Effects of Histone Deacetylase Inhibitor Oxamflatin on In Vitro Porcine Somatic Cell Nuclear Transfer Embryos
Effects of Histone Deacetylase Inhibitor Oxamflatin on In Vitro Porcine Somatic Cell Nuclear Transfer Embryos
复制标题
组蛋白脱乙酰酶抑制剂奥氨平对体外猪体细胞核移植胚胎的影响
DOI：
10.1089/cell.2013.0058
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发表时间：
2014-08-01
影响因子：
1.6
通讯作者：
Xiong, Yuanzhu
中科院分区：
文献类型：
--
作者：
Hou, Liming;Ma, Fanhua;Xiong, Yuanzhu
低克隆效率被认为是由体细胞核移植（SCNT）胚胎中分化供体细胞的不完全或异常表观遗传重编程引起的。Oxamflatin是一种新型的组蛋白去乙酰化酶抑制剂（HDACi），已被发现可以提高SCNT胚胎的体外和足月发育潜力。在本研究中，我们研究了oxamflatin处理对体外培养的猪SCNT胚胎的影响。我们的研究结果表明，在体外囊胚形成率的SCNT胚胎处理1 μ M oxamflatin为15小时postactivation是显着高于所有其他治疗。oxamflatin处理降低了克隆胚胎中的相对组蛋白去乙酰化酶（HDAC）活性，并导致组蛋白H3在赖氨酸9（AcH 3 K9）和组蛋白H4在赖氨酸5（AcH 4K 5）在原核，两细胞和四细胞阶段的超乙酰化水平，部分通过下调HDAC 1。通过oxamflatin抑制HDAC 6增加了早期SCNT胚胎有丝分裂细胞周期中α-微管蛋白的非组蛋白乙酰化水平。此外，我们证明，oxamflatin下调DNA甲基转移酶1（DNMT 1）的表达和全球DNA甲基化水平（5-甲基胞嘧啶）在两个细胞阶段的猪SCNT胚胎。研究发现，多能性相关基因POU 5 F1在奥沙利铂处理组中上调，其启动子区域的DNA甲基化趋势降低。处理oxamflatin没有改变位点特异性DNA甲基化水平的猪异染色质卫星DNA序列在囊胚阶段。同时，我们的研究结果表明，HDACi治疗可能有助于维持细胞凋亡相关元件的稳定状态，如乙酰化α-微管蛋白，这可能是早期SCNT胚胎中供体核重编程的关键决定因素。总之，奥沙姆黄素处理提高了猪SCNT胚胎的体外发育潜力。
Low cloning efficiency is considered to be caused by the incomplete or aberrant epigenetic reprogramming of differentiated donor cells in somatic cell nuclear transfer (SCNT) embryos. Oxamflatin, a novel class of histone deacetylase inhibitor (HDACi), has been found to improve the in vitro and full-term developmental potential of SCNT embryos. In the present study, we studied the effects of oxamflatin treatment on in vitro porcine SCNT embryos. Our results indicated that the rate of in vitro blastocyst formation of SCNT embryos treated with 1 mu M oxamflatin for 15 h postactivation was significantly higher than all other treatments. Treatment of oxamflatin decreased the relative histone deacetylase (HDAC) activity in cloned embryos and resulted in hyperacetylation levels of histone H3 at lysine 9 (AcH3K9) and histone H4 at lysine 5 (AcH4K5) at pronuclear, two-cell, and four-cell stages partly through downregulating HDAC1. The suppression of HDAC6 through oxamflatin increased the nonhistone acetylation level of alpha-tubulin during the mitotic cell cycle of early SCNT embryos. In addition, we demonstrated that oxamflatin downregulated DNA methyltransferase 1 (DNMT1) expression and global DNA methylation level (5-methylcytosine) in two-cell-stage porcine SCNT embryos. The pluripotency-related gene POU5F1 was found to be upregulated in the oxamflatin-treated group with a decreased DNA methylation tendency in its promoter regions. Treatment of oxamflatin did not change the locus-specific DNA methylation levels of Sus scrofa heterochromatic satellite DNA sequences at the blastocyst stage. Meanwhile, our findings suggest that treatment with HDACi may contribute to maintaining the stable status of cytoskeleton-associated elements, such as acetylated alpha-tubulin, which may be the crucial determinants of donor nuclear reprogramming in early SCNT embryos. In summary, oxamflatin treatment improves the developmental potential of porcine SCNT embryos in vitro.