ConProject-002
ConProject-002
基本信息
- 批准号:10649486
- 负责人:
- 金额:$ 14.87万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2019
- 资助国家:美国
- 起止时间:2019-09-16 至 2024-06-30
- 项目状态:已结题
- 来源:
- 关键词:3-DimensionalAMOT geneAccelerationAmniotic SacBMP4BedsBiological ModelsCell Differentiation processCell NucleusCellsCouplesCystDataDevelopmentEarly DiagnosisEmbryoEmbryonic DevelopmentEndodermEpiblastEpitheliumEquilibriumEthicsEventFertilityGelGene ActivationGenerationsGeneticGenetic EngineeringGenetic TranscriptionGoalsHumanHuman DevelopmentImageImplantInner Cell MassInvestigationKnowledgeLightLiquid substanceLocationManuscriptsMechanicsMembraneMesodermModelingModificationMolecularMolecular ProbesMonkeysMovementMusNatureNuclearPathway interactionsPatternPopulationPregnancyPrimatesProcessRodentRoleSideSignal TransductionSignaling MoleculeSiteStructureSupporting CellSystemTestingTight JunctionsUp-RegulationUterusValidationWorkamnionblastocystcell typegastrulationhuman embryonic stem cellhuman modelhuman pluripotent stem cellimplantationimprovedin vitro Modelinhibitormechanical signalmechanotransductionmouse modelnoveloverexpressionpluripotencyprogramsprospectiveself organizationsingle-cell RNA sequencingspecies differencethree dimensional cell culturetooltranscription factor
项目摘要
The amniotic membrane forms a tough fluid-filled sac that protects the developing embryo and is essential for
a successful pregnancy. Amniogenesis initiates early in human development as the embryo implants into the
uterine wall: the inner cell mass first polarizes to form a pluripotent cyst with central lumen and subsequently,
one half of this polarized cyst loses pluripotency markers and becomes squamous in nature (the prospective
amniotic ecotderm) while the other side (the epiblast) remains pluripotent. Gastrulation begins on the epiblast
side soon thereafter. These early post-implantation developmental steps are inaccessible to study in humans,
leaving an enormous gap in our knowledge about amnion fate determination and formation of the amniotic sac,
despite the central importance of these events to the survival of the developing embryo. A new in vitro model
can help to close that gap: human pluripotent stem cells (hPSC), cultured in specific 3D conditions, form
polarized pluripotent cysts that spontaneously self-organize into symmetric cysts composed entirely of amnion
cells (90-95%) as well as asymmetric cysts that resemble amniotic sac-like structures (5-10%). Asymmetrically
patterned cysts mirror Carnegie stage 5c human embryos and are called “post-implantation amniotic sac
embryoids” or “PASE”. Cyst formation occurs progressively over five days in culture. Live imaging shows that
asymmetric cysts arise from focal flattening at one pole of the cyst and laterally spreading of amnion fate;
symmetric cysts arise when flattening occurs in a multi-focal pattern. Mechanistically, the initial trigger for
amnion differentiation is mechanical and that this causes presumptive amnion cells to activate a BMP signaling
program that is both necessary and sufficient for amniogenesis. At 5 days of culture, PASE contain distinct
populations of amnion, epiblast and boundary cells; epiblast cells initiate EMT movements similar to
gastrulation. We will exploit this robust in vitro model to accomplish the following goals: Aim 1) Explore how
mechanical signals activate BMP signaling. Novel PiggyBac-based tools for genetic modification of hESC will
aid in these studies. Aim 2) Establish the hierarchy of gene activation that results in amniogenesis and
development of mature PASE. Single cell RNAseq will be used to dissect the transcriptional cascade that
accompanies symmetry breaking, spreading amniogenesis, boundary formation and initiation of epiblast EMT
movements. Aim 3) Functionally test transcription factors that control amnion fate. Genetic deletion and
overexpression studies will be used to explore the role of several potential master regulators of amnion fate.
Overall, the work proposed here will greatly accelerate the pace of discovery regarding critical but previously
inaccessible post-implantation events and thus will have enormous implications for understanding early
processes that impact embryonic development and human fertility.
