Targeting epigenetic nuclear reprogramming in aggregated cloned equine embryos

Abstract

Epigenetic perturbations during the reprogramming process have been described as the primary cause of the low efficiency of somatic cell nuclear transfer (SCNT). In this study, we tested three strategies targeting nuclear reprogramming to investigate effects on equine SCNT. First, we evaluated the effect of treating somatic cells with chetomin, a fungal secondary metabolite reported to inhibit the trimethylation on histone 3 lysine 9 (H3K9 me3). Second, caffeine was added to the culture medium during the enucleation of oocytes and before activation of reconstructed embryos as a protein phosphatase inhibitor to improve nuclear reprogramming. Third, we tested the effects of the histone deacetylase inhibitor trichostatin A (TSA) added during both activation and early embryo culture. Although none of these treatments significantly improved the developmental rates of the invitro aggregated cloned equine embryos, the first equine cloned foal born in Australia was produced with somatic cells treated with chetomin. The present study describes the use of chetomin, caffeine and TSA for the first time in horses, serving as a starting point for the establishment of future protocols to target epigenetic reprogramming for improving the efficiency of equine cloning. Cloning is an expensive and inefficient process, but has gained particular interest in the equine industry. In this study we explored different strategies to improve cloning efficiency and produced the first cloned foal born in Australia. Our data serve as a starting point for the establishment of future protocols for improving equine cloning efficiency.

Horse ooplasm supports in vitro preimplantation development of zebra ICSI and SCNT embryos without compromising YAP1 and SOX2 expression pattern

“…The low efficiency of SCNT has been primarily attributed to failures during nuclear reprogramming processes, which are necessary to restore the totipotency of the donor somatic cell nucleus [ 51 , 52 ]. In horses, several strategies were recently attempted to improve nuclear reprogramming, but non-significant differences were observed in development [ 53 ]. Moreover, failures during cell reprogramming could affect the expression pattern of proteins involved in cell-fate specification and compromise embryo viability [ 54 ].…”

State of the art of nuclear transfer technologies for assisting mammalian reproduction

“…Specifically, improvements in embryo developmental competence have been achieved using embryo aggregation. Our research group produced the first equine clone in Latin America in 2010 (Gambini et al, 2012(Gambini et al, , 2014 and the first equine clone born in Australia (Damasceno Teixeira et al, 2019). Embryo aggregation is currently used to produce equine clone's offspring on a large scale (Gambini & Maserati, 2017) and we estimate that about 1000 cloned horses have been produced worldwide.…”