Generation of the Human Pluripotent Stem-Cell-Derived Astrocyte Model with Forebrain Identity

Abstract

Astrocytes form functionally and morphologically distinct populations of cells with brain-region-specific properties. Human pluripotent stem cells (hPSCs) offer possibilities to generate astroglia for studies investigating mechanisms governing the emergence of astrocytic diversity. We established a method to generate human astrocytes from hPSCs with forebrain patterning and final specification with ciliary neurotrophic factor (CNTF). Transcriptome profiling and gene enrichment analysis monitored the sequential expression of genes determining astrocyte differentiation and confirmed activation of forebrain differentiation pathways at Day 30 (D30) and D60 of differentiation in vitro. More than 90% of astrocytes aged D95 in vitro co-expressed the astrocytic markers glial fibrillary acidic protein (GFAP) and S100β. Intracellular calcium responses to ATP indicated differentiation of the functional astrocyte population with constitutive monocyte chemoattractant protein-1 (MCP-1/CCL2) and tissue inhibitor of metalloproteinases-2 (TIMP-2) expression. The method was reproducible across several hPSC lines, and the data demonstrated the usefulness of forebrain astrocyte modeling in research investigating forebrain pathology.

Urokinase plasminogen activator mediates changes in human astrocytes modeling fragile X syndrome

“…Human pluripotent stem cells (hPSCs) were maintained on Matrigel-coated plates in Essential 8 medium (E8; Thermo Fisher Scientific Ltd., Vantaa, Finland). Astrocytes were differentiated as previously described (Peteri et al, 2020). Briefly, for differentiation, 80% confluent hPSCs were dissociated with 0.5 mM ethylene-…”

“…We used hPSC-derived forebrain astrocyte model (hASTRO) meaning astrocytes at D95 of differentiation in vitro (Peteri et al, 2020) to study putative alterations of human FXS astrocytes (Figure 1a). Differentiation of astrocytes followed the developmental processes of astrocytes from neuroepithelial induction and regional patterning using morphogens to progenitor expansion and astrocyte specification/maturation (Russ et al, 2021).…”

Delta Opioid Receptor in Astrocytes Contributes to Neuropathic Cold Pain and Analgesic Tolerance in Female Mice

“…However, significant differences were reported between human and mouse astrocytes, including larger size, increased morphological complexity as well as differing subtypes defined by their expression profiles amongst other techniques (Oberheim et al, 2009 ; Ji et al, 2019 ; Escartin et al, 2021 ). One way to address the role of DOR in human astrocytes would be the use of human astrocytes derived from induced pluripotent stem cells (iPSC) in vitro , although in vitro culture induces astrocytes that differ from in vivo astrocytes (Barbar et al, 2020 ; Franklin et al, 2021 ; Peteri et al, 2021 ). Of note, human iPSC-derived astrocytes could be grafted into the mouse brain and retain human-specific features (Preman et al, 2020 ).…”

Across Dimensions: Developing 2D and 3D Human iPSC-Based Models of Fragile X Syndrome

“…Very few studies have explored hiPSC-derived glial cells in the study of FXS. One group recently published a method to generate astrocytes with forebrain patterning from hPSCs (hASTRO) [ 144 ]. Using this method, the group found that the expression of urokinase plasminogen activator (uPA), a protease that regulates the degradation of the extracellular matrix, was increased in FXS hASTRO compared to control hASTRO [ 145 ].…”