Fig. 10

Effect of inhibition of neural stemness in tumorigenic cells on tumorigenicity. a–g Effect of inhibition of neural stemness in NE-4C cells via RA-induced neuronal differentiation on tumorigenicity. Compared with cells treated with DMSO, NE-4C cells treated RA at 1 μM for 72 h showed dramatic alterations in cell phenotype (a), the expression of neural stemness and neuronal proteins as assayed with IB (b), and colony formation in soft agar in a 10-day period (c, d). Significance in difference of colony formation (d) was calculated based on experiments in triplicate using two-tailed Student’s t-test. ***p < 0.001. Colonies larger than 50 μm in diameter were counted. e–g shows the difference in tumor formation of cells in nude mice. e Shows tumors in five of five mice injected with DMSO or RA treated cells, and f, g show difference in tumor volume (f) and weight (g) between the two groups. h–n Effect of inhibition of neural stemness in KO cells via RA-induced neuronal differentiation on tumorigenicity. Compared with KO cells treated with DMSO, cells treated with RA showed strong difference in cell morphology (h), protein expression as assayed with IB (i), and colony formation (j, k). Significance in difference of colony formation (k) was calculated based on experiments in triplicate using two-tailed Student’s t-test. ***p < 0.001. Colonies larger than 25 μm in diameter were counted. l–n shows the difference in tumor formation of cells treated with DMSO and RA in nude mice. l shows tumors in five of five mice injected with DMSO or RA treated cells, and m, n show difference in tumor volume (m) and weight (n) between the two groups. In f and m, significance of difference in tumor volume between two groups of mice was calculated using two-way ANOVA-Bonferroni/Dunn test. In g and n, significance of difference in tumor weight was calculated using two-tailed Student’s t-test. Data are shown as mean ± SEM. **p < 0.01, ***p < 0.001