Dataset Information

Analysis of extracelullar vesicles (EVs) protein cargo across the bovine estrous cycle

ABSTRACT: The success of the early reproductive events depends on an appropriate communication between gametes/embryos and the oviduct. Extracellular vesicles (EVs) contained in oviductal secretions have been suggested as new players in mediating this crucial cross-talk by transferring their cargo (e.g. proteins, mRNA and small ncRNA) from cell to cell. Therefore, the objective of the project was to analyze the protein content of EVs (exosomes and microvesicles) contained in the bovine oviductal fluid at different stages of the estrus cycle. For this purpose, pairs of oviducts with their attached ovaries were collected from cyclic cows at a local abattoir under different hormonal regulation along the estrous cycle. Oviducts were classified into one of the 4 stages of the estrous cycle according to the morphology of the corpus lutea(CL) and follicle populations: Stage 1 (S1): showed recently ovulated follicle (d 1 to 4 of estrous cycle); Stage 2 (S2): early luteal development with medium or large or both present (d 5 to 11); Stage 3 (S3): fully functional CL yellow or orange in color (d 11 to 17); or Stage 4 (S4): regressing CL with little vasculature and a large preovulatory follicle present (d 18 to 20). EVs were collected by ultracentrifugation and prepared for mass spectrometry analysis.

OTHER RELATED OMICS DATASETS IN: GSE110444

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Bos Taurus (bovine)

TISSUE(S): Oviductal Fluid

SUBMITTER: Lucie Combes-Soia

LAB HEAD: Carmen Alminana Brines

PROVIDER: PXD008851 | Pride | 2018-07-31

REPOSITORIES: Pride

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Publications

Deciphering the oviductal extracellular vesicles content across the estrous cycle: implications for the gametes-oviduct interactions and the environment of the potential embryo.

Almiñana C C Tsikis G G Labas V V Uzbekov R R da Silveira J C JC Bauersachs S S Mermillod P P

BMC genomics 20180822 1

<h4>Background</h4>The success of early reproductive events depends on an appropriate communication between gametes/embryos and the oviduct. Extracellular vesicles (EVs) contained in oviductal secretions have been suggested as new players in mediating this crucial cross-talk by transferring their cargo (proteins, mRNA and small ncRNA) from cell to cell. However, little is known about the oviductal EVs (oEVS) composition and their implications in the reproductive success. The aim of the study was ...[more]

PMID: 30134841

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Project description:The success of the early reproductive events depends on an appropriate communication between gametes/embryos and the oviduct. Extracellular vesicles (EVs) contained in oviductal secretions have been suggested as new players in mediating this crucial cross-talk by transferring their cargo (e.g. proteins, mRNA and small ncRNA) from cell to cell. However, little is known about the oviduct EVs composition and their implications in reproductive success. The objective of our study was to determine the changes of oviductal EVs small ncRNA content under the hormonal influence during the estrous cycle. Methods: EVs, exosomes and microvesicles, were isolated from bovine oviductal fluid at different stages of the estrous cycle (postovulatory-stage, early luteal phase, late luteal phase and pre-ovulatory stage). RNA enriched for short molecules was isolated and used for the preparation of small RNA-seq libraries. RNA-sequencing was performed on an Illumina HiSeq 2500. The obtained sequence reads that passed quality filters were annotated by BLAST alignment to various sets of non-coding and coding RNAs. Read counts were calculated based on the frequency of unique sequences after removal of PCR duplicates. Statistical analysis was done with EdgeR to identify differential small ncRNA abundance across the different stages of the estrous cycle. Results: Small RNA-Seq identified the presence of different types of ncRNAs (miRNAs, rRNA fragments, tRNA fragments, snRNA, snoRNA, and other ncRNAs), which were partially also under hormonal influence. Major differences were found between post-ovulatory and early luteal and the rest of the stages for small ncRNAs. Interesting miRNAs identified in oviductal EVs, miR-34b and miR-449a, showed differential abundance among stages. Dysregulation of these miRNAs has been associated with defective cilia in the oviduct and infertility. Conclusions: Our findings represent the first extensive oviductal signature of bovine oviductal EVs miRNA content and contribute to a better understanding of the role of EVs as modulators of gamete/embryo-maternal interactions.

Project description:Background: Acute coronary syndromes (ACS) are associated with aberrant gene expression and epigenetic mechanisms. In particular, de novo DNA methylation is typically linked to gene silencing, but its role in heart disease remains not fully understood. Extracellular vesicles (EVs) are active components in cellular communication for their ability to carry a plethora of signalling biomolecules, thus representing a promising new diagnostic/therapeutic approach in cardiovascular diseases (CVDs). Indeed, there is the need of novel biomarkers for ACS prediction and timely detection. Purpose: We hypothesized that specific epigenetic signals can be carried by EVs. In this regard, we isolated and characterized circulating EVs from ACS patients and evaluated their potential role to influence DNA methylation in target cells. Methods: Circulating EVs were recovered, by ultracentrifugation, from plasma samples of 19 ACS patients and 50 healthy subjects (HS). Nanoparticle tracking analysis (NTA) and western blot (WB) were used to confirm the EVs integrity and purity. Peripheral blood mononuclear cells (PBMCs) of volunteer donors were treated with both ACS and HS derived EVs and genomic DNA was extracted to perform epigenome wide analysis through Reduced Representation Bisulfite Sequencing. ShinyGO, PANTHER, and STRING tools were interrogated to perform GO and PPI network analyses. Flow Cytometry, qRT-PCR, and WB analysis were also performed to evaluate and validate both intra-vesicular and intra-cellular signals. Results: Plasma ACS-derived EVs showed a significant up-regulation of DNA methyltransferases (DNMTs) gene expression levels as compared to HS (P<0.001). Specifically, de novo methylation transcripts, as DNMT3A and DNMT3B were significantly increased in plasma ACS-EVs. DNA methylation analysis of PBMCs from volunteer donors treated with HS- and ACS-derived EVs showed that RNF166 and CCND3 genes resulted the most hyper- and hypo-methylated, respectively, after by ACS-EV treatment. In addition, PPI network analysis specifically evidenced the subnetwork with SRC, as a hub gene, connecting it to NOTCH1, FOXO3, CDC42, IKBKG, RXRA, DGKG, known as important genes already involved in the onset of CVDs. Surprising, other novel genes, BAIAP2, SYP, CHL1, and SHB, which were hypomethylated, were found significantly overexpressed in PBMCs (P<0.005), underlying the fundamental modulating properties of EV cargo in atherosclerosis. Conclusions: These findings support the significant role of ACS plasma-derived EVs, inducing de novo DNA methylation signals, and modulating specific signaling pathways in target cells.

Dataset Information

Analysis of extracelullar vesicles (EVs) protein cargo across the bovine estrous cycle

Publications

Deciphering the oviductal extracellular vesicles content across the estrous cycle: implications for the gametes-oviduct interactions and the environment of the potential embryo.

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