Mechanism of CDX2 gene expression regulated by miR-16-5p on proliferation and apoptosis of leukemia cells

Abstract

Objective To investigate the effect of caudal-type homeobox transcription factor 2 (CDX2) protein expression regulated by microRNA (miR) -16-5p on the proliferation and apoptosis of leukemia cells and its mechanism. Methods Real-time quantitative polymerase chain reaction (qRT-PCR) and Western blot were used to determine the expression levels of miR-16-5p and CDX2 in different leukemia cells and normal peripheral blood mononuclear cells (PBMC) , respectively. Dual-luciferase assay was used to determine the relationship between miR-16-5p and CDX2. MTT assay was used to determine the effect of miR-16-5p overexpression or inhibition of CDX2 expression on the proliferation of K562 cells. Flow cytometry was used to determine the effect of miR-16-5p overexpression or inhibition of CDX2 expression on the apoptosis of K562 cells. Western blot was used to determine the protein expression of CyclinD1, Bcl-2, Bax and p21. MTT assay and flow cytometry were used to determine the reversal of proliferation and apoptosis of K562 cells after overexpression of CDX2. Results Compared with PBMC cells, the expression level of miR-16-5p in different leukemia cells significantly decreased, whereas the expression level of CDX2 significantly increased (both P<0.05) . Dual-luciferase reporter assay system showed that miR-16-5p regulated the protein expression level of CDX2. MTT assay and flow cytometry showed that the A value of K562 cells in miR-16-5p group and si-CDX2 group was significantly lower than that in miR-NC group and si-NC group at 48 and 72 h (all P<0.05) . The apoptosis rate of K562 cells significantly increased [ (21.54±2.17) % vs (6.83±0.67) %, (19.38±1.85) % vs (7.24±0.71) %, both P<0.05]. The protein expression levels of CyclinD1 (0.32±0.04 vs 0.71±0.07, 0.38±0.04 vs 0.75±0.07) and Bcl-2 (0.33±0.03 vs 0.76±0.08, 0.36±0.04 vs 0.79±0.07) were significantly down-regulated (all P<0.05) . The protein expression levels of Bax (0.78±0.08 vs 0.32±0.03, 0.75±0.07 vs 0.26±0.03) and p21 (0.79±0.06 vs 0.28±0.03, 0.77±0.07 vs 0.24±0.03) were significantly up-regulated (all P<0.05) . Overexpression of CDX2 reversed the inhibiton and promotion of miR-16-5p on proliferation and apoptosis of leukemia cells, respectively. Conclusion miR-16-5p may regulate CDX2 protein to inhibit leukemia cell proliferation and induce apoptosis. Key words: Leukemia; miR-16-5p; CDX2; Proliferation; Apoptosis