RESUMEN
BACKGROUND Assisted reproductive technologies (ART) such as in vitro fertilisation (IVF) and intracytoplasmic sperm injection (ICSI) are believed to destabilise genomic imprints. An increased frequency of Beckwith-Wiedemann syndrome in children born after ART has been reported. Other, mostly epidemiological, studies argue against this finding. OBJECTIVE To examine the effect of ART on the stability of DNA methylation imprints, DNA was extracted from maternal peripheral blood (MPB), umbilical cord blood (UCB) and amnion/chorion tissue (ACT) of 185 phenotypically normal children (77 ICSI, 35 IVF, and 73 spontaneous conceptions). Using bisulfite based technologies 10 differentially methylated regions (DMRs) were analysed, including KvDMR1, H19, SNRPN, MEST, GRB10, DLK1/MEG3 IG-DMR, GNAS NESP55, GNAS NESPas, GNAS XL-alpha-s and GNAS Exon1A. RESULTS Methylation indices (MI) do not reveal any significant differences at nine DMRs among the conception groups in neither MPB, UCB nor in ACT. The only slightly variable DMR was that of MEST. Here the mean MI was higher in UCB and MPB of IVF cases (mean MI+/-SD: 0.41+/-0.03 (UCB) and 0.40+/-0.03 (MPB)) compared to the ICSI (0.38+/-0.03, p=0.003 (UCB); 0.37+/-0.04, p=0.0007 (MPB)) or spontaneous cases (0.38+/-0.03, p=0.003 (UCB); 0.38+/-0.04, p=0.02 (MPB)). Weak but suggestive correlations between DMRs were, however, found between MPB, UCB and ACT. CONCLUSION This study supports the notion that children conceived by ART do not show a higher degree of imprint variability and hence do not have an a priori higher risk for imprinting disorders.
Asunto(s)
Metilación de ADN , Genoma Humano/genética , Inestabilidad Genómica/genética , Técnicas Reproductivas Asistidas , Amnios/metabolismo , Proteínas de Unión al Calcio , Corion/metabolismo , Cromograninas , ADN/química , ADN/genética , ADN/aislamiento & purificación , Femenino , Sangre Fetal/metabolismo , Proteína Adaptadora GRB10/genética , Subunidades alfa de la Proteína de Unión al GTP Gs/genética , Humanos , Recién Nacido , Péptidos y Proteínas de Señalización Intercelular/genética , Masculino , Proteínas de la Membrana/genética , Canales de Potasio con Entrada de Voltaje/genética , Embarazo , Proteínas/genética , ARN Largo no Codificante , ARN no Traducido/genética , Análisis de Secuencia de ADN , Proteínas Nucleares snRNP/genéticaRESUMEN
Aberrant in sperm DNA methylation patterns and histone modification play a role in male fecundity decline. This study was prepared to determine whether sperm DNA methylation at CpG dinucleotides is different in oligospermic males compared to proven fertile males and then to evaluate the correlation between the changes in sperm DNA methylation patterns and semen parameters of oligospermic males. A total of 165 males (64 proven fertile males "controls" and 101 oligospermic males "cases") were included in the study. Three CpG sites have the highest difference in methylation levels (cg23081194, cg25750688, and cg04807108) were underwent to further analysis using deep bisulfite sequencing in 125 samples (44 controls and 81 cases). The results of a validation study showed that variation in methylation levels was found in more than one CpG site: there was a significant alteration in methylation levels at all CpGs tested within the UBE2G2 and cg25750688 site related amplicon (p≤0.0001), and at eight CpGs (CpG1, CpG3, CpG6, CpG8, CpG11, CpG13, CpG14, and CpG15) within the cg04807108 site related amplicon (p≤0.0001) in cases compared to controls. Besides, a significant correlation was found between the changes in the methylation levels at different CpGs and semen parameters of case group. In conclusion, this study showed that these sites have a significant alteration in sperm DNA methylation levels in oligospermic males compared to proven fertile males, and these changes correlated with semen parameters.