Zona pellucida from fertilised human oocytes induces a voltage-dependent calcium influx and the acrosome reaction in spermatozoa, but cannot be penetrated by sperm

BACKGROUND: The functions of three zona glycoproteins, ZP1, ZP2 and ZP3 during the sperm-zona pellucida (ZP) interaction are now well established in mice. The expression of an additional zona glycoprotein, ZPB/4, in humans, led us to reconsider the classical mouse model of gamete interaction. We investigated the various functions of human ZP (hZP) during the interaction of spermatozoa with fertilised and unfertilised oocytes. RESULTS: The hZP of fertilised oocytes retained their ability to bind sperm (albeit less strongly than that from unfertilised oocytes), to induce an intraspermatic calcium influx through voltage-dependent channels similar to that observed with hZP from unfertilised oocytes and to promote the acrosome reaction at a rate similar to that induced by the ZP of unfertilised oocytes (61.6 ± 6.2% vs60.7 ± 9.1% respectively). Conversely, the rate of hZP penetrated by sperm was much lower for fertilised than for unfertilised oocytes (19% vs 57% respectively, p < 0.01). We investigated the status of ZP2 in the oocytes used in the functional tests, and demonstrated that sperm binding and acrosome reaction induction, but not ZP penetration, occurred whether or not ZP2 was cleaved. CONCLUSION: The change in ZP function induced by fertilisation could be different in human and mouse species. Our results suggest a zona blocking to polyspermy based at the sperm penetration level in humans

Assessing the influence of distinct culture media on human pre-implantation development using single-embryo transcriptomics

The use of assisted reproductive technologies is consistently rising across the world. However, making an informed choice on which embryo culture medium should be preferred to ensure satisfactory pregnancy rates and the health of future children critically lacks scientific background. In particular, embryos within their first days of development are highly sensitive to their micro-environment, and it is unknown how their transcriptome adapts to different embryo culture compositions. Here, we determined the impact of culture media composition on gene expression in human pre-implantation embryos. By employing single-embryo RNA-sequencing after 2 or 5 days of the post-fertilization culture in different commercially available media (Ferticult, Global, and SSM), we revealed medium-specific differences in gene expression changes. Embryos cultured pre-compaction until day 2 in Ferticult or Global media notably displayed 266 differentially expressed genes, which were related to essential developmental pathways. Herein, 19 of them could have a key role in early development, based on their previously described dynamic expression changes across development. When embryos were cultured after day 2 in the same media considered more suitable because of its amino acid enrichment, 18 differentially expressed genes thought to be involved in the transition from early to later embryonic stages were identified. Overall, the differences were reduced at the blastocyst stage, highlighting the ability of embryos conceived in a suboptimal in vitro culture medium to mitigate the transcriptomic profile acquired under different pre-compaction environments

Assessing the influence of distinct culture media on human pre-implantation development using single-embryo transcriptomics

The use of assisted reproductive technologies is consistently rising across the world. However, making an informed choice on which embryo culture medium should be preferred to ensure satisfactory pregnancy rates and the health of future children critically lacks scientific background. In particular, embryos within their first days of development are highly sensitive to their micro-environment, and it is unknown how their transcriptome adapts to different embryo culture compositions. Here, we determined the impact of culture media composition on gene expression in human pre-implantation embryos. By employing single-embryo RNA-sequencing after 2 or 5 days of the post-fertilization culture in different commercially available media (Ferticult, Global, and SSM), we revealed medium-specific differences in gene expression changes. Embryos cultured pre-compaction until day 2 in Ferticult or Global media notably displayed 266 differentially expressed genes, which were related to essential developmental pathways. Herein, 19 of them could have a key role in early development, based on their previously described dynamic expression changes across development. When embryos were cultured after day 2 in the same media considered more suitable because of its amino acid enrichment, 18 differentially expressed genes thought to be involved in the transition from early to later embryonic stages were identified. Overall, the differences were reduced at the blastocyst stage, highlighting the ability of embryos conceived in a suboptimal in vitro culture medium to mitigate the transcriptomic profile acquired under different pre-compaction environments

