153 ZINC INSUFFICIENCY DURING PORCINE OOCYTES IN VITRO MATURATION CAUSED MEIOTIC BLOCK AND DEVELOPMENTAL FAILURE

Abstract

The objective was to investigate the effects of zinc (Zn) insufficiency during in vitro maturation (IVM) of porcine oocytes. Zinc insufficiency was induced by treatment of Zn chelator, N,N,N′,N′-tetrakis-(2-pyridylmethyl)-ethylendiamine (TPEN). In experiment 1, we investigated the effect of duration of Zn insufficiency in IVM on oocytes maturation and subsequent embryonic development after parthenogenetic activation (PA). First, 10 μM TPEN was added to the IVM medium for 0, 7, 15, or 22 h. After TPEN treatment, 10 μM Zn were supplemented on IVM medium except in the 0 h group. Reductions in the nuclear maturation rates were dependent on TPEN duration. The 0-h-treated oocytes showed 83.9 ± 3.9% metaphase II (MII) rate; the 7-h-treated oocytes had significantly lower MII rate (44.8 ± 3.0%) than 0-h-treated oocytes. The majority of 15- and 22-h-treated oocytes were arrested at metaphase I (MI rate: 98.0 ± 1.0 and 97.2 ± 1.7%, MII rate: 0 and 0%, respectively). Embryonic developmental competence was similar to maturation results. Reduction in cleavage and blastocyst (BL) rates were also dependent on duration of TPEN treatment (cleavage rate: 65.3 ± 1.4, 42.6 ± 4.8, 2.6 ± 0.1, and 3.0 ± 1.6%; BL formation rate: 29.3 ± 2.8, 9.2 ± 1.5, 0, and 0% for 0, 7, 15, and 22 h). Total cell number of BL was also significantly different. Total cell number of BL in the 0-h-treated group (51.4 ± 4.5) was significantly higher than that in the 7-h-treated group (23.2 ± 1.6). In experiment 2, to confirm that the Zn insufficiency caused oocyte immaturities and loss of developmental competence in TPEN-treated oocytes, we investigated nuclear maturation and subsequent embryonic development following 3 groups: (1) non treatment (control); (2) 10 μM TPEN treatment during 22 h of IVM; (3) 10 μM TPEN + 10 μM Zn treatment during 22 h of IVM. Only TPEN-treated oocytes and TPEN+Zn-treated oocytes showed contrasting results. Oocyte maturation rates and subsequent embryonic development competence in TPEN with Zn-treated oocytes were similar to control (MII rate: 93.0 ± 1.2 and 92.7 ± 1.8%, BL formation rate: 42.0 ± 6.7 and 40.0 ± 7.5% for TPEN+Zn-treated oocytes and control). These results were significantly different compared with only TPEN-treated oocytes’ results (MII rate: 0.61 ± 0.61%, BL formation rate: 0%). In conclusion, Zn is an essential element for successful oocyte maturation and embryo development in porcine. Zinc insufficiency caused meiotic block and had lasting effects on early embryo development. This work was supported, in part, by a grant from the Next-Generation BioGreen 21 Program (No. PJ00956901), Rural Development Administration, and the National Research Foundation of Korea Grant funded by the Korean Government (NRF-2012R1A1A4A01004885, NRF-2013R1A2A2A04008751), Republic of Korea.