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）：先兆子痫是妊娠的严重并发症，影响了母亲和胎儿，并且估计在全世界造成了14％的与妊娠有关的孕妇死亡。尽管公共卫生的影响很大，但其原因仍然很少了解，并且没有已知的治疗方法可以有效地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，已经提出，可溶性FMS样酪氨酸激酶1（SFLT-1）的循环水平异常升高，有助于系统性内皮功能障碍和通过疾病的临床表现其血管内皮生长因子（VEGF）活性的拮抗作用。结果，据信VEGF治疗可能会逆转孕产妇循环中高的SFLT-1水平引起的先兆子痫的表型。然而，在初步研究中，我们发现，令人惊讶的是，怀孕小鼠中VEGF的过表达导致血压升高，SFLT-1和可溶性恩格林（SENG）的血清水平升高以及肾脏中的组织学变化，类似于临床发现。人类和动物模型中的体育。 VEGF已被证明可以在多种环境中刺激几种细胞类型的SFLT-1产生。但是，胎盘VEGF在不同妊娠阶段的作用，通过SFLT-1的胎盘生产对其局部活动的重要性，以及其在调节胎盘中SFLT-1产生中的潜在作用，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活性在怀孕期间通过局部生产SFLT-1以胎盘发育的特异性调节，而胎盘VEGF水平可能是增加该组织中SFLT-1产生的主要刺激。该提案中的研究首先设计为开发一种新型，可诱导的胎盘特异性基因表达系统，并能够通过活体内成像监测整个前的基因表达。该技术将基于既定的基因递送的方法，专门针对胎盘组织，有效控制基因表达水平的一致性以及紧密控制的诱导启动子，该启动子可以迅速开关和关闭表达。然后，我们将使用此诱导系统在怀孕期间在怀孕期间各个时间点进行不同水平的胎盘VEGF表达的影响。为了定义SFLT-1在妊娠的不同阶段的生理作用，我们将使用针对SFLT-1的短发夹RNA（SHRNA）的胎盘特异性表达在胎盘中有选择地击倒SFLT-1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活性中的局部相互作用。此外，这种完全新颖的方法是使用新开发的，可诱导的，胎盘特异性的基因调节系统，将极大地促进开发新的动物模型，以研究胎盘疾病和正常胎盘发育，并将极大地促进疗法的发展怀孕并发症。 公共卫生相关性：这项工作涉及妊娠并发症前启示性的基本原因，该病房会影响全球5-7％的怀孕，并且是孕产妇和胎儿发病率和死亡率的主要原因。它开发了一种使用小鼠作为模型系统研究不同因素在胎盘功能和疾病中不同因素的作用的新方法，专门针对可溶性FMS类似FMS的酪氨酸激酶1（SFLT-1）的生理作用和机制，可溶性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发现高水平，被认为会引起该疾病的母性症状。了解该蛋白质的正常功能和异常功能将对保护母亲免受先兆子痫的不利后果的临床疗法的设计有直接而直接的影响，并将对如何设计这些疗法进行批判性理解，以进一步保护胎儿。