Skip to main page content
U.S. flag

An official website of the United States government

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2024 Jun 24;25(1):632.
doi: 10.1186/s12864-024-10532-7.

Deregulation of oxidative phosphorylation pathways in embryos derived in vitro from prepubertal and pubertal heifers based on whole-transcriptome sequencing

Milena Traut  1 , Ilona Kowalczyk-Zieba  1 , Dorota Boruszewska  1 , Joanna Jaworska  1 , Sandra Gąsiorowska  1 , Krzysztof Lukaszuk  2   3 , Katarzyna Ropka-Molik  4 , Katarzyna Piórkowska  4 , Tomasz Szmatoła  4   5 , Izabela Woclawek-Potocka  6
Affiliations

Deregulation of oxidative phosphorylation pathways in embryos derived in vitro from prepubertal and pubertal heifers based on whole-transcriptome sequencing

Milena Traut et al. BMC Genomics. .

Abstract

Background: Although, oocytes from prepubertal donors are known to be less developmentally competent than those from adult donors it does not restrain their ability to produce full-term pregnancies. The transcriptomic profile of embryos could be used as a predictor for embryo's individual developmental competence. The aim of the study was to compare transcriptomic profile of blastocysts derived from prepubertal and pubertal heifers oocytes. Bovine cumulus-oocyte complexes (COCs) were obtained by ovum pick- up method from prepubertal and pubertal heifers. After in vitro maturation COCs were fertilized and cultured to the blastocyst stage. Total RNA was isolated from both groups of blastocysts and RNA-seq was performed. Gene ontology analysis was performed by DAVID (Database for Annotation, Visualization and Integrated Discovery).

Results: A higher average blastocyst rate was obtained in the pubertal than in the pre-pubertal group. There were no differences in the quality of blastocysts between the examined groups. We identified 436 differentially expressed genes (DEGs) between blastocysts derived from researched groups, of which 247 DEGs were downregulated in blastocysts derived from pubertal compared to prepubertal heifers oocytes, and 189 DEGs were upregulated. The genes involved in mitochondrial function, including oxidative phosphorylation (OXPHOS) were found to be different in studied groups using Kyoto Encyclopedia of Genes (KEGG) pathway analysis and 8 of those DEGs were upregulated and 1 was downregulated in blastocysts derived from pubertal compared to prepubertal heifers oocytes. DEGs associated with mitochondrial function were found: ATP synthases (ATP5MF-ATP synthase membrane subunit f, ATP5PD- ATP synthase peripheral stalk subunit d, ATP12A- ATPase H+/K + transporting non-gastric alpha2 subunit), NADH dehydrogenases (NDUFS3- NADH: ubiquinone oxidoreductase subunit core subunit S3, NDUFA13- NADH: ubiquinone oxidoreductase subunit A13, NDUFA3- NADH: ubiquinone oxidoreductase subunit A3), cytochrome c oxidase (COX17), cytochrome c somatic (CYCS) and ubiquinol cytochrome c reductase core protein 1 (UQCRC1). We found lower number of apoptotic cells in blastocysts derived from oocytes collected from prepubertal than those obtained from pubertal donors.

Conclusions: Despite decreased expression of genes associated with OXPHOS pathway in blastocysts from prepubertal heifers oocytes, the increased level of ATP12A together with the lower number of apoptotic cells in these blastocysts might support their survival after transfer.

Keywords: Blastocyst; Cow; Mitochondria; Next-generation sequencing; Prepubertal heifer.

PubMed Disclaimer

Conflict of interest statement

The authors declare no competing interests.

Figures

Fig. 1
Fig. 1
The Principal Component Analysis (PCA) clustering of samples representing two groups pre-pubertal (red) and pubertal (blue)
Fig. 2
Fig. 2
Volcano plot showing the significant up and down regulated genes across experiment designed. The DEGs belonged to oxidative phosphorylation pathway were red marked
Fig. 3
Fig. 3
WebGestalt Analysis of the different Gene Ontology terms. Bar chart showing the number of genes from the nex-generation sequencing that are involved in the different Gene Ontology terms as predicted by the Gene Set Enrichment Analysis (GSEA) via WebGestalt. The graph is showing the number of genes involved in the different biological processes (Red), Cellular components (Blue) and Molecular functions (Green)
Fig. 4
Fig. 4
KEGG pathway diagram for oxidative phosphorylation. The functional pattern of the gene expression between pre- pubertal and pubertal groups. Boxes with red borders represent up- regulated genes and boxes with the yellow border represent down-regulated genes when the pre- pubertal group was compared with the pubertal group
Fig. 5
Fig. 5
Real-time PCR validation in in vitro obtained bovine blastocysts (n = 4 × 5 for pre- pubertal and n = 4 × 5 for pubertal group) obtained from pre- pubertal and pubertal oocytes). Different letters above the column mean a significant differences between groups (p < 0.05), as determined by Student’s test. The data as presented as arbitrary units and are expressed as the mean ± SEM
Fig. 6
Fig. 6
(A) TUNEL was used to asses the level of apoptosis in blastocysts derived from oocytes obtained from pre-pubertal and pubertal heifers. The number of individual cells that were TUNEL positive was counted in each blastocyst and is represented as the average number of cells that are TUNEL positive per blastocyst. Embryos derived from oocytes collected from pre-pubertal heifers displayed significant decrease the average number of TUNEL positive cells compared with blastocysts derived from oocytes obtained from pubertal heifers; mean ± SEM, P < 0.05 (B) Representative images of embryos from TUNEL assay. Negative: No fluorescence control; Positive: Embryos pretreated with DNAse to ensure TUNEL staining was successful; 5 embryos in each group
See this image and copyright information in PMC

Similar articles

See all similar articles

References

    1. Salilew-Wondim D, Tesfaye D, Rings F, Held-Hoelker E, Miskel D, Sirard MA, et al. The global gene expression outline of the bovine blastocyst: reflector of environmental conditions and predictor of developmental capacity. BMC Genomics. 2021;22(1):408. - PMC - PubMed
    1. Armstrong DT, Kotaras PJ, Earl CR. Advances in production of embryos in vitro from juvenile and prepubertal oocytes from the calf and lamb. Reprod Fertil Dev. 1997;9(3):333–9. doi: 10.1071/R96080. - DOI - PubMed
    1. Paramio MT, Izquierdo D. Current status of in vitro embryo production in sheep and goats. Reprod Domest Anim październik. 2014;49(Suppl 4):37–48. doi: 10.1111/rda.12334. - DOI - PubMed
    1. Herst PM, Rowe MR, Carson GM, Berridge MV. Functional Mitochondria in Health and Disease. Front Endocrinol (Lausanne) 2017;8:296. doi: 10.3389/fendo.2017.00296. - DOI - PMC - PubMed
    1. Van Blerkom J, Davis PW, Lee J. ATP content of human oocytes and developmental potential and outcome after in-vitro fertilization and embryo transfer. Hum Reprod luty. 1995;10(2):415–24. doi: 10.1093/oxfordjournals.humrep.a135954. - DOI - PubMed