Trichostatin A Improves Preimplantation Development of Bovine Cloned Embryos and Alters Expression of Epigenetic and Pluripotency Genes in Cloned Blastocysts

Abstract

ABSTRACT. We investigated the effects of exposure time and concentration of trichostatin A (TSA) on in vitro development and quality of bovine SCNT embryos. At multiple time points, the relative expression of genes related to pluripotency and reprogramming was analyzed in order to assess the quality of preimplantation embryos cultured in media with TSA using real-time PCR. Development into blastocysts was higher in 100 nM TSA than in controls (35.96 vs. 28.30%, P<0.05). Study of 100 nM TSA exposure time showed development into blastocysts was higher during both short-term and long-term exposure than in controls (36.17 and 34.04 vs. 23.45%), but there was no significant difference between TSA groups. Nanog expression in blastocysts after long-term TSA exposure was similar to that in IVF blastocysts and greater than in controls and short-term exposed embryos. The Oct4 levels in the short-term exposure group were similar to those of IVF blastocysts, while Oct4 expression in long-term exposed embryos was significantly higher than in other groups. Measurement of DNMT1 and HDAC1 in blastocysts showed a similar expression profile among IVF and TSA groups regardless of treatment duration. In conclusion, this study suggests that TSA treatment after SCNT in bovine embryos can improve in vitro development of embryos by increasing the blastocysts formation and positive reprogramming of the reconstructed embryo genome caused by downregulation of DNA methylation and up-regulation of pluripotency. The somatic cell nuclear transfer (SCNT) technique is promising for applications such as species preservation, livestock propagation and cell therapy for medical treatment [35,36]. Although cloning mammals by SCNT into oocyte cytoplasts have successfully produced offspring in a variety of species over the past decade, SCNT has low efficiency and a high frequency of developmental abnormalities [8,30]. One of the underlying problems in SCNT embryos is incomplete or incorrect status of epigenetic reprogramming such as DNA methylation or histone modification [28,44].For improving this acetylation reprogramming, SCNT studies have reported epigenetic reprogramming modification using histone deacetylase inhibitor in activation or culture of reconstructed oocytes. Epigenetic reprogramming represents an important feature of nuclear reprogramming in SCNT embryos [15,28]. Dynamic interactions exist between DNA methylation and acetylation in the aminoterminal domains of core histones in order to regulate DNA functions and control gene expression [22,34,42]. Indeed, the regulation of genes coordinating embryo and fetal development depends on important remodeling in chromatin structure and DNA [12,29].Trichostatin A (TSA) is a histone deacetylase inhibitor that reversibly binds the active catalytic site of HDACs causing acetylated histones to accumulate in cells [26] and reduces DNA methylation and expression of DNA methyltransferases (DNMTs) in reconstructed embryos [24,40]. Recently, treatment of SCNT cloned mouse embryos with TSA significantly ...

Supplement of autologous ooplasm into porcine somatic cell nuclear transfer embryos does not alter embryo development

“…The present study showed that the transcription level of OCT4 and CDX2 was significantly lower in both SCNT and AOT embryos than in IVF counterparts. In agreement with our results, the expression level of OCT4 was upregulated in IVF and in vivo blastocysts in comparison with SCNT counterparts (Kumar et al., ; Oh, Lee, Lee, & Lee, ; Xu et al., ). Epigenetic modifications play an important part in mammalian embryo development; appropriate modifications of both DNA methylation and histone acetylation are needed for normal development of embryos because the paternal and maternal genomes suffer apparent epigenetic DNA modifications upon fertilization to acquire totipotency for normal early embryo development (Cantone & Fisher, ).…”

“…Furthermore, it was suggested that DNA methylation pattern in SCNT embryos was closely correlated with lower embryo developmental potential because somatic nuclei did not suffer rapid DNA reorganization before the first mitosis (Beaujean et al., ). In regard to relevant gene expression for epigenetics, DNMT1 and HDAC1 were expressed higher in bovine SCNT embryos than in their IVF counterparts (Oh et al., ). In the present study, SCNT and AOT embryos showed apparently similar gene expression level regarding methylation and acetylation, but revealed higher expression of DNMT1 and DNMT3a than IVF counterparts.…”

Increasing of blastocyst rate and gene expression in co-culture of bovine embryos with adult adipose tissue-derived mesenchymal stem cells

“…Similarly, we also observed an increase in G6PDH expression, a gene related to carbohydrate metabolism and NADPH availability for steroid and fatty acid biosynthesis, as well as responsible for increasing the ribose 5-phosphate supply for nucleotide and nucleic acid synthesis [36][37][38]. Both genes are considered good markers of embryo quality short of the possibility of performing transfer and evaluating postimplantation development [39][40][41][42][43][44].…”

Combination of S‐adenosylhomocysteine and scriptaid, a non‐toxic epigenetic modifying reagent, modulates the reprogramming of bovine somatic‐cell nuclear transfer embryos

“…Another study reported that SPD has less deleterious effects than TSA on the development of SCNT embryos, even though it shares a common structure with TSA (Kim et al, ). A number of studies showed that the beneficial working concentration of TSA on bovine SCNT embryos was 1.0 µM (Jafarpour et al, ), 100 nM (Oh et al, ), 50 nM (Wang et al, ; Iager et al, ), and 5 nM (Cervera et al, ). On the other hand, the optimal concentration of SPD is 6 to 8 times higher than TSA (Su et al, ) versus 500 nM (Mao et al, ) and 5 nM (Akagi et al, ).…”