Changes in circulating microRNA levels can be identified as early as day 8 of pregnancy in cattle

Abstract

Poor reproductive performance remains a major issue in the dairy industry, with low conception rates having a significant impact on milk production through extended calving intervals. A major limiting factor is the lack of reliable methods for early pregnancy diagnosis. Identification of animals within a herd that fail to conceive within 3 weeks after insemination would allow early re-insemination and shorten calving intervals. In a previous study, we found an increase in plasma miR-26a levels in Day 16-pregnant relative to non-pregnant heifers, however changes in miRNA levels that early during pregnancy were very small which likely prevented the identification of robust biomarkers. In this study, we extended our analyses to a wider interval during pregnancy (Days 8 to 60, n = 11 heifers) with the rationale that this may facilitate the identification of additional early pregnancy miRNA biomarkers. Using small RNA sequencing we identified a total of 77 miRNAs that were differentially expressed on Day 60 relative to Day 0 of pregnancy. We selected 14 miRNAs for validation by RT-qPCR and confirmed significant differences in the expression of let-7f, let-7c, miR-30c, miR-101, miR-26a, miR-205 and miR-143 between Days 0 and 60. RT-qPCR profiling throughout Days 0, 8, 16 and 60 of pregnancy showed a distinct increase in circulating levels of miR-26a (3.1-fold, P = 0.046) as early as Day 8 of pregnancy. In summary, in contrast to earlier stages of pregnancy (≤ Day 24), marked differences in the levels of multiple miRNAs can be detected in circulation by Day 60 in cattle. Retrospective analyses showed miR-26a levels to be increased in circulation as early as Day 8, sooner than previously reported in any species, suggesting a biological role for this miRNA in the very early events of pregnancy.

The Dynamics of microRNA Transcriptome in Bovine Corpus Luteum during Its Formation, Function, and Regression

“…Although significant regulation could be confirmed for eight out of the ten selected miRNAs, the qPCR data show lower fold-change regulation than the fold-change regulation assessed by NGS. This is in agreement with results by different studies profiling miRNAs in tissue and plasma (Ioannidis and Donadeu, 2016 , 2017 ).…”

MiRNAs in the Peri-Implantation Period: Contribution to Embryo–Maternal Communication in Pigs

“…For example, miR-26a levels were higher in 3-week pregnant heifers compared with sham-inseminated counterparts [90]. Broadly conserved miRNAs (e.g., let-7c, miR-143) were found to be signatures of pregnancy in cows; however, elevated levels of these miRNAs were observed during late pregnancy [91,92]. Interestingly, circulating EV-derived miRNAs (e.g., miR-125b-2, let-7c) were associated with increased embryonic mortality on day 17 of pregnancy in cows [93].…”

Understanding the uterine environment in early pregnancy in cattle: How have the omics en-hanced our knowledge?

“…In the wider context of reproductive 'omics' an interesting recent development is the identification that miRNA molecules in circulation in bovine plasma can be used to identify pregnancy as early as day 8 (Ioannidis and Donadeu, 2017). Utilising such RNA sequencing capabilities for elucidating biomarkers for identifying potentially subfertile pathophysiologies in cattle, as in the human (Sathyapalan et al, 2015), will be an important development for both selectively mating, inseminating, or embryo transferring into heifers, in addition to enhancing our holistic understanding of fundamental reproductive events (Fig.…”