1. Fig. 1. Generation of forebrain progenitor cells with regional identity from human iPSCs.
Generation of forebrain progenitor cells with regional identity from human iPSCs. (A) Scheme of the RONA differentiation protocol showing generation of forebrain progenitors from human ESCs and iPSCs. Nog, noggin; SB, SB (B) Rosettes were immunopositive for FOXG1 and NES at day 9 after initiation of neural differentiation. DAPI, 4′,6-diamidino-2-phenylindole. (C and D) With the prolongation of neural differentiation, highly proliferative rosettes started to pile up, resulting in three-dimensional (3D) columnar cellular aggregates, which were positive for FOXG1 and NES. (D) Phase-contrast image of RONA. Scale bar, 50 μm (B to D). (E to G) FOXG1 immunoreactive RONAs were located in the center (E) and demarcated clearly with surrounding CK8-positive cells. (F) Cells underneath the RONAs were FOXG1−/CK8−. (G) The white inset box in (F) is magnified (×2.5). (H to L) Expression and quantification of the early forebrain regionalization markers FOXG1 (H), PAX6 (I), LHX2 (J), NKX2.1 (K), and GSX2 (L) after 9, 16, and 18 days of differentiation. Data are presented as means ± SEM (n = 3). Scale bar, 20 μm. (M and N) RNA sequencing gene expression profiling of RONAs derived from the human ESC H1 line showing the enrichment of forebrain progenitors with regional identity. Data show the average RPKM (reads per kilobase of transcript per million mapped reads) value of two biological replicates of human ESC H1 cell line RNA sequencing experiments. ESRRB, estrogen-related receptor β; SMA, smooth muscle actin; HNF1B, hepatocyte nuclear factor 1 homeobox B; MGE, medial ganglionic eminence; CGE, caudal ganglionic eminence. (O) Spontaneous neuronal differentiation after isolation of RONA neural progenitors without the application of exogenous mitogens. Nuclei were counterstained with DAPI (blue). Colors are indicated in the images. Scale bar, 20 μm. Data are from the human ESC H1 cell line in (B) to (O). DCX, doublecortin.
Jin-Chong Xu et al., Sci Transl Med 2016;8:333ra48
Published by AAAS