Pro-inflammatory regulation of angiogenic gene expression in human trophoblast

描述（由申请人提供）：许多产科并发症，包括先兆子痫和宫内生长迟缓（母体和胎儿发病/死亡的两个更重要原因），与滋养层功能障碍和异常血管功能相关。然而，负责这些功能缺陷的分子机制知之甚少。滋养层细胞通常产生高水平的血管生成生长因子，胎盘生长因子（PGF）。PGF与其在不同细胞类型上表达的膜相关受体（Flt 1）结合，所述细胞类型包括内皮细胞和滋养层，其中PGF促进血管舒张， 血管生成和细胞存活。临床研究证实，PGF的表达显着减少，而剪接变体和可溶性形式的Flt 1（sFlt 1）显着增加，在先兆子痫之前和期间。由此产生的“抗血管生成”环境有助于定义先兆子痫的临床症状：高血压和蛋白尿。虽然我们和其他人已经表明，氧张力可以调节滋养层细胞的PGF和sFlt 1的表达，我们现在有令人信服的新数据表明，这些基因在人类滋养层细胞的促炎途径激活，也是一种常见的先兆子痫发生类似的失调。重要的是，这些效应与氧张力无关。本研究的目的是阐明促炎通路（NFkB）激活诱导人滋养层细胞PGF和sFlt 1异常表达的分子机制。我们将使用与人胎盘离体研究相关的体外基因操作方法的组合，以实现以下特定目标。我们已经表明，GCM 1，胎盘特异性转录因子，是一个主要的调节fPGF在滋养层的表达。Aim 1将描述激活的NF κ B亚基通过改变GCM 1活性直接影响PGF转录的分子机制。目的2将确定转录和转录后调节机制介导的促炎途径激活，增加滋养层sFlt 1的表达。目的3将利用离体人胎盘培养物来概括促炎途径激活后正常绒毛中PGF和sFlt 1表达的表达差异。这些研究的结果将提供有关促炎信号传导（不依赖于氧张力）调节人类滋养层中PGF和sFlt 1表达平衡的能力的新机制信息。总的来说，这些方法将有助于更好地了解机制，有助于血管功能正常和灌注受损的怀孕，并可能提供新的治疗途径的治疗。我们还期望从这些研究中获得的信息将对血管生物学具有广泛的适用性，因为成人血管生成的部位与炎症相关。 公共卫生相关性：妊娠期间血管形成和/或功能不足会导致许多产科并发症，最常见的是先兆子痫。众所周知，胎盘促血管生成蛋白和抗血管生成蛋白的产生改变调节血管功能，并直接导致全身性疾病。这种不平衡是如何发生的尚不清楚。本申请将开始确定负责胎盘细胞产生的两种突出的促血管生成因子和抗血管生成因子的异常表达的新分子机制。

DESCRIPTION (provided by applicant): Many obstetrical complications, including preeclampsia and intrauterine growth retardation, two of the more significant causes of maternal and fetal morbidity/mortality, are associated with trophoblast dysfunction and aberrant vascular function. However, the molecular mechanisms responsible for these functional defects are poorly understood. Trophoblasts normally produce high levels of the angiogenic growth factor, placenta growth factor (PGF). PGF binds to its membrane associated receptor (Flt1) expressed on different cell types including endothelial cells and trophoblast where it promotes vasodilation, angiogenesis and cell survival. Clinical studies have confirmed that expression of PGF is significantly reduced; while a splice variant and soluble form of Flt1 (sFlt1) is dramatically increased, prior to and during preeclampsia. The resulting "anti-angiogenic" environment contributes to the clinical symptoms that define preeclampsia: hypertension and proteinuria. Although we and others have shown that oxygen tension can regulate trophoblast expression of PGF and sFlt1, we now have convincing novel data indicating similar dysregulation of these genes in human trophoblast by pro-inflammatory pathway activation, also a common occurrence in preeclampsia. Importantly, these effects are independent of oxygen tension. The goal of our studies is to delineate the molecular mechanisms by which pro-inflammatory pathway (NFkB) activation induces aberrant PGF and sFlt1 expression in human trophoblast. We will use a combination of in vitro gene manipulation approaches correlated with ex vivo studies of human placentae in the following specific aims to accomplish this goal. We have shown that GCM1, a placenta-specific transcription factor, is a principal regulator fPGF expression in trophoblast. Aim1will characterize the molecular mechanisms by which activated NFkB subunits influence PGF transcription directly by altering GCM1 activity. Aim 2 will determine transcriptional and post transcriptional regulatory mechanisms mediated via pro-inflammatory pathway activation that increases trophoblast sFlt1 expression. Aim 3 will utilize ex vivo human placentae cultures to recapitulate expression differences in PGFandsFlt1 expression in normal villi following pro-inflammatory pathway activation. Results from these studies will provide novel mechanistic information regarding the ability of pro- inflammatory signaling, independent of oxygen tension, to regulate the balance of PGF and sFlt1 expression in human trophoblast. Collectively, these approaches will facilitate a better understanding of mechanisms that contribute to vascular function during normal and perfusion compromised pregnancies and may provide new therapeutic avenues for treatment. We also expect that information gained from these studies will have broad applicability to vascular biology since sites of angiogenesis in adults are associated with inflammation. PUBLIC HEALTH RELEVANCE: Inadequate blood vessel formation and/or function during pregnancy contributes to a number of obstetrical complications, the most common being preeclampsia. It is well accepted that altered production of pro- and anti-angiogenic proteins by the placenta regulate blood vessel function and directly contributes to systemic disease. How this imbalance occurs is not known. This application will begin to determine the novel molecular mechanisms responsible for aberrant expression of two prominent pro- and anti-angiogenic factors produced by placental cells.