羊膜形成一个坚韧的充满液体的囊,保护发育中的胚胎,
成功怀孕。羊膜发生在人类发育的早期开始,胚胎植入子宫,
子宫壁:内细胞团首先极化形成具有中央腔的多能性囊肿,随后,
这种极化囊肿的一半失去了多能性标志物,并在性质上变成鳞状的(前瞻性研究)。
羊膜外胚层),而另一侧(外胚层)保持多能性。在上胚层开始
不久之后,这些早期植入后发育步骤无法在人类中进行研究,
在我们关于羊膜命运决定和羊膜囊形成的知识中留下了巨大的空白,
尽管这些事件对发育中的胚胎的存活至关重要。一种新的体外模型
可以帮助缩小这一差距:在特定的3D条件下培养的人类多能干细胞(hPSC),
自发自组织成完全由羊膜组成的对称囊肿的极化多能性囊肿
细胞(90-95%)以及类似羊膜囊样结构的不对称囊肿(5-10%)。不对称
图案化的囊肿反映了卡内基5c期人类胚胎,被称为“植入后羊膜囊
胚状体”或“PASE”。在培养物中,囊肿形成在五天内逐渐发生。现场成像显示,
不对称的囊肿是由于囊肿的一极局部变平和羊膜的侧向扩展造成的;
当扁平化以多病灶模式发生时,出现对称囊肿。从机制上说,
羊膜分化是机械的,这导致假定的羊膜细胞激活BMP信号传导
这是一个既必要又充分的计划。在培养5天时,PASE含有不同的
羊膜、上胚层和边界细胞群;上胚层细胞启动EMT运动,类似于
原肠胚形成我们将利用这个强大的体外模型来实现以下目标:
机械信号激活BMP信号传导。基于PiggyBac的新型hESC基因修饰工具将
帮助这些研究。目的2)建立导致肿瘤发生的基因激活层次,
发展成熟的PASE。单细胞RNAseq将用于剖析转录级联,
伴随着对称性破缺、扩展性分化、边界形成和上胚层EMT的启动
动作目的3)功能检测控制羊膜命运的转录因子。基因缺失和
将使用过表达研究来探索羊膜命运的几种潜在主调节因子的作用。
总的来说,这里提出的工作将大大加快发现关键但以前
难以接近的植入后事件,因此将有巨大的影响,了解早期
影响胚胎发育和人类生育能力的过程。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Kenichiro Taniguchi其他文献
Kenichiro Taniguchi的其他文献
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{{ truncateString('Kenichiro Taniguchi', 18)}}的其他基金
Mechanism of apicosome-driven lumen formation during human and mouse embryogenesis
人类和小鼠胚胎发生过程中顶端体驱动的管腔形成机制
- 批准号:
10424552 - 财政年份:2020
- 资助金额:
$ 14.87万 - 项目类别:
Mechanism of apicosome-driven lumen formation during human and mouse embryogenesis
人类和小鼠胚胎发生过程中顶端体驱动的管腔形成机制
- 批准号:
10650853 - 财政年份:2020
- 资助金额:
$ 14.87万 - 项目类别:
Mechanism of apicosome-driven lumen formation during human and mouse embryogenesis
人类和小鼠胚胎发生过程中顶端体驱动的管腔形成机制
- 批准号:
10249295 - 财政年份:2020
- 资助金额:
$ 14.87万 - 项目类别:
Mechanism of apicosome-driven lumen formation during human and mouse embryogenesis
人类和小鼠胚胎发生过程中顶端体驱动的管腔形成机制
- 批准号:
10029458 - 财政年份:2020
- 资助金额:
$ 14.87万 - 项目类别:
A self-organizing embryoid model of peri-implantation human development
人类植入前发育的自组织胚状体模型
- 批准号:
10019413 - 财政年份:2019
- 资助金额:
$ 14.87万 - 项目类别:
A self-organizing embryoid model of peri-implantation human development
人类植入前发育的自组织胚状体模型
- 批准号:
10649470 - 财政年份:2019
- 资助金额:
$ 14.87万 - 项目类别: