Figure 3.

IL33 promotes colon cancer stemness via its receptor ST2. A, Expression of ST2 protein in colon cancer cells. Primary colon cancer cells (#1 and #2) and HT-29 cells were stained with specific rabbit anti-human ST2 Ab and R-PE–conjugated goat anti-rabbit IgG. The expression of ST2 was determined by flow cytometer analyzer and expressed as the percentage of ST2⁺ cells in total colon cancer cells. One of four experiments is shown. B, Expression of ST2 mRNA in colon cancer cells. ST2 mRNA expression was determined by RT-PCR in human umbilical vascular endothelial cells (HUVEC), peripheral blood mononuclear cells (PBMC), primary colon cancer cells, and HT-29 cells. Primary colon cancer cells and HT-29 cells were cultured with 100 ng/mL IL33 for 24 hours. One of three experiments is shown. C, Effects of anti-ST2 on the role of IL33-mediated colon cancer sphere formation. Primary colorectal cancer cells were subject to sphere assay. IL33 (100 ng/mL) and/or anti-ST2 antibody (1 μg/mL) were added in the sphere culture. Results are expressed as the mean ± SEM; n = 4; ^**, P < 0.01. D–F, Effects of anti-ST2 on the role of IL33-stimulated colon cancer stem cell gene expression. Primary colorectal cancer cells were cultured with IL33 (100 ng/mL) and/or anti-ST2 antibody (1 μg/mL) for 24 hours. The expression of NANOG (D), NOTCH3 (E), and OCT3/4 (F) transcripts were quantified by real-time PCR. Results are expressed as the mean ± SEM; n = 4; ^*, P < 0.05; ^**, P < 0.01.