Abstract:

Objective To investigate the effect of silent information regulator 2（SIRT2）on the aerobic glycolysis， growth and proliferation of the colorectal cancer cells， and to clarify its related mechanism. Methods The HCT116 cells and SW480 cells were cultured， and Western blotting method was used to detect the expression levels of SIRT2 protein in two kinds of cells.The HCT116 cells with high expression of SIRT2 protein were selected for the following RNA interference experiment. Negative control group，SIRT2-siRNA#1 group， SIRT2-siRNA#2 group， and SIRT2-siRNA#3 group were established； the SW480 cells with low expression of SIRT2 protein were selected for the following SIRT2 overexpression experiment.The SIRT2 recombinant plasmids were constructed， and the SW480 cells at logarithmic growth phase were selected； blank group， empty vector plasmid transfection group， and SIRT2 plasmid transfection group were established；the recovery experiment was carried out in the HCT116 cells；control group， SIRT2-siRNA+glucose-6-phosphate dehydrogense（G6PD） plasmid co-transfection group and SIRT2-siRNA#3 group were established.The plasmid and siRNA were transfected into the SW480 cells and HCT116 cells by Lipofectamine 2000， respectively，the conterts of glucose in the culture medium in various groups were measured by glucose oxidase method after silencing SIRT2 and over-expression of SIRT2；the levels of lactic acid in the culture medium in various groups were measured by colorimetry after silencing SIRT2 and over-expression of SIRT2；the clone formation rates of the cells in various groups were determined by clonal formation assay； the proliferation activities of the cells in various groups were determined by CCK-8 assay；the expression levels of G6PD mRNA in the cells after silencing SIRT2 and over-expression of SIRT2 were determined by reverse transcriptionPCR（RT-PCR） method；the expression levels of G6PD protein in the cells in various groups after silencing SIRT2 and over-expression of SIRT2 were detected by Western blotting method；the expression levels of SIRT2 and G6PD proteins in colorectal cancer tissue were determined by immunohistochemistry.After co-transfection of SIRT2-siRNA and G6PD plasmids， the glucose levels，lactic acid levels，survival rates， and proliferation abilities of the cells were detected. Results Compared with negative control group， the level of glucose in the culture medium in SIRT2-siRNA#3 group was increased significantly（P<0.05）， the level of lactic acid was significantly decreased（P<0.05），the survival rate and proliferation ability of the cells were significantly decreased（P<0.05），the expression levels of G6PD mRNA and protein were decreased （P<0.05）；compared with empty vector plasmid transfection group，the level of glucose in the culture medium in SIRT2 plasmid transfection group was decreased significantly（P<0.05）， the level of lactic acid was significantly increased（P<0.05）， the proliferation ability and clone formation rate of the cells were significantly increased（P<0.05）.The expression levels of SIRT2 and G6PD proteins in colorectal cancer tissue were significantly higher than that in paracancerous tissue（P<0.05）. Compared with SIRT2-siRNA#3 group， the level of glucose in the culture medium in SIRT2-siRNA+G6PD plasmid co-transfection group was significantly decreased（P<0.05）， the level of lactic acid in the culture medium was significantly increased（P<0.05），and the proliferation ability and clone formation rate of the cells were significantly increased（P<0.05）. Conclusion SIRT2 promotes the aerobic glycolysis and growth and proliferation of the colorectal cancer cells，and its mechanism may be related to the expression of G6PD regulated by SIRT2.

Key words: Colorectal neoplasm, Silent information regulator 2, Glucose-6-phosphate dehydrogenase, Aerobic glycolysis, Growth and proliferation