An Efficient and Cost-Effective Approach to Generate Functional Human Inducible Pluripotent Stem Cell-Derived Astrocytes
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PROVIDER: PRJNA1060327 | ENA |
REPOSITORIES: ENA
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Project description:Human inducible pluripotent stem cell (hiPSC)-derived astrocytes (iAs) are critical to study astrocytes in health and disease. They provide several advantages over human fetal astrocytes in research, which include consistency, availability, disease modeling, customization, and ethical considerations. The generation of iAs is hampered by the requirement of Matrigel matrix coating for survival and proliferation. We provide a protocol demonstrating that human iAs cultured in the absence of Matrigel are viable and proliferative. Further, through a side-by-side comparison of cultures with and without Matrigel, we show significant similarities in astrocyte-specific profiling, including morphology (shape and structure), phenotype (cell-specific markers), genotype (transcriptional expression), metabolic (respiration), and functional aspects (glutamate uptake and cytokine response). In addition, we report that, unlike other CNS cell types, such as neuronal progenitor cells and neurons, iAs can withstand the absence of Matrigel coating. Our study demonstrates that Matrigel is dispensable for the culture of human iPSC-derived astrocytes, facilitating an easy, streamlined, and cost-effective method of generating these cells.
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2016-04-18 | E-MTAB-4553 | biostudies-arrayexpress
Project description:Direct reprogramming approaches offer an attractive alternative to stem-cell-derived models, allowing the retention of epigenetic information and age-associated cellular phenotypes. To explore such age-related phenotypes, several groups have generated multiple neuronal subtypes, neural progenitor cells, oligodendrocytes, and other cell types directly from fibroblasts. Other groups have had success at the efficient conversion of embryonic fibroblasts to astrocytes but have not yet achieved similar conversion efficiency for adult human fibroblasts. In order to generate astrocytes for the study of age-related diseases, we developed an improved direct conversion strategy employing a combination of small molecules to activate specific pathways that induce trans-differentiation of human adult fibroblasts to astrocytes. We demonstrate that this method produces mature GFAP+/S100β+ cells at high efficiency (40-45%), comparable to previous studies utilizing embryonic fibroblasts. Further, Fibroblast-derived induced Astrocytes (FdiAs) are enriched for markers of astrocyte functionality, including ion-channel buffering, gap-junction communication, and glutamate uptake; and exhibit astrocyte-like calcium signaling and neuroinflammatory phenotypes. RNA-Seq analysis indicates an adult rather than fetal astrocytic gene expression signature, with a greater correlation to temporal lobe astrocytes. Fibroblast-derived induced astrocytes provide a useful tool in understanding age-associated disease processes and complement existing in vitro models of induced neurons (iNs), providing an additional platform to study late-stage brain disorders.
2024-08-31 | GSE239320 | GEO
Project description:Direct reprogramming approaches offer an attractive alternative to stem-cell-derived models, allowing the retention of epigenetic information and age-associated cellular phenotypes, as well as a fast method to reach a target cell type. Several groups have previously generated multiple neuronal subtypes, neural progenitor cells, oligodendrocytes, and other cell types directly from fibroblasts. Other groups have had success at the efficient conversion of embryonic fibroblasts to astrocytes but have not yet achieved similar conversion efficiency for adult human fibroblasts. In order to generate astrocytes for the study of adult-stage disorders, we developed an improved direct conversion strategy employing a combination of small molecules to activate specific pathways that induce trans-differentiation of human adult fibroblasts to astrocytes. We demonstrate that this method produces mature GFAP+/S100β+ cells at high efficiency (40-45%), comparable to previous studies utilizing embryonic fibroblasts. Further, Fibroblast-derived induced Astrocytes (FdiAs) are enriched for markers of astrocyte functionality, including ion-channel buffering, gap-junction communication, and glutamate uptake; and exhibit astrocyte-like calcium signaling and neuroinflammatory phenotypes. RNA-Seq analysis indicates an adult rather than fetal astrocytic gene expression signature, with a greater correlation to temporal lobe astrocytes. Fibroblast-derived induced astrocytes provide a useful tool in studying the adult brain and complement existing in vitro models of induced neurons (iNs), providing an additional platform to study late-stage brain disorders.
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Project description:A generic, cost-effective and scalable cell lineage analysis platform
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