Department of Animal Science, Faculty of Veterinary Sciences, Universidad de Concepción, Campus Chillán, Avenida Vicente Méndez 595, Chillán, Chile.
Theriogenology. 2010 Jul 1;74(1):45-59. doi: 10.1016/j.theriogenology.2009.12.018. Epub 2010 Mar 1.
Somatic cloning in cattle is associated with impaired embryo development, caused by inappropriate epigenetic reprogramming during embryogenesis; however, there is a paucity of data regarding gene expression at the critical elongation and peri-implantation stages. The objective of the present study was to identify genes differentially expressed in bovine cloned embryos at Day 17 of development (Day 0=day of nucleus transfer or IVF). Day 7 blastocysts (Hand Made Cloned or IVP) were transferred to recipient cattle and collected at Day 17. The efficiency of recovery of elongated embryos was similar, however cloned embryos elongated less than IVP embryos (91.8+/-45.8 vs. 174+/-50mm) and fewer had embryonic discs (63 vs. 83%). Qualitative and quantitative PCR detected expression of OCT4, NANOG, IFNtau, EOMES, FGF4, SOX2, and CDX2 in all IVP embryos. In most cloned embryos, NANOG and FGF4 were absent (verified by qPCR); NANOG, EOMES, and FGF4 were underexpressed, whereas IFNtau was overexpressed in cloned embryos. Based on qPCRs, other genes, i.e., SPARC, SNRB1, and CBPP22, were down-regulated in cloned embryos, whereas HSP70 and TDKP1 were overexpressed. In bovine microarrays, 47 genes (3.6%) were deregulated in cloned embryos, including several involved in trophoblast growth and differentiation. In conclusion, we inferred that these data were indicative of incomplete epigenetic reprogramming after cloning; this could lead to aberrant gene expression and subsequently early pregnancy loss. There was an apparent association between incomplete morphological elongation and aberrant reprogramming of a subset of genes critical for early embryonic development.
牛的体细胞核移植克隆与胚胎发育受损有关，这是由于胚胎发生过程中表观遗传重编程不当所致；然而，关于在关键的延长和植入前阶段的基因表达，数据很少。本研究的目的是鉴定在牛克隆胚胎发育第 17 天（0 天=核转移或 IVF 日）差异表达的基因。第 7 天的囊胚（手工克隆或 IVP）被转移到受体牛中，并在第 17 天收集。伸长胚胎的回收率效率相似，但是克隆胚胎的伸长程度低于 IVP 胚胎（91.8+/-45.8 对 174+/-50mm），并且具有胚胎盘的胚胎较少（63 对 83%）。定性和定量 PCR 检测了所有 IVP 胚胎中 OCT4、NANOG、IFNtau、EOMES、FGF4、SOX2 和 CDX2 的表达。在大多数克隆胚胎中，NANOG 和 FGF4 缺失（qPCR 验证）；NANOG、EOMES 和 FGF4 表达下调，而 IFNtau 在克隆胚胎中表达上调。基于 qPCR，其他基因，即 SPARC、SNRB1 和 CBPP22，在克隆胚胎中下调，而 HSP70 和 TDKP1 表达上调。在牛微阵列中，47 个基因（3.6%）在克隆胚胎中失调，包括一些参与滋养层生长和分化的基因。总之，我们推断这些数据表明克隆后表观遗传重编程不完全；这可能导致异常的基因表达，随后导致早期妊娠丢失。在不完全的形态伸长和对一组对早期胚胎发育至关重要的基因的异常重编程之间存在明显的关联。
