Polyspermia is an adverse phenomenon during mammalian fertilization when more than one sperm fuses with a single oocyte. The egg cell is prepared to prevent polyspermia by, among other ways, producing cortical granules (CGs), which are specialized intracellular structures containing enzymes that aim to harden the zona pellucida and block the fusion of subsequent sperm. This work focused on exploring the expression profile of genes that may be associated with cortical reactions, and evaluated the distribution of CGs in immature oocytes and the peripheral density of CGs in mature oocytes. Oocytes were isolated and then processed for in vitro maturation (IVM). Transcriptomic analysis of genes belonging to five ontological groups has been conducted. Six genes showed increased expression after IVM (ARHGEF2, MAP1B, CXCL12, FN1, DAB2, and SOX9), while the majority of genes decreased expression after IVM. Using CG distribution analysis in immature oocytes, movement towards the cortical zone of the oocyte during meiotic competence acquisition was observed. CGs peripheral density decreased with the rise in meiotic competence during the IVM process. The current results reveal important new insights into the in vitro maturation of oocytes. Our results may serve as a basis for further studies to investigate the cortical reaction of oocytes.

• Keywords
• cortical granule, microarray, molecular markers, oocyte maturation, pig,
• MeSH
• Cell Differentiation * MeSH
• Cytoplasmic Granules metabolism   MeSH
• In Vitro Oocyte Maturation Techniques methods   MeSH
• Cells, Cultured MeSH
• Oocytes cytology   metabolism   MeSH
• Swine MeSH
• Transcriptome * MeSH
• Animals MeSH
• Check Tag
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

This paper aims to identify and describe new genetic markers involved in the processes of protein expression and modification reflected in the change of mitochondrial activity before and after in vitro maturation of the oocyte. Porcine oocytes collected from the ovaries of slaughtered landrace gilts were subjected to the process of in vitro maturation. Transcriptomic changes in the expression profile of oocyte genes involved in response to hypoxia, the transmembrane protein receptor serine threonine kinase signaling pathway, the "transforming growth factor β receptor signaling pathway", "response to protein stimulus", and "response to organic substance" were investigated using microarrays. The expression values of these genes in oocytes was analyzed before (immature) and after (mature) in vitro maturation, with significant differences found. All the significantly altered genes showed downregulation after the maturation process. The most changed genes from these gene ontologies, FOS, ID2, VEGFA, BTG2, CYR61, ESR1, AR, TACR3, CCND2, CHRDL1, were chosen to be further validated, described and related to the literature. Additionally, the mitochondrial activity of the analyzed oocytes was measured using specific dyes. We found that the mitochondrial activity was higher before the maturation process. The analysis of these results and the available literature provides a novel insight on the processes that occur during in vitro oocyte maturation. While this knowledge may prove to be useful in further research of the procedures commonly associated with in vitro fertilization procedures, it serves mostly as a basic reference for further proteomic, in vivo, and clinical studies that are necessary to translate it into practical applications.

• Keywords
• microarray, mitochondrial activity, oocyte maturation, pig,
• MeSH
• Cell Hypoxia genetics   MeSH
• In Vitro Oocyte Maturation Techniques MeSH
• Cells, Cultured MeSH
• Mitochondria genetics   metabolism   MeSH
• Oocytes cytology   metabolism   MeSH
• Oogenesis genetics   MeSH
• Swine MeSH
• Signal Transduction MeSH
• Transforming Growth Factor beta metabolism   MeSH
• Transcriptome * MeSH
• Receptor Protein-Tyrosine Kinases metabolism   MeSH
• Animals MeSH
• Check Tag
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Transforming Growth Factor beta MeSH
• Receptor Protein-Tyrosine Kinases MeSH

The growth and development of oocyte affect the functional activities of the surrounding somatic cells. These cells are regulated by various types of hormones, proteins, metabolites, and regulatory molecules through gap communication, ultimately leading to the development and maturation of oocytes. The close association between somatic cells and oocytes, which together form the cumulus-oocyte complexes (COCs), and their bi-directional communication are crucial for the acquisition of developmental competences by the oocyte. In this study, oocytes were extracted from the ovaries obtained from crossbred landrace gilts and subjected to in vitro maturation. RNA isolated from those oocytes was used for the subsequent microarray analysis. The data obtained shows, for the first time, variable levels of gene expression (fold changes higher than |2| and adjusted p-value < 0.05) belonging to four ontological groups: regulation of cell proliferation (GO:0042127), regulation of cell migration (GO:0030334), and regulation of programmed cell death (GO:0043067) that can be used together as proliferation, migration or apoptosis markers. We have identified several genes of porcine oocytes (ID2, VEGFA, BTG2, ESR1, CCND2, EDNRA, ANGPTL4, TGFBR3, GJA1, LAMA2, KIT, TPM1, VCP, GRID2, MEF2C, RPS3A, PLD1, BTG3, CD47, MITF), whose expression after in vitro maturation (IVM) is downregulated with different degrees. Our results may be helpful in further elucidating the molecular basis and functional significance of a number of gene markers associated with the processes of migration, proliferation and angiogenesis occurring in COCs.

