Cyclic adenosine monophosphate (cAMP) modulators have been used to avoid spontaneous oocyte maturation and concomitantly improve oocyte developmental competence. The current work evaluated the effects of the addition of cAMP modulators forskolin, 3-isobutyl-1-methylxanthine (IBMX) and cilostamide during in vitro maturation on the quality and yields of blastocysts. The following experimental groups were evaluated: (i) slicing or (ii) aspiration and maturation in tissue culture medium (TCM)199 for 24 h (TCM24slicing and TCM24aspiration, respectively), (iii) aspiration and maturation in the presence of cAMP modulators for 30 h (cAMP30aspiration) and in vivo-produced blastocysts. In vitro-matured oocytes were fertilized and presumptive zygotes were cultured in vitro to assess embryo development. Cleavage, blastocyst formation, blastocyst cell number, mRNA abundance of selected genes and global methylation profiles were evaluated. Blastocyst rate/zygotes for the TCM24aspiration protocol was improved (32.2 ± 2.1%) compared with TCM24slicing and cAMP30aspiration (23.4 ± 1.2% and 23.3 ± 2.0%, respectively, P 0.05), while those from the other groups were significantly elevated. It is concluded that retrieval, collection systems and addition of cAMP modulators can affect oocyte developmental competence, which is reflected not only in blastocyst rates but also in global DNA methylation and gene expression patterns.

<p>La producción de embriones in vitro (PIV) es una biotecnología reproductiva que impacta los sistemas ganaderos favoreciendo el mejoramiento genético y que también se ha utilizado como modelo de investigación del desarrollo embrionario en otras especies. Para el desarrollo de ésta técnica, se colectan oocitos in vivo por medio de laparoscopia y aspiración folicular guiada por ultrasonografía ó a través de ovarios recolectados en plantas de sacrificio. Los oocitos son sometidos a procesos de maduración in vitro, utilizando diversos medios, adición de gonadotropinas y factores de crecimiento. Los espermatozoides son sometidos a tratamientos de capacitación espermática in vitro, donde adquieren capacidad fertilizante y se separan los espermatozoides vivos de los componentes seminales y crio-protectores. Para la fertilización in vitro se realiza un cocultivo de espermatozoides y oocitos en un medio que favorece la actividad espermática y penetración de la zona pelúcida. Posteriormente, los embriones obtenidos son sometidos a cultivo en medios que contengan fuentes de energía, aminoácidos y factores de crecimiento, con el fin de mantener su sobrevivencia y desarrollo. En la actualidad la técnica de PIV en bovinos se ha masificado y se trabaja en los procesos de estandarización en otras especies. A partir de la PIV se dio el desarrollo de nuevas biotecnologías como la clonación y transgénesis. El objetivo de esta revisión es describir las fases de la PIV y su relación con los eventos fisiológicos de la fertilización y desarrollo embrionario temprano.</p>

Resveratrol (3,4′,5-trihydroxystilbene) is a phytoalexin identified in various plant species, particularly in grapevine peel. It is a strong antioxidant, induces mitochondrial biogenesis and enhances Sirtuin 1 (SIRT1) activity by inhibiting phosphodiesterase. SIRT1 belongs to the family of NAD+-dependent histone deacetylates and has been shown to regulate several key cellular processes, including transcriptional silencing, aging, chromatin remodeling, and genomic stability, via deacetylation of p53, FoxO transcription factors, and nuclear factor kappa B (NF-κB). The aim of this study was to determine whether supplementation of the maturation and fertilisation medium with resveratrol influences bovine oocyte maturation and subsequent embryonic development and whether these effects are mediated via SIRT1 pathway. Three different resveratrol concentrations were used during in vitro maturation (IVM) and IVF. Cumulus-oocyte complexes (n = 2878) were collected from slaughterhouse ovaries and subjected to IVM medium supplemented with 0.2 µM, 1 µM, or 20 µM resveratrol® (Sigma-Aldrich, Buchs, Switzerland) for 24 h followed by IVF with the same concentrations of resveratrol for 19 h. The IVM and IVF medium without resveratrol (controls) and dimethyl sulfoxide supplementation as vehicle control were also included. Presumptive zygotes were cultured in vitro until Day 8 to assess embryo development, and maturation rates, cleavage, and blastocyst formation were evaluated. Maturation rates as determined by polar body extrusion (0.2 µM: 64.2% ± 7; 1 µM: 82.3% ± 4; 20 µM: 68.8% ± 2; control: 74.6% ± 5 and vehicle control: 70.2% ± 6, respectively; P ≤ 0.05) did not differ dramatically. Oocytes in 1 µM resveratrol supplemented maturation medium showed distinct detachment of cumulus cells in comparison with those in the other treatment and control groups. Cleavage rates were reduced in the 0.2 µM and 20 µM group compared with controls (0.2 µM: 44.21% ± 2; 1 µM: 58.4% ± 3; 20 µM: 40.9% ± 5; control: 56.6% ± 2 and vehicle control: 55.2% ± 6, respectively; P ≤ 0.05). Blastocyst rates were impaired in the low and high resveratrol concentration groups compared to all other groups (0.2 µM: 11.3% ± 1; 1 µM: 33.4% ± 3; 20 µM: 8.2% ± 4; control: 26.7% ± 4 and vehicle control: 20.8% ± 2, respectively; P ≤ 0.05). Relative mRNA abundance of SIRT1 in matured oocytes from the 1 µM group did not differ significantly compared to the controls. Results so far indicate that very low and high concentrations of resveratrol impair development to the blastocyst stage. In conclusion, a 1 µM resveratrol supplementation during IVM and IVF seems to improve the developmental competence of oocytes, which is reflected not only in the elevated blastocyst rates but also in higher degree of expansion of cumulus cells after IVM and maturation rates.