BackgroundMaternal effect mutations in the components of the subcortical maternal complex (SCMC) of the human oocyte can cause early embryonic failure, gestational abnormalities and recurrent pregnancy loss. Enigmatically, they are also associated with DNA methylation abnormalities at imprinted genes in conceptuses: in the devastating gestational abnormality biparental complete hydatidiform mole (BiCHM) or in multi-locus imprinting disease (MLID). However, the developmental timing, genomic extent and mechanistic basis of these imprinting defects are unknown. The rarity of these disorders and the possibility that methylation defects originate in oocytes have made these questions very challenging to address.MethodsSingle-cell bisulphite sequencing (scBS-seq) was used to assess methylation in oocytes from a patient with BiCHM identified to be homozygous for an inactivating mutation in the human SCMC component KHDC3L. Genome-wide methylation analysis of a preimplantation embryo and molar tissue from the same patient was also performed.ResultsHigh-coverage scBS-seq libraries were obtained from five KHDC3Lc.1A>G oocytes, which revealed a genome-wide deficit of DNA methylation compared with normal human oocytes. Importantly, germline differentially methylated regions (gDMRs) of imprinted genes were affected similarly to other sequence features that normally become methylated in oocytes, indicating no selectivity towards imprinted genes. A range of methylation losses was observed across genomic features, including gDMRs, indicating variable sensitivity to defects in the SCMC. Genome-wide analysis of a pre-implantation embryo and molar tissue from the same patient showed that following fertilisation methylation defects at imprinted genes persist, while most non-imprinted regions of the genome recover near-normal methylation post-implantation.ConclusionsWe show for the first time that the integrity of the SCMC is essential for de novo methylation in the female germline. These findings have important implications for understanding the role of the SCMC in DNA methylation and for the origin of imprinting defects, for counselling affected families, and will help inform future therapeutic approaches.

ObjectiveIn order to increase the number of mature oocytes usable for intracytoplasmic sperm injection (ICSI), we aimed to investigate the effect of co-culturing granulosa cells (GCs) on human oocyte maturation in vitro, the fertilization rate, and embryo development.MethodsA total of 133 immature oocytes were retrieved and were randomly divided into two groups; oocytes that were cultured with GCs (group A) and oocytes that were cultured without GCs (group B). After in vitro maturation, only oocytes that displayed metaphase II (MII) underwent the ICSI procedure. The maturation and fertilization rates were analyzed, as well as the frequency of embryo development.ResultsThe mean age of the patients, their basal levels of follicle-stimulating hormone, and the number of oocytes recovered from the patients were all comparable between the two study groups. The number of oocytes that reached MII (mature oocytes) was 59 out of 70 (84.28%) in group A, compared to 41 out of 63 (65.07%) in group B (p=0.011). No significant difference between fertilization rates was found between the two study groups (p=0.702). The embryo development rate was higher in group A (33/59, 75%) than in group B (12/41, 42.85%; p=0.006). The proportion of highest-quality embryos and the blastocyst formation rate were significantly lower in group B than in group A (p=0.003 and p<0.001, respectively).ConclusionThe findings of the current study demonstrate that culturing immature human oocytes with GCs prior to ICSI improves the maturation rate and the likelihood of embryo development.

BACKGROUND:Modern life prompted man to increasingly generate, transmit and use electricity that leads to exposure to different levels of electromagnetic fields (EMFs). Substantial evidence indicates that exposure to common sources of EMF such as mobile phones, laptops or wireless internet-connected laptops decreases human semen quality. In some countries, mobile jammers are occasionally used in offices, shrines, conference rooms and cinemas to block the signal.AIMS:To the best of our knowledge, this is the first study to investigate the effect of short term exposure of human sperm samples to radiofrequency (RF) radiations emitted by common mobile jammers.SUBJECTS AND METHODS:Fresh semen samples were collected by masturbation from 30 healthy donors who had referred to Infertility Treatment Center at the Mother and Child Hospital with their wives. Female problem was diagnosed as the reason for infertility in these couples.STATISTICAL ANALYSIS:T-test and analysis of variance were used to show statistical significance.RESULTS:The motility of sperm samples exposed to jammer RF radiation for 2 or 4 h were significantly lower than those of sham-exposed samples. These findings lead us to the conclusion that mobile jammers may significantly decrease sperm motility and the couples’ chances of conception.CONCLUSION:Based on these results, it can be suggested that in countries that have not banned mobile jammer use, legislations should be urgently passed to restrict the use of these signal blocking devices in public or private places.

Infertility affects up to 15% of reproductive-aged couples worldwide, with male factor being detected in 40%-50% of the cases. Proper sperm production is associated with the establishment of appropriate epigenetic marks in developing germ cells. Several studies have demonstrated the association between abnormal spermatogenesis and epigenetic disturbances with the major focus on DNA methylation. Imprinted genes are expressed in a parent-of-origin-specific manner, and the role of their DNA methylation in proper spermatogenesis has been documented recently. The existing evidence along with the absence of relevant data in south of Iran prompted us to study the methylation of H19 imprinted gene in spermatozoa of idiopathic infertile patients (males with abnormalities in sperm parameters) and healthy controls by Combined Bisulfite Restriction Analysis. According to our results, the lowest methylation percentage of H19 imprinted gene belongs to three cases with sperm characteristics under normal range (two cases Oligoasthenoteratozoospermia and one case Oligoteratozoospermia). However, our results show that the median of methylation percentage for H19 is not statistically significant between case and control groups. Our results and those of others introduce DNA methylation as a potential marker of fertility and should be investigated with more patients in future studies.

Abstract. This study was carried out to screen the GDF9 gene and evaluate the polymorphism effect on litter size of four Iranian sheep breeds using the PCR-RFLP and PCR-SSCP methods. First, sequencing of the GDF9 gene in 16 twin-birth, 4 triplet-birth, and 2 infertile ewes showed that, in addition to G2, G3, G4, G5, and G6 mutations that have been previously reported in other breeds, a new G0 mutation, called C25T, exists in the GDF9 sequence of 1 out of 22 ewes and causes L9F substitution in the signal peptide region. None of the triplet-birth or infertile ewes carried G1, G4, G7, FecGE, G8, or FecGT mutations. In the second experiment, a large dataset was used: 605 individuals including 496 ewes (145 Afshari, 54 Shal, 126 Ghezel, and 171 Lori-Bakhtyari sheep), and 109 rams (26 Afshari, 23 Shal, 10 Ghezel, and 50 Lori-Bakhtyari sheep. There were no sheep carrying the G7, G8, or Thoka mutations. Among all 109 rams that were used in this study, none of them were homozygous for the G1 mutation. Moreover, abundance of heterozygote rams (G1/G+) varied from 0.0 (Afshari) to 28.6 % (Lori-Bakhtyari and Ghezel). The highest and the lowest frequencies of the G4 mutation were 30.6 and 3.0 % in Shal and Afshari breeds, respectively. Moreover, G4 abundance varied from 0.0 to 42.3 %, from 3.0 to 26.9, and from 3.0 to 30.6 % in rams, ewes, and overall, respectively. There was a significant difference in the abundance of G1 and G4 mutations between breeds. However, neither the G1 nor the G4 mutation was associated with litter size in Afshari, Ghezel, Lori-Bakhtyari, or Shal sheep breeds. In conclusion, the results of this study showed that GDF9 G1 and G4 mutations are not the reason for higher litter size in Iranian sheep. Moreover, the GDF9 G0 and G6 mutations do not cause triplet births or infertility in Iranian ewes. Therefore, it is unlikely that variant GDF9 mRNA induces larger litter size or infertility in Iranian ewes.