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      HEPES buffer in ovary-transportation medium influences developmental competence of cattle oocytes

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          Abstract

          This study was conducted to investigate the effects of ovary transportation in a semi-complex medium containing HEPES at different temperatures on the developmental competence and the quality of in vitro-produced embryos. The cattle ovaries were transported in normal saline (NS), phosphate buffer saline (PBS), K simplex optimization medium (KSOM), Chatot-Ziomek-Bavister medium (CZB) and Charles Rosenkrans medium (CRl) at various temperatures (38 °C, 25 °C and 4 °C). The developmental competence of retrieved cumulus oocyte complexes (COCs) was evaluated by maturation, fertilization, morula and blastocyst formation and numbers of inner cell mass (ICM) and trophectoderm (TE) cells. The COC maturation rate was affected by medium and temperature. It was found that 4 °C resulted in a higher maturation (81.0 ± 4.75) rate than other transportation temperatures. The CR1 (80.5 ± 6.66) and KSOM (80.2 ± 6.15) gave a better maturation rate than the others. Fertilization rate, which was evaluated by cleavage rate, was not affected by transportation temperature. However, the transporting medium had a significant effect on the fertilization rate. Moreover, CR1 (43.6 ± 4.60), KSOM (43.2 ± 4.86) and CZB (41.1 ± 4.86) media gave higher percentages of cleaved embryos. There was no significant difference in morula and blastocyst formation rate or in ICM and TE cell counts regarding transportation factors. In conclusion, the transport of ovaries in CR1 at 4 °C is effective for maintaining early developmental competence of cattle oocytes.

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          Expression of Bcl-2 and Bax proteins in relation to quality of bovine oocytes and embryos produced in vitro.

          The mechanisms underlying the visual assessment and selection of immature oocytes resulting in optimum embryonic development following in vitro maturation, fertilization and culture (in vitro maturation (IVM)/in vitro fertilization (IVF)/in vitro embryo culture (IVC)) are unknown. Also, the reasons for the more frequent occurrence of cytoplasmic fragmentation in in vitro produced bovine embryos, resulting in poor survival following cryopreservation and decreased pregnancy rates following embryo transfer are not clear. The objectives of this study are: (1) to investigate whether differences in the quality of immature oocytes and embryo fragmentation are associated with apoptosis; and (2) to study the pattern of Bcl-2 and Bax expression in oocytes and embryos to help elucidate their potential roles in the regulation of apoptosis during development. Bovine oocytes were obtained from slaughterhouse ovaries and divided into four grades (grades I-IV) based on their morphology. Oocytes of different grades were cultured in serum-free medium for 48h. Embryos were produced only from grade I oocytes (highest quality) via IVM, IVF and IVC procedures. The morphological analysis of apoptosis in oocytes and embryos was carried out using propidium iodide staining and terminal deoxynucleotidyl transferase mediated dUTP nick end labeling. The expression of Bcl-2 and Bax in oocytes and embryos of different qualities and stages was determined using western blotting. The results showed that the number of morphologically abnormal oocytes with shrinkage and/or fragmentation of the ooplasm, which are typical features of apoptosis, was significantly higher in grade IV oocytes (denuded oocytes, the lowest quality) than in grade I oocytes after 48h in vitro culture (P<0.05). DNA fragmentation, a hallmark of the biochemical changes seen in apoptotic cell death, was observed in morphologically fragmented oocytes and embryos. The expression of Bcl-2 was high in good quality oocytes and embryos, low in fragmented embryos, and hardly detectable in denuded oocytes. In contrast, the expression of Bax was found in all types of oocytes and embryos with the highest expression in the denuded oocytes. This implies that the ratio of Bcl-2 to Bax may be used to gauge the tendency of oocytes and embryos towards either survival or apoptosis. Overall, our results show that apoptosis appears to be an underlying mechanism of bovine oocyte degeneration and embryo fragmentation. Interactions between the Bcl-2 family of proteins may play a critical role in pre-implantation embryo development. These findings could have important implications for improving IVF and related techniques.
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            Simplified technique for differential staining of inner cell mass and trophectoderm cells of mouse and bovine blastocysts

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              Cryopreservation of ova and embryos from livestock: Current status and research needs

               H. Niemann (1991)
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                Author and article information

                Contributors
                Role: ND
                Role: ND
                Role: ND
                Journal
                sajas
                South African Journal of Animal Science
                S. Afr. j. anim. sci.
                The South African Society for Animal Science (SASAS)
                2221-4062
                2015
                : 45
                : 5
                : 538-546
                Affiliations
                [1 ] University of Ankara Turkey
                Article
                S0375-15892015000500009
                10.4314/sajas.v45i5.11

                This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.

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                Product Information: website
                Categories
                Agriculture, Dairy & Animal Science
                Genetics & Heredity
                Nutrition & Dietetics
                Physiology

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