Low oxygen tension during in vitro maturation of porcine follicular oocytes improves parthenogenetic activation and subsequent development to the blastocyst stage

Abstract

To establish a reliable in vitro maturation system for activation and subsequent development as nuclear recipients for the effective production of pig clones, we assessed maturation, activation and parthenogenetic development in response to the following: (1) type of immature oocytes (cumulus–oocyte complexes (COCs) or parietal granulosa plus cumulus–oocyte complexes (GCOCs)); (2) oxygen (O 2) tension (5 or 20%); and (3) maturation period (36–60 h). The rate of nuclear maturation to metaphase-II (M-II) in the GCOC group (73.0 ± 3.1%) was higher than that in the COC group ( P < 0.05, 60.6 ± 3.5%), but the rates did not differ between the 5 and 20% O 2 tension groups. M-II rate increased ( P < 0.05) to about 70% after 42 h and then remained constant until 60 h of culture. When oocytes were matured under 5% O 2 tension and stimulated, the rate of normal oocyte activation (a female pronucleus formation and emission of the second polar body) was higher ( P < 0.05, 38.5 ± 3.9%) than when oocytes were matured under 20% O 2 tension (24.5 ± 3.9%). On the other hand, the rate of normal activation was not significantly different between the COC and GCOC groups, and the highest ( P < 0.05) normal activation rate was obtained in oocytes cultured for 48 and 54 h (48.4 ± 5.5% and 47.9 ± 8.2%, respectively). When COC and GCOC matured for 48 h under 5 and 20% O 2 tension were stimulated and subsequently cultured in vitro for 6 days, the rate of blastocyst formation did not differ between the oocyte types nor between the O 2 tension groups. However, blastocyst quality, as measured by mean total cell number, was significantly higher in the 5% O 2 group ( P < 0.05, 34.6 ± 2.0 for COC; 33.8 ± 1.8 for GCOC) compared with the 20% O 2 group (25.9 ± 1.8 for COC; 27.0 ± 2.0 for GCOC). In conclusion, low O 2 tension (5%) during in vitro maturation of porcine oocytes promoted their ability to be activated normally and improved the quality of parthenogenetic blastocysts developed in vitro in modified NCSU-37 solutions. This knowledge may be applicable for preparation of in vitro matured oocytes with good quality as recipient oocytes for generating pig clones.