Modeling esophageal/respiratory birth defects in human pluripotent stem cell (PSC)-derived fetal tissues
Modeling esophageal/respiratory birth defects in human pluripotent stem cell (PSC)-derived fetal tissues
批准号:
10174986
负责人:
James M Wells
金额:
$32.3万
依托单位国家:
美国
项目类别:
财政年份:
2017
资助国家:
美国
项目状态:
已结题
起止时间:
2017-08-15 至 2022-05-31
关键词:
AchalasiaAffectAutologousBarrett EsophagusBiochemicalBiological AssayBiological ModelsCell LineChIP-seqCollaborationsColonComplexCongenital AbnormalityDefectDevelopmentDiseaseDominant-Negative MutationDorsalEmbryoEndodermEnteric Nervous SystemEosinophilic EsophagitisEpithelialEsophageal AtresiaEsophageal TissueEsophagusFailureFetal TissuesFoodGastroesophageal reflux diseaseGenesGenetic TranscriptionGerm LayersHMG-Box DomainsHumanHuman CharacteristicsImpairmentLeadMalignant neoplasm of esophagusMegaesophagusModelingMolecularMorphogenesisMorphologyMovementMusMuscleMutationOperative Surgical ProceduresOral cavityOrganoidsPathologyPathway interactionsPatientsPhysiologicalPluripotent Stem CellsPrimitive foregut structureProcessRegulator GenesReporterResortRespiratory SystemRoleSignal TransductionSmooth MuscleStomachStratified EpitheliumSurgeonTissue EngineeringTissuesTracheaTracheoesophageal FistulaTranscriptional ActivationTubebasebeta catenincell motilitycell typeconstrictionembryo tissuehuman pluripotent stem cellhuman tissueinduced pluripotent stem cellloss of functionmolecular modelingmotility disordernerve supplynervous system developmentnovelprotein Bprotein protein interactionreconstructionrespiratorysegregationstem cell based approachtissue reconstructiontranscription factortranscriptome sequencingvertebrate embryos
中文摘要
总结:在人PSC-E4衍生的胚胎组织中建模EA/TEF
在脊椎动物胚胎的发育过程中,一个共同的前肠管产生食道,
这涉及一系列复杂的分子和形态学过程。 背
前肠管形成食道,腹侧区域形成呼吸道,如果不能这样做,
导致气管食管出生缺陷,如食管闭锁和气管食管瘘(EA/TEF)。
如项目2中所讨论的,关于Wnt和BMP信号传导如何通过以下途径促进呼吸命运,
转录因子Nkx2.1的激活。 相比之下,对促食管因素知之甚少。 鼠标
和人类研究表明,HMG-1盒转录因子Sox 2参与了细胞的分离,
然而,Sox 2是否促进食管命运或作用于
主要抑制呼吸抑制诱导途径的背侧前肠尚不清楚。我们假设
机制参与正常的食管发育。
在人类中,大多数导致EA/TEF的基因仍然未被识别。 然而,SOX 2中的杂合突变
可导致EA和TEF,这与Sox 2杂合缺失的小鼠相反,这些小鼠是正常的。
然而,小鼠胚胎前肠内胚层Sox 2的完全缺失导致食管发育不全,
Sox 2也在食管的肠神经系统(ENS)发育期间表达。鉴于
EA患者可能存在运动障碍,我们推测EA相关基因可能影响ENS
发展然而,一项关于EA-ENA相关突变如何不同地影响上皮和/或ENS的研究表明,
从来没有在任何物种身上做过更别说人类了 我们提出了几种新的PSC-100
基于研究Sox 2和其他EA-E2相关基因如何影响人食管特化的方法,
人多能干细胞来源的食管上皮细胞形态发生和功能性神经支配
具有肠神经系统的类器官
在这个项目中,我们的目标是确定食管特异性的机制,
通过首先关注关键的食管因子Sox 2来研究人类的发展。我们假设SOX 2
通过一个未知的基因,既抑制呼吸系统,又促进食管的命运
监管网络。我们将使用人PSC-E2衍生的前肠模型与SOX 2获得性表达和丧失性表达相结合,
识别被SOX 2抑制的呼吸GRN和被SOX 2抑制的食管GRN
依赖。相反,我们将确定NKX2.1是否抑制食管命运。我们将利用
人前肠培养物鉴定人SOX 2的直接转录靶标的可扩展性,
NKX2.1使用RNA-测序和ChIP-测序。然后我们将研究TEF的发病机制
和由Sox 2突变引起的EA。我们将产生携带基于患者的突变的PSC系,
SOX 2,并研究这些如何影响食管和呼吸系统谱系的形成。 我们将
确定SOX 2突变对Wnt和BMP信号传导的影响,以及Sox 2是否通过直接蛋白-BMP蛋白作用
与效应蛋白B-β连环蛋白/TCF和Smads的相互作用。 最后,我们将探讨如何EA
突变对食管的不同细胞类型有不同的影响;
和ENS。考虑到一些EA患者具有相关的运动障碍,包括失弛缓症3,
收缩4和巨食管5,我们将研究Sox 2突变是否也有ENS缺陷。我们将使用
来源于项目1和2中鉴定的EA/TEF患者的iPSC系,以模拟分子缺陷
使用我们的人PSC-GFP衍生的类器官模型来解释这种出生缺陷。
英文摘要
Summary: Modeling EA/TEF In Human PSC-Derived Embryonic Tissues
During development of the vertebrate embryo, a common foregut tube gives rise to the esophagus and
respiratory tract and this involves an array of complex molecular and morphological processes. The dorsal
foregut tube forms the esophagus and the ventral domain forms the respiratory tract, and failure to do so can
result in tracheaesophageal birth defects such as esophageal atresia and tracheoesophageal fistula (EA/TEF).
