Endocrine Abstracts

JOINT183

Objectives: Adipokines regulate reproduction in dependence of metabolic conditions. Visfatin, upregulated in obesity, improves oocyte maturation in obese and old individuals. Our previous studies showed that lack of visfatin leads to decrease in blastocyst number in obese mice. The aims of this study were to determine the effect of visfatin on the blastocyst transcriptome in normal weight and high-fat-diet induced obese mice, as well as blastocyst implantation rate and ovarian function of offspring.

Methods: We knockdowned the mRNA expression of Nampt at the 1 cell stage embryo (siRNA electroporation) and cultured it until the blastocyst stage, then we isolated RNA and evaluated blastocyst transcriptome (RNASeq, padj<0.05), DEGs ontology and pathways were analyzed in qProfiler and DAVID software. As a next step the blastocyst implantation rate/offspring number were calculated and offsprings (17.5 dpi) ovaries were collected to measure steroidogenesis (Star, Cyp11a1, Cyp17a1, Hsd3b, Cyp19a1), folliculogenesis/oogenesis (Gdf-9, Bmp-15, Figla, Dnmt1) and proliferation/apoptosis (Pcna, Bax, Bcl2) related genes level (real-time PCR, n = 5). Statistical analysis was performed in GraphPad Prism software.

Results: Transcriptomic analysis showed 73 upregulated and 24 downregulated genes in the blastocyst from normal weight mice after Nampt silencing. Ontology and pathways analysis classified these genes to biological processes (cell death, glycolytic process, metabolic process, anatomical structure development) and cellular components (cytoplasm, cytoskeleton, cell junction). According to KEGG we noted estrogen signaling, p53 and progesterone mediated oocyte maturation pathways. In obese mice group only 2 genes were upregulated (Gm10052, Tspan7) and 1 (Tmem²54c) downregulated. Visfatin silencing does not affect the implantation rate and offspring number in normal weight mice. However, we observed the changes in the expression of marker genes of proper ovarian function in offspring including stimulation in Bmp-15 and Cyp17a1 and inhibition in Hsd3b transcripts.

Conclusion: Visfatin is an important factor in early embryogenesis, and the effect of its absence in early embryonic life is long-lasting and affects the fertility of female offspring. It also indicates that visfatin may play a compensatory role in reproduction during obesity.

Keywords: visfatin, early embryogenesis, siRNA induced gene knockdown, transcriptome.