Neuronal differentiation on N-cadherin substrate without exogenous growth factors or inhibitors.

<p><b>(A-B)</b> ESCs cultured in suspension for 5 days with and without CKI-7 and SB-431542 (CK/SB) was further induced on N-cadherin substrate for 48 h in the absence of exogenous growth factors. The degree of neural conversion was analyzed by <b>(A)</b> neurite extension index and <b>(B)</b> qRT-PCR for relative mRNA expression of nestin, neurogenin 1 (Ngn1), and βIII-tubulin (Tuj). <b>(C)</b> Cellular phenotype of differentiated cells on N-cad-Fc and gelatin substrates at 20 days of differentiation was assessed by bright-field images. Scale bar: 200 μm. <b>(D)</b> Characterization of neural cells at 20 days of differentiation by the expression of neuronal progenitor cell markers, including N-cad, Nestin, Ngn1, Mitf and Tuj. <b>(E)</b> The relative mRNA expression level of neurotransmitter and subtype-specific genes (Nurr, CHAT, SLC, GAD2 and GFAP) in ESC- and iPSC-derived neuronal cells differentiated for 20 days on N-cadherin substrate. Abbreviation: ESCs, embryonic stem cells; iPSCs, induced pluripotent stem cells; Ngn1- Neurogenin 1; Mitf- Microphthalmia-associated transcription factor; N-cad, N-cadherin; Nurr- nuclear receptor-related factor; CHAT- choline acetyltransferase; SLC- solute carrier; GAD- glutamate decarboxylase; GFAP- glial fibrillary acidic protein.