The placenta facilitates the transport of nutrients to the fetus, removal of waste products from the fetus, immune protection of the fetus and functions as an endocrine organ, thereby determining the environment for fetal growth and development. Additionally, the placenta is a highly metabolic organ in itself, utilizing a majority of the oxygen and glucose derived from maternal circulation. Consequently, optimal placental function is required for the offspring to reach its genetic potential in utero. Among ruminants, pregnant sheep have been used extensively for investigating pregnancy physiology, in part due to the ability to place indwelling catheters within both maternal and fetal vessels, allowing for steady-state investigation of blood flow, nutrient uptakes and utilization, and hormone secretion, under non-stressed and non-anesthetized conditions. This methodology has been applied to both normal and compromised pregnancies. As such, our understanding of the in vivo physiology of pregnancy in sheep is unrivalled by any other species. However, until recently, a significant deficit existed in determining the specific function or significance of individual genes expressed by the placenta in ruminants. To that end, we developed and have been using in vivo RNA interference (RNAi) within the sheep placenta to examine the function and relative importance of genes involved in conceptus development (PRR15 and LIN28), placental nutrient transport (SLC2A1 and SLC2A3), and placenta-derived hormones (CSH). A lentiviral vector is used to generate virus that is stably integrated into the infected cell’s genome, thereby expressing a short-hairpin RNA (shRNA), that when processed within the cell, combines with the RNA Induced Silencing Complex (RISC) resulting in specific mRNA degradation or translational blockage. To accomplish in vivo RNAi, day 9 hatched and fully expanded blastocysts are infected with the lentivirus for 4 to 5 h, and then surgically transferred to synchronized recipient uteri. Only the trophectoderm cells are infected by the replication deficient virus, leaving the inner cell mass unaltered, and we often obtain ~70% pregnancy rates following transfer of a single blastocyst. In vivo RNAi coupled with steady-state study of blood flow and nutrient uptake, transfer and utilization can now provide new insight into the physiological consequences of modifying the translation of specific genes expressed within the ruminant placenta.

中文翻译：

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反刍动物胎盘功能和生理学的体内研究——综述


胎盘有利于向胎儿输送营养物质、清除胎儿体内的废物、保护胎儿的免疫以及内分泌器官的功能，从而决定胎儿生长发育的环境。此外，胎盘本身就是一个高度代谢的器官，利用来自母体循环的大部分氧气和葡萄糖。因此，后代需要最佳的胎盘功能才能在子宫内发挥其遗传潜力。在反刍动物中，怀孕的绵羊已被广泛用于研究怀孕生理学，部分原因是能够在母体和胎儿血管内放置留置导管，从而可以对血流、营养吸收和利用以及激素分泌进行稳态研究。在无压力和非麻醉的条件下。该方法已应用于正常妊娠和受损妊娠。因此，我们对绵羊怀孕体内生理学的了解是任何其他物种都无法比拟的。然而，直到最近，在确定反刍动物胎盘表达的单个基因的特定功能或意义方面还存在显着的缺陷。为此，我们开发并一直在绵羊胎盘内使用体内 RNA 干扰 (RNAi) 来检查参与孕体发育（PRR15 和 LIN28）、胎盘营养转运（SLC2A1 和 SLC2A3）以及相关基因的功能和相对重要性。胎盘源性激素（CSH）。 慢病毒载体用于生成稳定整合到受感染细胞基因组中的病毒，从而表达短发夹 RNA (shRNA)，当在细胞内加工时，该病毒与 RNA 诱导沉默复合物 (RISC) 结合，产生特定的 mRNA降解或翻译受阻。为了完成体内 RNAi，第 9 天孵化并完全扩张的囊胚用慢病毒感染 4 至 5 小时，然后通过手术转移到同步受体子宫。只有滋养外胚层细胞被复制缺陷型病毒感染，内细胞团保持不变，并且在单个囊胚移植后我们通常获得约 70% 的妊娠率。体内 RNAi 与血流和营养吸收、转移和利用的稳态研究相结合，现在可以为改变反刍动物胎盘内表达的特定基因的翻译的生理后果提供新的见解。