title: Expression and regulatory function of miRNA-34a in targeting survivin in gastric cancer cells.
abstract: We aimed to investigate the expression of microRNA-34a (miR-34a) in human gastric cancer cells and to evaluate the effects of miR-34a, acting via its gene survivin, on gastric cancer cell HGC-27 to provide potential new strategies for treating gastric cancer. In vitro cultures of the human gastric cancer cell lines MGC80-3, HGC-27, NCI-N87, and SGC-7901 and the normal human gastric epithelial cell line GES-1 were established. The expression of miR-34a in each gastric cancer cell line and GES-1 normal human gastric epithelial cell line was detected using quantitative real-time polymerase chain reaction (qRT-PCR). After the HGC-27 cells were transfected with a miR-34a mimic for 48 h, the changes in the expression levels of miR-34a were detected using qRT-PCR. The effect of miR-34a on HGC-27 cell viability was measured using a tetrazolium-based colorimetric [-(4,5)-dimethylthiahiazo-(-z-y1)-3,5-di-phenytetrazoliumromide (MTT)] assay. Flow cytometry was used to analyze the effects of miR-34a on HGC-27 cell proliferation. Annexin V/propidium iodide double staining and flow cytometry were used to analyze the effects of miR-34a on HGC-27 cell apoptosis. A Transwell invasion chamber was used to detect the effects of miR-34a on HGC-27 cell invasion. Finally, western blotting was used to analyze the effects of miR-34a on survivin protein expression. The qRT-PCR test determined that miR-34a expression in gastric cancer cells was significantly reduced compared to the normal gastric epithelial cell line GES-1 (p < 0.01). Compared to the control group, cellular miR-34a expression levels were significantly increased in HGC-27 human gastric carcinoma cells after transfection with a miR-34a mimic for 48 h (p < 0.01). The MTT assay demonstrated that after overexpressing miR-34a in HGC-27 cells, cellular viability was significantly reduced (p < 0.05). Flow cytometry analysis determined that upon miR-34a overexpression, the proliferation index decreased significantly (p < 0.05), and cellular apoptosis was significantly increased (p < 0.01). The Transwell invasion chamber assay illustrated that after increasing the expression of miR-34a, the number of cells passing through the Transwell chamber was significantly reduced (p < 0.01). Based on western blotting, compared with the control group, survivin protein expression levels were significantly decreased in the HGC-27 cells transfected with the miR-34a mimic for 48 h (p < 0.01). In conclusion, the expression level of miR-34a was downregulated in human gastric cancer cell lines. miR-34a can negatively regulate survivin protein expression and inhibit gastric cancer cell proliferation and invasion. Therapeutically enhancing miR-34a expression or silencing the survivin gene may benefit patients with gastric cancer.
authors.affiliation: Department of Oncology, Ruijin Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China.
authors.forename: Weiguo
authors.initials: W
authors.lastname: Cao
authors.forename: Rong
authors.initials: R
authors.lastname: Fan
authors.forename: Lifu
authors.initials: L
authors.lastname: Wang
authors.forename: Shidan
authors.initials: S
authors.lastname: Cheng
authors.forename: Hao
authors.initials: H
authors.lastname: Li
authors.forename: Jinsong
authors.initials: J
authors.lastname: Jiang
authors.forename: Mei
authors.initials: M
authors.lastname: Geng
authors.forename: Yening
authors.initials: Y
authors.lastname: Jin
authors.forename: Yunlin
authors.initials: Y
authors.lastname: Wu
mesh_terms: D017209:Apoptosis; D015153:Blotting, Western; D002465:Cell Movement; D049109:Cell Proliferation; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000235:genetics / Q000378:metabolism*; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D013274:Stomach Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D000077022:Survivin; D014407:Tumor Cells, Cultured
chemical_list: C107489:BIRC5 protein, human; D051034:Inhibitor of Apoptosis Proteins; C522425:MIRN34 microRNA, human; D035683:MicroRNAs; D000077022:Survivin