Abstract
BackgroundDespite its role in increasing the number of offspring during the lifetime of an individual animal, controlled ovarian hyperstimulation (COH) may have detrimental effects on oocyte development, embryo quality and endometrial receptivity. Circulating miRNAs in bio-fluids have been shown to be associated with various pathological conditions including cancers. Here we aimed to investigate the effect of COH on the level of extracellular miRNAs in bovine follicular fluid and blood plasma and elucidate their mode of circulation and potential molecular mechanisms to be affected in the reproductive tract.MethodTwelve simmental heifers were estrous synchronized and six of them were hyperstimulated using FSH. Follicular fluid samples from experimental animals were collected using ovum pick up technique at day 0 of the estrous cycle and blood samples were collected at day 0, 3 and 7 of post ovulation. The expression profile of circulatory miRNAs in follicular fluid and blood plasma were performed using the human miRCURY LNA™ Universal RT miRNA PCR array system. A comparative threshold cycle method was used to determine the relative abundance of the miRNAs.ResultsA total of 504 and 402 miRNAs were detected in both bovine follicular fluid and blood plasma, respectively. Of these 57 and 21 miRNAs were found to be differentially expressed in follicular fluid and blood plasma, respectively derived from hyperstimulated versus unstimulated heifers. Bioinformatics analysis of those circulating miRNAs indicated that their potential target genes are involved in several pathways including TGF-beta signaling pathway, MAPK signaling pathway, pathways in cancer and Oocyte meiosis.Moreover, detail analysis of the mode of circulation of some candidates showed that most of the miRNA were found to be detected in both exosomal and Ago2 protein complex fraction of both follicular fluid and blood plasma.ConclusionOur data provide the consequence of hyperstimulation induced changes of extracellular miRNAs in bovine follicular fluid and blood plasma, which may have a potential role in regulating genes associated not only with bovine ovarian function but also involved in altering various physiological in bovine oocytes, embryos and modulating reproductive tract environment.
Highlights
Despite its role in increasing the number of offspring during the lifetime of an individual animal, controlled ovarian hyperstimulation (COH) may have detrimental effects on oocyte development, embryo quality and endometrial receptivity
Our data provide the consequence of hyperstimulation induced changes of extracellular miRNAs in bovine follicular fluid and blood plasma, which may have a potential role in regulating genes associated with bovine ovarian function and involved in altering various physiological in bovine oocytes, embryos and modulating reproductive tract environment
Detection of miRNAs in bovine follicular fluid and blood plasma Here, we examined the abundance of mature miRNAs in follicular fluid and blood plasma of hyperstimulated and unstimulated groups using ready-to-use Human miRNA Panels (I + II) (Exiqon, Denmark) covering 748 mature human miRNAs
Summary
Despite its role in increasing the number of offspring during the lifetime of an individual animal, controlled ovarian hyperstimulation (COH) may have detrimental effects on oocyte development, embryo quality and endometrial receptivity. A large number of gene products are involved in this process, which are expressed in cell type and time dependent manner Any deviation in the expression of these genes due to environmental factors could lead to abnormal oocyte development, apoptosis, and poor cellular communication with the companion somatic cells [11]. In this context the role of miRNAs as post transcriptional regulaters of gene expression can not be ruled out as these regulatory molecules are abundantly expressed in follicular cells and in extracellular space in follicular microenvironment. The function of exosomes and shedding vesicles are believed to be cell-to-cell communication and platforms for multi signalling processes [23, 24]
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