Circulating miRNAs in maternal plasma as potential biomarkers of early pregnancy in sheep

Abstract

MicroRNA (miRNA) plays an important role in the control of gene expression and is implied in many biological functions, including embryo implantation and development. The aim was to assess plasma miRNA profiles during the peri-implantation and ascertain potential candidate miRNA markers for early pregnancy diagnosis in ovine plasma. The plasma samples were obtained from a total of 24 ewes on days 12 (pre-implantation; P12, n = 4), 16 (implantation; P16, n = 4) and 22 (post-implantation; P22, n = 4) after mating, and on their corresponding days of 12 (Pre-C; C12, n = 4), 16 (Imp-C; C16, n = 4) and 22 (Post-C; C22, n = 4) of the estrous cycle. The miRNA profiles in plasma were assessed by microarray technology. We detected the presence of 60 ovine-specific miRNAs in plasma samples. Of these miRNAs, 22 demonstrated a differential expression pattern, especially between the estrous cycle and early pregnancy, and targeted 521 genes. Two miRNAs (oar-miR-218a and oar-miR-1185-3p) were confirmed using RT-qPCR in the ovine plasma samples. Protein-protein interaction (PPI) network of target genes established six functional modules, of which modules 1 and 3 were enriched in the common GO terms, such as inflammatory response, defense response, and regulation of immune response. In contrast, module 2 was enriched in the developmental process involved in reproduction, embryo development, embryonic morphogenesis, and regulation of the developmental process. The results indicate that miRNAs profiles of plasma seemed to be modulated during the peri-implantation stage of pregnancy in ewes. Circulating miRNAs could be promising candidates for diagnosis in early ovine pregnancy.

Expression of circulating oar-miR-485-5p and oar-miR-493-5p during the estrous cycle and early pregnancy in ovine plasma

“…In recent years, numerous studies that uncovered many miRNAs have identified circulating plasma miRNAs ( Hitit et al, 2022a ). Villous trophoblast-derived miRNAs are transported through maternal blood in extracellular vesicles or connected to them ( Kotlabova et al, 2011 ; De los Santos Funes et al, 2023 ).…”

“…To thaw the plasma, we used a dry bath set to 20 degrees. Subsequently, 250 ul of plasma was processed through the miRCURY RNA Isolation Kit—Biofluids (Exiqon #300112 Vedbaek- Denmark) in compliant with the manufacturer's instructions to isolate total cell-free RNA ( Hitit et al, 2022a ). DNA contamination was removed from samples using on-column DNase according to manufacturer's instructions.…”

“…As a result, miRNAs were shown to be differently expressed during the peri-implantation phase ( Chakrabarty et al, 2007 ). Their expression patterns differ between implanted and non-implanted locations the extracellular vesicle, microRNAs that are either derived from the placenta or the embryo have the potential to enter the bodily fluids ( Hitit et al, 2022a ; Tan et al, 2020 ). Because of their sturdy structure, miRNAs are able to withstand damaging effects such as freezing and thawing as well as high temperatures; as a result, using them to identify early pregnancy is the most cutting-edge method currently available ( De los Santos Funes et al, 2023 ).…”

miRNA expression profiles of peripheral white blood cells from beef heifers with varying reproductive potential

“…Among the traits, phenotypes associated with reproductive outcome and fertility are of major interest to cattle producers, mainly because of the high production loss from a heifer of low reproductive potential. Specifically in reproductive biology, miRNAs have been demonstrated to be important regulators of embryonic development in humans and livestock, such as predicting pregnancy outcomes ( Ioannidis and Donadeu, 2016 ; Hitit et al, 2022 ), embryo viability ( Pohler et al, 2017 ) and implantation ( Kose et al, 2022 ), and endometrial receptivity ( Zang et al, 2021 ). Moreover, miRNAs regulate the gene targets by repressing or stimulating their expression ( Orang et al, 2014 ).…”

Infection by the lungwormRhabdias pseudosphaerocephalaaffects the expression of immune-related microRNAs by its co-evolved host, the cane toadRhinella marina

“…We deemed genes as putative targets of a particular miRNA only if predicted so by all three tools (Oliveira et al, 2017). To refine our results, we filtered out putative target genes that had an energy score >-15 kcal/mol (all three tools), and a sequence complementary score <150 (miRanda) (Hitit et al, 2022).…”