Development to the blastocyst stage of parthenogenetically activated in vitro matured porcine oocytes after solid surface vitrification (SSV)

Abstract

The effects of solid surface vitrification (SSV) on viability and parthenogenetic development of in vitro matured (IVM) porcine oocytes was investigated in the present study. Cumulus-free IVM porcine oocytes were subjected either to SSV or SSV combined with a cytochalasin B (CB) pre-treatment (SSV + CB) or all steps of SSV but without cooling (toxicity control = TC; toxicity control with CB pre-treatment = TC + CB). Oocyte viability was evaluated by plasma membrane integrity and esterase activity measured by a combined staining with fluorescein diacetate, propidium iodide and Hoechst 33342. Surviving oocytes were parthenogenetically activated then cultured in vitro (IVC) for 6 days. The proportion of live oocytes after vitrification was significantly lower than that of the TC, TC + CB and the control groups, regardless of the CB pre-treatment. Treatment of oocytes with cryoprotectants did not decrease the rates of surviving oocytes. After activation of oocytes, the proportion of cleaved embryos was significantly higher in the SSV + CB ( P < 0.05) than that of the SSV group. Nevertheless, significantly more oocytes cleaved ( P < 0.05) in the TC, TC + CB and the control groups. On Day 6, the rate of blastocysts in the SSV and SSV + CB groups did not differ significantly. The number of oocytes developing to blastocyst and the mean number of blastomeres per embryo were significantly higher ( P < 0.05) in the TC, TC + CB and the control compared with that of the SSV and SSV + CB groups. To our knowledge, this is the first report on parthenogenetic development to blastocysts of porcine oocytes vitrified at the metaphase-II stage. Results indicate that the high concentrations of cryoprotectants were not harmful for in vitro development, and that CB pre-treatment may increase survival and development of SSV vitrified porcine oocytes.