Dissecting gene dysregulation at the maternal-fetal interface in preeclampsia

描述（由申请人提供）：我们正在检验一种假设，即先兆子痫（PE）患者胎盘主要滋养层（TB）亚群的整体基因表达谱将导致鉴定在该综合征中起重要作用的新分子。这一策略的动力是发现PE始终与某些胎盘病理学相关。细胞滋养层细胞（CTB）的间质成分通常仅限于浅表蜕膜。同样地，血管内侵犯在涉及的螺旋小动脉的数量和它们被修改的程度方面受到限制。来自受影响患者胎盘的合胞体滋养细胞（STB）也有明显的变化，如合胞体结。这些改变伴随着分子变化，其中许多已经通过分析PE中通常在TB功能方面发挥重要作用的分子的表达来鉴定。然而，我们已经开始应用无偏的方法，以获得更全面的了解肺结核基因表达的变化，发生在PE。这种方法承认我们对这种综合征的胎盘成分的理解有限。最近，我们使用转录组学方法来分析CTB基因在各种重度PE（sPE）中的表达。具体地，将从sPE患者和对照女性的胎盘分离的CTB培养48小时以允许分化/侵袭。微阵列分析揭示了一套CTB基因的sPE相关上调，在培养期结束时，恢复到对照水平。它们包括以前与PE相关的因子和许多新分子，包括血管生成调节因子SEMA 3B。我们继续表明，这种神经纤毛蛋白-1和-2配体的水平升高表型模拟了PE对CTB的许多作用，其机制包括通过PI 3 K/AKT和GSK 3 α/β途径下调VEGF信号传导。在sPE CTB中观察到相同的变化。我们的数据支持的理论，在PE中，在体内环境损害CTB分化/侵袭，差异表达的分子有助于机制，和临床症状是由患者特异性因素决定。在确定了sPE中侵袭性CTB的基因特征后，我们现在提出对其他三种受影响的TB人群（STB、合胞结和血管内CTB）进行无偏倚分析。具体而言;我们将使用激光显微切割从PE、sPE和对照胎盘中分离这些细胞，并使用全局转录谱分析方法来鉴定失调基因（Aim 1）。我们将使用我们的新细胞培养模型，人TB祖细胞，以确定所观察到的变化的功能意义（目的2）。我们认为，这些实验将产生新的信息胎盘缺陷的分子基础，在PE。这些发现也可能具有重要的转化潜力，例如，失调分子的一个子集可能是循环生物标志物和/或改善胎盘功能的治疗靶点，我们的数据表明这是可能的。

DESCRIPTION (provided by applicant): We are testing the hypothesis that global gene expression profiling of the major trophoblast (TB) subpopulations of placentas from preeclampsia (PE) patients will lead to the identification of novel molecules that play important roles in this syndrome. The impetus for this strategy is the finding that PE is consistently associated with certain placental pathologies. The interstitial component of cytotrophoblast (CTB) invasion is frequently restricted to the superficial decidua. Likewise, endovascular invasion is constrained in terms of the number of spiral arterioles that are involved and the extent to which they are modified. Syncytiotrophoblasts (STBs) from placentas of affected patients also have overt changes such as syncytial knots. These alterations are accompanied by molecular changes, many of which have been identified by profiling, in PE, the expression of molecules that normally play important roles in terms of TB functions. However, we have begun to apply unbiased approaches to gain a more comprehensive understanding of changes in TB gene expression that occur in PE. This approach acknowledges our limited understanding of the placental component of this syndrome. Recently, we used a transcriptomics approach to profile CTB gene expression in various severe forms of PE (sPE). Specifically, CTBs that were isolated from the placentas of sPE patients and control women were cultured for 48 h to allow differentiation/invasion. Microarray analyses revealed sPE-associated upregulation of a suite of CTB genes that, by the end of the culture period, returned to control levels. They included factors previously associated with PE and many novel molecules, including the angiogenic regulator, SEMA3B. We went on to show that elevating levels of this neuropilin-1 and -2 ligand phenocopied many of the effects of PE on CTBs by mechanisms that include downregulating VEGF signaling through the PI3K/AKT and GSK3α/β pathway. The same changes were observed in sPE CTBs. Our data support the theory that, in PE, the in vivo environment impairs CTB differentiation/invasion, the differentially expressed molecules contribute to the mechanisms, and that the clinical signs are determined by patient-specific factors. Having identified a genetic signature for invasive CTBs in sPE, we now propose an unbiased analysis of the three other affected TB populations, STBs, syncytial knots and endovascular CTBs. Specifically; we will use laser microdissection to isolate these cells from PE, sPE and control placentas and a global transcriptional profiling approach to identify the dysregulated genes (Aim 1). We will employ our new cell culture model, human TB progenitor cells, to determine the functional significance of the observed changes (Aim 2). We think that these experiments will yield new information about the molecular bases of placental defects in PE. The findings could also have significant translational potential, e.g., a subset of the dysregulated molecules could be circulating biomarkers and/or therapeutic targets for improving placental function, which our data suggest is possible.