Project 1: Early Events in Reprogramming Somatic Cells

项目 1：体细胞重编程的早期事件

• 批准号: 8381785
• 负责人: SHERMAN Morton WEISSMAN
• 金额: $ 58.3万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8381785
• 关键词: Adult; Affect; Biology; Biopsy Specimen; Cell Count; Cell Culture Techniques; Cell Fraction; Cell Proliferation; Cell Separation; Cell Therapy; Cells; Chromatin; Chromosomal Stability; Chromosomes; Commit; DNA; DNA Methylation; DNA Sequence; Deoxyribonucleases; Development; Differentiation Antigens; Disease model; Elderly; Embryo; Epigenetic Process; Epithelial Cells; Equilibrium; Event; Gene Expression; Genes; Genetic Transcription; Genomics; Germ Layers; Global Change; Heterogeneity; Human; Hypersensitivity; In Vitro; Mesenchymal; Modification; Mutation; Needles; Nucleic Acids; Oncogenic; Pattern; Population; Procedures; Process; Production; Punch Biopsy; RNA analysis; RNA methylation; Regenerative Medicine; Reporting; Repression; Retroviral Vector; Site; Somatic Cell; Staging; Surface Antigens; System; Variant; Vertebral column; Virus; base; embryonic stem cell; epigenomics; histone modification; human disease; human embryonic stem cell; induced pluripotent stem cell; interest; novel; overexpression; pluripotency; programs; research study; response; stem; tool; transcription factor; vector

Basic biology of reprogramming IPSC There has been great interest in the production of pluripotent iPSCs from adults, because of their biologic interest and potential value for studying mechanisms of human disease, and potentially ultimately for regenerative medicine. However, the induction requires several weeks and is generally very inefficient, in that only a small fraction of the cells are induced. On prolonged cultivation with inducing factors, almost all cells can give rise to progeny among which are iPSCs, so that conversion appears to involve some stochastic event that can occur in many cells rather than being limited to a few pre-determined cells in any culture. However, cell populations may be heterogeneous with respect to their ease of conversion. Conversion of somatic cells to IPSCs may follow a defined sequence of events, possibly involving early repression of differentiation markers and morphologic changes reminiscent of a transition from mesenchymal to epithelial cells, followed by the expression of ES-like markers of dedifferentiation, and finally by the activation of endogenous Nanog. Also, not all emerging colonies develop into fully pluripotent cells, suggesting that the conversion process requires multiple separable steps. Even after development of iPSCs they may retain markers such as sites of DNA methlylation that retain traces of their cell of origin, and progressive passage of the cells may cause them to be progressively more like embryonic stem (ES) cells, Epigenetic reprogramming of iPS cells: When induced to pluripotency, somatic cells acquire the features of ES cell-like states in their gene expression and epigenetic status. Human ES cells have histone modification states that allow them to differentiate into all three germ layers when instructed. So-called bivalent H3K4methylation and H3K27methylation on developmentally important genes are conserved in pluripotent cells. In addition, IPSCs acquire an ES cell-like global DNA methylation pattern. However, current factor-based reprogramming does not completely reprogram somatic epigenetic state to the ES cell state, and cells retain s significant number of iPSC specific differential methylated regions (DMRs) distinct from those of human ES cells. The aberrant epigenetic reprogramming affects the differentiation potential of IPSCs. Due to the extremely low efficiency of human somatic cell reprogramming. the elucidation of epigenetic changes during reprogramming induction has been challenging. By combining the technical advancement in analyzing global gene expression. DNA methylation analysis, and histone modification with isolation of cells of intermediate stages of reprogramming. we will reveal the sequence of epigenetic changes. Considering the importance of application of iPSCs in cell therapy and novel in vitro disease modeling, dissecting the epigenetic modification of reprogramming has high impact.

重编程IPSC的基本生物学 由于其生物学兴趣和研究人类疾病机制的生物学兴趣和潜在价值，并且最终有可能最终用于再生医学，因此人们对成年人的多能IPSC产生非常感兴趣。但是，归纳需要几个 几周，通常非常低效率，因为只能诱导一小部分细胞。在诱导因子的长时间培养时，几乎所有细胞都可以引起后代，因此iPSCS中的转化似乎涉及一些随机事件，这些随机事件可能发生在许多细胞中，而不是仅限于任何培养物中的少数预定细胞。但是，细胞种群在易于转化的方面可能是异质的。体细胞向IPSC的转化可能遵循定义的事件序列，可能涉及早期抑制分化标记物以及形态学的变化，让人联想到从间质细胞到上皮细胞的过渡，然后表达脱发的ES样标记物，最后通过内构纳米的激活。同样，并非所有新兴菌落都会发展为完全多能细胞，这表明转化过程需要多个可分离的步骤。 Even after development of iPSCs they may retain markers such as sites of DNA methlylation that retain traces of their cell of origin, and progressive passage of the cells may cause them to be progressively more like embryonic stem (ES) cells, Epigenetic reprogramming of iPS cells: When induced to pluripotency, somatic cells acquire the features of ES cell-like states in their gene expression and epigenetic status.人ES细胞具有组蛋白的修饰状态，可以在指导时将其分化为所有三个细菌层。在多能细胞中，所谓的二价H3K4-甲基化和H3K27M甲基化在发育重要的基因上是保守的。此外，IPSC还获得了ES细胞样全局DNA甲基化模式。但是，基于因子的重编程并未完全重新编程到ES细胞状态的体细胞表观遗传态，并且细胞保留了与人类ES细胞不同的IPSC特异性差异甲基化区（DMR）。异常表观遗传重编程会影响IPSC的分化潜力。由于人类体细胞重编程的效率极低。在重编程诱导过程中阐明表观遗传变化的挑战是具有挑战性的。通过结合分析全球基因表达的技术进步。 DNA甲基化分析和组蛋白修饰，分离重编程中间阶段的细胞。我们将揭示表观遗传变化的顺序。考虑到IPSC在细胞疗法和新型体外疾病建模中的应用的重要性，解剖重编程的表观遗传修饰具有很大的影响。