In Vitro Fertilization and Embryo Culture Strongly Impact the Placental Transcriptome in the Mouse Model

BACKGROUND: Assisted Reproductive Technologies (ART) are increasingly used in humans; however, their impact is now questioned. At blastocyst stage, the trophectoderm is directly in contact with an artificial medium environment, which can impact placental development. This study was designed to carry out an in-depth analysis of the placental transcriptome after ART in mice. METHODOLOGY/PRINCIPAL FINDINGS: Blastocysts were transferred either (1) after in vivo fertilization and development (control group) or (2) after in vitro fertilization and embryo culture. Placentas were then analyzed at E10.5. Six percent of transcripts were altered at the two-fold threshold in placentas of manipulated embryos, 2/3 of transcripts being down-regulated. Strikingly, the X-chromosome harbors 11% of altered genes, 2/3 being induced. Imprinted genes were modified similarly to the X. Promoter composition analysis indicates that FOXA transcription factors may be involved in the transcriptional deregulations. CONCLUSIONS: For the first time, our study shows that in vitro fertilization associated with embryo culture strongly modify the placental expression profile, long after embryo manipulations, meaning that the stress of artificial environment is memorized after implantation. Expression of X and imprinted genes is also greatly modulated probably to adapt to adverse conditions. Our results highlight the importance of studying human placentas from ART

The hypomethylation of imprinted genes in IVF/ICSI placenta samples is associated with concomitant changes in histone modifications

Although more and more children are born by Assisted Reproductive Technologies (ART), ART safety has not fully been demonstrated. Notably, ART could disturb the delicate step of implantation, and trigger placenta-related adverse outcomes with potential long-term effects, through disrupted epigenetic regulation. We have previously demonstrated that placental DNA methylation was significantly lower after IVF/ICSI than following natural conception at two differentially methylated regions (DMRs) associated with imprinted genes (IGs): H19/IGF2 and KCNQ1OT1. As histone modifications are critical for placental physiology, the aim of this study was to profile permissive and repressive histone marks in placenta biopsies to reveal a better understanding of the epigenetic changes in the context of ART. Utilizing chromatin immunoprecipitation (ChIP) coupled with quantitative PCR, permissive (H3K4me3, H3K4me2, and H3K9ac) and repressive (H3K9me3 and H3K9me2) post-translational histone modifications were quantified. The analyses revealed a significantly higher quantity of H3K4me2 precipitation in the IVF/ICSI group than in the natural conception group for H19/IGF2 and KCNQ1OT1 DMRs (P = 0.016 and 0.003, respectively). Conversely, the quantity of both repressive marks at H19/IGF2 and SNURF DMRs was significantly lower in the IVF/ICSI group than in the natural conception group (P = 0.011 and 0.027 for H19/IGF2; and P = 0.010 and 0.035 for SNURF). These novel findings highlight that DNA hypomethylation at imprinted DMRs following ART is linked with increased permissive/decreased repressive histone marks, altogether promoting a more permissive chromatin conformation. This concomitant change in epigenetic state at IGs at birth might be an important developmental event because of ART manipulations

Optimal Timing for Oocyte Denudation and Intracytoplasmic Sperm Injection

Objectives. To analyze the impact of oocyte denudation and microinjection timings on intracytoplasmic sperm injection (ICSI) outcomes. Study Design. We included ICSI cycles with the following parameters: rank 1 or 2, female age <36 years, male factor infertility, long protocol using GnRH agonist and rFSH for ovarian stimulation, and use of freshly ejaculated sperm (=110). Several ICSI parameters were analyzed according to the time between oocyte retrieval and denudation (1) and the time between denudation and ICSI (2) using a statistical logistic regression analysis. Results. Neither 1 nor 2 had a significant influence on the Metaphase II (MII) rate but the fertilisation rate (FR) showed a significant improvement when 1 was longer (optimal results at 1=3 hours) while FR significantly decreased with the increase of 2. Optimal implantation (IR) and pregnancy (PR) rates were obtained when 1 was around 2 hours. Conclusion. Incubation of oocytes around 2 hours between retrieval and denudation may not increase MII rate but appears to lead to the optimal combination of FR and IR

