Fig. 2

NSCs undergo early entrance into replicative senescence in the presence of misfolded hSOD. A. Expression of the WT hSOD1 and the hSOD1 G93A mutation in NSCs results in diminished proliferation. Cell proliferation was visualized by immunostaining with Ki67 (Green) and neural stem cell marker SOX2 (Red) in P5 NSCs^WT. a) fluorescence image, scale bar: 20 μm. b) quantification of Ki-67 positive cell count. One-way ANOVA followed by Bonferroni’s Multiple Comparison Post-Test. B. P5 NSCs^WT were treated with mCT4 and CT4 for 24 h, and cell proliferation was detected by immunostaining with antibodies to Ki67 (Green) and to the neural stem cell marker SOX2 (Red). a) fluorescence image, scale bar: 50 μm. Quantification of b) Ki-67 and c) SOX2 + positive cell count. Student’s t-test. C. Western blot detection of P53 at different passages of NSCs^WT and NSCs^G93A. Full-length blots are presented in Additional file 1: Fig. 2C. a) gel image; b) densitometry. Two-way ANOVA followed by Sidak’s multiple comparisons test was performed. All in vitro analysis was done in triplicate from three independent cultures. D. Quantitative detection of hSOD1 thiol in the spinal cord of the WT hSOD1 mice at 135 days of age. Student’s t-test. E. a) Representative immunoblot and quantitative data of b) P16 and c) P53 expression in the spinal cord of the WT hSOD1 mice (n = 5 in control, and n = 6 in AAV-CT4), Student’s t-test. Full-length blots are presented in Additional file 1: Fig. 2E