Ventayol-Guirado Marc, Hernandez-Rodriguez Jessica, Florit Joana, Llull-Alberti Maria Victoria, Barragan Raquel, Ferragut Joana Francesca, Martorell Jaume, Heine-Suñer Damian, Martinez Iciar, Picornell Antonia, Torres-Juan Laura, Peralta Laura
Genomics of Health Research Group, Health Research Institute of the Balearic Islands (IdISBa), 07120, Palma, Balearic Islands, Spain.
Molecular Diagnostics and Clinical Genetics Unit, Hospital Universitari Son Espases, 07120, Palma, Balearic Islands, Spain.
J Assist Reprod Genet. 2025 May 22. doi: 10.1007/s10815-025-03511-8.
Cell-free mitochondrial DNA (cf-mtDNA) has emerged as a promising non-invasive method to predict embryo implantation potential. This study presents the first sequencing of cf-mtDNA in follicular fluid, aiming to explore the impact of ovarian stimulation treatment on the accumulation of mtDNA mutations and assess their implications for IVF outcomes.
We selected 24 women aged 27 to 35 from the Assisted Reproduction Unit based on inclusion criteria requiring them to be undergoing their first IVF cycle without known infertility issues. For each participant, blood samples were collected prior to ovarian stimulation, and follicular fluid samples were obtained at the time of oocyte retrieval. Long-PCR amplification and next-generation sequencing were used to identify mtDNA mutations.
Ovarian stimulation-induced mtDNA mutations in follicular fluid were identified primarily in the D-loop region of the mitochondrial genome. These mutations were correlated with low oocyte counts and poor embryo quality, ultimately reducing IVF success rates. The accumulation of these mutations followed a time-dependent pattern, with longer stimulation durations resulting in higher mutation rates and lower pregnancy outcomes. Prolonged stimulations may exacerbate oxidative stress, negatively impacting oocyte quality.
By introducing the first sequencing of follicular fluid cf-mtDNA, this study establishes a direct link between ovarian stimulation-induced mtDNA mutations and reduced IVF efficacy, highlighting them as non-invasive biomarkers for predicting IVF outcomes. Future research may focus on optimizing stimulation protocols or incorporating antioxidants to minimize mtDNA mutations, improving oocyte quality and overall IVF success rates for patients.
游离线粒体DNA（cf-mtDNA）已成为一种有前景的非侵入性方法，用于预测胚胎着床潜力。本研究首次对卵泡液中的cf-mtDNA进行测序，旨在探讨卵巢刺激治疗对mtDNA突变积累的影响，并评估其对体外受精（IVF）结局的影响。
我们从辅助生殖科选取了24名年龄在27至35岁之间的女性，入选标准要求她们正在进行首次IVF周期且无已知的不孕问题。对于每位参与者，在卵巢刺激前采集血样，并在取卵时获取卵泡液样本。采用长链聚合酶链反应（Long-PCR）扩增和下一代测序来鉴定mtDNA突变。
在卵泡液中鉴定出的卵巢刺激诱导的mtDNA突变主要位于线粒体基因组的D环区域。这些突变与低卵母细胞数量和胚胎质量差相关，最终降低了IVF成功率。这些突变的积累呈现出时间依赖性模式，刺激持续时间越长，突变率越高，妊娠结局越低。长时间刺激可能会加剧氧化应激，对卵母细胞质量产生负面影响。
通过首次对卵泡液cf-mtDNA进行测序，本研究建立了卵巢刺激诱导的mtDNA突变与IVF疗效降低之间的直接联系，突出了它们作为预测IVF结局的非侵入性生物标志物的作用。未来的研究可能集中在优化刺激方案或加入抗氧化剂以尽量减少mtDNA突变，改善卵母细胞质量和患者的总体IVF成功率。
