Epigenetic safety of in vitro maturation in PCOS: genome-wide DNA methylation profiling of cord blood from a randomized controlled trial

BACKGROUND: In vitro maturation (IVM) provides a safer alternative to conventional in vitro fertilization (IVF) for women with polycystic ovary syndrome (PCOS) by mitigating the risk of ovarian hyperstimulation. However, concerns persist regarding whether IVM perturbs epigenetic reprogramming in the offspring. Current evidence is constrained by candidate-gene approaches or a lack of parental controls. This study aimed to evaluate the genome-wide DNA methylation safety of IVM compared with conventional IVF using a rigorous trio-based design. METHODS: This secondary epigenetic analysis was nested within a randomized controlled trial (RCT) (ClinicalTrials.gov: NCT03463772). We included 10 nuclear families (trios), comprising five IVM-conceived and five IVF-conceived singleton offspring alongside their biological parents. Both groups utilized a uniform freeze-only single-blastocyst transfer strategy to minimize hormonal confounding. Genomic DNA from umbilical cord blood (UCB) and parental peripheral blood was analyzed using reduced representation bisulfite sequencing (RRBS). Genome-wide methylation patterns and differentially methylated regions (DMRs) were subsequently compared between the groups. RESULTS: Clinical characteristics were comparable between the IVM and IVF groups. Genome-wide analyses demonstrated high concordance in UCB methylation patterns, revealing no significant differences in global CpG methylation levels or distributions across key genomic features (promoters, CpG islands, and gene bodies). Only three rare DMRs were identified in UCB (representing ~ 0.0001% of the genome), none of which mapped to imprinted or developmentally critical loci. Furthermore, methylation variability remained consistent between the groups. CONCLUSIONS: Our findings provide robust mechanistic evidence supporting the epigenetic safety of IVM. The remarkable stability of the neonatal methylome confirms that specific IVM conditions do not compromise early developmental programming, thereby endorsing IVM as a safe and viable alternative for women with PCOS. TRIAL REGISTRATION: ClinicalTrials.gov registry, NCT03463772. Registered on March 13, 2018.