Abstract:

Objective To investigate the inhibitory effect of p16 gene on the growth of colon cancer cell line SW480. Methods p16 gene was cloned into eukaryotic express vector pEGFP-N3 using subclone technique, and the recombinant plasmid was transfected into SW480 cells using Lipofectamine method. MTT assay was performed to analyse the proliferation of cells without transfection (SW480 group), cells transfected with blank plasmid (SW480-GFP group) and cells transfected with p16 (SW480-p16 group) after culture for 24, 48 and 72 h. Real-Time PCR and Western blotting were employed to detect the expression of p16, CDK4 and cyclin D1 mRNA and protein respectively.ResultsCompared with SW480 group and SW480-GFP group, the expression of p16 gene in SW480-p16 group was significantly increased (P<0.01). MTT assay indicated that the cell proliferation in SW480-p16 group was significantly lower than that in SW480 group and SW480GFP group after culture for 48 h and 72 h (P<0.01). Real-Time PCR analysis revealed the expression of p16 mRNA in SW480-p16 group which was 2.59 times of that in SW480, and 2.38 times in SW480-GFP (P<0.01). The expression of cyclin D1 mRNA in SW480-p16 group was significantly lower than that in SW480 group and SW480-GFP group （P<0.05）, and the expression of CDK4 mRNA in SW480-p16 group was slightly lower than that in SW480 group and SW480-GFP group (P>0.05). Western blotting demonstrated that the expression of p16 protein in SW480-p16 group was significantly higher than that in SW480-GFP group and SW480 group （P<0.01), while the expression of CDK4 and cyclin D1 protein in SW480-p16 group was significantly lower than that in SW480-GFP group and SW480 group （P<0.01). Conclusion Colon cancer cells transfected with p16 gene can resume the function of inhibiting tumor growth.

Key words: p16 gene, clone, colon cancer cells, cyclin D1, cyclin dependent kinase 4 p16