Effect of oocyte source and transport time on rates of equine oocyte maturation and cleavage after fertilization by ICSI, with a note on the validation of equine embryo morphological classification

Abstract

Production of equine embryos using intracytoplasmic sperm injection (ICSI) is rapidly gaining interest in the horse industry. Due to increasing client demand, equine practitioners with limited experience in embryology are attempting to set up clinical ICSI programs, with little success. We report here studies performed with the purpose of establishing an equine in vitro maturation (IVM)/ICSI program. We addressed three objectives: to determine (1) the effect of oocyte source (transvaginal follicle aspiration [TVA] vs. abattoir) on maturation, cleavage and blastocyst rates; (2) the impact of time of oocyte recovery (soon after death vs. delayed [median time 7.5 h]) on these parameters in abattoir-derived ovaries; and (3) the correlation of post-ICSI embryo morphology with histologically-confirmed nuclear status. Maturation rates were greater for TVA-derived than for abattoir-derived oocytes (67% vs. 32%, respectively; P <0.01). Duration of ovary transport did not affect oocyte maturation (34-48%) or cleavage rates (70-73%). Chromatin staining revealed that light-microscopic evaluation was accurate in determining oocyte maturation to metaphase II (14/15, 93%), but was not accurate in classification of either normal embryo cleavage (8/31 cleaved embryos possessed normal nuclei) or blastocyst formation (7/15 embryos classified as blastocysts were verified on staining). Early embryo and blastocyst viability was confirmed by transfer of some embryos into recipient mares. These findings indicate that oocyte source (TVA vs. abattoir) can affect results in an equine IVM/ICSI system, and suggest that new laboratories should use a systematic approach of comparison of nuclear chromatin staining with morphological classification to validate embryo development after ICSI.