Figure 5.
HOXA1 Is a Downstream Target of HotairM1 in CSCs
(A) The correlation between HOXA1 and HotairM1 determined through TCGA analysis. (B) HOXA1 expression after HotairM1 was stably knocked down. (C) Expression of HOXA1 was detected in tumor tissues and paired adjacent peri-tumor tissues. (D) HOXA1 was silenced in tumor cells by two independent siRNAs, and silencing efficiency was detected using real-time PCR. (E) HotairM1 expression after HOXA1 was silenced was detected. (F) Western blot of Nanog expression in DLD1 cells and OCM1 cells transfected with HOXA1 siRNAs. (G) Sphere formation assay was performed to detect the sphere-forming capacities of HOXA1-silenced cells and control cells. Representative images of tumor spheres at a dose of 1,000 cells per well are shown. (H) Statistical analysis of spheres per 1,000 cells was performed. (I) Images of colonies formed from the plate-cloning assay are shown. (J) Colonies formed in the dish were calculated and analyzed. (K) Migration abilities of HOXA1-silenced and control tumor cells were analyzed through Transwell assay. Images of cells on the outside of the Transwell are shown. (L) The OD630 absorbance values of stained migrated cells were obtained to calculate the migrating rate. (M) Tumorigenesis ability was determined using a soft agar assay. Representative images of stained colonies formed in the upper agar are shown. (N) The colony number in the upper agar was counted, and the relative colonies per site were calculated. ∗p < 0.05; ∗∗p < 0.01, by two-tailed Student’s t test.