CircEPSTI1 Regulates Cisplatin Resistance of Gastric Cancer Cells by Targeting miR-216b-5p
GUO Hailong, MA Ye, WANG Xiaoyun, SHI Peidong, WAN Boshun*
The objective was to observe the effect of circEPSTI1 on cisplatin resistance of gastric cancer cells by targeting miR-216b-5p and to explore its possible mechanism. The qRT-PCR (quantitative real-time polymerase chain reaction) was used to detect the expression of circEPSTI1 and miR-216b-5p in gastric cancer tissues, adjacent tissues, human gastric mucosal cells GES-1, human gastric cancer cells HGC27 and gastric cancer drug-resistant cells HGC27/DDP. The dual luciferase reporting assay was used to verify the targeted regulatory relationship between circEPSTI1 and miR-216b-5p. HGC27/DDP cells were selected as the research object and divided into four groups: the si-NC group, the si-circEPSTI1 group, the si-circEPSTI1+anti-miR-NC group, and the si-circEPSTI1+anti-miR-216b-5p group. Cell proliferation, clonal formation, apoptosis, migration, and invasion were detected by CCK-8 assay, plate clonal formation assay, flow cytometry, and Transwell assay, respectively. Western blot was used to detect the protein expression of Cleavea-caspase 3, Cleavea-caspase 9, E-cadherin, and N-cadherin. Compared with adjacent tissues, the results showed that circEPSTI1 expression in gastric cancer tissues was up-regulated (P<0.05) and miR-216b-5p expression was down-regulated (P<0.05). Compared with GES-1 cells, circEPSTI1 expression in HGC27 cells and HGC27/DDP cells was up-regulated (P<0.05), while miR-216b-5p expression was down-regulated (P<0.05). Compared with HGC27 cells, circEPSTI1 expression in HGC27/DDP cells was up-regulated (P<0.05) and miR-216b-5p expression was down-regulated (P<0.05). Overexpression of miR-216b-5p could reduce the luciferase activity of wt-circEPSTI1 (P<0.05), but did not affect the luciferase activity of mut-circEPSTI1. Compared with the si-NC group, cell proliferation inhibition rate, apoptosis rate, Cleavea-caspase 3, Cleavea-caspase 9, and E-cadherin protein expression in the si-circEPSTI1 group were up-regulated (P<0.05). The clone formation numbers, migration, and invasion cells were decreased (P<0.05), and the expression of N-cadherin protein was down-regulated (P<0.05). Compared with the si-circEPSTI1+anti-miRNC group, cell proliferation inhibition rate, apoptosis rate, Cleavea-caspase 3, Cleavea-caspase 9, and E-cadherin expression were down-regulated in the si-circEPSTI1+anti-miR-216b-5p group (P<0.05). The number of clone formation, migration, and invasion cells increased (P<0.05), and the expression of N-cadherin protein was upregulated (P<0.05). The results showed that circEPSTI1 promoted the proliferation, clone formation, migration, and invasion, and inhibited the apoptosis in drug-resistant gastric cancer cells by targeting the expression of miR216b-5p, thus promoting the drug resistance of gastric cancer cells to DDP. This mechanism may be related to the change of apoptosis-related protein expression and the reversal of the epithelial-mesenchymal transformation process.
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