Modeling preeclampsia in mice with inducible placenta-specific gene expression

具有可诱导胎盘特异性基因表达的小鼠先兆子痫模型

• 批准号: 8191562
• 负责人: Nihar R Nayak
• 金额: $ 24.05万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-01 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8191562
• 关键词: Address; Adenoviruses; Adverse effects; Affect; Animal Model; Animals; Biological Markers; Biological Models; Bioluminescence; Blood; Blood Circulation; Blood Pressure; Cells; Cessation of life; Clinical; Clinical Research; Data; Development; Disease; Doxycycline; Endoglin; Etiology; Fetus; Firefly Luciferases; Functional disorder; Gene Delivery; Gene Expression; Gene Expression Profiling; Gene Expression Regulation; Genes; Giant Cells; Growth Factor Overexpression; Hemorrhage; Human; Image; Imaging Techniques; Kidney; Laboratories; Lentivirus Vector; Life; Mediating; Mediator of activation protein; Methods; Modeling; Monitor; Morbidity - disease rate; Mothers; Mus; Pathology; Phenotype; Physiological; Placenta; Placenta Diseases; Placentation; Play; Pre-Eclampsia; Pregnancy; Pregnancy Complications; Production; Public Health; Regulation; Reporting; Risk; Rodent; Role; Serum; Site; Staging; Stimulus; Subfamily lentivirinae; Symptoms; System; Techniques; Termination of pregnancy; Testing; Tetanus Helper Peptide; Thinking; Time; Tissues; Transgenes; Up-Regulation; Vascular Endothelial Growth Factor Receptor-1; Vascular Endothelial Growth Factors; Woman; Work; adverse outcome; angiogenesis; base; blastocyst; cell type; clinical phenotype; design; enhanced green fluorescent protein; fetal; in vivo; knock-down; molecular imaging; mortality; novel; novel strategies; overexpression; pregnant; prevent; promoter; protein function; research study; small hairpin RNA; therapy development; transgene expression; trophoblast; viral gene delivery

DESCRIPTION (provided by applicant): Preeclampsia is a serious complication of pregnancy, affecting both the mother and fetus and causing an estimated 14% of pregnancy-related maternal deaths worldwide. Despite its large public health impact, however, its causes are still poorly understood, and there are no known treatments that effectively alleviate the risks for both mother and fetus. Based on clinical studies in humans and experimental studies in rodents, it has been suggested that the abnormal elevation of the circulating levels of soluble fms-like tyrosine kinase 1 (sFlt- 1) contributes to the systemic endothelial dysfunction and clinical manifestations of the disease through its antagonism of vascular endothelial growth factor (VEGF) activity. As a result, it is believed that VEGF treatment may reverse the preeclamptic phenotype caused by high sFlt-1 levels in the maternal circulation. However, in preliminary studies, we found, surprisingly, that overexpression of VEGF in pregnant mice results in elevated blood pressure, increased serum levels of sFlt-1 and soluble endoglin (sEng), and histological changes in the kidneys, similar to the clinical findings of PE in humans and animal models. VEGF has been shown to stimulate sFlt-1 production in several cell types and placental explant cultures in multiple contexts. However, the roles of placental VEGF at different stages of pregnancy, the significance of regulation of its local activity by placental production of sFlt-1, and its potential role in regulating sFlt-1 production in the placenta have never been examined, primarily due to lack of suitable animal models to test the placenta-specific effects of VEGF. We hypothesize that VEGF activity in the placenta is regulated in a placental developmental stage- specific manner during pregnancy through local production of sFlt-1, and placental VEGF levels may be a primary stimulus for increased production of sFlt-1 in this tissue. Studies in this proposal are designed, first, to develop a novel, inducible, placenta-specific gene expression system, with the ability to monitor gene expression throughout pregancy by live in vivo imaging. This technique will be based on established methods for viral delivery of genes specifically to placental tissue, effective control of consistency in gene expression levels, and a tightly controlled inducible promoter which allows expression to be rapidly switched both on and off. We will then examine the effects of different levels of placental VEGF expression at various time points during pregnancy using this inducible system. To define the physiological role of sFlt-1 during different stages of pregnancy, we will selectively knock down sFlt-1 expression in the placenta using placenta-specific expression of a short hairpin RNA (shRNA) targeted against sFlt-1. The results of these experiments will delineate, for the first time, the local interactions of VEGF and sFlt-1 in regulating VEGF activity in the placenta. Additionally, this entirely novel approach, using a newly-developed, inducible, placenta-specific gene regulation system, will greatly facilitate development of new animal models for the study of both placental disease and normal placental development and will greatly enable the development of therapies for complications of pregnancy. PUBLIC HEALTH RELEVANCE: This work addresses the fundamental causes of the pregnancy complication preeclampsia, which affects 5-7% of pregnancies worldwide and is a leading cause of maternal and fetal morbidity and mortality. It develops a new method to study the roles of different factors in placental functions and diseases, using mice as a model system, specifically addressing the physiological role and mechanism of regulation of soluble fms-like tyrosine kinase 1 (sFlt-1), a soluble form of the vascular endothelial growth factor receptor 1 (Flt-1), which is found at high levels in the blood of women with preeclampsia and is believed to cause the maternal symptoms in this disease. Understanding the normal and abnormal functions of this protein will have a direct and immediate impact on the design of clinical therapies for protecting the mother from the adverse consequences of preeclampsia and will provide critical understanding of how these therapies can be designed to further protect the fetus.

