| Optimization of culture systems for in vitro production of animal oocytes is achieved by supplementation with various antioxidants. Previous research has shown that epigallocatechin gallate (EGCG) has positive effects on in vitro maturation and in vitro fertilization of animal oocytes, and is thought to act by decreasing the concentrations of reactive oxygen species (ROS) as a result of its antioxidant properties. This project has investigated the effects of addition of various concentrations of EGCG to the maturation medium on the maturation of buffalo oocytes, their maturation quality, and early embryo development in vitro.Experiment 1 investigated the effects of different concentrations of EGCG (0,5,10,20 and 30 ?mol/L) on in vitro maturation of buffalo oocytes. The results showed:(i) the cumulus expansion index (CEI) in was higher in groups receiving 10, or 20 ?mol/L EGCG than that in the control (2.49,2.42 vs.2.22, P<0.05); the CEI values for the 5 and 30?mol/L EGCG treatment groups were also higher than that of the control but the differences were not significant (2.36 and 2.28 vs.2.22, P>0.05); (?) the maturation rate in the 10 and 20 ?mol/L EGCG treatment group was significantly higher than that in the control (59.31%,50.98%vs.45.26%, P<0.05), whereas the maturation rate of the 5 and 30 ?mol/L EGCG treatment groups were not significantly different from the control (47.04%,42.81% vs.45.26% P>0.05); (iii) the cleavage and blastocyst rates of the 10 ?mol/L EGCG treatment group were better than with the other groups, and significantly higher than those of the control group after parthenogenetic activation (67.07% vs.57.53%,31.69% vs.20.66%, p<0.05, respectively). Also, after in vitro fertilization, the cleavage and blastocyst rates for the 10 ?mol/L EGCG treatment group were significantly higher than those of the control group (59.64% vs.44.65%,24.68% vs.15.02%, p<0.05, respectively). These results demonstrate that EGCG can help promote maturation and early embryo development of buffalo oocytes in vitro, and the most effective concentration in this study was 10 ?mol/LExperiment 2 investigated the effects of different concentrations of EGCG (0,5,10,20 and 30 ?mol/L) on the antioxidant capacity of suspensions of buffalo oocytes. The results showed:(i) for the 20 and 30 ?mol/L EGCG treatment groups, the DCF fluorescence intensity of ROS excreted by MII phase pocytes was significantly lower than that in the control (18.77,25.39 vs.38.04, P<0.05); (ii) the concentrations of H2O2 excreted by MII phase oocytes were not significantly different among these treatment groups (4.68,4.33,4.16,4.51 vs 4.90, P>0.05); (?) addition of 5,10 or 20 ?mol/L EGCG to maturation medium resulted in significantly higher excretion of GSH by MII phase oocytes than for the control group (3.03,3.21,3.25 vs 2.59, P<0.05), whereas addition of 30 ?mol/L EGCG to maturation medium did not result in any significant difference from the control group (2.99 vs.2.59, P>0.05). (?) overall the results show that EGCG influences antioxidant processes involved in in vitro maturation.Experiment 3 investigated the effects of addition of 10 ?mol/L EGCG to the maturation medium, fertilization medium, or both media on the in vitro fertilization of buffalo oocytes. The results showed that (i) for each group, the cleavage and blastocyst rates were significantly higher than that in the control (61.31%,59.14% and 58.57 vs.45.71%, P<0.05, and 23.61%,22.77% and 23.56% vs.16.74%, P<0.05, respectively). Thus EGCG can promote in vitro fertilization and early embryo development of buffalo oocytes, and at the same time improve the quality of the blastocysts. In this study, optimal results were achieved by adding 10 ?mol/L EGCG to either the maturation or fertilization medium.EGCG is a powerful antioxidant and has been shown to be able to promote the maturation of buffalo oocytes, their fertilization and early embryo development in vitro. This study also showed that its most effective concentration was 10 ?mol/L. |
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