Pluripotent human stem cells as models for normal and diseased trophoblast

多能人类干细胞作为正常和患病滋养层的模型

• 批准号: 8206784
• 负责人: R. MICHAEL ROBERTS
• 金额: $ 33.97万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-12-16 至 2015-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8206784
• 关键词: 2-methoxyestradiol; Abbreviations; Activins; Affect; Angiogenesis Inhibitors; Apoptosis; BMP4; Blood Vessels; Blood flow; Cell Culture Techniques; Cells; Cessation of life; Characteristics; Child; Child health care; Chorionic villi; Collagen Type I; Decidua; Development; Disease; Edema; Embryo; Event; Exhibits; Extracellular Matrix; Failure; Fetus; Fibroblasts; First Pregnancy Trimester; Glycine; Goals; Growth; HIF1A gene; HLA G antigen; Homologous Gene; Human; Hypertension; In Vitro; Infant; Inflammatory Response; Invaded; Lead; Life; Matrix Metalloproteinases; Minor; Mission; Modeling; Mothers; Mus; National Institute of Child Health and Human Development; Natural Killer Cells; Nodal; Oxygen; Pathology; Perfusion; Phenotype; Placenta; Pluripotent Stem Cells; Population; Pre-Eclampsia; Pregnancy; Production; Property; Proteins; Proteinuria; Reaction; Signal Transduction; Spiral Artery of the Endometrium; Stress; Study models; Symptoms; Syncytiotrophoblast; Syndrome; Term Birth; Time; Transformed Cell Line; Umbilical cord structure; Up-Regulation; Uterus; Vascular Diseases; Villous; Woman; Women' s Health; base; blood perfusion; bone morphogenic protein; cell type; crosslink; cytokine; cytotrophoblast; design; early onset; human embryonic stem cell; human stem cells; immune resistance; in vitro Model; induced pluripotent stem cell; inhibitor/antagonist; innovation; matrigel; migration; myometrium; novel strategies; obesity risk; public health relevance; stem; transcription factor; trend; trophoblast; vascular inflammation

DESCRIPTION (provided by applicant): The main goal of this proposal is to generate extravillous trophoblast (EVT) from human embryonic stem cells (hESC) after they have been treated with BMP4 (BMP-hESC) and use them as a model to study EVT emergence, migration, and invasiveness, as well as intrinsic and extrinsic factors that influence these properties. This project has high significance because EVT ultimately become the invasive trophoblast (TR) cells of human placenta that penetrate maternal blood vessels and enhance blood flow. While invasion of the uterine decidua and the proximate inner third of the myometrium by EVT is a characteristic feature of a healthy pregnancy, shallow invasion is associated with placental pathologies, among them pre-eclampsia (PE) and intra-uterine growth retardation (IUGR). PE is a pregnancy-specific syndrome that affects ~ 3 % of pregnancies and is responsible for 15 % of pre-term births and an estimated 50,000 deaths per year worldwide. It is characterized by maternal hypertension, edema, and proteinuria, conditions that in the more serious, early onset disease can manifest as soon as 20 weeks of pregnancy. Aim 1 will establish and characterize a hESC-derived cell culture model for the study of EVT. This aim is based on the hypothesis that the sub-population of HLA-G positive BMP-hESC that migrates through a Matrigel-coated matrix under high oxygen (O2) conditions are EVT homologs. HLA-G positive cells that remain associated with the BMP-hESC colonies under low O2 conditions represent a precursor population of non- invasive EVT whose properties will be distinct from the invasive population captured under high O2 conditions. Aim 2 will examine the ability of the EVT derived from BMP-hESC to invade through cross-linked, relatively stiff matrices, such as Type I collagen, and to interact with microcapillaries cultured within such matrices. Aim 3 will examine a possible model for PE in which up-regulation of stable HIF1 alpha (HIF1A) protein is hypothesized to counteract the effects of high O2 during in vitro development of BMP-hESC and retard development into invasive EVT. If time permits or the HIF1A hypothesis proves incorrect, we would plan to study the effects of knockdown of at least one transcription factor that has been implicated in EVT emergence and differentiation, namely ASCL2. Aim 4 will attempt to re-create EVT from infants born to mothers with PE by generating induced pluripotent stem cells (iPSC) from discarded umbilical cord and converting these pluripotent cells to CT, EVT and other lineages by the BMP4 approach. The properties of these cells can then be compared with EVT generated from cord cells from pregnancies not complicated by PE. The hypothesis underpinning this aim is that some forms of PE are initiated by genetically-based placental pathology, and these abnormalities will be manifested in the phenotype of EVT generated from umbilical cord-derived iPSC. Together, these aims will bring innovative and new approaches to the study of human TR cells and to examining the basis of placental pathologies, especially PE. They will create the first in vitro model that makes possible the study of both the emergence and invasiveness of EVT cells, and the first model to study these early events directly in TR from pregnancies complicated by PE. PUBLIC HEALTH RELEVANCE: This project is designed to establish a new model for studying extravillous trophoblast (EVT), a placental cell type that invades the wall of the womb during the first trimester of pregnancy and whose failure to invade properly can lead to serious consequences for mother and child, including a condition called pre-eclampsia. Instead of isolating the cells from placentae, the goal is to generate EVT from human pluripotent stem cells, thereby allowing us to study factors that control their invasiveness. Also by generating such pluripotent cells from umbilical cords of babies born to mothers that developed pre-eclampsia, we hope to recreate the cell type that caused the disease in the first place.

