Zhou Chi, Dobrinsky John, Tsoi Stephen, Foxcroft George R, Dixon Walter T, Stothard Paul, Verstegen John, Dyck Michael K
Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, Alberta, Canada.
International Center for Biotechnology, Minitube of America, Mount Horeb, Wisconsin, United States of America.
PLoS One. 2014 Mar 14;9(3):e91728. doi: 10.1371/journal.pone.0091728. eCollection 2014.
The in vitro production of early porcine embryos is of particular scientific and economic interest. In general, embryos produced from in vitro Assisted Reproductive Technologies (ART) manipulations, such as somatic cell chromatin transfer (CT) and parthenogenetic activation (PA), are less developmentally competent than in vivo-derived embryos. The mechanisms underlying the deficiencies of embryos generated from PA and CT have not been completely understood. To characterize the altered genes and gene networks in embryos generated from CT and PA, comparative transcriptomic analyses of in vivo (IVV) expanded blastocysts (XB), IVV hatched blastocyst (HB), PA XB, PA HB, and CT HB were performed using a custom microarray platform enriched for genes expressed during early embryonic development. Differential expressions of 1492 and 103 genes were identified in PA and CT HB, respectively, in comparison with IVV HB. The "eIF2 signalling", "mitochondrial dysfunction", "regulation of eIF4 and p70S6K signalling", "protein ubiquitination", and "mTOR signalling" pathways were down-regulated in PA HB. Dysregulation of notch signalling-associated genes were observed in both PA and CT HB. TP53 was predicted to be activated in both PA and CT HB, as 136 and 23 regulation targets of TP53 showed significant differential expression in PA and CT HB, respectively, in comparison with IVV HB. In addition, dysregulations of several critical pluripotency, trophoblast development, and implantation-associated genes (NANOG, GATA2, KRT8, LGMN, and DPP4) were observed in PA HB during the blastocyst hatching process. The critical genes that were observed to be dysregulated in CT and PA embryos could be indicative of underlying developmental deficiencies of embryos produced from these technologies.
早期猪胚胎的体外生产具有特殊的科学和经济意义。一般来说，通过体外辅助生殖技术（ART）操作产生的胚胎，如体细胞染色质转移（CT）和孤雌生殖激活（PA），其发育能力低于体内来源的胚胎。PA和CT产生的胚胎发育缺陷的潜在机制尚未完全明确。为了表征CT和PA产生的胚胎中基因和基因网络的变化，我们使用了一个定制的微阵列平台对体内（IVV）扩张囊胚（XB）、IVV孵化囊胚（HB）、PA XB、PA HB和CT HB进行了比较转录组分析，该平台富集了早期胚胎发育过程中表达的基因。与IVV HB相比，分别在PA和CT HB中鉴定出1492个和103个基因的差异表达。PA HB中“eIF2信号传导”、“线粒体功能障碍”、“eIF4和p70S6K信号传导的调节”、“蛋白质泛素化”和“mTOR信号传导”通路下调。在PA和CT HB中均观察到Notch信号相关基因的失调。预测TP53在PA和CT HB中均被激活，因为与IVV HB相比，TP53的136个和23个调控靶点在PA和CT HB中分别显示出显著的差异表达。此外，在囊胚孵化过程中，PA HB中观察到几个关键的多能性、滋养层发育和着床相关基因（NANOG、GATA2、KRT8、LGMN和DPP4）的失调。在CT和PA胚胎中观察到失调的关键基因可能表明这些技术产生的胚胎存在潜在的发育缺陷。
