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标题： Closed culture system can reduce human embryo gene expression and DNA methylation changes caused by assisted reproductive technology

讲者：李竞宇

单位：重庆市妇幼保健院

关键词：

human embryo time-lapse RNA-seq DNA methylation assisted reproductive technology

播放： 3699

论文摘要： Study question: Do embryos cultured in closed system and standard incubator have different gene expression and DNA methylation profiles?

Summary answer: Embryos cultured in a closed system display a more stable gene expression and less epigenetic disorders profiles compared with the embryos cultured in a standard incubation system.

What is known already: Although many studies investigated the effect of time-lapse monitoring (TLM) and standard incubation (SI) system on embryonic development or clinical outcomes, whether culture of human embryos in the two system will result in different gene expression and DNA methylation have not been described.

Study design, size, duration: This study included 60 women, year range 25-30, without any ovarian pathology, undergoing IVF treatment. Surplus MII oocytes and Day 3 embryos were donated after written informed consent. Only one oocyte or embryo was donated from one couple. Pools of 5 oocytes or embryos for each sample were prepared. Every group for RNA-seq had three independent technical replicates. Thus, a total of 15 MII oocytes, 30 embryos (15 embryos from TLM and 15 embryos from SI) were processed for RNA-Seq analysis. For methylation sequencing., only biological replication were performed. A total of 5 MII oocytes, 10 embryos (5 embryos from TLM and 5 embryos from SI) were processed for methylation analysis. Recruitment took place from Dec 2016 to September 2017.

Participants/materials, setting, methods: Embryos with the similar quality from TLM and SI were prepared for this experiment. RNA-Seq libraries were loaded on Illumina Hiseq 4000 platform for 150-bp paired-end sequencing. DNA methylation libraries were prepared for 125-bp paired-end sequencing on HiSeq2500 platform. After sequencing, bioinformatics analysis (hierarchical cluster, principal component analysis, DESeq2, SeqMonk, Gene ontology analysis) were performed.

Main results and the role of chance: RNA-Seq data showed that the SI group have higher variability of gene expression patterns than TLM. In addition, more genes involved in RNA splicing and DNA repair were activated in SI group. We analyzed the DNA methylation data and found global demethylation from zygotes to 8-cell embryos cultured in SI and TLM. Relatively higher methylation was observed in TLM group. Furthermore, differentially methylated regions analysis identified several genes whose methylation could be critical, such as IGF2R and PEG10.

Limitations, reasons for caution: The embryo may correct several alterations in gene expression and methylation with its recovered ability during a longer culture time. Pooling embryos may lead to the loss of information on inter-embryo heterogeneity. No technical replicates were performed in the methylation sequencing. Finally, further research should be undertaken to investigate whether these differential transcripts and methylated regions contribute directly to human embrynic development or birth defects.

Wider implications of the findings : This study provides the first comprehensive comparison between SI and TLM via transcriptome and DNA methylation analysis, and will serve as a basis for assessing the safety of TLM application in assisted reproductive technology.