Research Paper Volume 13, Issue 8 pp 11170—11187

The critical role of peroxiredoxin-2 in colon cancer stem cells

PRDX2 knockdown sensitizes CCSCs to chemotherapeutics. (A) Effect of 24 hours exposure of control- and shPRDX2-HT29-CD133+CD44+ CCSCs to 500 μg/mL of 5-FU or 100 μM oxaliplatin, as determined by flow cytometry apoptosis analysis with PE-labeled Annexin V containing 7-AAD staining. (B) Intracellular ROS levels of control- and shPRDX2-HT29-CD133+CD44+ CCSCs were measured before drug exposure and 24 hours after 500 μg/mL 5-FU or 100 μM oxaliplatin exposure. ROS levels are presented as the means ± SD of fluorescence intensity measured by flow cytometry with a DCFH-DA probe. Three independent experiments were performed in triplicate. Statistical analysis: Student’s t-test, ***p ****p C) Survival curves obtained from clonogenic assays of control- and shPRDX2-HT29-CD133+CD44+ CCSCs exposed to different H2O2 doses. The data are presented as the means ± SD of three independent experiments performed in triplicate for each dose. Statistical analysis: Student’s t-test, ***p ****p D) DNA damage of control- and shPRDX2-HT29-CD133+CD44+ CCSCs was measured before drug exposure and 24 hours after exposure to 500 μg/mL of 5-FU or 100 μM oxaliplatin by alkaline comet assay. The data are presented as the means of median tail moments ±SD of three independent experiments performed in triplicate. Statistical analysis: Student’s t-test, *p ***p

Figure 6. PRDX2 knockdown sensitizes CCSCs to chemotherapeutics. (A) Effect of 24 hours exposure of control- and shPRDX2-HT29-CD133+CD44+ CCSCs to 500 μg/mL of 5-FU or 100 μM oxaliplatin, as determined by flow cytometry apoptosis analysis with PE-labeled Annexin V containing 7-AAD staining. (B) Intracellular ROS levels of control- and shPRDX2-HT29-CD133+CD44+ CCSCs were measured before drug exposure and 24 hours after 500 μg/mL 5-FU or 100 μM oxaliplatin exposure. ROS levels are presented as the means ± SD of fluorescence intensity measured by flow cytometry with a DCFH-DA probe. Three independent experiments were performed in triplicate. Statistical analysis: Student’s t-test, ***p < 0.001, and ****p < 0.0001. (C) Survival curves obtained from clonogenic assays of control- and shPRDX2-HT29-CD133+CD44+ CCSCs exposed to different H2O2 doses. The data are presented as the means ± SD of three independent experiments performed in triplicate for each dose. Statistical analysis: Student’s t-test, ***p < 0.001, and ****p < 0.0001 vs. shPRDX2. (D) DNA damage of control- and shPRDX2-HT29-CD133+CD44+ CCSCs was measured before drug exposure and 24 hours after exposure to 500 μg/mL of 5-FU or 100 μM oxaliplatin by alkaline comet assay. The data are presented as the means of median tail moments ±SD of three independent experiments performed in triplicate. Statistical analysis: Student’s t-test, *p < 0.05, and ***p < 0.001.