Comparison of proliferative capacity between F3 and F3.Olig2 human NSCs. (A) Cell proliferation assay measured by Cell Counting Kit-8. Cells were grown in a 96-well plate and the viability was measured at different time points after initial culture. ** = p < 0.01, *** = p < 0.001 by unpaired T test. Error bars indicate mean ± SD. N = 4 replicate experiments. (B) BrdU incorporation assay. Cells were grown on a 9 mm coverslip for 36 hours and BrdU was added for 2 hours. *** = p < 0.001 by unpaired T test. Error bars indicate mean ± SD. N = 3 coverslips for each condition. (C-H) Representative images of transverse spinal cord sections doubly stained with human mitochondria (red) and Ki67 (green) at 2 weeks after transplantation. A majority of grafted F3 cells (C-E) did not express Ki67, whereas the nuclei of F3.Olig2 grafted cells (F-H) were frequently colocalized with Ki67 (arrows). The nuclei were visualized by DAPI (blue) (E, H). Scale bar = 20 μm. (I) Quantification of the percent grafted cells containing Ki67 positive nuclei. Error bars indicate mean ± SD. ** = p < 0.01 by unpaired T test. N = 3 and 4 animals for F3 and F3.Olig2 groups, respectively.