Combined Chaetocin/Trichostatin A Treatment Improves the Epigenetic Modification and Developmental Competence of Porcine Somatic Cell Nuclear Transfer Embryos

Abstract

Developmental defects in somatic cell nuclear transfer (SCNT) embryos are principally attributable to incomplete epigenetic reprogramming. Small-molecule inhibitors such as histone methyltransferase inhibitors (HMTi) and histone deacetylase inhibitors (HDACi) have been used to improve reprogramming efficiency of SCNT embryos. However, their possible synergistic effect on epigenetic reprogramming has not been studied. In this study, we explored whether combined treatment with an HMTi (chaetocin) and an HDACi (trichostatin A; TSA) synergistically enhanced epigenetic reprogramming and the developmental competence of porcine SCNT embryos. Chaetocin, TSA, and the combination significantly increased the cleavage and blastocyst formation rate, hatching/hatched blastocyst rate, and cell numbers and survival rate compared to control embryos. In particular, the combined treatment improved the rate of development to blastocysts more so than chaetocin or TSA alone. TSA and combined chaetocin/TSA significantly reduced the H3K9me3 levels and increased the H3K9ac levels in SCNT embryos, although chaetocin alone significantly reduced only the H3K9me3 levels. Moreover, these inhibitors also decreased global DNA methylation in SCNT embryos. In addition, the expression of zygotic genome activation- and imprinting-related genes was increased by chaetocin or TSA, and more so by the combination, to levels similar to those of in vitro-fertilized embryos. These results suggest that combined chaetocin/TSA have synergistic effects on improving the developmental competences by regulating epigenetic reprogramming and correcting developmental potential-related gene expression in porcine SCNT embryos. Therefore, these strategies may contribute to the generation of transgenic pigs for biomedical research.

Oroxin A reduces oxidative stress, apoptosis, and autophagy and improves the developmental competence of porcine embryos in vitro

“…Furthermore, for the aforementioned reasons, the desirable OA‐mediated impacts might be reflected in the enhancements of ex vivo developmental potential, molecular quality, mitochondrial functions and incidence of genomic DNA‐inherited genes to undergo reprogramming of their transcriptional activities in porcine and other mammalian in vitro‐produced embryos that have been created using a variety of advanced assisted reproductive technologies (ARTs). Among those ARTs, special importance should be assigned to in vitro fertilization (IVF) by standard gamete co‐incubation or intracytoplasmic sperm injection (Arraztoa et al, 2017; Casillas et al, 2018; Kagawa et al, 2022; Thongkittidilok et al, 2022) and cloning by somatic cell nuclear transfer (Deng et al, 2020; Gorczyca et al, 2021; Jeong et al, 2021; Wiater et al, 2021).…”

Luteolin supplementation during porcine oocyte maturation improves the developmental competence of parthenogenetic activation and cloned embryos

“…SCNT was performed as described previously, with modifications ( Jeong et al, 2020b , 2021 , 2017 ). MII oocytes in DPBS supplemented with 4 mg/mL BSA, 75 µg/mL penicillin G, 50 µg/mL streptomycin sulfate, and 7.5 µg/mL cytochalasin B were cut using a sharp pipette and then the first polar body and cytoplasm-containing chromosomes were removed using the squeezing method under an inverted microscope (DMI 3000B; Leica Microsystems, Wetzlar, Germany) equipped with a micromanipulator (NT-88-V3; Nikon Narishige, Tokyo, Japan).…”

“…Indirect immunofluorescence assay was performed as described previously ( Jeong et al, 2021 ). Oocytes or blastocysts were fixed in 4% paraformaldehyde overnight at 4 °C and washed three times for 10 min each in DPBS supplemented with 1 mg/mL PVA (DPBS-PVA).…”

— Invited Review — Reproductive technologies needed for the generation of precise gene-edited pigs in the pathways from laboratory to farm

“…Recently, by adding chaetocin to IVC medium for 24 h after activation, Jeong et al [ 77 ] improved reconstructed embryonic development, including the cleavage rate, blastocyst formation rate and cell number, and increased Oct4 , Nanog , Sox2 , Cdx2 , Bcl2 , and Bcl-x1 gene expression, but SUV39h1 and SUV39h2 (DNA methyltransferase genes) expression and H3K9me3 content were decreased. By supplying chaetocin and TSA at the same time, the reconstructed embryo development and the levels of H3K9me3 and H3K9ac in SCNT embryos were further improved; moreover, the expression of ZGA- and imprinting-related genes was also increased [ 78 ].…”