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Impact of inhibition of miRNA-21 on biological functions of colorectal cancer cells
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Abstract:
Objective : To determine the effects of microRNA-21 (miR-21) inhibition on the various functional aspects (e.g., proliferation, apoptosis, cell cycle and invasion and migration) of colon cancer HCT116 cells in vitro. Methods: HCT116 cells were transfected with an miR-21 inhibitor, a non-sequence specific inhibitor as a negative control and the transfection reagent as a mock control, respectively. The level of miR-21 in transfected HCT116 was determined by Real-time PCR. The Proliferation, apoptosis, cell cycle progression, invasion and migration of the transfectants were evaluated by methyl thiazolyl tetrazolium (MTT) assay, flow cytometry (FCM), transwell invasion and migration assays, respectively. Protein level of phosphatase and tensin homolog (PTEN) and PTEN promoter activity in HCT116 cells after transfection were evaluated by Western blotting analysis and the luciferase reporter assay, respectively. Results: The level of miR-21 was significantly lower in HCT116 cells transfected with the miR-21 inhibitor than in cells transfected with non-specific inhibitor or mock. At 72 hours after transfection, the miR-21 inhibitor significantly suppressed HCT116 cell proliferation (1.05±0.45 vs 1.43±0.02 and 1.45±0.01, P<0.001), significantly increased HCT116 cell apoptosis (\[16.30±1.00\]% vs \[1.87±0.53\]% and \[1.86±0.12\]%, P<0.0001) and HCT cell cycle arrest in G0/G1 phase, and significantly decreased the invasion (50±2.0 vs 115±3.0 and 111±3.0, P<0.0001) and migration abilities (22±20 vs 52.3±2.5 and 53.0±1.0, P<0.0001), as compared with the two controls. Moreover, miR-21 inhibition resulted in remarkable increases in protein levels and activity of PTEN in HCT116 cells. Conclusion: MicroRNA21 may regulate proliferation/apoptosis, invasion and migration of colorectal cancer cells, possibly through modulating the expression and activity of PTEN.
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Project Supported:
Project supported by the Natural Science Foundation of Chongqing (No. csts2012JJA0038)
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