The effect of miR-224 down-regulation on SW80 cell proliferation and apoptosis and weakening of ADM drug resistance.

ABSTRACT

OBJECTIVE: Glycogen synthase kinase-3ß (GSK-3ß) can phosphorylate and degrade ß-catenin, and negatively regulates Wnt/ß-catenin signal pathway. MiR-224 up-regulation is associated with colorectal cancer (CRC). Bioinformatics analysis showed complementary binding sites between miR-224 and GSK-3ß. This study investigated if miR-224 plays a role in mediating GSK-3ß expression, Wnt/ß-catenin pathway activity, CRC cell proliferation, apoptosis as well as drug sensitivity of Adriamycin (ADM). MATERIALS AND METHODS: Dual luciferase gene reporter assay demonstrated the regulatory relationship between miR-224 and GSK-3ß. Expression of miR-224, GSK-3ß, ß-catenin, and Survivin was measured in normal colon epithelium NCM460, CRC cell line SW480, and drug-resistant SW480/ADM cell line. Flow cytometry measured apoptosis under ADM with an IC50 concentration of SW480 cells, followed by CCK-8 analysis of cell proliferation. SW480/ADM cells were treated with miR-224 inhibitor and/or pSicoR-GSK-3ß, followed by analysis of the expressions of GSK-3ß, ß-catenin and Survivin, cell apoptosis, and cell proliferation by EdU staining. RESULTS: MiR-224 targeted and inhibited GSK-3ß expression. In SW480/ADM cells, GSK-3ß expression and cell apoptosis rate were lower than those in SW480 cells, whilst miR-224, ß-catenin, and Survivin expression or proliferation were higher than those in SW480 cells. Transfection of miR-224 inhibitor and/or pSicoR-GSK-3ß significantly increased GSK-3ß expression in SW480/ADM cells, and decreased ß-catenin and Survivin expression, leading to reduced proliferation potency, enhanced cell apoptosis and suppressed ADM resistance. CONCLUSIONS: MiR-224 up-regulation is associated with ADM resistance of CRC cells. Suppression of miR-224 expression up-regulated GSK-3ß expression, inhibited Wnt/ß-catenin signal pathway activity and Survivin expression, as well as reduced ADM resistance of CRC SW480 cells.