Effects of Guaiazulene on In Vitro Bovine Embryo Production and on mRNA Transcripts Related to Embryo Quality
AuthorDovolou, E.; Clemente, M.; Amiridis, G. S.; Messinis, I.; Kallitsaris, A.; Gutierrez-Adan, A.; Rizos, D.
Contents: Reactive oxygen species (ROS) are between the major contributors for the reduced rate of in vitro bovine embryo production. It is believed that they can cause abnormal meiosis of oocytes, developmental arrest or cell death of embryos. Reports on the effectiveness of various antioxidants on embryo yield are rather conflicting mainly due to the nature and the concentration of the substances used. Here we report the effects of guaiazulene - an exogenous antioxidant, without known properties that could interfere with the biological process of IVF - on embryo development and on the quality of the produced blastocysts. Bovine cumulus oocyte complexes (COCs) were aspirated from abattoir ovaries and COCs were matured in TCM199 with FCS and EGF at 39°C under an atmosphere of 5% CO 2 in air, with maximum humidity. After 24h oocytes were inseminated with frozen/thawed semen and co-incubated for further 24h. Zygotes were cultured in groups of 25 in 25μl of SOF with 5% FCS at 39°C under an atmosphere of 5% CO 2, 5% O 2 in air with maximum humidity. In the first experiment the maturation medium was modified with addition of 0.1mm of G (n=497), or 0.01mm of guaiazulene (n=468), 0.05% DMSO-the guaiazulene diluent (Control +, n=467), and 459 oocytes were used as Control -. In the second experiment, the culture medium was modified with the addition of 0.1mm of guaiazulene (n=344), 0.01mm of guaiazulene (n=345), 0.05% DMSO (Control +, n=347) and 355 were the Control -. Blastocyst yield was recorded on days 6, 7, 8 and 9. Day 7 blastocysts from each experiment and group were snap frozen and stored for mRNA extraction. Quantification of transcripts for mRNA of genes related to metabolism (AKR1B1, PTGS2, GADPH, SLC2A5, G6PD); oxidation (GPX1); and implantation (PLAC8) was carried out by real time quantitative RT-PCR. Data for embryo development and on transcript abundance were analysed by χ 2 and anova respectively. In the first experiment no differences were found between groups in terms of cleavage rate (Control -: 74.20%; Control +: 74.58%; 0.01mm: 71.61%; 0.1mm: 71.63%) or day 9 blastocyst yield (Control -: 28.26%; Control +: 25.80%; 0.01mm: 25.86%; 0.1mm: 25.25%). In the second experiment, cleavage rate tended to be higher in 0.01mm group than in Control - (77.87% vs 71.41% respectively, p=0.07). No other differences were detected in cleavage rate (Control +: 71.32%; 0.1mm: 72.75%) or in the overall blastocyst yield on day 9 (Control -: 25.50%; Control +: 26.71%; 0.01mm: 29.58%; 0.1mm: 25.75%). In both experiments the relative abundance of genes studied varied between groups but these differences were not statistically significant. Our results imply that oxidation has minimal effect on the in vitro embryo production. Guaiazulene, a compound possessing no biological properties other than those of a strong antioxidant, while it increased cleavage rate, it failed to improve either the blastocyst formation rate, or the quality of the produced embryos under 5% O 2. © 2011 Blackwell Verlag GmbH.
Showing items related by title, author, creator and subject.
UROKINASE-TYPE PLASMINOGEN ACTIVATOR DOES NOT AFFECT IN VITRO BOVINE EMBRYO DEVELOPMENT AND QUALITY Krania, F.; Dovolou, E.; Rekkas, C. A.; Heras, S.; Pappas, I.; Van Soom, A.; Amiridis, G. S. (2015)The effects of modification of the in vitro embryo culture media (IVC) with the addition of urokinase-type plasminogen activator (u-PA) on the yield and/or quality of bovine embryos were examined in two experiments. In ...
Amiridis, G. S.; Cseh, S. (2012)In modern agriculture, assisted reproductive technologies are being used for out of season oestrus induction, enhancement of reproductive performance and genetic improvement. In addition, they can have substantial contribution ...
A light and electron microscopic study of the limb long bones perichondral ossification in the quail embryo (Coturnix coturnix japonica) Pourlis, A. F.; Antonopoulos, J.; Magras, I. N. (2006)The perichondral ossification of the limb long bones in the quail embryo is investigated, in this study, by means of light and electron microscopy. Longitudinal sections of the humerus, radius, ulna, femur, tibia and fibula ...