Comparison of oocyte vitrification using a semi-automated or a manual closed system in human siblings: survival and transcriptomic analyses

Abstract

Background Indications of oocyte vitrification increased substantially over the last decades for clinical and ethical reasons. A semi-automated vitrification system was recently developed making each act of vitrification reproducible. In this study, we evaluated the efficiency of the semi-automated technique of oocyte vitrification by survival rate, morphometric assessment and resistance to empty micro-injection gesture as compared with a manual method. Additionally, we intended to evaluate transcriptomic consequences of both techniques using single-cell RNA-seq technology. Results Post-warming survival rate, oocyte surfaces and resistance to empty micro-injection were comparable between semi-automated and manual vitrification groups. Both oocyte vitrification techniques showed limited differences in the resulting transcriptomic profile of sibling oocytes since only 5 differentially expressed genes were identified. Additionally, there was no difference in median transcript integrity number or percentage of mitochondrial DNA between the two groups. However, a total of 108 genes were differentially expressed between fresh and vitrified oocytes (FDR < 0.05) and showed over-represented of genes related to important cellular process. Conclusions Our results provide reassurance about the influence of semi-automation as compared with the manual vitrification method. Concerning oocyte vitrification itself, no tight common transcriptomic signature associated has been observed across studies. Trial registration NCT03570073.

Transcriptomics Reveal Molecular Differences in Equine Oocytes Vitrified before and after In Vitro Maturation

“…All the altered pathways in our study are directly related to the cell cycle, meiosis, and cell division [ 84 , 85 , 86 ], which are the main pathways distorted in vitrified oocytes in humans [ 75 ] and cows [ 52 , 53 ]. Other pathways related to mRNA and RNA catabolic processes and membrane-bound and intracellular organelles were affected after the vitrification of human [ 49 ] and bovine oocytes [ 53 ], which coincide with the clusters found in VIM vs. VMAT, where the metabolic process and transcription initiation in the biological function and the polymerase complex and organelle membrane in the cell component were affected. Remarkably, pathways related to calcium and ion transport, which were common in our three comparisons, have not been mentioned in other studies, except in cumulus cells after vitrification in porcine oocytes [ 55 ].…”

“…Whereas a first study in mice [ 51 ] showed minor or no changes in the transcriptome, a recent study reported 4747 DE genes [ 50 ]. Similarly, in humans, Monzo et al (2012) [ 48 ] found 608 DE genes using GeneChip arrays, and later, Huo et al (2021) [ 75 ] and Barberet et al (2022) [ 49 ] found notable differences, with a total of 1987 and 108 DE genes, respectively, using single-cell RNA-seq. In pigs and cattle, comparisons were made between fresh in vitro matured and oocytes vitrified at the GV stage followed by in vitro maturation.…”

“…Comparing different studies of the same species only revealed an alteration in a limited number of common genes. In humans, only seven DE genes were common in studies by Huo et al (2021) [ 75 ] and Barberet et al (2022) [ 49 ]. Additionally, they displayed an opposite variation of expression.…”

“…Many approaches, such as the use of antioxidants [ 8 ], ice-blockers [ 41 ], different mixtures of cryoprotectants [ 7 , 42 ], and an increased cooling rate [ 3 ] have been tried to optimize oocyte vitrification, but the achieved outcomes are still unsatisfactory. Different studies have reported differential expression in development-related genes after vitrification [ 43 , 44 , 45 , 46 ] and the transcriptomes of vitrified vs. fresh oocytes have been reported in humans [ 47 , 48 , 49 ], mice [ 50 , 51 ], cattle [ 52 , 53 , 54 ], and porcine [ 55 ]. To the best of our knowledge, no transcriptomics studies have been performed on the vitrified oocytes of horses.…”

Overview of Gene Expression Dynamics during Human Oogenesis/Folliculogenesis

“…In addition, throughout growth and maturation, oocytes are exposed to multiple environmental stressors, which can alter the sequential gene expression changes governing oocyte development [ 56 ]. Our identification of transcripts expressed along oocyte development that are highly susceptible to those factors is of major importance, as they may be the key to understanding the apparition of oocyte defects and deficiencies and overcoming these outcomes [ 27 , 28 , 29 , 57 , 58 , 59 , 60 , 61 , 62 , 63 , 64 , 65 , 66 , 67 ]. For oocyte in vitro maturation and cryoconservation, regaining expected levels of expression of those transcripts by adapting assisted reproductive protocols may become a path of research to increase mature oocyte rates in the future.…”