Production of piglets from in vitro-produced blastocysts by ultrasound-guided ovum pick-up from live donors

Abstract

The objective of this research was to develop a system for piglet production by transvaginal ultrasound-guided ovum pick-up (OPU), in vitro production (IVP) of embryos and embryo transfer. First, to establish a culture system for a small number of oocytes or embryos, we evaluated the effect of different incubation volumes and culture densities on fertilizing ability and developmental competence in vitro. Porcine oocytes derived from slaughterhouse ovaries were matured, fertilized and then cultured in vitro in groups as follows: 50 oocytes in 500 μL medium for IVM, 20 oocytes in 100 μL medium for IVF and 20 embryos in 40 μL medium for IVC (Group I); 20 in 100 μL for IVM, 20 in 100 μL for IVF and 20 in 40 μL for IVC (Group II); and 10 in 100 μL for IVM, 10 in 100 μL for IVF and 10 in 40 μL for IVC (Group III). Percentages of sperm penetration, cleavage and blastocyst formation did not differ among the groups. Second, to increase the collection efficiency of porcine oocytes by transvaginal ultrasound-guided OPU, the effects of aspiration pressure on follicular oocyte collection were assessed. Oocytes were aspirated from ovaries of live sows using 80 or 100 mmHg. The recovered oocytes were divided into four categories according to the surrounding cumulus cells and quality of oocytes. The number of oocytes recovered using 100 mmHg pressure was significantly higher than with 80 mmHg pressure. However, there were no significant differences in the population of oocytes grouped by the morphological criteria, number of blastocysts per session and the total cell number in blastocysts between the two vacuum pressures. Finally, 81 oocytes obtained by OPU from five donor sows were subjected to IVP and 47 transferable embryos (9.4 ± 4.0 [mean ± SD] morulae/blastocysts per session) were obtained at 5 days after IVF. When they were transferred into five recipient gilts (5–16 embryos per recipient), three of five recipients became pregnant and farrowed a total of 12 live piglets. The present results demonstrate that porcine blastocysts can be produced by OPU-IVP and develop to full term after embryo transfer.