Fig. 9

Characterization of tumorigenicity and differentiation potential of WT and Myod1 gene KO cells. a, b Soft agar colony formation assay of WT and KO cells during a 10-day period (a). Significance in difference of colony formation (b) was calculated based on experiments in triplicate using two-tailed Student’s t-test. **p < 0.01. Colonies larger than 25 μm in diameter were counted. c Difference in invasion and migration of WT and KO cells during time periods as indicated. d–f Tumor formation of WT and KO cells in nude mice. (D) shows tumor formation in five of five mice injected with KO cells, and e, f show difference in tumor volume (e) and weight (f) between WT and KO cells. Significance of difference in tumor volume between two groups of mice was calculated using two-way ANOVA-Bonferroni/Dunn test, and 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. g–i Tissue differentiation in tumor formed by KO cells, as examined by tissue-specific gene expression (g, h) and marker expression (i). g, h show difference in expression of genes representing neuronal differentiation and neural stemness (g), and mesodermal and endodermal tissues (h) between KO cells and KO cell xenograft tumors, as detected with RT-qPCR. Significance in expression change was calculated based on experiments in triplicate using two-tailed Student’s t-test. Data are shown as mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. NS: not significant. i IHC detection of cell/tissue markers in tumor sections. Below the panels for marker detection are corresponding sections stained with HE. Objective magnification for sections: 20×; insets: 40×