Implications of miRNA expression pattern in bovine oocytes and follicular fluids for developmental competence

Abstract

24Developmental competence determines the oocyte capacity to support initial embryo 25 growth, but the molecular mechanisms underlying this phenomenon are still ill-26 defined. Changes in microRNA (miRNA) expression pattern have been described 27 during follicular growth in several species. Therefore, aim of this study was to 28 investigate whether miRNA expression pattern in cow oocyte and follicular fluid (FF) 29 is associated with the acquisition of developmental competence. Samples were 30 collected from ovaries with more than, or fewer than, 10 mid-antral follicles (H-and 31 L-ovaries) because previous studies demonstrated that this parameter is a reliable 32 predictor of oocyte competence. After miRNA deep sequencing and bioinformatic 33 data analysis, we identified 58 miRNAs in FF and 6 in the oocyte that were 34 differentially expressed between H-and L-ovaries. Overall, our results indicate that 35 miRNA levels both in FF and in the ooplasm must remain within specific thresholds 36 and that changes in either direction compromise oocyte competence. Some of the 37 miRNAs found in FF (miR-769, miR-1343, miR-450a, miR-204, miR-1271 and miR-38 451) where already known to regulate follicle growth and their expression pattern 39 indicate that they are also involved in the acquisition of developmental competence. 40Some miRNAs were differentially expressed in both compartments but with opposite 41 patterns, suggesting that miRNAs do not flow freely between FF and oocyte. Gene 42Ontology analysis showed that the predicted gene targets of most differentially 43 expressed miRNAs are part of a few signalling pathways. Regulation of maternal 44 mRNA storage and mitochondrial activity seem to be the processes more 45 functionally relevant in determining oocyte quality. In conclusion, our data identified a 46 few miRNAs in the follicular fluid and in the ooplasm that modulate the oocyte 47Recently, microRNAs (miRNAs), which regulate gene expression at the mRNA level, 65 have been associated with folliculogenesis and oogenesis [10, 11]. MiRNAs, which 66 range in size from 18 to 25 nucleotides (nt), have been found in the different 67 compartments of ovarian follicles, including granulosa cells [12, 13], theca cells [14], 68 follicular fluid and the oocyte itself [15]. Studies on the role of miRNAs during follicle 69 development in humans [16][17][18], mice [19, 20], cattle [10, 21, 22], pigs [23] and 70horses [24] suggest that they regulate the cellular differentiation processes which 71 occur during follicular development. 72 Follicular fluid and germinal vesicle oocyte collection 111Ovaries were collected at a commercial abattoir and were transported to the 112 laboratory in warmed (27-30°C) Dulbecco Phosphate Buffered Saline (PBS). Ovaries 113 were classified into low and high antral follicle count categories according to the 114 methods used in previous works [28, 29]. Briefly, the ovaries were assigned to high 115 antral follicle count ovaries (H ovaries) when more than 10 mid-antral follicles (2-5 116 mm in dia...

Inhibition of miR-152 During in Vitro Maturation Enhances the Developmental Potential of Porcine Embryos

“…We mainly found that a suppressed expression of miR-152 during oocyte maturation signi cantly improved the blastocyst rate. Recently, it has been reported that the miRNAs expression pattern is highly correlated with oocyte quality and developmental competence [25]. Moreover, stage-speci c and dynamic changes in the expression pattern of several miRNAs were observed during oocyte [26][27][28] and embryo development [29,30], indicating a regulatory role of miRNAs in oogenesis and embryogenesis.…”

A Nuclear and Cytoplasmic Characterization of Bovine Oocytes Reveals That Cysteamine Partially Rescues the Embryo Development in a Model of Low Ovarian Reserve

“…The relative fluorescence of MTG and MTO was analyzed using NIH ImageJ 1.53e software [69] and expressed as mean grey values after background subtraction. The oocyte MTO/MTG ratio was calculated as an indicator of the MMP independent of the mitochondrial mass as proposed by [76,78]. Oocytes having an undetectable fluorescent signal were excluded from the quantitative image analyses.…”

“…Conversely, the mitochondrial activity was higher in GV oocytes compared to MII (Figure 6B), indicating a reduced mitochondrial membrane potential in MII oocytes, which is more evident when normalizing the MTO signal by the MTG signal (Figure 6C). Indeed, such analysis has been proposed as a relative measure of the mitochondrial activity that is independent of the mitochondrial mass [76,78].…”

Oocyte Selection for <em>in vitro</em> embryo Production in Bovine Species: Non-Invasive Approaches for the New Challenges of Oocyte Competence

“…The bovine FF contains free miRNAs as well as some associated with exosomes [171,172]. Recently, the study of Pasquariello et al, [173] showed by first time, the miRNA content of different populations of oocytes categorized according to their competence. Interestingly, they discovered that the most differentially expressed miRNAs (miR-24, miR-10a and miR-320a) in FF found in high competent follicles were part of the regulation of neurotrophin signaling pathway, which supports follicle formation and development, as well as the TGF-beta signaling pathways that controls the production of ovarian peptide hormones.…”