Although our knowledge regarding oocyte quality and development has improved significantly, the molecular mechanisms that regulate and determine oocyte developmental competence are still unclear. Therefore, the objective of this study was to identify and analyze the transcriptome profiles of porcine oocytes derived from large or small follicles using RNA high-throughput sequencing technology. RNA libraries were constructed from oocytes of large (LO; 3-6 mm) or small (SO; 1.5-1.9 mm) ovarian follicles and then sequenced in an Illumina HiSeq4000. Transcriptome analysis showed a total of 14,557 genes were commonly detected in both oocyte groups. Genes related to the cell cycle, oocyte meiosis, and quality were among the top highly expressed genes in both groups. Differential expression analysis revealed 60 up- and 262 downregulated genes in the LO compared with the SO group. BRCA2, GPLD1, ZP3, ND3, and ND4L were among the highly abundant and highly significant differentially expressed genes (DEGs). The ontological classification of DEGs indicated that protein processing in endoplasmic reticulum was the top enriched pathway. In addition, biological processes related to cell growth and signaling, gene expression regulations, cytoskeleton, and extracellular matrix organization were among the highly enriched processes. In conclusion, this study provides new insights into the global transcriptome changes and the abundance of specific transcripts in porcine oocytes in correlation with follicle size.

• MeSH
• genové regulační sítě fyziologie   MeSH
• oocyty metabolismus   MeSH
• oogeneze genetika   MeSH
• ovariální folikul cytologie   MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• prasata genetika   růst a vývoj   MeSH
• signální transdukce genetika   MeSH
• stanovení celkové genové exprese MeSH
• transkriptom * MeSH
• vysoce účinné nukleotidové sekvenování MeSH
• vývojová regulace genové exprese fyziologie   MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Endocrine Disrupting Chemicals (EDCs) are molecules able to interfere with the vertebrate hormonal system in different ways, a major one being the modification of the activity of nuclear receptors (NRs). Several NRs are expressed in the vertebrate brain during embryonic development and these NRs are suspected to be responsible for the neurodevelopmental defects induced by exposure to EDCs in fishes or amphibians and to participate in several neurodevelopmental disorders observed in humans. Known EDCs exert toxicity not only on vertebrate forms of marine life but also on marine invertebrates. However, because hormonal systems of invertebrates are poorly understood, it is not clear whether the teratogenic effects of known EDCs are because of endocrine disruption. The most conserved actors of endocrine systems are the NRs which are present in all metazoan genomes but their functions in invertebrate organisms are still insufficiently characterized. EDCs like bisphenol A have recently been shown to affect neurodevelopment in marine invertebrate chordates called ascidians. Because such phenotypes can be mediated by NRs expressed in the ascidian embryo, we review all the information available about NRs expression during ascidian embryogenesis and discuss their possible involvement in the neurodevelopmental phenotypes induced by EDCs.

• MeSH
• biologické modely MeSH
• embryo nesavčí účinky léků   MeSH
• embryonální vývoj účinky léků   MeSH
• endokrinní disruptory toxicita   MeSH
• nervový systém * účinky léků   embryologie   růst a vývoj   MeSH
• neurotoxiny toxicita   MeSH
• receptory cytoplazmatické a nukleární metabolismus   MeSH
• Urochordata * účinky léků   embryologie   růst a vývoj   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

Oocyte developmental competence is acquired during folliculogenesis and regulated by complex molecular mechanisms. Several molecules are involved in these mechanisms, including microRNAs (miRNAs) that are essential for oocyte-specific processes throughout the development. The objective of this study was to identify the expression profile of miRNAs in porcine oocytes derived from follicles of different sizes using RNA deep sequencing. Oocytes were aspirated from large (LO; 3-6 mm) or small (SO; 1.5-1.9 mm) follicles and tested for developmental competence and chromatin configurations. Small RNA libraries were constructed from both groups and then sequenced in an Illumina NextSeq. 500. Oocytes from the LO group exhibited higher developmental competence and different chromatin configuration compared with oocytes from the SO group. In total, 167 and 162 known miRNAs were detected in the LO and SO groups, respectively. MiR-205, miR-16, miR-148a-3p, and miR-125b were among the top 10 highly expressed miRNAs in both groups. Eight miRNAs were differentially expressed (DE) between both groups. Target gene prediction and pathway analysis revealed 46 pathways that were enriched with miRNA-target genes. The oocyte meiosis pathway and signaling pathways including FoxO, PI3K-Akt, and cAMP were predictably targeted by DE miRNAs. These results give more insights into the potential role of miRNAs in regulating the oocyte development.

