We next examined the differentiation potential of these NPCs. The NPCs were cultured in a neuron differentiation media system (N2B27 + 20 ng bdnf + 1 μM dibutyryl-cAMP) for 21 days. The cells were then stained for TuJ1, a neuronal cell marker, and GFAP, an astrocyte marker. We found that both the neuronal marker and the astrocyte marker were expressed in the cultured cells (Fig. 1). These data indicated that NPCs derived from hiPSCs could differentiate into neuronal cells as well as astrocytes, and could be used as an in vitro model of neural differentiation. Furthermore, the neuronal cells stained positive for GABA, Glu1R, tyrosine hydroxylase (TH), and synapsin 1, indicating that the NPCs can differentiate into different types of mature neurons (Supplementary Fig. S1). Further analyses found that in differentiated cells, 54.9% were gabaergic neurons, 17.3% were TH-positive neurons, and 10.7% were glutamatergic neurons (Supplementary Fig. S1). The composition of neuronal cells did not change over the 15-day differentiation period.