Differences in expression rather than methylation at placenta-specific imprinted loci is associated with intrauterine growth restriction

Background: genome-wide studies have begun to link subtle variations in both allelic DNA methylation and parent-of-origin genetic effects with early development. Numerous reports have highlighted that the placenta plays a critical role in coordinating fetal growth, with many key functions regulated by genomic imprinting. With the recent description of wide-spread polymorphic placenta-specific imprinting, the molecular mechanisms leading to this curious polymorphic epigenetic phenomenon is unknown, as is their involvement in pregnancies complications. Results: profiling of 35 ubiquitous and 112 placenta-specific imprinted differentially methylated regions (DMRs) using high-density methylation arrays and pyrosequencing revealed isolated aberrant methylation at ubiquitous DMRs as well as abundant hypomethylation at placenta-specific DMRs. Analysis of the underlying chromatin state revealed that the polymorphic nature is not only evident at the level of allelic methylation, but DMRs can also adopt an unusual epigenetic signature where the underlying histones are biallelically enrichment of H3K4 methylation, a modification normally mutually exclusive with DNA methylation. Quantitative expression analysis in placenta identified two genes, GPR1-AS1 and ZDBF2, that were differentially expressed between IUGRs and control samples after adjusting for clinical factors, revealing coordinated deregulation at the chromosome 2q33 imprinted locus. Conclusions: DNA methylation is less stable at placenta-specific imprinted DMRs compared to ubiquitous DMRs and contributes to privileged state of the placenta epigenome. IUGR-associated expression differences were identified for several imprinted transcripts independent of allelic methylation. Further work is required to determine if these differences are the cause IUGR or reflect unique adaption by the placenta to developmental stresses

Comment on Cannarella et al. DNA Methylation in Offspring Conceived after Assisted Reproductive Techniques: A Systematic Review and Meta-Analysis. <i>J. Clin. Med.</i> 2022, <i>11</i>, 5056

We read the study by Cannarella et al. [...

Analyse rétrospective de la prise en charge de l'infertilité des hommes bléssés médullaires au CECOS de Dijon

DIJON-BU Médecine Pharmacie (212312103) / SudocSudocFranceF

Facteurs de succès en insémination intra-utérine (influence du temps entre la fin de la préparation spermatique et l'insémination : une étude multicentrique)

Objectif : Évaluer l influence du délai entre la fin de la préparation spermatique et l insémination sur le taux de succès défini par la présence d un sac gestationnel intra-utérin un mois après cette AMP. Matériel et Méthode : D avril à juin 2012, 490 cycles d insémination ont été inclus dans une étude prospective multicentrique. Différents paramètres cliniques et biologiques ont été analysés en analyse uni et multivariée. Résultats : Le délai entre la fin de préparation et l insémination a une influence sur le taux de grossesse tant en analyse univariée que multivariée avec une relation non linéaire. Les 3 autres facteurs prédictifs de succès retrouvés sont l âge de la femme (relation linéaire), le diagnostic d infertilité non idiopathique, et la numération de spermatozoïdes mobiles inséminés (relation non linéaire). Les chances maximales de grossesse sont obtenues dans la population de référence suivante : femme de 25 ans, diagnostic non idiopathique, numération comprise entre 1 et 10 millions et temps fin de préparation-insémination entre 40 et 80 minutes. Le simple fait de respecter ce délai pour un cycle compense l effet négatif de 16 ans d âge féminin. Conclusion : Notre étude montre pour la première fois dans la littérature que le fait de respecter un temps compris entre 40 et 80 min de la fin de préparation spermatique à l insémination augmente de façon significative les chances de grossesse en IIU. Cet élément organisationnel reproductible est très facile à mettre en place dans l organisation des centres et ne demande aucun investissement financier. Notre étude est poursuivie pour validation interne et pourrait devenir une recommandation de bonnes pratiques cliniques.DIJON-BU Médecine Pharmacie (212312103) / SudocSudocFranceF