Cui Xiang-Shun, Xu Yong-Nan, Shen Xing-Hui, Zhang Li-Qun, Zhang Jia-Bao, Kim Nam-Hyung
Center for Laboratory Animal, Jilin University, 5333 Xi'an Road, Changchun, Jilin Province, People's Republic of China.
Cell Reprogram. 2011 Apr;13(2):179-89. doi: 10.1089/cell.2010.0060.
Low efficiency of somatic cell nuclear transfer (SCNT) is attributed to incomplete reprogramming of transferred nuclei into oocytes. Trichostatin A (TSA), a histone deacetylase inhibitor, has been used to enhance nuclear reprogramming following SCNT. However, the molecular mechanism of TSA for the improvement of the preimplantation embryo and fetal development following SCNT is not known. The present study investigates embryo viability and gene expression of cloned bovine preimplantation embryos in the presence and absence of TSA compared to embryos produced by in vitro fertilization or parthenogenetic activation. Our results indicated that TSA treatment significantly improved total and inner cell mass (ICM) cell number and ratio of ICM:trophectoderm (TE) and also decreased the apoptotic index including total, ICM, and ratio of ICM:TE. Four apoptotic-related genes, Bcl-xL, survivin, Bcl2-associated X protein (Bax), and caspase 3 (Casp3), and four pluripotency/differentiation related genes, Oct4, SRY (sex determining region Y)-box 2 (Sox2), Cdx2, and colony-stimulating factor 1 receptor (Csf1r), were measured by real-time RT-PCR. TSA treatment resulted in the high expression of antiapoptotic gene Bcl-xL and low expression of pro-apoptotic gene Bax compared to untreated NT embryos, fertilized embryos, or parthenotes. Furthermore, mRNA expression of Cdx2 was higher in NT-TSA embryos than in NT and in vitro fertilization (IVF) counterparts. Additionally, low expression of microRNA (mir)-21 in NT embryos was enhanced following TSA treatment. These results suggest that TSA positively regulates nuclear reprogramming, and TSA may increased resistance or reduced signal for induction of apoptosis.
体细胞核移植（SCNT）效率低下归因于移植到卵母细胞中的细胞核重编程不完全。曲古抑菌素A（TSA）是一种组蛋白脱乙酰酶抑制剂，已被用于增强SCNT后的核重编程。然而，TSA改善SCNT后着床前胚胎和胎儿发育的分子机制尚不清楚。本研究调查了与体外受精或孤雌激活产生的胚胎相比，在有或没有TSA的情况下克隆牛着床前胚胎的胚胎活力和基因表达。我们的结果表明，TSA处理显著提高了总细胞数和内细胞团（ICM）细胞数以及ICM：滋养外胚层（TE）的比例，同时也降低了包括总细胞、ICM以及ICM：TE比例的凋亡指数。通过实时RT-PCR检测了四个凋亡相关基因，Bcl-xL、survivin、Bcl2相关X蛋白（Bax）和半胱天冬酶3（Casp3），以及四个多能性/分化相关基因，Oct4、性别决定区Y框2（Sox2）、Cdx2和集落刺激因子1受体（Csf1r）。与未处理的核移植胚胎、受精胚胎或孤雌胚胎相比，TSA处理导致抗凋亡基因Bcl-xL的高表达和促凋亡基因Bax的低表达。此外，NT-TSA胚胎中Cdx2的mRNA表达高于NT胚胎和体外受精（IVF）胚胎。此外，TSA处理后，NT胚胎中微小RNA（mir）-21的低表达得到增强。这些结果表明，TSA正向调节核重编程，并且TSA可能增加了抗凋亡能力或降低了诱导凋亡的信号。
