Department of Animal Science, North Carolina State University, Raleigh, NC, USA.
Theriogenology. 2010 Oct 15;74(7):1288-95. doi: 10.1016/j.theriogenology.2010.05.035. Epub 2010 Aug 5.
The objective of this study was to determine the effects of in vitro embryo production on physical development and levels of expression of mRNAs for insulin-like growth factor (IGF) ligands (IGF1, IGF2), their receptors (IGF1R, IGF2R), and IGF binding protein-2 (IGFBP2) in bovine fetuses during early gestation. In vivo embryos were recovered from superovulated Holstein cows. For production of embryos in vitro, Holstein oocytes were matured, fertilized, and subsequently cultured in M199 with 10% serum to 168 hpi. On Day 70 of gestation, fetuses (in vivo, n = 14; in vitro, n = 13) were recovered, serum samples collected, and physical measurements recorded. Semi-quantitative RT-PCR assays were used to determine the levels of expression of mRNAs for IGF1, IGF2, IGF1R, and IGF2R in fetal liver and skeletal muscle. Western blots were used to assess levels of IGFBP2 in fetal serum. Fetal body weight did not differ with treatment; however, production of embryos in vitro was associated with decreased crown-nose length and a tendency for increased paired kidney weight, which became significant when expressed on a per bodyweight basis. There was no effect of treatment on levels of IGFBP2 in fetal serum. Levels of IGF1 mRNA in fetal liver were decreased (P < 0.001) in the in vitro group. Levels of IGF2R mRNA in both liver and skeletal muscle were also decreased (P < 0.01) in fetuses from the in vitro group. In summary, fetuses at Day 70 of gestation from embryos produced in vitro had shortened crown-nose length and increased kidney weight on a per bodyweight basis, as well as decreased expression of mRNAs for IGF1 in liver and IGF2R in both liver and skeletal muscle, compared with fetuses from embryos produced in vivo. In conclusion, in vitro embryo culture was associated with subtle changes in fetal development as well as altered expression of both imprinted and non-imprinted genes.
本研究旨在确定体外胚胎生产对牛胎儿早期妊娠中物理发育和胰岛素样生长因子 (IGF) 配体 (IGF1、IGF2)、其受体 (IGF1R、IGF2R) 和 IGF 结合蛋白-2 (IGFBP2) mRNA 表达水平的影响。体内胚胎从超排荷斯坦奶牛中回收。为了进行体外胚胎生产，将荷斯坦牛卵母细胞成熟、受精，随后在含有 10%血清的 M199 中培养至 168 hpi。在妊娠第 70 天，回收胎儿（体内，n=14；体外，n=13），采集血清样本并记录物理测量值。半定量 RT-PCR 检测用于确定胎儿肝脏和骨骼肌中 IGF1、IGF2、IGF1R 和 IGF2R mRNA 表达水平。Western blot 用于评估胎儿血清中 IGFBP2 的水平。胎儿体重与处理无关；然而，体外胚胎生产与冠鼻长度缩短和双侧肾脏重量增加有关，当按体重表示时，这种增加具有统计学意义。处理对胎儿血清中 IGFBP2 的水平没有影响。体外组胎儿肝脏中 IGF1 mRNA 水平降低（P<0.001）。体外组肝脏和骨骼肌中 IGF2R mRNA 水平也降低（P<0.01）。总之，与体内胚胎生产的胎儿相比，体外生产的胚胎 70 日龄胎儿的冠鼻长度缩短，体重基础上肾脏重量增加，肝脏中 IGF1 mRNA 表达减少，肝脏和骨骼肌中 IGF2R mRNA 表达减少。总之，体外胚胎培养与胎儿发育的细微变化以及印记和非印记基因的表达改变有关。
