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Archive > Volume 27 Issue 11 > 2020,27(11):1213-1219. DOI:10.3872/j.issn.1007-385X.2020.11.003 Prev Next
Basic Research
siRNA knocking down the expression of NEK2 gene enhances the chemosensitivity of colorectal carcinoma cells to 5-FU
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Abstract:
Objective：To investigate the effects of RNA interfering NEK2 (NIMA-related kinase 2) gene on the sensitivity of colorectal carcinoma (CRC) cell lines to 5-fluorouracil (5-FU) and the possible mechanisms. Methods: The mRNA and protein expressions of NEK2 in CRC cell lines were detected by quantitative polymerase chain reaction (qPCR) and Western blotting assay,respectively. siRNAs targeting NEK2 were constructed and transfected into CRC HCT116 and SW620 cells. The experiments were divided into positive interference group 1 (transfection with NEK2 siRNA1), positive interference group 2 (transfection with NEK2 siRNA2) and negative control group (transfection with si-NC), all the CRC cells were treated with 5-FU. CCK-8 assay, Flow cytometry and V-FICT/PI Annexin staining analysis were used to observe the effects of NEK2 gene knockdown on proliferation, cell cycle distribution and apoptosis of CRC cells under 5-FU treatment. Western blotting was used to detect the effects of NEK2 gene knockdown on the expression of Wnt/β-catenin signaling pathway related proteins in CRC cells under 5-FU treatment. Results: NEK2 was highly expressed in CRC HCT116 and SW620 cells at both protein and mRNA levels (all P<0.05). siRNA NEK2 transfection could effectively inhibit the protein and mRNA expressions of NEK2 in HCT116 and SW620 cells (all P<0.01). After treatment with various concentrations of 5-FU, the cell survival rate and IC50, as well as the expression levels of β-catenin (cytoblasts), C-MYC and Cyclin D1,in the positive interference group 1 and 2 were significantly decreased, while cell cycle blockage at G0/G1 phase, the apoptosis rate and the expression level of β-catenin (cytoplasm) were significantly increased (all P<0.01), as compared with the negative control group (all P<0.01). Conclusion: Silencing NEK2 gene can effectively improve the sensitivity of human colorectal cancer cells to 5-FU, which may be achieved by regulating the expression of Wnt/β-catenin signaling pathway related proteins.
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Project Supported:
Project supported by the Medical Science and Technology Research Program of Henan Province (No. 201702246)
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