MicroRNA-32 promotes gastric cancer cell proliferation and migration by inhibiting gene expression of Kruppel-like factor 4

Abstract

Objective To investigate the potential role of microRNA (miRNA, miR)-32 in the regulation of gastric carcinoma cell proliferation and invasion, as well as the regulatory mechanism. Methods The level of miR-32 in gastric cancer cells and normal gastric cell line was measured by real-time fluorescent quantitative polymerase chain reaction (FQ-PCR). Gastric carcinoma cell line MGC8-03 cells were divided into four groups: miR-32 overexpression group transfected with miR-32 mimics, scramble group transfected with scramble, lent-shmiR-32 group transfected with lent-shmiR-32 and lent-vector group transfected with lent-vector by lipofectamine 2000 or lentiviral vector. The proliferation ability was measured and compared by xCELLigence. The invasion ability was measured by Transwell. The expression level of Kruppel-like factor 4 (KLF4) protein was detected by Western blotting. Results Real-time PCR results indicated that, compared to the gene expression level in the gastric normal epithelial cell GES-1, miR-32 RNA level in the five gastric cancer cell liness, including AGS, KATO-Ⅲ, MGC8-03, NCI-N87 and SGC-7901, was increased significantly (0.18±0.03 vs. 12.40±1.40, 35.80±2.10, 27.10±3.60, 6.20±1.60 and 8.50±3.50 respectively, P<0.01). The number of MGC8-03 cells in miR-32 overexpression group was significantly increased at 48 or 72 h as compared with scramble group [(160.3±5.4)% vs. (104.2±3.8)% at 48 h, P<0.01; (310.2±5.4)% vs. (107.2±2.9)% at 72 h, P<0.01]. As compared with scramble group, the number of invasion cells in miR-32 overexpression group was significantly increased as well [(327±18) vs. (113±10), P<0.01]. The cell number in lent-shmiR-32 group was less than lent-vector group at 48 or 72 h after culture [(46.5±3.9)% vs. (102.3±2.9)% at 48 h, P<0.01; (46.5±3.9)% vs. (102.3±2.9)% at 72 h, P<0.01]. Transwell results indicated that the number of invasive cells in lent-shmiR-32 group was significantly less than in lent-vector group [(41±9) vs. (99±13), P<0.01]. The expression level of KLF4 mRNA and protein in lent-shmiR-32 group was significantly higher than in lent-vector group [(10.3±1.9) vs. (1.8±0.3), P<0.01; (3.79±0.54) vs. (1.35±0.21), P<0.01]. The expression level of KLF4 protein and mRNA in miR-32 overexpression group was significantly reduced as comparing with scramble group [(0.27±0.05) vs. (1.00±0.14), P<0.01; (1.16±0.13) vs. (3.26±0.65), P<0.01]. Conclusion miR-32 promotes gastric carcinoma cell proliferation and invasion by inhibiting gene expression of KLF4. Key words: MicroRNA-32; Gastric carcinoma; Proliferation; Invasion; Kruppel-like factor 4