A human iPS cell line for ready-to-use human iAstrocytes that support human neurons

Abstract Human iPSC-derived neuronal networks are increasingly being employed in basic and applied research to enhance translation. Astrocytes are essential for neuronal network function, but are often not included, or replaced with mouse astrocytes, which compromises translation. Current protocols produce hiPSC-derived astrocytes by stepwise differentiation using small molecules and cytokines, or by forward programming by inducing transcription factors introduced by lentiviral transduction. Here we created a stable, inducible hiPSC line capable of producing iAstrocytes by introducing the transcription factors NFIB and SOX9 into the AAVS1 locus of the BIHi005-A hiPSC line. iAstrocytes induced from this line upregulated astrocytic genes over four weeks in culture, expressed GFAP and S100B and exhibited spontaneous calcium waves and responses to ATP and CPA. In co-cultures, iAstrocytes supported the growth and function of mature iNeuron networks. Pre- and post-synaptic markers and synchronous neuronal activity measured by high-density multi-electrode array recordings and neuronal calcium imaging, appeared by four weeks. The use of iAstrocytes will help to standardize the use of human astrocytes to support human neural networks and enhance translation.