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Archive > Volume 31 Issue 2 > 2024,31(2):135-145. DOI:10.3872/j.issn.1007-385X.2024.02.004 Prev Next
Basic Research
lncRNA NEAT1 promotes the expression of EZH2 in gastric cancer cells and improves cell proliferation and migration through inhibiting hsa-miR-450b-5p
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Abstract:
Objective: To screen the upstream miRNAs and lncRNAs of EZH2 gene, analyze their expressions in gastric cancer cells,verify the targeting relationship between them, and discuss their effects on the proliferation, migration and apoptosis of gastric cancer cells. Methods: The upstream miRNA (has-miR-450b-5p) of EZH2 was queried, analyzed and screened by ENCORI, miRDB and Target Scan databases, and the upstream lncRNA (lncRNA NEAT1) of hsa-miR-450b-5p was screened by ENCORI database and DAINA database. The binding sites between hsa-miR-450b-5p, lncRNA NEAT1 and EZH2 were predicted. Dual-luciferase reporter assay was used to verify the binding relationship between hsa-miR-450b-5p and lncRNA NEAT1. The expression levels of lncRNA NEAT1 and EZH2 in normal gastric epithelial cells (GES-1) and gastric cancer cells (MGC-803, SGC-7901 and MKN-28) were detected by qPCR and WB. MGC-803 and SGC-7901 cells were divided into the hsa-miR-450b-5p-mimic group, the mimic-NC group,the si-NEAT1 group and the si-NC group according to different transfections, and the overexpression and knockdown effects were verified by qPCR 36~48 h after transfection. qPCR and WB were used to detect and observe the effects of overexpression of hsa-miR-450b-5p on the protein expressions of lncRNA NEAT1 and EZH2mRNA, in cells, and the effect of knockdown of lncRNA NEAT1 on the expressions of hsa-miR-450b-5p and EZH2 mRNA. CCK-8 method, scratch healing assay and flow cytometry were used to detect the effects of knockdown of EZH2 or knockdown of lncRNA NEAT1 on cell proliferation, migration, and apoptosis, respectively. Results: The upstream miRNA and lncRNA of EZH2 obtained through bioinformatic analysis and screening were has-miR-450b-5p and lncRNA NEAT1, and the targeting relationship between the two was verified by double luciferase reporter gene assay. lncRNA NEAT1 and EZH2 mRNA 、protein were highly expressed in gastric cancer cells (both P<0.05). Compared with the mimic-NC group,the levels of miR-450b-5p in MGC-803 and SGC-7901 cells in the hsa-miR-450b-5p-mimic group increased significantly, while the expressions of EZH2 mRNA, protein and lncRNA NEAT1 decreased significantly (P<0.05 or P<0.01). Compared with the si-NC group, the expressions of lncRNA NEAT1 and EZH2 mRNA in MGC-803 and SGC-7901 cells decreased significantly in the si-NEAT1 group (both P<0.01), and the expression of hsa-miR-450b-5p in SGC-7901 cells increased significantly (P<0.05). The proliferation and migration abilities of MGC-803 and SGC-7901 cells were significantly reduced after knockdown of EZH2 or lncRNA NEAT1 (both P<0.01). Conclusion: lncRNA NEAT1 and EZH2 were highly expressed in gastric cancer cells, and lncRNA NEAT1 promoted the expression of EZH2 and improved the proliferation and migration abilities of gastric cancer MGC-803 and SGC-7901 cells through hsa-miR-450b-5p.
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