• Keywords
• cellular competence, microarray, oocytes, pig,
• MeSH
• Apoptosis genetics   MeSH
• Down-Regulation MeSH
• Gene Regulatory Networks MeSH
• In Vitro Oocyte Maturation Techniques MeSH
• Cumulus Cells metabolism   pathology   MeSH
• Oocytes growth & development   metabolism   pathology   MeSH
• Cell Movement genetics   MeSH
• Swine MeSH
• Cell Proliferation genetics   MeSH
• RNA genetics   metabolism   MeSH
• Oligonucleotide Array Sequence Analysis MeSH
• Gene Expression Profiling MeSH
• Transcriptome * MeSH
• Up-Regulation MeSH
• Animals MeSH
• Check Tag
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• RNA MeSH

Proper maturation of the mammalian oocyte is a compound processes determining successful monospermic fertilization, however the number of fully mature porcine oocytes is still unsatisfactory. Since oocytes' maturation and fertilization involve cellular adhesion and membranous contact, the aim was to investigate cell adhesion ontology group in porcine oocytes. The oocytes were collected from ovaries of 45 pubertal crossbred Landrace gilts and subjected to two BCB tests. After the first test, only granulosa cell-free BCB⁺ oocytes were directly exposed to microarray assays and RT-qPCR ("before IVM" group), or first in vitro matured and then if classified as BCB⁺ passed to molecular analyses ("after IVM" group). As a result, we have discovered substantial down-regulation of genes involved in adhesion processes, such as: organization of actin cytoskeleton, migration, proliferation, differentiation, apoptosis, survival or angiogenesis in porcine oocytes after IVM, compared to oocytes analyzed before IVM. In conclusion, we found that biological adhesion may be recognized as the process involved in porcine oocytes' successful IVM. Down-regulation of genes included in this ontology group in immature oocytes after IVM points to their unique function in oocyte's achievement of fully mature stages. Thus, results indicated new molecular markers involved in porcine oocyte IVM, displaying essential roles in biological adhesion processes.

• Keywords
• gamete biology, in vitro maturation (IVM), molecular biology, oocytes, pig,
• MeSH
• Apoptosis MeSH
• Cell Adhesion MeSH
• Cell Differentiation MeSH
• Down-Regulation * MeSH
• Gene Regulatory Networks * MeSH
• In Vitro Oocyte Maturation Techniques veterinary   MeSH
• Oocytes cytology   metabolism   MeSH
• Oogenesis MeSH
• Swine MeSH
• Cell Proliferation MeSH
• Animals MeSH
• Check Tag
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

BACKGROUND: The full maturational capability of mammalian oocytes is accompanied by nuclear and cytoplasmic modifications, which are associated with proliferation and differentiation of surrounding cumulus cells. These events are regulated on molecular level by the expression of target genes involved in signal transduction pathways crucial for folliculogenesis and oogenesis. Transforming growth factor beta signaling includes several molecules that are involved in the regulation of oogenesis and embryo growth, including bone morphogenetic protein (BMP). However, the BMP-related gene expression profile in oocytes at different maturational stages requires further investigation. METHODS: Oocytes were isolated from pubertal crossbred Landrace gilts follicles, selected with a use of BCB staining test and analyzed before and after in vitro maturation. Gene expression profiles were examined using an Affymetrix microarray approach and validated by RT-qPCR. Database for Annotation, Visualization, and Integrated Discovery (DAVID) software was used for the extraction of the genes belonging to a BMP-signaling pathway ontology group. RESULTS: The assay revealed 12,258 different transcripts in porcine oocytes, among which 379 genes were down-regulated and 40 were up-regulated. The DAVID database indicated a "BMP signaling pathway" ontology group, which was significantly regulated in both groups of oocytes. We discovered five up-regulated genes in oocytes before versus after in vitro maturation (IVM): chordin-like 1 (CHRDL1), follistatin (FST), transforming growth factor-beta receptor-type III (TGFβR3), decapentaplegic homolog 4 (SMAD4), and inhibitor of DNA binding 1 (ID1). CONCLUSIONS: Increased expression of CHRDL1, FST, TGFβR3, SMAD4, and ID1 transcripts before IVM suggested a subordinate role of the BMP signaling pathway in porcine oocyte maturational competence. Conversely, it is postulated that these genes are involved in early stages of folliculogenesis and oogenesis regulation in pigs, since in oocytes before IVM increased expression was observed.

• Keywords
• In vitro maturation, Microarray, Oocytes, Pig,
• MeSH
• In Vitro Oocyte Maturation Techniques * MeSH
• Bone Morphogenetic Proteins genetics   metabolism   MeSH
• Cumulus Cells cytology   metabolism   physiology   MeSH
• Microarray Analysis MeSH
• Oocytes cytology   metabolism   physiology   MeSH
• Oogenesis genetics   MeSH
• Swine genetics   metabolism   MeSH
• Signal Transduction genetics   MeSH
• Gene Expression Profiling MeSH
• Transcriptome MeSH
• Gene Expression Regulation, Developmental MeSH
• Animals MeSH
• Check Tag
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Bone Morphogenetic Proteins MeSH