• MeSH
• chromatin genetika   metabolismus   MeSH
• mikro RNA biosyntéza   MeSH
• oocyty cytologie   metabolismus   MeSH
• oogeneze fyziologie   MeSH
• prasata MeSH
• sekvenční analýza RNA * MeSH
• vysoce účinné nukleotidové sekvenování * MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The purpose of the study was to analyze seminal plasma composition, sperm production, and sperm motility over the course of the spawning season in the common carp (Cyprinus carpio) and rainbow trout (Oncorhynchus mykiss). The highest mean percent of motile sperm (carp, 98.3 ± 1.6%; rainbow trout, 92.8 ± 5.6%) and highest spermatozoon velocity (carp, 286.3 ± 7.8 μm sec-1; rainbow trout, 245.3 ± 8.3 μm sec-1) were observed in the middle phase of the spawning period, at 5 sec post-activation. Sperm volume and concentration increased in the early spawning period, then decreased significantly toward the end of the season in both species. Seminal plasma osmolality was 262-270 mOsmol kg-1from carp and 232-248 mOsmol kg-1from rainbow trout, with little variation over the spawning period. Protein concentration in carp seminal plasma was stable throughout the reproductive season, whereas the highest protein level (2.1 ± 0.3 mg mL-1) in rainbow trout was observed in the middle phase of the spawning period. Seminal plasma composition in both species was analyzed using two-dimensional polyacrylamide gel electrophoresis combined with mass spectrometry. Thirteen differentially expressed protein spots in carp seminal plasma and sixteen spots in rainbow trout seminal plasma varied over the course of the reproductive season. The unique proteins identified are involved in the regulation of spermatogenesis, sperm motility, maintenance of sperm membrane lipid stability, and antioxidant protection. These results provide a deeper understanding of the role of fish seminal plasma proteins as well as a list of novel markers of sperm quality. Mol. Reprod. Dev. 83: 968-982, 2016 © 2016 Wiley Periodicals, Inc.

• MeSH
• kapři metabolismus   MeSH
• Oncorhynchus mykiss metabolismus   MeSH
• proteiny semenné plazmy metabolismus   MeSH
• rybí proteiny metabolismus   MeSH
• sperma metabolismus   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

The effect of reactive oxygen species production on the motility parameters, DNA integrity, acid phosphatase activity, and protein tyrosine phosphorylation in spermatozoa of the common carp (Cyprinus carpio) was investigated. Spermatozoa were exposed to different concentrations of xanthine and xanthine oxidase (X-XO) either in the presence or absence of antioxidants for 15 and 60 min. A dose- and time-dependent reduction in spermatozoa motility and velocity was observed. Comet assays showed a dramatic increase in DNA fragmentation after 15 min. Changes in tyrosine phosphorylation of spermatozoa proteins were observed by Western blotting with anti-phosphotyrosine antibodies, and proteins of interest were identified by mass spectrometry. After a 60 min exposure to X-XO, O-linked N-acetylglucosamine transferase, isoform 4 was phosphorylated and septin-8-A was dephosphorylated. Acid phosphatase activity also decreased in a dose-dependent manner after a 60 min exposure to oxidative stress. The results demonstrate that oxidative stress impaired functional variables (sperm motility, velocity, DNA integrity) of carp spermatozoa, and altered intracellular signalling pathways through changes in tyrosine phosphorylation and acid phosphatase activity.

