Dynamic expression of <i>Thyrotropin‐releasing hormone</i> in the mouse definitive endoderm

McKnight, Kristen D.; Hou, Juan; Hoodless, Pamela A.

doi:10.1002/dvdy.21313

Cited by 23 publications

(21 citation statements)

References 45 publications

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“…S3B). Interestingly, expression of Embigin, previously identified as a potential Lhx1 target in the AME (Shimono and Behringer 1999), and Trh, another DE marker (McKnight et al 2007), are both dramatically reduced.…”

Section: Transcriptional Profiling Identifies Lhx1 Downstream Target mentioning

confidence: 92%

Lhx1 functions together with Otx2, Foxa2, and Ldb1 to govern anterior mesendoderm, node, and midline development

et al. 2015

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Gene regulatory networks controlling functional activities of spatially and temporally distinct endodermal cell populations in the early mouse embryo remain ill defined. The T-box transcription factor Eomes, acting downstream from Nodal/Smad signals, directly activates the LIM domain homeobox transcription factor Lhx1 in the visceral endoderm. Here we demonstrate Smad4/Eomes-dependent Lhx1 expression in the epiblast marks the entire definitive endoderm lineage, the anterior mesendoderm, and midline progenitors. Conditional inactivation of Lhx1 disrupts anterior definitive endoderm development and impedes node and midline morphogenesis in part due to severe disturbances in visceral endoderm displacement. Transcriptional profiling and ChIP-seq (chromatin immunoprecipitation [ChIP] followed by high-throughput sequencing) experiments identified Lhx1 target genes, including numerous anterior definitive endoderm markers and components of the Wnt signaling pathway. Interestingly, Lhx1-binding sites were enriched at enhancers, including the Nodal-proximal epiblast enhancer element and enhancer regions controlling Otx2 and Foxa2 expression. Moreover, in proteomic experiments, we characterized a complex comprised of Lhx1, Otx2, and Foxa2 as well as the chromatin-looping protein Ldb1. These partnerships cooperatively regulate development of the anterior mesendoderm, node, and midline cell populations responsible for establishment of the left-right body axis and head formation.

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Section: Transcriptional Profiling Identifies Lhx1 Downstream Target mentioning

confidence: 92%

Lhx1 functions together with Otx2, Foxa2, and Ldb1 to govern anterior mesendoderm, node, and midline development

et al. 2015

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“…RNA probes were purified using G-50 Sephadex columns (Amersham) and stored at -80°C. Whole mount in situ hybridization was performed as described in [18]. Embryos were incubated in BM purple alkaline phosphatase substrate (Roche) at 4°C or room temperature.…”

Section: Whole Mount In Situ Hybridizationmentioning

confidence: 99%

A Molecular Signature for Purified Definitive Endoderm Guides Differentiation and Isolation of Endoderm from Mouse and Human Embryonic Stem Cells

Wang

McKnight

Wong

et al. 2012

Stem Cells and Development

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Embryonic definitive endoderm (DE) generates the epithelial compartment of vital organs such as liver, pancreas, and intestine. However, purification of DE in mammals has not been achieved, limiting the molecular "definition" of endoderm, and hindering our understanding of DE development and attempts to produce endoderm from sources such as embryonic stem (ES) cells. Here, we describe purification of mouse DE using fluorescence-activated cell sorting (FACS) and mice harboring a transgene encoding enhanced green fluorescent protein (eGFP) inserted into the Sox17 locus, which is expressed in the embryonic endoderm. Comparison of patterns of signaling pathway activation in native mouse DE and endoderm-like cells generated from ES cells produced novel culture modifications that generated Sox17-eGFP⁺ progeny whose gene expression resembled DE more closely than achieved with standard methods. These studies also produced new FACS methods for purifying DE from nontransgenic mice and mouse ES cell cultures. Parallel studies of a new human SOX17-eGFP ES cell line allowed analysis of endoderm differentiation in vitro, leading to culture modifications that enhanced expression of an endoderm-like signature. This work should accelerate our understanding of mechanisms regulating DE development in mice and humans, and guide further use of ES cells for tissue replacement.

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“…We find that CER1 is upregulated in both putative hDE and mDE ( Figure 3C). Thyrotropinreleasing hormone (Trh), here upregulated in both putative mDE and hDE ( Figure 3C), transiently marks the entire DE population and is not expressed in extra embryonic endoderm [36]. However, Sox7, expressed in the mouse primitive, parietal and VE, but not in DE during early embryogenesis [28], is expressed in hDE and mDE cultures.…”

Section: Resultsmentioning

confidence: 99%

A Functional Assay for Putative Mouse and Human Definitive Endoderm using Chick Whole-Embryo Cultures

Johannesson¹

2012

J Stem Cell Res Ther

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Introduction: Embryonic stem cells (ESCs) represent a prospective cell source for treating degenerative diseases such as diabetes. Several studies have addressed the generation of definitive endoderm (DE) from this cell source by attempting to recapitulate the signaling events occurring during embryogenesis. However, the subsequent differentiation of DE has failed to generate functional insulin-producing beta cells. To assure that we have the correct starting material, we need to fully characterize ESC-derived DE, by assessing whether the cells are functionally equivalent to the in vivo counterpart. Thus, the purpose of this study is to describe a method whereby the in vivo functionality of DE derived from ESCs can be assessed.Methods: By directed differentiation, putative DE was derived from human and mouse ESCs. This putative DE was subsequently transplanted into the endoderm of chick embryos to determine any occurrence of integration. Putative DE was analyzed by gene and protein expression prior to transplantation and 48 h post transplantation.Results: Putative DE, derived from mouse and human ESCs, was successfully integrated within the chick endoderm. Endoderm-specific genes were expressed in the putative DE prior to integration and endoderm-specific proteins were assessed 48 h post transplantation. Conclusions:We describe the detailed methodological procedure for transplanting putative DE derived from ESCs, and the subsequent analysis of the migration and development of the grafted cells. Our result show that putative DE integrates with the chick endoderm and participate in the development of the chicken gut, indicating the generation of functional DE from ESCs. This functional assay can be used to assess the generation of functional DE derived from both human and mouse ESCs and provides a valuable tool for cell characterization. This is an important initial step in the differentiation process towards fully functional beta cells.

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Dynamic expression of Thyrotropin‐releasing hormone in the mouse definitive endoderm

Cited by 23 publications

References 45 publications

Lhx1 functions together with Otx2, Foxa2, and Ldb1 to govern anterior mesendoderm, node, and midline development

Lhx1 functions together with Otx2, Foxa2, and Ldb1 to govern anterior mesendoderm, node, and midline development

A Molecular Signature for Purified Definitive Endoderm Guides Differentiation and Isolation of Endoderm from Mouse and Human Embryonic Stem Cells

A Functional Assay for Putative Mouse and Human Definitive Endoderm using Chick Whole-Embryo Cultures

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