Placental Function in Preeclampsia

描述(申请人提供)：过度炎症反应和氧化应激增加是先兆子痫(PE)妇女内皮功能改变的两个主要病理生理表现，PE是一种人类妊娠特有的多系统疾病。虽然PE的病因尚不清楚，但人们认为源于胎盘滋养层细胞(TCS)的“有毒”因子进入母体循环并导致EC功能障碍。在本项目的初始资助阶段，我们发现胎盘TCS来源的糜蛋白酶样蛋白酶(Chymase)负责上调EC P-选择素和E-选择素的表达，这两个分子是增强PE中EC炎症反应的关键分子。在这次续期拨款申请中，我们将进一步研究胎盘糜酶的作用，以及该酶在EC功能调节中的潜在细胞和分子机制，从而解释与PE相关的EC功能改变。我们的中心假设是胎盘TCS产生的糜乳酶不仅启动了EC的炎症表型，而且还诱导了EC的氧化应激和血管激活剂的释放，这是PE中EC功能障碍的特征。这一假说直接将胎盘滋养层细胞异常与内皮细胞功能障碍联系起来，将通过以下4个特定目标进行验证：1)鉴定正常和PE妊娠胎盘中的糜乳酶(类糜蛋白酶)，2)确定胎盘糜酶是否负责激活内皮细胞并诱导PE中的炎症表型，3)研究胎盘来源的糜乳酶是否诱导PE中的内皮氧化应激，以及4)探索糜乳酶是否诱导EC血管收缩刺激PE中的血管收缩。拟议的工作将使用子宫肌测长血管内皮细胞(UtMVECs)、脐带内皮细胞、TCS和VSMCs。用TC或TC条件培养液培养的内皮细胞将被用来研究胎盘糜酶在PE过程中引起EC功能改变的机制。血管内皮细胞与血管平滑肌细胞的共培养将被用来研究内皮细胞和循环因素对血管平滑肌细胞收缩能力的影响，模拟与PE相关的血管反应。还将研究乳糜酶诱导的EC功能障碍所涉及的蛋白质、基因和信号转导途径。这项拟议工作的结果应该会增加我们的认识，并建立胎盘TC活性和EC功能障碍在PE中的直接联系。

DESCRIPTION (provided by applicant): Exaggerated inflammatory response and increased oxidative stress are the two major pathophysiological manifestations of the altered endothelial function in women with preeclampsia (PE), a multi-system disorder unique to human pregnancy. Although the etiology of PE is not known, it is believed that 'toxic' factors derived from placental trophoblasts (TCs) enter the maternal circulation and induce EC dysfunction. In the initial funding period of this project, we identified that chymotrypsin-like protease (chymase) derived from placental TCs is responsible for upregulation of EC P-selectin and E-selectin expression, key molecules of increased EC inflammatory response in PE. In this renewal grant application, we will further investigate the role of placental chymase and the potential cellular and molecular mechanisms of this protease in regulation of EC function that account for the altered EC function associated with PE. Our central hypothesis is that chymase produced by placental TCs not only initiates an inflammatory phenotype in ECs but also induces EC oxidative stress and vasoactivator release that characterize the EC dysfunction in PE. This hypothesis directly links placental trophoblast aberration and EC dysfunction, which will be tested by experiments outlined under 4 specific aims: 1) to characterize the chymase (chymotrypsin-like protease) in placentas from normal and PE pregnancies, 2) to determine if placental chymase is responsible for activating ECs and inducing an inflammatory phenotype in PE, 3) to address whether placenta-derived chymase induces endothelial oxidative stress in PE, and 4) to explore if chymase-induced EC vasoconstrictor release that stimulates vasoconstriction in PE. The proposed work will employ uterine myometric vascular ECs (UtMVECs), umbilical cord ECs, TCs, and VSMCs. ECs cultured with TC or TC conditioned medium will be used to study mechanisms underlying the placental chymase-induced alterations in EC function during PE. Co-culture of ECs with VSMCs will be employed to study effects of ECs and circulating factors on VSMC contractility that mimic a vascular response of relevance to PE. The proteins, genes and signal transduction pathways involved in chymase-induced EC dysfunction will also be studied. Results obtained from the proposed work should enhance our knowledge and establish a direct link of placental TC activity and EC dysfunction in PE.