• MeSH
• fosforylace MeSH
• fragmentace DNA * MeSH
• kapři metabolismus   MeSH
• kyselá fosfatasa metabolismus   MeSH
• motilita spermií * MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• rybí proteiny metabolismus   MeSH
• spermie metabolismus   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

This study evaluated physiological and functional sperm parameters and the seminal plasma proteome of Eurasian perch (Perca fluviatilis) over the course of their reproductive season. Spermatozoa velocity (169.56 ± 6.53 to 158.5 ± 7.4 µm sec(-1)), percent motility (95.89 ± 4.28% to 89.55 ± 4.5%), and osmolality of seminal plasma (290 ± 5 to 297 ± 12 mOsmol kg(-1)) remained stable throughout the reproductive season. Milt volume and protein concentration of seminal plasma gradually increased and reached the highest values late in the reproductive period. Spermatozoa concentration peaked in the mid-reproductive season (66.90 ± 13 × 10(9)  spermatozoa ml(-1)) and decreased towards the end (54 ± 10 × 10(9)  spermatozoa ml(-1)). A proteomic analysis of seminal plasma using two-dimensional polyacrylamide gel electrophoresis revealed 10 protein spots significantly altered over the course of the reproductive season. Subsequent protein characterization suggested that time in the reproductive season predominantly affected proteins involved in membrane trafficking, organization, cell motility, and oxido-reductase activity. This study provides new data on physiological properties of sperm and protein patterns of seminal plasma over the course of the reproductive season that should be considered in the development of methods for artificial reproduction of perch.

• MeSH
• 2D gelová elektroforéza MeSH
• genetická variace MeSH
• motilita spermií * genetika   MeSH
• okounovití MeSH
• oxidoreduktasy metabolismus   MeSH
• proteiny semenné plazmy genetika   MeSH
• proteom analýza   MeSH
• roční období MeSH
• sperma fyziologie   MeSH
• spermie fyziologie   MeSH
• transport proteinů MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Global transcription silencing occurs in the oocyte during its final phase of growth. The particular mechanism of this silencing is not well understood. Here, we investigated the silencing of RNA polymerase II transcription in porcine oocytes. First, we investigated the transcriptional activity of germinal vesicle oocytes derived from stimulated and non-stimulated gilts, but no transcriptional activity was observed. Second, we focused on the fate of RNA polymerase II in growing and fully grown oocytes. Active and inactive forms of RNA polymerase II were detected in growing oocytes by immunofluorescence and Western blots. In contrast, only the inactive form of RNA polymerase II was detected in fully grown oocytes. To evaluate if the inactive form of RNA polymerase II is released from DNA, the oocytes were subsequently permeabilized and fixed in one step. After this modified fixation protocol, the immunofluorescent labeling was negative in fully grown oocytes, but remained unchanged (positive) in growing oocytes. These results indicate that the inactive form of RNA polymerase II is not bound to DNA during the oocyte growth. Finally, based on Western blot analysis of different stages of oocyte maturation, the inactive form of RNA polymerase II was detected in metaphase I but not in metaphase II. Our study confirmed the global transcription silencing of fully grown oocytes. Compared with other mammalian species (e.g., mouse), the mechanism of RNA polymerase II silencing in porcine oocytes seems to be similar, despite some differences in dynamics.

• MeSH
• adenosin chemie   metabolismus   MeSH
• autoradiografie MeSH
• fosforylace MeSH
• genetická transkripce MeSH
• gonadotropiny metabolismus   MeSH
• imunohistochemie MeSH
• izotopové značení MeSH
• myši MeSH
• oocyty chemie   růst a vývoj   metabolismus   fyziologie   MeSH
• prasata MeSH
• RNA-polymerasa II chemie   genetika   metabolismus   MeSH
• umlčování genů * MeSH
• uridin chemie   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Several lines of evidence suggest that in mice the activation of SMAD2/3 signaling by oocyte secreted factors, together with epidermal growth factor receptor (EGFR) activation, is essential to induce cumulus expansion. Here we show that inhibition of EGFR kinase in follicle stimulating hormone (FSH)-stimulated porcine oocyte-cumulus cell complex (OCCs) strongly decreases hyaluronan (HA) synthesis and its retention in the matrix, as well as progesterone synthesis. Although porcine cumulus cells undergo expansion independently of oocytes, we use biochemical and gene expression analyses to show that they do require activation of SMAD2/3 for optimal stimulation of HA synthesis and proteins involved in the organization of this polymer in the expanded matrix. Furthermore, FSH-induced progesterone synthesis by porcine cumulus cells was increased by blocking SMAD2/3 activation. In conclusion, these results support the hypothesis that an FSH-EGF autocrine loop is active in porcine OCCs, and provide the first evidence that the SMAD2/3 signaling pathway is induced by paracrine/autocrine factors in porcine cumulus cells and is involved in the control of both cumulus expansion and steroidogenesis.

