Association between promoter methylation of MLH1 and MSH2 and reactive oxygen species in oligozoospermic men-A pilot study

Abstract

MLH1 and MSH2 are important genes for DNA mismatch repair and crossing over during meiosis and are implicated in male infertility. Therefore, the methylation patterns of the DNA mismatch repair genes MLH1 and MSH2 in oligozoospermic males were investigated. Ten oligozoospermic patients and 29 normozoospermic donors were analysed. Methylation profiles of the MLH1 and MSH2 promotors were analysed. In addition, sperm motility and seminal reactive oxygen species (ROS) were recorded. Receiver operating characteristic (ROC) analysis was conducted to determine the accuracy of the DNA methylation status of MLH1 and MSH2 to distinguish between oligozoospermic and normozoospermic men. In oligozoospermic men, MLH1 was significantly (p = .0013) more methylated compared to normozoospermic men. Additionally, there was a significant positive association (r = .384; p = .0159) between seminal ROS levels and MLH1 methylation. Contrary, no association between MSH2 methylation and oligozoospermia was found. ROC curve analysis for methylation status of MLH1 was significant (p = .0275) with an area under the curve of 61.1%, a sensitivity of 22.2% and a specificity of 100.0%. This pilot study indicates oligozoospermic patients have more methylation of MLH1 than normozoospermic patients. Whether hypermethylation of the MLH1 promoter plays a role in repairing relevant mismatches of sperm DNA strands in idiopathic oligozoospermia warrants further investigation.

Chromosomal and Y‐chromosome microdeletion analysis in 1,300 infertile males and the fertility outcome of patients with AZFc microdeletions

“…Infertility affects about 15% of couples and 12% of men during their reproductive years, and male infertility is responsible for half of these infertility cases (Agarwal et al, ; Gunes, Arslan, Hekim, & Asci, ). The aetiology is unknown in about 30% of male infertility cases (Gunes et al, ). Genetic factors are highlighted as the most frequent male infertility causes.…”

Microarray meta‐analysis identifies candidate genes for human spermatogenic arrest

“…Some of the p53‐related genes in our results were previously studied in humans or animals. In humans, the level of MLH1 methylation in NOA patients is significantly higher than that in healthy subjects and is positively correlated with the level of reactive oxygen species (ROS; Gunes et al, ). Moreover, a recent human study indicated that single nucleotide polymorphisms (SNPs) of PARP1 are highly associated with oligospermia (Chen, Pu, Zhang, & A, ).…”

Investigating the relationship between BRCA1 and BRCA2 genes methylation profile and sperm DNA fragmentation in infertile men

“…For somatic cell lysis, first an aliquot of liquefied ejaculate sample containing <10 × 10 6 /ml spermatozoa was added into a test tube and centrifuged for 8 min at 1,500 × g to remove the seminal plasma. After washing this pellet with 1 ml of 1× PBS, the pellet was treated with 12 ml of somatic cell lysis buffer (0.5% Triton X‐100 and 0.1% sodium dodecyl sulphate [SDS] in 200 ml of distilled water) on ice for 30 min (Gunes et al, ). Subsequently, the supernatant was removed by centrifugation for 15 min at 300 × g .…”

“…After the complete elimination of somatic cells, the sperm DNA was isolated with QIAamp DNA Mini kit (Qiagen) according to the manufacturer's instructions (Gunes et al, ). DNA samples were stored at −80ºC until bisulphite modification step.…”

“…Bisulphite modification of sperm DNA samples was carried out using the EZ DNA Methylation kit (Zymo Research; Gunes et al, ).…”