Effect of miR-106a expression negatively regulated by KLF4 on the invasive activity of human gastric cancer cells BGC-823 / 西安交通大学学报(医学版)

ABSTRACT

Objective: To investigate the effect of Krüppel-like factor 4 (KLF4) on the expression of miR-106a and their interaction on the invasive activity of human gastric cancer cells. Methods: The JASPAR database was used to screen the transcriptional factor combined with miR-106a promoter. KLF4 over-expression vector KLF4-pcDNA and miR-106a reporter gene pmiR-106a-WT/MUT-luc were both constructed, and the dual luciferase reporter assay was used to detect the effect of KLF4 on the activity of miR-106a promoter. Forty specimens of human gastric cancer and their adjacent formalin-fixed paraffin-embedded (FFPE) specimens were collected, and Real-time PCR and immunohistochemistry were both used to detect the expression of KLF4. Human gastric cancer cell line BGC-823 was divided into four groups miR-106a mimic, mimic NC, KLF4+pcDNA, and pcDNA basic. Transwell assay was employed to detect the invasive ability of the gastric cancer cells after four different treatments. Results: Dual luciferase reporter assay indicated that KLF4-pcDNA could antagonize the activity of miR-106a promoter, which suggested that the activity of pmiR-106a-WT-luc luciferase was inhibited (pmiR-106a-WT+pcDNA-basic compared with pmiR-106a-WT+pcDNA-KLF4, P0.05). FFPE samples showed that the relative expression of KLF4 in gastric cancer was 0.69±0.59, which significantly differed from that in adjacent paracancerous tissues (P<0.001). The expression of KLF4 was correlated with differentiation degree, lymph node metastasis, and infiltration depth of human gastric cancer (P<0.05). The positive expression of KLF4 was mainly located in the nucleus and cytoplasm of gastric mucosal epithelial cells. Transwell essay showed that the number of invasive cells in the four different treatment groups was not entirely the same (P<0.001) 89.00±14.85 for miR-106a mimic, 50.90±17.94 for mimic NC, 37.70±12.60 for KLF4+pcDNA, and 66.20±3.19 for pcDNA basic. Conclusion: Transcription factor KLF4 at least has direct binding with miR-106a promoter in vitro, which may be involved in the invasion and metastasis process of gastric cancer cell by negatively regulating the expression of miR-106a at the upstream transcription level.