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. The extant methods for generating functional astrocytes required long culture periods and there remained much ambiguity on whether such paradigms follow 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 in hPSC-derived neural precursor cells 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 the pathophysiology of neurological diseases concerning astrocytes.

A Novel Vitronectin Peptide Facilitates Differentiation of Oligodendrocytes from Human Pluripotent Stem Cells (Synthetic ECM for Oligodendrocyte Differentiation)

“…We introduced SOX10 in OPCs using a DOX-inducible lentiviral system [ 17 ]. To optimize gene transduction, various titers of EGFP-lentivirus were infected.…”

“…In this study, we used two hPSC lines, an ESC line (WA09, commonly known as H9) obtained from WiCell (Madison, WI, USA) and an iPSC line that we previously established (NL1) [ 17 ]. The majority of the differentiation data in this study were obtained with the ESC line (H9), unless indicated otherwise.…”

“…For oligodendrocyte differentiation, OPCs (at passage 3–5 before cryopreservation) were seeded at a density of 2.0 × 10 5 cells/cm 2 on a culture plate pre-coated with various matrices in OPC medium. One day later, the cells were infected with a lentivirus encoding SOX10 (Addgene, clone ID number 45843, Watertown, MA, USA) and a lentivirus of reverse tetracycline-controlled transactivator ( rtTA ) (Addgene, clone ID number 20342) for doxycycline (DOX)-induced SOX10 expression and incubated for 14–16 h. Viral vectors and packaging were constructed as described elsewhere [ 17 ]. After incubation with viruses, the culture medium was replenished with fresh medium containing 2.5 µg/mL (DOX), included in the media for the entire differentiation period.…”

Navigating Human Astrocyte Differentiation: Direct and Rapid one-step Differentiation of Induced Pluripotent Stem Cells to Functional Astrocytes Supporting Neuronal Network development

“…Many different hiPSC-astrocyte differentiation protocols have been described in literature 8,[16][17][18][19][20][21][22][23][24][25][26][27][28] . Generally these protocols can be categorized into two main approaches: differentiation using gene overexpression of transcription factor(s) such as SOX9, NFIA and NFIB 25,26,28 , or differentiation using an intermediate differentiation step towards neural progenitor cells (NPCs) 16,17,27 . Direct differentiation using gene overexpression often results in homogenous cultures of astrocytes 25,26,28 .…”

“…Generally these protocols can be categorized into two main approaches: differentiation using gene overexpression of transcription factor(s) such as SOX9, NFIA and NFIB 25,26,28 , or differentiation using an intermediate differentiation step towards neural progenitor cells (NPCs) 16,17,27 . Direct differentiation using gene overexpression often results in homogenous cultures of astrocytes 25,26,28 . In line with the large scale of astrocytic functions, increasing evidence shows a wide molecular, morphological, and functional astrocytic diversity in vivo [29][30][31] , which is considered to also be important for in vitro neuronal cultures 32,33 .…”