Figure 1: p53 attenuation enhances the conversion of human fibroblasts to iDA neurons.

(a–e) Using the protocol in a, MRC5 cells were reprogrammed to iDA neurons in neural induction media with lentiviruses expressing Ascl1, Nurr1 and Lmx1a (ANL) (b), ANL plus p53 shRNA (ANLp) (c), ANL plus miR124 (ANLm) (d) or ANL plus miR124 and p53 shRNA (ANLmp) (e). Scale bar, 100 μm. (f,g) Percentage of TH⁺ or Tuj1⁺ cells in all cells (DAPI⁺) at day 9 (f). *^,#P<0.05, unpaired, two-tailed Student’s t-tests versus TH⁺ or Tuj1⁺ in ANL, respectively. ^{‡, †}P<0.05, unpaired, two tailed Student’s t-tests versus TH⁺ or Tuj1⁺ in ANLm, respectively; n=8 wells from four independent experiments for each condition. Average numbers of TH⁺, Tuj1⁺ or DAPI⁺ cells per frame under × 10 lenses were plotted in g. *P<0.05, unpaired, two tailed Student’s t-tests versus ANL for TH⁺ or Tuj1⁺ cells, respectively; n=8 wells from four independent experiments for each condition. The scale for DAPI⁺ cells is according to the y axis on the right. (h) Western blotting of p53 and α-tubulin (α-Tub) in total cell lysates at day 1. Total neurite length (i) and number of neurites (j) per TH⁺ neurons. *P<0.05, unpaired, two tailed Student’s t-tests versus ANL; ^#P<0.05, unpaired, two tailed Student’s t-tests versus ANLm; n=30–60 TH⁺ neurons from three independent experiments for each condition. (k,l) The reprogramming efficiency with ANLm or ANLm plus nutlin-3a (k) or with ANLm plus MDM2 (l). *^{, #}P<0.05, unpaired, two tailed Student’s t-tests versus TH⁺ or Tuj1⁺, respectively; n=6 wells from 3 independent experiments for each condition. Insets, western blotting of p53 and α-tubulin. Error bars denotes standard error of measurement.