As discussed in project 2, much is known about how Wnt and BMP signaling promote a respiratory fate by
activation of the transcription factor Nkx2.1. In contrast, little is known about pro-esophageal factors. Mouse
and human studies demonstrate that the HMG-box transcription factor Sox2 is involved in segregation of the
esophageal and respiratory lineages, however whether Sox2 promotes an esophageal fate or acts
predominantly to repress respiratory-inducing pathways the dorsal foregut is unclear. We hypothesize that both
mechanisms are involved in normal esophageal development.
In humans, most genes that cause EA/TEF remain unidentified. However, heterozygous mutations in SOX2
can cause of EA and TEF, which is in contrast to mice with heterozygous loss of Sox2, which are normal.
Complete loss of Sox2 from the foregut endoderm of mouse embryos results in esophageal agenesis, however
Sox2 is also expressed during development of the enteric nervous system (ENS) of the esophagus. Given that
patients with EA can have motility defects, we hypothesize some EA-associated genes may affect ENS
development. However, a study of how EA-associated mutations differentially affect the epithelium and/or ENS
of the esophagus has never been done in any species, let alone humans. We propose several novel PSC-
based approaches to study how Sox2 and other EA-associated genes impact Human esophagus specification,
epithelial morphogenesis, and functional innervation using human pluripotent stem cell-derived esophageal
organoids with an enteric nervous system.
In this project we aim to identify the mechanisms underlying esophageal specification and
development in humans by first focusing on the key esophageal factor Sox2. We hypothesize that SOX2
acts both to repress the respiratory lineage, and promote an esophageal fate via an unidentified gene
regulatory network. We will use a human PSC-derived foregut model in combination with SOX2 gain- and loss-
of-function to identify a respiratory GRN that is repressed by SOX2 and an esophageal GRN that is SOX2-
dependant. Conversely we will determine if NKX2.1 represses the esophageal fate. We will take advantage of
the expandable nature of human foregut cultures to identify direct transcriptional targets of human SOX2 and
NKX2.1 using RNA-seq and ChIP-seq. We will then investigate the disease mechanisms underlying TEF
and EA that are caused by Sox2 mutations. We will generate PSC lines harboring patient-based mutations in
SOX2 and investigate how these impact the formation of the esophageal and respiratory lineages. We will
identify the impact of SOX2 mutations on Wnt and BMP signaling and if Sox2 acts by direct protein-protein
interactions with the effector proteins b-catenin/TCF and Smads. Lastly we will investigate how EA
mutations differentially effect the different cell types of the esophagus;; the epithelial, smooth muscle
and ENS. Given that some patients with EA have associated motility disorders including achalasia 3,
constrictions 4 and megaesophagus 5, we will investigate if Sox2 mutations also have ENS deficits. We will use
iPSC lines derived from EA/TEF patients identified in projects 1 and 2 to model the molecular deficits
underlying this birth defect using our human PSC-derived organoid model.
