0588478 - ÚŽFG 2025 RIV GB eng J - Journal Article
Dvořan, Michal - Iyyappan, Rajan - Mašek, T. - Pospíšek, M. - Kubelka, Michal - Šušor, Andrej
Assessment of active translation in cumulus-enclosed and denuded oocytes during standard in vitro maturation and early embryo development.
Human Reproduction. Roč. 39, č. 8 (2024), č. článku deae126. ISSN 0268-1161. E-ISSN 1460-2350
R&D Projects: GA ČR(CZ) GA22-27301S; GA MŠMT EF15_003/0000460
Institutional support: RVO:67985904
Keywords : in vitro maturation * oocyte * embryo * art * reproduction * mouse * human
OECD category: Developmental biology
Impact factor: 6, year: 2023 ; AIS: 1.911, rok: 2023
Method of publishing: Limited access
Result website:
https://academic.oup.com/humrep/article-abstract/39/8/1752/7692003?redirectedFrom=fulltext&login=trueDOI: https://doi.org/10.1093/humrep/deae126

STUDY QUESTION Which actively translated maternal transcripts are differentially regulated between clinically relevant in vitro and in vivo maturation (IVM) conditions in mouse oocytes and zygotes?SUMMARY ANSWER Our findings uncovered significant differences in the global transcriptome as well as alterations in the translation of specific transcripts encoding components of energy production, cell cycle regulation, and protein synthesis in oocytes and RNA metabolism in zygotes.WHAT IS KNOWN ALREADY Properly regulated translation of stored maternal transcripts is a crucial factor for successful development of oocytes and early embryos, particularly due to the transcriptionally silent phase of meiosis.STUDY DESIGN, SIZE, DURATION This is a basic science study utilizing an ICR mouse model, best suited for studying in vivo maturation. In the treatment group, fully grown germinal vesicle oocytes from stimulated ovaries were in vitro matured to the metaphase II (MII) stage either as denuded without gonadotropins (IVM DO), or as cumulus-oocyte complexes (IVM COC) in the presence of 0.075 IU/ml recombinant FSH (rFSH) and 0.075 IU/ml recombinant hCG (rhCG). To account for changes in developmental competence, IVM COC from non-stimulated ovaries (IVM COC-) were included. In vivo matured MII oocytes (IVO) from stimulated ovaries were used as a control after ovulation triggering with rhCG. To simulate standard IVM conditions, we supplemented media with amino acids, vitamins, and bovine serum albumin. Accordingly, in vitro pronuclear zygotes (IMZ) were generated by IVF from IVM DO, and were compared to in vivo pronuclear zygotes (IVZ). All experiments were performed in quadruplicates with samples collected for both polyribosome fractionation and total transcriptome analysis. Samples were collected over three consecutive months.PARTICIPANTS/MATERIALS, SETTING, METHODS All ICR mice were bred under legal permission for animal experimentation (no. MZE-24154/2021-18134) obtained from the Ministry of Agriculture of the Czech Republic. Actively translated (polyribosome occupied) maternal transcripts were detected in in vitro and in vivo matured mouse oocytes and zygotes by density gradient ultracentrifugation, followed by RNA isolation and high-throughput RNA sequencing. Bioinformatic analysis was performed and subsequent data validation was done by western blotting, radioactive isotope, and mitotracker dye labelling.MAIN RESULTS AND THE ROLE OF CHANCE Gene expression analysis of acquired polysome-derived high-throughput RNA sequencing data revealed significant changes (RPKM >= 0.2, P <= 0.005) in translation between in vitro and in vivo matured oocytes and respectively produced pronuclear zygotes. Surprisingly, the comparison between IVM DO and IVM COC RNA-seq data of both fractionated and total transcriptome showed very few transcripts with more than a 2-fold difference. Data validation by radioactive isotope labelling revealed a decrease in global translation bof20% in IVM DO and COC samples in comparison to IVO samples. Moreover, IVM conditions compromised oocyte energy metabolism, which was demonstrated by both changes in polysome recruitment of each of 13 mt-protein-coding transcripts as well as by validation using mitotracker red staining.LARGE SCALE DATA The data discussed in this publication have been deposited in NCBI's Gene Expression Omnibus and are accessible through GEO Series accession number GSE241633 (https://www.ncbi.nlm.nih.gov/geo/query/acc.
Permanent Link: https://hdl.handle.net/11104/0355455