Improved fertilization and embryo development resulting in birth of live piglets after intracytoplasmic sperm injection and in vitro culture in a cysteine-supplemented medium

Abstract

The effects of cysteine treatment on fertilization rate, intracellular concentration of glutathione, and embryo development in vitro and after embryo transfer were examined following intracytoplasmic sperm injection (ICSI) of in vitro-matured porcine oocytes using a piezo drive unit. Culture of presumed zygotes after ICSI with 1.71–3.71 mM cysteine for 3–12 h improved ( P < 0.05) fertilization rates as compared to treatment with 0.57 mM cysteine or to controls (0 mM) (56 to 68%, 48%, 35%, respectively). Extension of treatment time with cysteine beyond 3 h did not further increase fertilization rates, suggesting that cysteine promoted early developmental events after ICSI (e.g. decondensation of sperm chromatin). There was no effect of cysteine supplementation on oocyte glutathione levels after ICSI. Pretreatment of spermatozoa for 3 h with 1.71 mM cysteine did not improve fertilization rates. The incidence of blastocysts formation when cultured in 1.71 mM cysteine for 3 h after ICSI was 31%, which was higher ( P < 0.05) than controls (18%). Transfer of 20–38 embryos cultured with 1.71 mM cysteine for 3 h after ICSI to each of seven recipients yielded three deliveries with an average litter size of 4.0. We concluded that cysteine supplementation for the first 3 h after ICSI improved fertilization and embryo development rates, with no influence on glutathione levels in oocytes, and that the cysteine-treated ICSI embryos developed to full term. The study also showed that porcine oocytes matured in a chemically defined medium had the ability for full-term development after piezo-ICSI without additional treatments for oocyte activation.