A defined culture system for in vitro studies on trophoblast differentiation
用于滋养层分化体外研究的明确培养系统
基本信息
- 批准号:9765344
- 负责人:
- 金额:$ 7.6万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2018
- 资助国家:美国
- 起止时间:2018-09-01 至 2022-02-28
- 项目状态:已结题
- 来源:
- 关键词:ActivinsBiological ModelsBone Morphogenetic ProteinsCell Culture TechniquesCellsConditioned Culture MediaDevelopmentDown-RegulationEmbryoEnvironmentEpiblastEpidermal Growth FactorEpigenetic ProcessEthicsFetal healthFibroblastsFoundationsHeterogeneityHistocompatibility Antigens Class IHumanHypermethylationIn VitroLegalMaternal HealthMediatingModelingMolecularMusNodalPathologyPlacentaPlacentationPregnancyPromoter RegionsProtocols documentationReportingResearchRoleSamplingSerumSignal PathwaySignal TransductionSignaling ProteinSupplementationSyncytiotrophoblastSystemTestingValidationVillousVitronectinWorkadverse outcomebasebiomarker panelblastocystcell typeculture platescytotrophoblastembryonic stem cellhuman embryonic stem cellinhibitor/antagonistinterestpromoterrhosphingosine 1-phosphatestem cell differentiationtissue culturetooltrophoblast
项目摘要
PROJECT SUMMARY/ABSTRACT
The trophectoderm layer of the blastocyst stage embryo is the precursor for all trophoblast (TB) cell types
in the placenta – the villous cytotrophoblasts (vCTBs), syncytiotrophoblast (STB), and extravillous trophoblasts
(EVTs) comprising column cytotrophoblasts (cCTBs) and invasive cytotrophoblasts (iCTBs). Abnormalities in
TB development result in placental pathology, and are associated with adverse outcomes during pregnancy. Yet,
despite being critically important to maternal and fetal health, human TB development
remains poorly understood. Mechanistic studies on human TB development are difficult due to challenges
in research with human embryos, and limited availability of placental samples from early gestation. Further,
human placental samples are associated with heterogeneity, containing multiple TB and non-TB cell types, and
have likely undergone adaptation to the maternal environment. To overcome these limitations, we propose to
use TB derived from human embryonic stem cells (hESCs) as a model system for studies on human TB
development. However, the molecular mechanisms underlying the acquisition of TB fate in hESCs
remain unclear. This lack of understanding is a major impediment for widespread use of hESC-derived TB as
a model system for early human TB development. A major challenge in this context is the lack of defined
culture conditions for initiating TB differentiation of hESCs and/or differentiation of hESC-derived TB to
specialized TB subtypes. Therefore, here we propose to validate a completely defined culture system
for TB differentiation of hESCs, including selective differentiation to STB or iCTBs.
Accordingly, in Aim 1, we will validate a culture system based on the completely defined E6/7/8 medium,
and vitronectin-coated tissue culture plates. Specifically, we will investigate the expression of a panel of
biomarkers to confirm that bona fide TB sub-types can be obtained using these conditions. In Aim 2, we will
investigate whether TB differentiation of hESCs to STB and iCTBs can occur in defined conditions where
exogenous bone morphogenetic protein (BMP) is not added, or where BMP signaling through the SMAD1/5/8
signaling pathway is inhibited. Finally, in Aim 3, we will test the hypothesis that Rho/ROCK signaling
downstream of S1P activation mediates TB differentiation of hESCs in the proposed culture system.
The significance of our work relates to its relevance to our understanding of early human TB
development. Validation of defined culture conditions for TB differentiation of hESCs will enable mechanistic
studies on TB differentiation of hESCs, and differentiation of hESC-derived TB to specialized TB sub-types. This
in turn will enable routine acceptance use of hESC-derived TB a model system for early TB development. Such a
model system will be a powerful tool for advancing understanding of early human placental development.
项目总结/摘要
胚泡期胚胎的滋养外胚层是所有滋养层细胞类型的前体
在胎盘中-绒毛细胞滋养层(vCTB)、合体滋养层(STB)和绒毛外滋养层
在一些实施方案中,本发明涉及包含柱状细胞滋养层(cCTB)和侵袭性细胞滋养层(iCTB)的EVT。异常
TB的发展导致胎盘病理学,并与妊娠期间的不良结局相关。然而,
尽管对孕产妇和胎儿健康至关重要,但人类结核病的发展
仍然知之甚少。人类结核病发展的机制研究由于挑战而困难
在人类胚胎的研究中,早期妊娠的胎盘样本有限。此外,本发明还
人胎盘样品与异质性相关,含有多种TB和非TB细胞类型,
很可能已经适应了母体环境为了克服这些限制,我们建议
使用源自人类胚胎干细胞(hESC)的TB作为研究人类TB的模型系统
发展然而,在hESC中获得TB命运的分子机制,
仍然不清楚。这种缺乏了解是广泛使用hESC衍生的TB作为治疗的主要障碍。
人类结核病早期发展的模型系统。这方面的一个主要挑战是缺乏明确的
用于启动hESC的TB分化和/或hESC衍生的TB分化为
专门的结核病亚型。因此,在这里,我们建议验证一个完全定义的文化系统
用于hESC的TB分化,包括选择性分化为STB或iCTB。
因此,在目标1中,我们将验证基于完全确定的E6/7/8培养基的培养系统,
和玻连蛋白包被的组织培养板。具体来说,我们将研究一组
生物标志物,以确认使用这些条件可以获得真正的TB亚型。在目标2中,我们将
研究hESC向STB和iCTB的TB分化是否可以在确定的条件下发生,
不添加外源性骨形态发生蛋白(BMP),或其中BMP通过SMAD 1/5/8信号传导
信号传导通路被抑制。最后,在目标3中,我们将检验Rho/ROCK信号传导
S1 P激活下游的T-B介导了所提出的培养系统中hESC的TB分化。
我们工作的意义在于它与我们对早期人类结核病的理解相关
发展用于hESC的TB分化的确定的培养条件的验证将使得能够进行机制性的研究。
关于hESC的TB分化和hESC衍生的TB分化为特化TB亚型的研究。这
反过来将使hESC衍生TB的常规接受使用成为早期TB开发的模型系统。这样的
模型系统将成为促进对人类早期胎盘发育了解的有力工具。
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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Balaji M Rao其他文献
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{{ truncateString('Balaji M Rao', 18)}}的其他基金
Generating in vitro models of trophectoderm formation
生成滋养外胚层形成的体外模型
- 批准号:
10468196 - 财政年份:2021
- 资助金额:
$ 7.6万 - 项目类别:
Synthetic matrices for studies on trophoblast differentiation in 3D culture
用于研究 3D 培养中滋养层分化的合成基质
- 批准号:
10288194 - 财政年份:2021
- 资助金额:
$ 7.6万 - 项目类别:
Generating in vitro models of trophectoderm formation
生成滋养外胚层形成的体外模型
- 批准号:
10287887 - 财政年份:2021
- 资助金额:
$ 7.6万 - 项目类别:
Synthetic matrices for studies on trophoblast differentiation in 3D culture
用于研究 3D 培养中滋养层分化的合成基质
- 批准号:
10457418 - 财政年份:2021
- 资助金额:
$ 7.6万 - 项目类别:
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