描述（由申请人提供）：该提案的主要目的是在接受BMP4（BMP-HESC）处理后，从人类胚胎干细胞（HESC）中产生额外的滋养细胞（EVT），并将它们用作研究EVT的出现，迁移，迁移和侵入性和固有和胚外因素的模型。该项目具有很高的意义，因为EVT最终成为人胎盘的侵入性滋养细胞（TR）细胞，该细胞穿透了母体血管并增强血液流动。虽然EVT的子宫dec骨入侵和肌层的近端内部三分之一是健康妊娠的特征，但浅侵袭与胎盘病理相关，其中包括胎盘症状（PE）前（PE）（PE）和乌顿内生长迟缓（IUGR）。 PE是一种妊娠特异性综合征，影响约3％的怀孕，并导致15％的孕期前出生，全球估计每年50,000例死亡。它的特征是孕产妇高血压，水肿和蛋白尿，在更严重的早期发作疾病中可能会在怀孕20周后表现出来。 AIM 1将建立并表征EVT研究的hESC衍生细胞培养模型。该目标是基于以下假设：HLA-G阳性BMP-HESC的亚群在高氧（O2）条件下通过基质胶涂层的基质迁移是EVT同源物。在低O2条件下与BMP-HESC菌落保持相关的HLA-G阳性细胞代表了非入侵EVT的前体群体，其性质将与在高O2条件下捕获的侵入性种群不同。 AIM 2将检查源自BMP-HESC通过交联，相对僵硬的矩阵（例如I型胶原蛋白）侵入的EVT的能力，并与在此类矩阵中培养的微毛细管相互作用。 AIM 3将检查一个可能的PE模型，在该模型中，假设稳定的HIF1α（HIF1A）蛋白的上调可以抵消BMP-HESC体外发育过程中高O2的影响，并将其延迟发育成侵入性EVT。如果时间允许或HIF1A假设证明不正确，我们计划研究至少一个转录因子的敲低的效果，这与EVT出现和分化有关，即ASCL2。 AIM 4将尝试通过从丢弃的脐带产生诱导的多能干细胞（IPSC）来从出生于患有PE的母亲的婴儿中创建EVT，并通过BMP4方法将这些多能细胞转换为CT，EVT和其他谱系。然后，可以将这些细胞的特性与从PE并不复杂的怀孕产生的EVT进行比较。该目标的基础的假设是，某些形式的PE是由基于遗传的胎盘病理学引发的，这些异常将体现在脐带衍生的IPSC产生的EVT表型中。这些目标共同为研究人类TR细胞的研究带来创新和新的方法，并检查胎盘病理，尤其是PE的基础。他们将创建第一个体外模型，从而使EVT细胞的出现和侵入性研究成为可能，以及第一个直接研究这些早期事件的模型。 公共卫生相关性：该项目旨在建立一种新的模型，用于研究胎盘滋养细胞（EVT），这是一种胎盘细胞类型，在怀孕的头三个月中侵入子宫壁并且未能适当入侵可能会对母亲和儿童造成严重后果，包括一种称为pre-eclampsia的疾病。目的不是从胎盘中隔离细胞，而是从人多能干细胞中产生EVT，从而使我们能够研究控制其侵入性的因素。同样，通过从出生的母亲的脐带中产生这种多能细胞，这些母亲出生的母亲，我们希望首先重现引起该疾病的细胞类型。