Fig. 5
The expression of CSC markers is increased in cisplatin-resistant NOY-1 CisR cells. a Increased expression of CD133 marker in NOY-1 CisR cells compared to parental NOY-1 cells was shown in the flow cytometric analysis. Chemoresistant HT-29/EGFP/FUR cells with high CD133 expression were used for the antibody titration and assay setup as a control. b The flow cytometry analysis by Aldefluor Assay revealed 4.5-fold increased ALDH activity in NOY-1 CisR cells when compared to parental cells. The number shown in each panel determined the percentage of ALDH + cells. HT-29/EGFP/FUR were used for the assay setup as a positive control. c NOY-1 CisR showed higher expression of ALDH1A3 and ABCG2 in expression analysis by qRT-PCR. MSC were used as a positive control for the OCT4 and ABCG2 expression, HT-29/EGFP were used as a positive control for the CD133 and Nanog expression. Chemoresistant HT-29/EGFP/FUR cells were used as a positive control for the ALDH1A3 expression, chemosensitive HT-29/EGFP cells missing ALDH1A3 expression were used as a negative control. d NOY-1 CisR cells are significantly resistant to ALDH inhibitors—DEAB and disulfiram, and to inhibitor of STAT3 signaling—napabucasin. Relative viability was determined by luminescent viability assay on day 6. Values are expressed as the averages of quadruplicates ± SD. e DEAB (100 µg/ml), disulfiram (200 ng/ml) and napabucasin (4 µM) effectively inhibited overall ALDH activity in NOY-1 CisR cells. The number shown in each panel was determined the percentage of ALDH + cells in Aldefluor Assay. **P < 0.01, ***P < 0.001, ****P < 0.0001