Integration of systems and synthetic biology to advance development of human tissues ex vivo

系统与合成生物学的整合促进人体组织离体发育

• 批准号: 10020408
• 负责人: Mo Reza Ebrahimkhani
• 金额: $ 41.5万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-30 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10020408
• 关键词: Address; Adult; Advanced Development; Animal Experiments; Animals; Biliary; Biology; Biomedical Engineering; Biotechnology; Cells; Clustered Regularly Interspaced Short Palindromic Repeats; Computational Biology; Computer Analysis; Data; Dependence; Development; Disease; Endothelium; Engineered Gene; Engineering; Epigenetic Process; Event; FGF2 gene; Face; Fetal Liver; GATA6 transcription factor; Gene Activation; Genetic; Genetic Engineering; Genetic Transcription; Germ Layers; Goals; Human; Human Development; Human Engineering; In Vitro; Liver; Methods; Mus; Names; Organ; Organizational Culture; Organogenesis; Organoids; Patients; Phenotype; Population; Regulator Genes; Source; Stromal Cells; Synthetic Genes; System; Systems Biology; Systems Integration; Technology; Therapeutic; Tissue Engineering; Tissues; Training; Transcriptional Activation; Transforming Growth Factor beta; Vascular Endothelial Cell; analog; base; cell fate specification; cell type; computational platform; drug development; fetal; genetic signature; genome-wide; human adult stem cell; human disease; human model; human stem cells; human tissue; improved; in vivo; induced pluripotent stem cell; insight; inter-individual variation; intercellular communication; liver injury; mouse model; novel; overexpression; precision medicine; programs; self organization; spatiotemporal; stem cells; synthetic biology; tool; transcription factor

The rapidly evolving field of stem cell bioengineering and organoids technology face key challenges: a) stem cell derived tissues are stalled developmentally and show fetal-stage phenotypes b) they often lack key subsets of vascular endothelial and stromal cells derived from different germ layers c) There is a lack of toolset to quantitatively assess cell or tissue identity and d) to guide morphogenetic events towards their native adult phenotypes. In this proposal, we will address these issues by undertaking an integrative synthetic biology and systems biology approach. We recently generated novel human fetal liver organoids ex vivo using human iPSCs. Our approach entails genetic engineering of human iPSCs via overexpression of GATA6 transcription factor in pluripotent media with bFGF and TGF-β. Through this strategy, we showed development of human endodermal and mesodermal, intercellular communications, co-differentiation and self-organization of cultures into a multi- cell type fetal liver organoid. We will employ this tissue as a unique testbed to develop and address biotechnology challenges for in vitro maturation, assessment and engineering of organoids. Through aim 1 of this proposal we will develop and validate a set of genetic toolset to drive cell-fate reprogramming of the multicellular tissue. Through aim 2, we will establish a computational platform to quantitatively assess liver organoids and to identify transcriptional regulators of stage specific development IMPACT: We tackle improvement and assessment of human organoids ex vivo, two key challenges in stem cell bioengineering. The successful completion of this study will result in a hypothesis-driven framework for rational engineering and advancement of stem cell-derived tissues ex vivo. Our study will also generate synthetic liver tissues with close proximity to adult human liver. It reduces dependence on animal experiments and increases access to refined human tissues.

快速发展的干细胞生物工程和类器官技术领域面临着关键挑战： 衍生的组织在发育上停滞并显示胎儿阶段表型B）它们通常缺乏关键的 血管内皮细胞和基质细胞来源于不同的胚层c）缺乏工具集， 定量评估细胞或组织身份，以及d）引导形态发生事件朝向它们的天然成体 表型在本提案中，我们将通过开展综合合成生物学来解决这些问题， 系统生物学方法我们最近使用人iPSC离体产生了新的人胎肝类器官。 我们的方法需要通过在细胞中过表达GATA 6转录因子对人iPSC进行基因工程改造。 含有bFGF和TGF-β的多能培养基。通过这种策略，我们展示了人类内胚层的发育， 和中胚层，细胞间通讯，共分化和自组织的文化，成为一个多- 细胞型胎肝类器官。我们将使用这种组织作为一个独特的试验平台，以开发和解决生物技术 类器官体外成熟、评估和工程化的挑战。通过本提案的目标1， 将开发和验证一套遗传工具集，以驱动多细胞组织的细胞命运重编程。 通过aim2，我们将建立一个计算平台来定量评估肝脏类器官并识别 阶段特异性发展的转录调节因子影响：我们解决了改善和评估 离体人类类器官，干细胞生物工程的两个关键挑战。成功完成本 研究将产生一个假设驱动的框架，用于合理的工程和干细胞衍生的 离体组织。我们的研究还将产生与成人肝脏非常接近的合成肝脏组织。它 减少了对动物实验的依赖，并增加了获得精制人体组织的机会。