IMCD phosphodiesterase in pregnancy; role in ECFVE

描述(由申请人提供)：大的、持续的血浆容量扩张(PVE)是正常、成功怀孕的必要特征。不理想的PVE与不良的胎儿结局有关，并且是先兆子痫的一个组成部分。该项目的目标是确定正常妊娠PVE的主要机制，以便最终能够预防与PVE失败相关的妊娠并发症。这一建议的动机是正常妊娠的肾脏和血流动力学反应与不适当的PVE状态下的肾脏和血流动力学反应有几个明显的相似之处，如肾病或肝硬变。病理性钠滞留的主要肾脏机制是依赖cGMP作为第二信使的利钠机制失效，这是由于磷酸二酯酶(PDE)增加了cGMP的水解率。在这里，我们将检验一种假设，即妊娠期间肾小管PDE系统(特别是内髓集合管)选择性上调，并导致对钠尿刺激的广泛不耐受，如急性容量扩张和应用心钠素，从而导致PVE。我们还将确定PDE的异构体和在肾脏中的位置，这些PDE参与了对正常P的肾脏反应，并确定妊娠期间肾脏PDE活性增加是翻译事件还是翻译后事件造成的。我们将研究血管紧张素II和肾神经活动(主要的肾脏钠保留系统)是如何参与的；我们将检验妊娠PVE的“不足”假说，我们将确定是否母体激素孕酮、催乳素或松弛素是导致P的肾脏PDE活性和PVE增加的主要原因。最后，我们将在妊娠期长期局部抑制相关的PDE(S)，以确定对母体血流动力学和胎儿结局的影响。所有研究都将在大鼠身上进行，使用体内技术来测量肾脏的钠处理能力，以及对分离的肾小球、近端小管和内髓集合管细胞进行体外技术，包括酶活性分析、Western印迹、免疫细胞化学、Northern印迹或核糖核酸酶保护分析和原位杂交。

DESCRIPTION (provided by applicant): A large, sustained plasma volume expansion (PVE) is a necessary feature of normal, successful pregnancy. Suboptimal PVE is associated with adverse fetal outcome, and is a component of preeclampsia. The goal of this project is to determine the primary mechanism of the normal gestational PVE, so that ultimately, complications of pregnancy associated with failure to PVE can be prevented. This proposal is motivated by several marked similarities between the renal and hemodynamic responses to normal pregnancy, and those seen in states of inappropriate PVE, such as nephrosis or cirrhosis. The primary renal mechanism in pathological sodium retention is a failure of natriuretic mechanisms that rely on cGMP as second messenger, due to increased cGMP hydrolysis by phosphodiesterase (PDE). Here, we will test the hypothesis that selective upregulation of the renal tubular PDE system (particularly the inner medullary collecting duct, IMCD) occurs in pregnancy and causes widespread refractoriness to natriuretic stimuli, such as acute volume expansion and administered atrial natriuretic peptide, leading to PVE. We will also characterize the isoforms and location in the kidney of the PDEs involved in the renal response to normal P and determine whether increased renal PDE activity in pregnancy results from translational or posttranslational events. We will investigate how angiotensin II and renal nerve activity (major renal sodium retaining systems) are involved; we will test the "underfill" hypothesis of gestational PVE and we will determine whether the maternal hormones progesterone, prolactin or relaxin are primarily responsible for the increased renal PDE activity and PVE of P. Finally, we will chronically and locally inhibit the relevant PDE(s) during pregnancy to establish the impact on maternal hemodynamics and fetal outcome. All studies will be in rat, using a combination of in vivo techniques to measure renal sodium handling capacity and in vitro techniques on isolated glomeruli, proximal tubules and inner medullary collecting duct cells, to include enzyme activity assays, Western blot, immunocytochemistry, Northern blot or RNAse protection assay and in situ hybridization.