ArticleCells, tissues, organs2026
Rapid, Growth Factor-Reduced Induction of Functional Neurons from hiPSCs.
Article in Cells, tissues, organs, 2026. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.
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Abstract
introductionHuman induced pluripotent stem cells (hiPSCs) can be rapidly converted into neurons via Neurogenin-2 (NGN2) overexpression, but many protocols require costly reagents during the initial induction phase that may limit adoption by labs without routine neuronal differentiation experience. We developed a simplified, low-cost protocol using a tetracycline-inducible (TET-on) NGN2 system in minimal media to generate cortical neurons in as few as 6 days.
methodsKOLF2.1J hiPSCs were stably transfected with a TET-on NGN2 cassette using the nonviral PiggyBac system and induced with doxycycline in Essential 6 media with or without the Notch inhibitor DAPT. Neurogenesis was evaluated with immunocytochemistry (ICC) and qRT-PCR, and cultures matured in defined conditions were characterized by multielectrode array (MEA) recordings to assess functional maturation.
resultsDAPT markedly improved hiPSC-to-neuron conversion efficiency, and yielded glutamatergic neurons expressing cortical markers. MEA recordings showed spontaneous activity by day 14 and synchronous network firing by day 35. Secondary PB transfection enabled Td-Tomato labelling of KOLF2.1J:PB-TO-NGN2 hiPSCs, allowing 24 h live imaging of neurite outgrowth.
conclusionThis streamlined, growth-factor-reduced workflow provides an accessible route for generating functional neurons from patient-derived hiPSCs, including in labs with limited hiPSC or neuronal culture experience.
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