methylation changes are at a higher risk for IVF failure. These aberrant epigenetic changes may also have serious health implications for children conceived using such sperm by IVF.