N- and AD-iPSCs were firstly cultured in the mTeSR1 medium, and then the neural differentiation was proceeded as reported with a modification [26, 27]. Briefly, iPSCs were dissociated into single cells by accutase and replated onto Matrigel-coated dishes. When cells reached 95% full in the dish, the cultural medium was changed gradually from the KSR medium containing 500 ng/ml rmNoggin (R&D System, Minneapolis, MN, USA), 10 μM SB431542 (Tocris Bioscience, Bristol, UK) and sodium butyrate (STEMCELL Technologies, Vancouver, BC, Canada), to the N2 medium (DMEM/F12 supplemented with 1% L-Glutamine, 1% NEAA, 1% P/S and 1% N2 supplement) containing 500 ng/ml rmNoggin, 10 μM SB431542 and 0.1 mM sodium butyrate. At day 10 of induction, cells were passaged en bloc with dispase digestion and cultured in the N2B27 medium containing 20 ng/mL bFGF. Upon the appearance of polarized cells after being passaged for about one week, rosettes were mechanically picked up and dissociated into single cells by 0.05% trypsin. All later passages of NPCs were maintained in the N2B27 medium with 20 ng/ml of bFGF.