期刊论文(0)
专著(0)
科研奖励(0)
会议论文
Project-3: Modeling EA/TEF in human organoids
-
批准号:10458162
-
项目类别:
-
资助金额:$39.75万
-
财政年份:2017
-
负责人:James M Wells
-
依托单位:
Project-3: Modeling EA/TEF in human organoids
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批准号:10647838
-
项目类别:
-
资助金额:$39.75万
-
财政年份:2017
-
负责人:James M Wells
-
依托单位:
Human Endocrine Cell Development
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批准号:8814212
-
项目类别:
-
资助金额:$45.5万
-
财政年份:2012
-
负责人:James M Wells
-
依托单位:
Generating Human Intestinal Organoids with an ENS.
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批准号:8415736
-
项目类别:
-
资助金额:$34.43万
-
财政年份:2012
-
负责人:James M Wells
-
依托单位:
Generating Human Intestinal Organoids with an ENS.
-
批准号:8516139
-
项目类别:
-
资助金额:$32.82万
-
财政年份:2012
-
负责人:James M Wells
-
依托单位:
Generating Human Intestinal Organoids with an ENS.
-
批准号:8665593
-
项目类别:
-
资助金额:$7.65万
-
财政年份:2012
-
负责人:James M Wells
-
依托单位:
Mammalian Foregut and Liver Development
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批准号:8204678
-
项目类别:
-
资助金额:$36.09万
-
财政年份:2009
-
负责人:James M Wells
-
依托单位:
Mammalian Foregut and Liver Development
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批准号:7895193
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项目类别:
-
资助金额:$10.0万
-
财政年份:2009
-
负责人:James M Wells
-
依托单位:
Mammalian Foregut and Liver Development
-
批准号:7753860
-
项目类别:
-
资助金额:$40.22万
-
财政年份:2009
-
负责人:James M Wells
-
依托单位:
Mammalian Foregut and Liver Development
-
批准号:7582986
-
项目类别:
-
资助金额:$39.44万
-
财政年份:2009
-
负责人:James M Wells
-
依托单位:
Mammalian Foregut and Liver Development
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批准号:8018155
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项目类别:
-
资助金额:$36.09万
-
财政年份:2009
-
负责人:James M Wells
-
依托单位:
Stem Cell/Organoid and Genome Editing Core
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批准号:10620719
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项目类别:
-
资助金额:$14.99万
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财政年份:2007
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负责人:James M Wells
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依托单位:
Pluripotent Stem Cell and Organoid Core
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批准号:9535292
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项目类别:
-
资助金额:$13.95万
-
财政年份:2007
-
负责人:James M Wells
-
依托单位:
Pluripotent Stem Cell and Organoid Core
-
批准号:9312950
-
项目类别:
-
资助金额:$14.34万
-
财政年份:2007
-
负责人:James M Wells
-
依托单位:
Stem Cell/Organoid and Genome Editing Core
-
批准号:10442029
-
项目类别:
-
资助金额:$14.99万
-
财政年份:2007
-
负责人:James M Wells
-
依托单位:
Mechanisms of endoderm specification along the A-P axis.
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批准号:7092306
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项目类别:
-
资助金额:$27.54万
-
财政年份:2006
-
负责人:James M Wells
-
依托单位:
Mechanisms of endoderm specification along the A-P axis
-
批准号:7410109
-
项目类别:
-
资助金额:$27.67万
-
财政年份:2006
-
负责人:James M Wells
-
依托单位:
Mechanisms of endoderm specification along the A-P axis.
-
批准号:7800390
-
项目类别:
-
资助金额:$27.4万
-
财政年份:2006
-
负责人:James M Wells
-
依托单位:
Mechanisms of endoderm specification along the A-P axis
-
批准号:7615114
-
项目类别:
-
资助金额:$27.67万
-
财政年份:2006
-
负责人:James M Wells
-
依托单位:
Mechanisms of endoderm specification along the A-P axis
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批准号:7222756
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项目类别:
-
资助金额:$27.67万
-
财政年份:2006
-
负责人:James M Wells
-
依托单位:
海外基金