IGFBP-1 hyperphosphorylation in IUGR: Role of mTOR and CK2

胎儿宫内发育迟缓(IUGR)增加围产期并发症的风险，并易患上成人 疾病。胰岛素样生长因子I(IGF-I)是胎儿生长的关键调节因子，其生物利用度为 受与胰岛素样生长因子结合蛋白-1(IGFBP-1)的调节。我们和其他人已经提供了证据 IGFBP-1的磷酸化显著增加其与IGF-I和IGFBP-1的亲和力 过度磷酸化可能是IUGR生长受阻的重要机制。然而， 引起IUGR中IGFBP-1磷酸化的分子机制尚不清楚。中环 这一机械性建议中的假设是抑制mTOR信号转导和激活蛋白激酶 胎肝中的CK2构成营养缺乏和IGFBP-1升高之间的关键分子联系 体外和体内IUGR的分泌和磷酸化。我们的假设是基于我们的 令人信服的初步数据，包括证明(1)mTOR抑制诱导显著的IGFBP-1 蛋白激酶CK2的磷酸化和沉默取消IGFBP-1的过度磷酸化 (2)mTOR活性降低，而CK2表达和IGFBP-1表达降低 IUGR狒狒肝脏中的磷酸化水平增加。为了验证我们的假设，我们将研究HepG2细胞， 原代胎肝细胞，对照组和胎儿宫内发育迟缓(IUGR) 具体目标：在目标1中，我们将在HepG2细胞中使用基因沉默方法和药物抑制剂 和胎儿原代肝细胞机械地将mTOR信号与蛋白激酶的调节联系起来 CK2和IGFBP-1的磷酸化。此外，我们将检验增加的IGFBP-1的假设 氨基酸缺失引起的磷酸化是通过抑制mTOR和激活CK2来实现的。在AIM 2我们将使用一种成熟的胎儿宫内发育迟缓(IUGR)的狒狒模型，包括母体营养限制。我们会 检测胎肝组织中mTOR和CK2、IGFBP-1的表达及磷酸化 IGFBP-1在正常和IUGR狒狒胎儿中的血清浓度和磷酸化。意义：这是 这项工作有可能确定IUGR发生的新的分子机制。 此外，我们将利用高度相关的非人类灵长类动物模型，提供独特的胎儿 通过人体研究得不到的样本。创新：这项工作将提供一个创新的机制链接 在mTOR和IGF-I信号之间，调节细胞生长的两条关键通路。这两种技术的结合 体外培养的人肝癌细胞株和原代胎猴肝细胞的机制及肝组织的研究 从狒狒IUGR模型是翻译的和创新的。

Intrauterine growth restriction (IUGR) increases the risk for perinatal complications and predisposes for adult disease. Insulin-like growth factor I (IGF-I) is a key regulator of fetal growth, and IGF-I bioavailability is modulated by binding to IGF binding protein-1 (IGFBP-1). We and others have provided evidence that phosphorylation of IGFBP-1 markedly increases its affinity to bind IGF-I and that IGFBP-1 hyperphosphorylation may constitute an important mechanism by which growth is reduced in IUGR. However, the molecular mechanisms causing IGFBP-1 phosphorylation in IUGR are largely unknown. The central hypothesis in this mechanistic proposal is that inhibition of mTOR signaling and activation of protein kinase CK2 in the fetal liver constitutes a key molecular link between nutrient deprivation and increased IGFBP-1 secretion and phosphorylation in vitro and in IUGR in vivo. Our hypothesis has been formulated based on our compelling preliminary data including the demonstration that (1) mTOR inhibition induces marked IGFBP-1 phosphorylation and silencing of protein kinase CK2 abolishes IGFBP-1 hyperphosphorylation induced by mTOR inhibition in HepG2 cells and (2) mTOR activity is decreased whereas CK2 expression and IGFBP-1 phosphorylation are increased in the liver of IUGR baboons. To test our hypothesis, we will study HepG2 cells, primary fetal hepatocytes from baboons, and blood and liver tissue of control and IUGR baboon fetuses in two specific aims: In Aim 1 we will use gene silencing approaches and pharmacological inhibitors in HepG2 cells and fetal baboon primary hepatocytes to mechanistically link mTOR signalling to regulation of protein kinase CK2 and IGFBP-1 phosphorylation. In addition, we will test the hypothesis that the increased IGFBP-1 phosphorylation due to amino acid deprivation is mediated by inhibition of mTOR and activation of CK2. In Aim 2 we will use a well-established baboon model of IUGR involving maternal nutrient restriction. We will determine the activity of mTOR and CK2, IGFBP-1 expression and phosphorylation in fetal liver samples and IGFBP-1 serum concentrations and phosphorylation in control and IUGR baboon fetuses. Significance: This work has the potential to identify a novel molecular mechanism underlying the development of IUGR. Furthermore, we will utilize a highly relevant non-human primate model that provides unique access to fetal samples not available through human studies. Innovation: This work will provide an innovative mechanistic link between mTOR and IGF-I signaling, two critical pathways in the regulation of cell growth. The combination of mechanistic approaches in cultured HepG2 and primary fetal baboon hepatocytes with studies of liver tissue from a baboon IUGR model is translational and innovative.