Project 4: The Placenta-specific Glucose Transporter Modulation: obesity, metabol

项目摘要/摘要--项目4 肥胖症的增加，特别是在工业化国家，已经达到了流行病的程度。在 美国三分之二的成年人(20岁及以上)超重或肥胖，并有身体质量指数或BMI 25公斤/平方米以上。这种流行病蔓延到怀孕人口，超过一半的美国人 女性怀孕时体重指数增加，因此肥胖是最常见的原因之一。 高危产科综合征。母亲超重和肥胖与胎儿发育障碍有关 发育导致新生儿结局不良，易患心血管疾病和代谢疾病 在以后的生活中会有障碍。胎盘是胎儿和母体循环之间的接口，是一种 它的主要功能之一是为胎儿提供营养。因为胎儿可获得的营养素的量是 胎盘是胎儿生长发育的重要决定因素，与胎儿生长过度有关。其作用机制 关于子宫内营养过剩、胎儿过度生长和晚年疾病发展之间的联系，人们知之甚少。 并被认为包括胎盘营养物质运输的变化，遗传和表观遗传变化。我们 建议开发一种基因疗法来解决这个问题，方法是使用一种独特的iggyBac转座子- 该系统是我所在本中心一期工程中开发的。这些精灵 质粒能够有效地将基因导入宿主基因组。此外，这种非病毒式方法 与基于病毒的系统相比，它具有许多优点，包括低毒和低免疫反应。我们 最近采用了一种体内微创方法(超声波靶向微泡破坏， UTMD)用于运送精灵载体，实现报告基因在小鼠肝脏中的长期表达 老鼠。UTMD可以介导生物活性分子的部位特异性递送到超声波可以到达的靶器官 比如胎盘。具体地说，我们将首先尝试通过稳定地敲除小鼠中Glut1的表达 通过UTMD将shRNAmir结构导入胎盘基因组。然后我们将确定 Glut1基因敲除对胎盘葡萄糖转运能力及胎儿生长和代谢的影响 子代的综合症。我们还将确定Glut1基因敲除对初级细胞功能的影响 人滋养细胞(PHT)。因为胎盘是一种生命周期有限的组织，对胎儿来说是必不可少的 发育但在出生后被丢弃，它非常适合通过基因改善胎儿的健康 修改，而不会引起与直接向宿主基因组转移基因相关的问题。这个 我们将在此应用程序中测试的假设是胎盘特异性的Glut1表达减少和 随之而来的葡萄糖转运到胎盘的减少将减轻胎儿过度生长和随后 喂食高脂肪食物的母鸡后代中的代谢综合征。此外，胎盘基因转移可能会起到 作为减少胎儿过度生长的早期干预策略。

PROJECT SUMMARY/ABSTRACT - Project 4 The increase in obesity, particularly in industrialized countries has taken on epidemic proportions. In the US two-thirds of adults (20 years of age and older) are overweight or obese, with a body mass index or BMI above 25kg/m2. This epidemic extends to the pregnant population where more than half of all American women enter pregnancy with an increased BMI and as a consequence obesity is one of the most common high-risk obstetric syndromes. Maternal overweight and obesity are associated with disturbances in fetal growth leading to poor neonatal outcomes and a predisposition for cardiovascular disease and metabolic disorders later in life. The placenta serves as an interface between the fetal and maternal circulation and one of its key functions is to supply nutrients to the fetus. As the quantity of nutrients available to the fetus is a significant determinant of fetal growth, the placenta has been implicated in fetal overgrowth. The mechanisms linking in utero nutrient excess, fetal overgrowth and disease development later in life are poorly understood and are thought to include alterations in placental nutrient transport, genetic and epigenetic changes. We propose to develop a gene therapy approach to address this problem using a unique piggyBac transposon- based transfection system that was developed at our institute in Phase I of this COBRE. These GENIE plasmids are able to efficiently introduce genes into the host genome. Moreover, this non-viral approach presents many advantages over virus-based systems, including low toxicity and low immune response. We have recently employed a minimally invasive in vivo method (Ultrasound Targeted Microbubble Destruction, UTMD) for the delivery of GENIE plasmids achieving long-term expression of a reporter gene in the liver of mice. UTMD can mediate site-specific delivery of bioactive molecules to ultrasound-accessible target organs such as the placenta. Specifically, we will first attempt to knockdown the Glut1 expression in mice by stably introducing shRNAmir constructs into the genome of the placenta via UTMD. We will then determine the effects of Glut1 knockdown on placental glucose transport capacity on fetal growth and the metabolic syndrome in the offspring. We will also determine the effects of Glut1 knockdown on the function of primary human trophoblast (PHT) cells. Since the placenta is a tissue with a finite life span, essential for fetal development but discarded after birth, it is well suited for improving fetal wellbeing through genetic modifications, without causing problems associated with gene transfer directly into the host genome. The hypothesis that we will test in this application is that a placenta-specific reduction in Glut1 expression and consequent decrease of glucose transport into the placenta will attenuate fetal overgrowth and subsequent metabolic syndrome in offspring from dams fed a high-fat diet. Furthermore, placental gene transfer may serve as an early intervention strategy to reduce fetal overgrowth.