Expression of microRNAs in bovine and human pre-implantation embryo culture media

Abstract

MicroRNAs (miRNA) are short non-coding RNAs which act to regulate expression of genes driving numerous cellular processes. These RNAs are secreted within exosomes from cells into the extracellular environment where they may act as signaling molecules. In addition, they are relatively stable and are specifically expressed in association to certain cancers making them strong candidates as biological markers. Moreover, miRNAs have been detected in body fluids including urine, milk, saliva, semen, and blood plasma. However, it is unknown whether they are secreted by embryonic cells into the culture media. Given that miRNAs are expressed throughout embryonic cellular divisions and embryonic genome activation, we hypothesized that they are secreted from the embryo into the extracellular environment and may play a role in the developmental competence of bovine embryos. To test this hypothesis, bovine embryos were cultured individually from day 5 to day 8 of development in an in vitro fertilization system and gene expression of 5 miRNAs was analyzed in both embryos and culture media. Differential miRNA gene expression was observed between embryos that developed to the blastocyst stage and those that failed to develop from the morula to blastocyst stage, deemed degenerate embryos. MiR-25, miR-302c, miR-196a2, and miR-181a expression was found to be higher in degenerate embryos compared to blastocyst embryos. Interestingly, these miRNAs were also found to be expressed in the culture media of both bovine and human pre-implantation embryos. Overall, our results show for the first time that miRNAs are secreted from pre-implantation embryos into culture media and that miRNA expression may correlate with developmental competence of the embryo. Expression of miRNAs in in vitro culture media could allow for the development of biological markers for selection of better quality embryos and for subsequent successful pregnancy.

Increasing of blastocyst rate and gene expression in co-culture of bovine embryos with adult adipose tissue-derived mesenchymal stem cells

“…MSC-derived exosomal microvesicles have been recently documented and may provide a potential explanation for the micromodulation potential observed with stem cells. Moreover, it was recently shown that human and cattle preimplantation embryos also secreted exosomes throughout the in vitro culture period [31,32]. Therefore, one may also speculate a potential reciprocal effect between b-ATMSCs and embryos in culture that may explain the observed effects.…”

Literature review of baseline information to support the risk assessment of RNAi‐based GM plants

“…The same year, placental miRNAs were found circulating in maternal plasma , which was one of the discoveries leading to the notion that miRNAs could be a mobile regulating molecule (Iguchi et al, 2010) and that could even mediate transgenerational epigenetic heritance (Sharma, 2015) or be transmitted across species Buck et al, 2014). Since then, extracellular miRNAs were identified in a broad range of biological fluids, including plasma, aqueous humour, cerebrospinal fluid, nasal mucus, or milk (Pegtel et al, 2011;Huang et al, 2013b;Kropp et al, 2014;Baglio et al, 2015;Dismuke et al, 2015;Wu et al, 2015a). miRNAs were identified in the cargo of exosomes, membranous vesicles 40 to 100 nm in diameter, which are constitutively released by almost all cell type and are found essentially in every biological fluid (reviewed, for example, in (Rak, 2013;Yoon et al, 2014) However, extracellular miRNAs do not need to be necessarily encapsulated in extracellular vesicles, as two studies showed that 95-99% of extracellular miRNA are not in extracellular vesicles but associated with AGO proteins in serum and cell culture media (Arroyo et al, 2011;Turchinovich et al, 2011).…”

Human Embryos Created by Embryo Splitting Secrete Significantly Lower Levels of miRNA-30c

“…Aberrant miRNA profiles in transferable blastocysts have been linked with male factor infertility and polycystic ovaries [5]. Since miRNAs are known to be secreted [18], attempts have been made to link specific miRNAs detected in the SBM with embryo ploidy status and reproductive competence [2][3][4][5]. miRNAs detected in the spent medium from early embryos might be the output of paracrine circuits, reflecting normal communication among cells of healthy embryos; alternatively, they might be a component of the communication pathway between the embryo and the uterine epithelium, influencing epithelial readiness for embryo apposition and implantation.…”

“…For example, miR-30b is involved in development of embryonic ectoderm [22], miR-373 promotes mesendoderm differentiation [23], and miR-24 is required for hematopoietic differentiation [24]. However, only miR-24 [25,26] and miR-25 [3] were detected in preimplantation embryo culture media.…”

“…They are secreted and can be found in embryo culture media. Attempts have been made to link specific miRNAs detected in the medium with embryo ploidy status and reproductive competence to predict in vitro fertilization (IVF) outcome, and several candidates have been identified [2][3][4][5]. Although further studies are warranted, a comprehensive profile of miRNAs secreted by human preimplantation embryos in spent culture media is taking shape [6].…”