Extracellular vesicle-coupled miRNA profiles in follicular fluid of cows with divergent post-calving metabolic status

Tsige Hailay,Michael Hoelker,Mikhael Poirier,Karl Schellander,Mohammed Saeed-Zidane,Franca Rings,Samuel Gebremedhn,Christiane Neuhoff,Dawit Tesfaye,Dessie Salilew-Wondim,Christina Dauben,Ernst Tholen

doi:10.1038/s41598-019-49029-9

Abstract

Most high-yielding dairy cows enter a state of negative energy balance (NEB) during early lactation. This, in turn, results in changes in the level of various metabolites in the blood and follicular fluid microenvironment which contributes to disturbed fertility. Extracellular vesicles (EVs) are evolutionarily conserved communicasomes that transport cargo of miRNA, proteins and lipids. EV-coupled miRNAs have been reported in follicular fluid. However, the association between postpartum NEB and EV-coupled miRNA signatures in follicular fluid is not yet known. Energy balance analysis in lactating cows shortly after post-calving revealed that the majority of the cows exhibited transiently negative energy balance levels, whereas the remaining cows exhibited either consistently negative or consistently positive energy levels. Metabolic status was associated with EV-coupled miRNA composition in the follicular fluid. Cows experiencing NEB showed reduced expression of a large number of miRNAs while cows with positive energy balances primarily exhibited elevated expression of EV-coupled miRNAs. The miRNAs that were suppressed under NEB were found to be involved in various metabolic pathways. This is the first study to reveal the presence of an association between EV-coupled miRNA in follicular fluid and metabolic stress in dairy cows. The involvement of differentially expressed miRNAs in various pathways associated with follicular growth and oocyte maturation suggest the potential involvement of specific follicular miRNAs in oocyte developmental competence, which may partially explain reduced fertility in cows due to post-calving metabolic stress.

Highlights

Selection for milk yield in dairy cows in the last 70 years has resulted in a significant increase in milk yield and concomitant reduction in fertility
Cows were considered to be in negative energy balance when non-esterified fatty acids (NEFA) and ß-OHB concentrations in serum were >0.55 mmol/L and >0.65 mmol/L, respectively, as applied previously[30,31]
Our results revealed that out of 38 differentially expressed miRNAs, 31 miRNAs were upregulated and only seven miRNAs were downregulated in Always positive cows (APCs) or lactating cows compared to heifers (Table 2)

Summary

Introduction

Selection for milk yield in dairy cows in the last 70 years has resulted in a significant increase in milk yield and concomitant reduction in fertility. During NEB, cows have a shortage of circulating glucose and their body starts to metabolize fat reserves through lipogenesis and/ketogenesis This leads to an increase of undesired substances in the blood such as non-esterified fatty acids (NEFA) and beta-hydroxybutyrate, which are known to have a negative effect on folliculogenesis[5,6]. Biochemical reactions like brilliant cresyl blue staining[13] and morphological parameters are utilized to select oocytes invasively Molecules such as DNA, mRNA, miRNAs, lipids, and proteins that are released into follicular fluid from surrounding follicular cells via extracellular vesicles during the cell-to-cell communication could serve as non-invasive molecular markers for oocyte competence. We aimed to investigate the effect of lactation physiology on the expression of EV-coupled miRNAs in follicular fluid by comparing metabolically non-stressed lactating cows versus heifers

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Methods

Results

Discussion

Conclusion