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NFIB induces functional astrocytes from human pluripotent stem cell-derived neural precursor cells mimicking in vivo astrogliogenesis

ABSTRACT: The ability to generate astrocytes from human pluripotent stem cells (hPSCs) offers a promising cellular model to study the development and physiology of human astrocytes. However, the extant methods for generating functional astrocytes required long culture periods and remained much ambiguity whether their paradigm follows the innate developmental program. In this report, we provided an efficient and rapid method for generating physiologically functional astrocytes from hPSCs. Overexpressing the nuclear factor IB (NFIB) in hPSC-derived neural precursor cells (NPCs) induced a highly enriched astrocyte population in 2 weeks. RNA sequencing and functional analyses demonstrated progressive transcriptomic and physiological changes in the cells, resembling in vivo astrocyte development. Further analyses substantiated previous results and established the MAPK pathway necessary for astrocyte differentiation. Hence, this differentiation paradigm provides a prospective in vitro model for human astrogliogenesis studies and pathophysiology of neurological diseases concerning astrocytes.

ORGANISM(S): Homo sapiens

PROVIDER: GSE157768 | GEO | 2020/09/11

REPOSITORIES: GEO

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Project description:The gene regulatory mechanisms that steer neural stem cells towards astrocytes during brain development are largely unknown. Here we set out to comprehensively identify transcription factors that drive the developmental trajectory of astrogliogenesis from mouse embryonic stem cells. To attain this we performed a genomewide analysis of gene expression and epigenetic signature hallmarks of primed and active enhancers through the distinct stages of astrogliogenesis. We revealed distal regulatory elements that become activated in a stage-specific manner which confer the transcriptional program underlying generation and maturation of astrocytes. By searching for lineage-specific transcription factors functioning at these elements, we identified ATF3 and NFIA as drivers of differentiation of neural precursor cells to astrocytes, while RUNX2 promotes their maturation. These transcription factors are crucial for switching the chromatin state of these elements from primed to active, thereby driving the gene expression program underlying astrocyte differentiation and maturation.Method: RNA was extracted from embryonic stem cells (ES) differentiated into neural progenitor cells (aNPC) and subsequently differentiated for 1 day into early astrocytes (eA) and further progressively for 5 and 21 days into mature astrocytes (lA_1 and lA_2). These samples are further subjected for RNA-sequencing to reveal the genome wide changes at transcriptional level at each stage. aNPC, eA and lA_1 (further called as lA) were subjected for H3K27ac and H3K4me1 ChIP and sequenced genome wide (ChIP-seq) to reveal the active and primed enhancers regions during astrogliogenesis, respectively. In quest to identify the TF required for early and late astrogliogeneis, we depleted Nfia (SiNfia) and Atf3 (siAtf3) in eA and Stat3 (siStat3), Prdm9 (siPrdm9) and Runx2 (siRunx2) at lA by using siRNAs and perfomed H3K27ac ChIP and sequence them. We also extracted RNA after depletion (only after siNfia, siAtf3 and siRunx2) and perform the RNA-seq.

Project description:MartínJiménez2017 - Genome-scale reconstruction of the human astrocyte metabolic network This model is described in the article: Genome-Scale Reconstruction of the Human Astrocyte Metabolic Network. Martín-Jiménez CA, Salazar-Barreto D, Barreto GE, González J. Front Aging Neurosci 2017; 9: 23 Abstract: Astrocytes are the most abundant cells of the central nervous system; they have a predominant role in maintaining brain metabolism. In this sense, abnormal metabolic states have been found in different neuropathological diseases. Determination of metabolic states of astrocytes is difficult to model using current experimental approaches given the high number of reactions and metabolites present. Thus, genome-scale metabolic networks derived from transcriptomic data can be used as a framework to elucidate how astrocytes modulate human brain metabolic states during normal conditions and in neurodegenerative diseases. We performed a Genome-Scale Reconstruction of the Human Astrocyte Metabolic Network with the purpose of elucidating a significant portion of the metabolic map of the astrocyte. This is the first global high-quality, manually curated metabolic reconstruction network of a human astrocyte. It includes 5,007 metabolites and 5,659 reactions distributed among 8 cell compartments, (extracellular, cytoplasm, mitochondria, endoplasmic reticle, Golgi apparatus, lysosome, peroxisome and nucleus). Using the reconstructed network, the metabolic capabilities of human astrocytes were calculated and compared both in normal and ischemic conditions. We identified reactions activated in these two states, which can be useful for understanding the astrocytic pathways that are affected during brain disease. Additionally, we also showed that the obtained flux distributions in the model, are in accordance with literature-based findings. Up to date, this is the most complete representation of the human astrocyte in terms of inclusion of genes, proteins, reactions and metabolic pathways, being a useful guide for in-silico analysis of several metabolic behaviors of the astrocyte during normal and pathologic states. This model is hosted on BioModels Database and identified by: MODEL1608180000. To cite BioModels Database, please use: BioModels Database: An enhanced, curated and annotated resource for published quantitative kinetic models. To the extent possible under law, all copyright and related or neighbouring rights to this encoded model have been dedicated to the public domain worldwide. Please refer to CC0 Public Domain Dedication for more information.

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NFIB induces functional astrocytes from human pluripotent stem cell-derived neural precursor cells mimicking in vivo astrogliogenesis

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