12. Study on the function and mechanism of miR- 301a in the carcinogenesis and development of gastric cancer

Aims: We discovered that miR-301a is one the the most upregulated miRNAs in gastric cancer by using miRNA array. The main purpose of this study is to explore the expression levels, the function of miR-301a , and its regulatory mechanisms in gastric cancer.
Methods: The overexpression of miR-301a in gastric cancer cell lines and gastric cancer tissues were examined by qRTPCR. The biological effects, including proliferation, apoptosis, cell cycle, migration, invasion and subcutaneous tumorigenesis, were investigated in SGC-7901 cells, as miR-301a was upregulated or downregulated. Downstream target genes of miR-301a were predicted by bioimformatics and further validated by luciferase reportor system, qRT-PCR and immunoblot.
Results: The expression of miR-301a was higher in gastric cancer cell lines than immortalized gastric epithelial cell line GES-1 and normal gastric mucosa by qRT-PCR. Elevation of miR-301a expression was observed in 51 gastric cancer tissues compared with their matched non-tumorous tissues. Further analysis showed that the expression level of miR-301a was negatively correlated with tumor differentiation. Overexpression of miR- 301a in SGC-7901 gastric cancer cells promoted in vitro cell growth, inhibited apoptosis, and increased migration/invasion and in vivo tumorigenicity. On the other hand, knockdown miR- 301a in SGC-7901 cells impaired the maliganant behavior and arrested cell cycle progression at G2/M phase. Bioinformatics analysis discovered putative binding sequences of miR-301a located at 3’UTR of transcription factor RUNX3. The regulation was validated by using dual-luciferase reporter assay. qRT-PCR and Western blotting analysis verified that miR-301a downregulated RUNX3 expression through post-transcriptional way.
Conclusions: The expression of miR-301a was significantly elevated in gastric cancer tissues compared with matched nontumor tissues. The level of miR-301a was negatively correlated with tumor differentiation. Overexpression of miR-301a in gastric cancer cells promoted their malignant behavior and vise versa. Signaling pathway analysis discovered RUNX3 as one of its major targets which mediated its effects on gastric cancer cells.