Differential effects of oxygen on hatching of fertilized and parthenogenetic embryos

Abstract

Embryos exposed to in vitro culture are prone to oxidative damage. Low oxygen tension during early preimplantation development appears beneficial, but is less studied during later development. Moreover, the effects of oxygen on development of parthenogenetic embryos remain unclear. Laboratory studies of fertilized and parthenogenetic mouse embryos, cultured in low (5%) and high (20%, atmospheric) O2, respectively, their rates of development to blastocyst, hatching, and expression profiles of apoptosis, imprinted, oxidation and protease genes. Parthenogenetic embryos (PEs) were achieved by activation of oocytes with strontiuim and cyctochalasin D. Embryos were cultured in KSOM at 37°C under 6% CO2 and 5% or 20% O2. Some embryos were processed for Oct4 expression and apoptosis by TUNEL analysis. Real-time PCR was used for gene expression analysis. Blastocysts developed from zygotes under conditions of low or high O2 were transferred into psuedopregnant recipients for term development. Both PEs and zygotes cultured under 5% or 20% O2 exhibited similar high rates for development to blastocysts (>96%). Apoptosis did not differ between parthenogenetic and fertilized blastocysts, regardless of O2 tension. Notably, blastocysts (n = 77) cultured under 5% O2 showed higher rates of implantation (71% vs. 47%) and live pups (44% vs. 20%), compared to those (n = 89) cultured under 20% O2. Total cell number of PE blastocysts was not affected by O2 tension, but the number of ICM was increased under 5% O2. Zygotes cultured under 5% O2 had markably lower rates of hatching (21%) vs. those under 20% O2 (57%). By contrast, the hatching rate (91%) of PEs cultured under 5% O2 was higher than under 20% O2 (80%). Reduced expression of ERK/MAPK but increased Sod1 was observed for PE blastocysts, with no effects of oxygen tension. High O2 reduced expression of Sod2 in both types of embryos. As expected, expression of the imprinted genes H19 was increased but Snrpn decreased in PE blastocysts, compared to fertilized blastocysts, but neither was affected by oxygen tension. Oxygen tension does not affect development to blastocyst of either fertilized or PEs, but low oxygen tension promotes embryo survival following postimplantation development. Low oxygen inhibits hatching of fertilized blastocysts, but promotes hatching of parthenogenetic blastocysts. The mechanism of oxygen's differential effects on fertilized vs. parthenogenic blastocysts remains to be determined.