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Published on: March 30, 2019
MicroRNA-215 Regulates the Apoptosis of HCT116 Colon Cancer Cells by Inhibiting X-Linked Inhibitor of Apoptosis
Chuanhui Lu1, Ming Hong1, Bo Chen1
1Department of Gastrointestinal Surgery, Xiamen Cancer Hospital, The First Affiliated Hospital of Xiamen University, Xiamen, P.R. China.
Abstract:
X-linked inhibitor of apoptosis protein (XIAP) is the strongest member of the family of inhibitor of apoptosis protein. Studies found that the expression of XIAP in colon cancer tissue was significantly higher than that in adjacent tissues. Studies have shown that the expression of microRNA-215 (miR-215) was significantly lower than that of the adjacent tissues. This study investigated whether dysregulated miR-215 and XIAP play important roles in colon cancer cell apoptosis and the incidence of colon cancer. Forty-two patients with colorectal cancer (CRC) diagnosed and treated in the authors' hospital were selected. Human CRC cell line HCT116 and normal colonic mucosal epithelial cells (CMECs) were used. Luciferase reporter gene vector was constructed and dual-luciferase reporter gene assay was performed. HCT116 cells were cultured in vitro and divided into five groups: mimic normal control (NC) group, miR-215 mimic group, si-NC group, si-XIAP group, and miR-215 mimic + si-XIAP group. Western blot and polymerase chain reaction were conducted to examine XIAP and caspase-3. Apoptosis was detected by flow cytometry and cell proliferation was detected by cell counting kit-8 assay. Compared with the adjacent tissues, the expression of miR-215 in colon cancer tissue was significantly lower, whereas the expression of XIAP in colon cancer tissue was significantly higher. The apoptosis rate and miR-215 expression level of HCT116 cells were lower than that of normal CMECs, whereas XIAP expression was significantly higher than that in normal colon mucosa epithelial cells. MiR-215 targeted the 3'-untranslated regions of XIAP and inhibited its expression. Overexpressing miR-215 and (or) silencing XIAP expression could significantly enhance the activity of caspase-9 and caspase-3, and promote the apoptosis of HCT116 cells. MiR-215 inhibited the expression of XIAP and promoted the apoptosis of HCT116 cells.
Insights
MicroRNA-215 (miR-215) suppresses colon cancer by inhibiting X-linked inhibitor of apoptosis protein (XIAP). Restoring miR-215 or reducing XIAP promotes colon cancer cell apoptosis.
Area of Science:
- Oncology
- Molecular Biology
- Gene Regulation
Background:
- X-linked inhibitor of apoptosis protein (XIAP) is upregulated in colon cancer tissues.
- MicroRNA-215 (miR-215) expression is downregulated in colon cancer tissues.
- Dysregulation of miR-215 and XIAP may influence colon cancer cell apoptosis and incidence.
Purpose of the Study:
- To investigate the role of dysregulated miR-215 and XIAP in colon cancer cell apoptosis.
- To determine the relationship between miR-215 and XIAP expression in colon cancer.
- To explore the therapeutic potential of modulating miR-215 and XIAP in colon cancer.
Main Methods:
- Analysis of miR-215 and XIAP expression in patient tissues and cell lines.
- Luciferase reporter gene assay to confirm direct targeting of XIAP by miR-215.
- In vitro experiments using HCT116 cells with miR-215 mimics and XIAP inhibitors.
- Western blot, PCR, flow cytometry, and cell counting kit-8 assay to assess apoptosis and proliferation.
Main Results:
- Colon cancer tissues showed significantly lower miR-215 and higher XIAP expression compared to adjacent tissues.
- HCT116 cells exhibited lower apoptosis and miR-215 levels, and higher XIAP expression than normal CMECs.
- MiR-215 directly targets XIAP's 3'-UTR, inhibiting its expression.
- Overexpression of miR-215 and/or silencing XIAP significantly increased caspase-3/9 activity and promoted HCT116 cell apoptosis.
Conclusions:
- MiR-215 acts as a tumor suppressor in colon cancer by inhibiting XIAP expression.
- Modulating miR-215 and XIAP levels offers a potential therapeutic strategy for promoting colon cancer cell apoptosis.
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