Dissect formative pluripotency using cultured pluripotent stem cells

使用培养的多能干细胞剖析形成性多能性

基本信息

批准号：
10297495
负责人：
Jun Wu
金额：
$ 34.04万
依托单位：
UT SOUTHWESTERN MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-07-22 至 2026-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10297495
关键词：
Adult Affinity Chromatography Cell Differentiation process Cell Lineage Cells ChIP-seq Characteristics Chemicals Chimera organism Clustered Regularly Interspaced Short Palindromic Repeats Competence Derivation procedure Development Developmental Biology Embryo Epiblast Fibroblast Growth Factor Fibroblast Growth Factor Receptors Foundations Gastrula Gene Expression Profile Generations Genetic Germ Cells Goals Homeodomain Proteins Human In Situ In Vitro Infertility Laboratories Maintenance Methods Modeling Molecular Mus Names Oocytes Oogonia Pathway interactions Phase Phenotype Play Pluripotent Stem Cells Proteins Proteomics Regenerative Medicine Regulation Regulatory Element Reproductive Biology Resources Role Signal Transduction Somatic Cell Structure of primordial sex cell Study models Testing Time Tissues Transforming Growth Factor beta Transgenes Validation austin base blastocyst cell type embryonic stem cell established cell line experimental study implantation in vivo induced pluripotent stem cell insight novel permissiveness pluripotency regeneration potential stem cell model stem cells tool transcription factor transcriptome

项目摘要

PROJECT SUMMARY/ABSTRACT Derivation of pluripotent stem cells (PSCs) has revolutionized developmental biology and regenerative medicine. To stably maintain PSCs in culture and guide them to differentiate with high efficiency and fidelity into a variety of cell types, it is important to understand the molecular mechanisms governing pluripotency (the ability of a cell to generate any tissues in the body). Two phases of pluripotency, naïve and primed, have been defined and studied in detail thanks to the successful derivation of mouse embryonic stem cells (ESCs) and epiblast stem cells (EpiSCs), respectively. Mouse ESCs most closely resemble epiblast from a 4-day-old mouse blastocyst (~embryonic day 4, or E4), while “primed” EpiSCs display a global gene expression signature similar to the E7 epiblast of a post-implantation mouse embryo. Despite these advances, however, however, there is lack of a well-established PSC model that resembles E5-6 early post-implantation epiblast, which corresponds to the formative phase of pluripotency. Formative pluripotency exists within a time window during which naïve pluripotency is reconfigured to prepare for multilineage competency, including germ cells. Functionally, formative pluripotency is characterized by both chimera competency and permissiveness for direct primordial germ cell (PGC) induction. Several recent studies have attempted to define this state by transient epiblast-like cells (EpiLCs) differentiated from ESCs. To date, however, stable formative PSCs have not yet been generated. By modulating the FGF, TGF-β and WNT pathways, we recently derived PSCs from both mice and humans (referred to as FTW-PSCs) that are permissive for direct PGC-like cell induction in vitro and are capable of contributing to intra- or inter-species chimeras in vivo. FTW-PSCs harbor molecular, cellular and phenotypic features characteristic of formative pluripotency. The overall objective of this proposal is to use these newly established cell lines to comprehensively dissect the formative state across species. The proposed studies will elucidate the roles of several transcription factors in regulating mouse and human formative pluripotency, as well as demonstrate that FTW-PSCs are a robust platform for dissecting the molecular mechanisms underlying human and mouse PGC specification. In addition, we will establish an in vitro platform for the generation of functional mouse oocytes and human oogonia based on formative FTW-PSCs, thereby providing an invaluable resource for studying germ cell development and human infertility. Our proposal has tremendous potential to revolutionize regenerative medicine and reproductive biology.

项目概要/摘要多能干细胞 (PSC) 的衍生彻底改变了发育生物学和再生学在培养物中稳定维持PSC并引导其高效、保真地分化为药物。多种细胞类型，了解控制多能性的分子机制（能力已经定义了多能性的两个阶段：幼稚期和启动期。并通过小鼠胚胎干细胞（ESC）和外胚层的成功衍生进行了详细研究小鼠 ESC 与 4 日龄小鼠的外胚层最为相似。囊胚（〜胚胎第 4 天，或 E4），而“启动”EpiSC 则显示出类似的全局基因表达特征然而，尽管取得了这些进展，但仍存在着对植入后小鼠胚胎的 E7 外胚层的影响。缺乏类似于 E5-6 早期植入后外胚层的完善 PSC 模型，其对应多能性的形成阶段存在于幼稚的时间窗口内。多能性被重新配置，为多谱系能力做好准备，包括功能性的、形成性的。多能性的特征是嵌合体能力和直接原始生殖细胞的许可性 (PGC) 诱导。最近的几项研究试图通过瞬时外胚层样细胞来定义这种状态。然而，迄今为止，尚未产生稳定的形成性 PSC。通过调节 FGF、TGF-β 和 WNT 通路，我们最近从小鼠和人类中提取了 PSC（称为作为 FTW-PSC），它们允许在体外直接诱导 PGC 样细胞，并且能够促进 FTW-PSC 体内或种间嵌合体具有分子、细胞和表型特征。该提案的总体目标是利用这些新的特征。建立细胞系来全面剖析跨物种的形成状态。研究将阐明几种转录因子在调节小鼠和人类形成中的作用多能性，并证明 FTW-PSC 是解剖分子的强大平台此外，我们将建立一个体外平台。基于形成性 FTW-PSC 生成功能性小鼠卵母细胞和人卵原细胞，从而我们的提案为研究生殖细胞发育和人类不育症提供了宝贵的资源。彻底改变再生医学和生殖生物学的巨大潜力。