描述（由申请人提供）：先兆子痫是一种严重的妊娠并发症，会影响母亲和胎儿，并造成全球约 14% 的妊娠相关孕产妇死亡。然而，尽管其对公共健康影响巨大，但其原因仍知之甚少，并且没有已知的治疗方法可以有效减轻母亲和胎儿的风险。基于人类的临床研究和啮齿动物的实验研究表明，可溶性 fms 样酪氨酸激酶 1 (sFlt-1) 循环水平的异常升高通过以下方式导致全身内皮功能障碍和疾病的临床表现：其拮抗血管内皮生长因子（VEGF）活性。因此，人们认为 VEGF 治疗可以逆转母体循环中高 sFlt-1 水平引起的先兆子痫表型。然而，在初步研究中，我们令人惊讶地发现，怀孕小鼠中 VEGF 的过度表达会导致血压升高、血清 sFlt-1 和可溶性内皮糖蛋白 (sEng) 水平升高以及肾脏组织学变化，与临床发现类似。人类和动物模型中的PE。 VEGF 已被证明可以在多种细胞类型和多种情况下的胎盘外植体培养物中刺激 sFlt-1 的产生。然而，胎盘 VEGF 在妊娠不同阶段的作用、胎盘产生 sFlt-1 调节其局部活性的意义及其在调节胎盘 sFlt-1 产生中的潜在作用尚未得到研究，主要是由于缺乏合适的动物模型来测试 VEGF 的胎盘特异性作用。我们假设胎盘中的 VEGF 活性在妊娠期间通过局部产生 sFlt-1 以胎盘发育阶段特异性方式受到调节，并且胎盘 VEGF 水平可能是该组织中 sFlt-1 产量增加的主要刺激因素。该提案中的研究首先旨在开发一种新颖的、可诱导的、胎盘特异性基因表达系统，能够通过活体成像监测整个妊娠期的基因表达。该技术将基于已建立的病毒递送基因特异性至胎盘组织的方法、基因表达水平一致性的有效控制以及允许表达快速打开和关闭的严格控制的诱导型启动子。然后，我们将使用这种诱导系统检查妊娠期间不同时间点不同水平的胎盘 VEGF 表达的影响。为了确定 sFlt-1 在妊娠不同阶段的生理作用，我们将使用针对 sFlt-1 的短发夹 RNA (shRNA) 的胎盘特异性表达，选择性地敲低胎盘中 sFlt-1 的表达。这些实验的结果将首次描述 VEGF 和 sFlt-1 在调节胎盘中 VEGF 活性方面​​的局部相互作用。此外，这种全新的方法，使用新开发的、可诱导的、胎盘特异性基因调控系统，将极大地促进用于研究胎盘疾病和正常胎盘发育的新动物模型的开发，并将极大地促进治疗方法的开发妊娠并发症。 公共健康相关性：这项工作解决了妊娠并发症先兆子痫的根本原因，该疾病影响全球 5-7% 的妊娠，是孕产妇和胎儿发病和死亡的主要原因。它开发了一种新方法来研究不同因素在胎盘功能和疾病中的作用，以小鼠为模型系统，专门研究可溶性 fms 样酪氨酸激酶 1 (sFlt-1) 的生理作用和调节机制。血管内皮生长因子受体 1 (Flt-1) 的形式，在先兆子痫女性的血液中含量很高，据信是导致这种疾病的母体症状的原因。了解这种蛋白质的正常和异常功能将对保护母亲免受先兆子痫不良后果的临床疗法的设计产生直接和直接的影响，并将为如何设计这些疗法以进一步保护胎儿提供重要的理解。