• MeSH
• benzamidy farmakologie   MeSH
• C-reaktivní protein metabolismus   MeSH
• chinazoliny farmakologie   MeSH
• dioxoly farmakologie   MeSH
• epidermální růstový faktor metabolismus   MeSH
• erbB receptory antagonisté a inhibitory   metabolismus   MeSH
• folikuly stimulující hormon metabolismus   MeSH
• glukuronosyltransferasa antagonisté a inhibitory   metabolismus   MeSH
• isochinoliny farmakologie   MeSH
• kumulární buňky metabolismus   MeSH
• kyselina hyaluronová biosyntéza   MeSH
• meióza účinky léků   MeSH
• molekuly buněčné adheze antagonisté a inhibitory   metabolismus   MeSH
• myši MeSH
• oocyty enzymologie   fyziologie   MeSH
• prasata MeSH
• progesteron biosyntéza   MeSH
• protein Smad2 antagonisté a inhibitory   metabolismus   MeSH
• protein Smad3 antagonisté a inhibitory   metabolismus   MeSH
• pyridiny farmakologie   MeSH
• pyrroly farmakologie   MeSH
• sérový amyloidový protein metabolismus   MeSH
• signální transdukce účinky léků   MeSH
• tyrphostiny farmakologie   MeSH
• vývojová regulace genové exprese účinky léků   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

In our study, we have examined the pattern of global histone modification changes in somatic cell nuclei after their transfer into mouse oocytes at different stages of maturation or after their parthenogenetic activation. While germinal vesicle (GV) staged immature oocytes are strongly labeled with anti-acetylated histone H3 and H4 antibodies, the signal is absent in both metaphase I and metaphase II oocytes (MI, MII). In contrast, the oocytes of all maturation stages show a presence of trimethylated H3/K4 in their chromatin. When somatic cells were fused to intact or enucleated GV oocytes, both the GV and the somatic cell nucleus showed a very strong signal for all the antibodies used. On the other hand, when somatic cells nuclei that are AcH3 and AcH4 positive before fusion are introduced into either intact or enucleated MI or MII oocytes, their acetylation signal decreased rapidly and was totally absent after a prolonged culture. This was not the case when anti-trimethyl H3/K4 antibody was used. The somatic cell chromatin showed only a slight decrease in the intensity of labeling after its transfer into MI or MII oocytes. This decrease was, however, evident only after a prolonged culture. These results suggest not only a relatively higher stability of the methylation modification but also some difference between the oocyte and somatic chromatin. The ability to deacetylate the chromatin of transferred somatic nuclei disappears rapidly after the oocyte activation. Our results indicate that at least some reprogramming activity appears in the oocyte cytoplasm almost immediately after GV breakdown (GVBD), and that this activity rapidly disappears after the oocyte activation. Copyright 2007 Wiley-Liss, Inc.

• MeSH
• fertilizace in vitro MeSH
• financování organizované MeSH
• histonacetyltransferasy metabolismus   MeSH
• histonlysin-N-methyltransferasa metabolismus   MeSH
• histony metabolismus   MeSH
• hybridní buňky metabolismus   MeSH
• metylace MeSH
• myši MeSH
• oocyty metabolismus   MeSH
• oogeneze fyziologie   MeSH
• proteinmethyltransferasy MeSH
• techniky jaderného přenosu MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH

• MeSH
• donory oxidu dusnatého farmakologie   MeSH
• finanční podpora výzkumu jako téma MeSH
• oocyty cytologie   metabolismus   účinky léků   MeSH
• oxid dusnatý MeSH
• penicilamin analogy a deriváty   farmakologie   MeSH
• synthasa oxidu dusnatého antagonisté a inhibitory   metabolismus   MeSH
• techniky in vitro MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH