Vitamin C (Vc) is a natural compound supplemented to culture media to guarantee the appropriate reactive oxygen species (ROS) level, as well as protect cells from oxidative damage and apoptosis. The current study was conducted to determine the effects of Vc (0, 2.5, 5, 10, 20 and 40 μg/ml) on the ROS production, developmental ability and quality of in vitro produced porcine parthenotes. The results show that: (i) the ROS levels in the embryos significantly decrease in the Vc-treated groups compared with the control (p < 0.05), (ii) the rates of blastocyst formation and total cell numbers in each blastocyst are significantly higher in the Vc-treated groups than in the control (p < 0.05); the optimum concentration of Vc is 20 μg/ml, (iii) the relative expression of Bcl-xL significantly increases and that of Bax is downregulated after Vc treatment. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling analysis indicates that the ratio of apoptotic cells in the blastocyst is also significantly lower in Vc-treated groups (p < 0.05) and (iv) Vc treatment can also increase the expression of the Nanog gene in porcine embryos, with a fivefold increase in 20 μg/ml Vc treatment compared with the control (p < 0.05). Therefore, Vc improves the development of porcine embryos by reducing the ROS levels. Vc addition in PZM-3 medium can decrease the number of apoptotic cells and increase the cell numbers in blastocysts to produce high-quality porcine embryos in vitro.
The objective of this study was to determine if insulin-transferrin-selenium (ITS) promoted a nuclear and cytoplasmic maturation of porcine oocytes that better supports subsequent embryonic development. The rate of oocyte in vitro maturation (IVM) in an experimental group treated with hormones for 42 h was significantly increased compared with that in a control group without hormone treatment (47.8% vs. 11.7%, respectively, p < 0.05). Following reduction of the hormone treatment period from 42 h to 21 h, which included both the first 21 h period of hormones treatment (45.4%) and the second 21 h period of hormone treatment (44.8%), the rate of oocyte IVM was still higher than that of the control group (p < 0.05). To improve porcine oocyte nuclear maturation, 1% ITS was added to medium supplemented with hormones. The rate of nuclear maturation in the ITS-treated group was significantly higher than in the ITS-untreated group (78.6% vs. 54.4%, respectively, p < 0.05). ITS treatment also significantly reduced the per cent of oocytes with type I and type III cortical granule (CG) distribution, respectively, and significantly increased the per cent of oocytes with type II CG distribution (85.3%). These observations indicated that the synchronization rates of nuclear and ooplasmic maturation reached 67.04% (78.56 × 85.33%). In conclusion, the combination of modified Tissue Culture Medium-199 (mM199) + 10 ng/ml epidermal growth factor (EGF) + 10 IU/ml pregnant mare serum gonadotrophin (PMSG) + 10 IU/ml human chorion gonadotrophin (hCG) + 2.5 IU/ml follicle stimulating hormone (FSH) + 1% ITS is suitable for culturing porcine oocytes in vitro, and effectively enhances porcine oocyte nuclear and cytoplasmic maturation.
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