Defined System for In Vitro Production of Porcine Embryos Using a Single Basic Medium

TLDR

Porcine blastocysts can be generated by IVF and IVC in chemically defined media, prompting investigation of basic media and macromolecular additives for IVM to establish a single‑medium system. The study aims to develop a defined in‑vitro embryo production system that uses one basic medium for IVM, IVF, and IVC. Immature oocytes were matured in either POM or mNCSU37 supplemented with 10 % porcine follicular fluid or 3 mg ml⁻¹ polyvinyl alcohol, then activated parthenogenetically or fertilized, and cultured in PZM‑5 for blastocyst development. Maturation in mNCSU37+PVA yielded lower metaphase‑II, cleavage, and blastocyst rates, whereas POM+PVA increased normal fertilization but reduced blastocyst cell numbers, yet the defined single‑medium protocol successfully produced porcine blastocysts.

Abstract

We have previously indicated that porcine blastocysts can be produced by in vitro fertilization (IVF) and culture (IVC) in chemically defined porcine gamete medium (PGM) and porcine zygote medium (PZM)-5, respectively, In the present study, the effects of basic media and macromolecular components on in vitro maturation (IVM) were investigated to develop a defined system for in vitro embryo production using a single basic medium through IVM, IVF and IVC. Porcine immature oocytes were matured in porcine oocyte medium (POM) or modified North Carolina State University (mNCSU) 37, which were supplemented with either 10% (v/v) porcine follicular fluid (pFF) or 3 mg/ml polyvinyl alcohol (PVA) as a macromolecular component (designated POM+pFF, POM+PVA, mNCSU37+pFF and mNCSU37+PVA). In the maturation with mNCSU37+PVA, the percentages of oocytes that reached the metaphase II stages were significantly lower than those in the other treatments. Following IVM with the above media, oocytes were treated with an electrical stimulus and cycloheximide for parthenogenetic activation and were cultured in PZM-5 for 5 days. The rates of cleavage and blastocyst formation of parthenogenetic oocytes were significantly lowered for maturation with mNCSU37+PVA compared with the other treatments, while there were no significant differences in the total numbers of cells in blastocysts among the treatments. Following IVF and IVC, the rates of penetration, male pronucleus formation, cleavage and blastocyst formation were significantly lower when oocytes were matured in mNCSU37+PVA than in other maturation media. The normal fertilization rate was significantly higher in POM+PVA compared with the other treatments, although the total number of cells in blastocysts was reduced with the addition of PVA to both POM and mNCSU37 compared with pFF supplementation. These results demonstrate that porcine blastocysts can be produced by the defined system using a single basic medium.