miR-320通过靶向E2F1基因抑制结直肠癌细胞的糖代谢

miR-320 inhibits glycometabolism in colorectal cancer by targeting E2F1 gene

目的研究微小RNA（miR320）靶向E2F1基因对结直肠癌的肿瘤糖代谢的影响。方法使用实时荧光定量聚合酶链反应（qRTPCR）检测miR320在结直肠癌细胞株和癌组织中的表达水平。生物信息学预测miR320与E2F1的结合位点。荧光素酶实验检测miR320是否可靶向调节E2F1。从mRNA水平及蛋白水平验证E2F1与miR320相互关系，使用葡萄糖/葡萄糖氧化酶法试剂盒和乳酸检测试剂盒分析在SW480和LOVO细胞中过表达miR320及干扰E2F1表达后，肿瘤细胞糖代谢的变化。结果qRTPCR测得miR320在结直肠癌细胞株（F=42.327，P＜0.001）和癌组织（t=4.345，P=0.023）中低表达，荧光素酶报告基因检测miR320可靶向负调节E2F1的表达（t=4.716，P=0.042），mRNA水平（t=4.780，P=0.041；t=5.506，P=0.031）和蛋白水平证实在LOVO和SW480细胞株中E2F1与miR320可相互作用，在SW480和LOVO中过表达miR320可使细胞上清中的葡萄糖（t=5.262，P=0.034；t=21.079，P=0.002）和乳酸（t=9.609，P=0.011；t=18.582，P=0.003）含量降低，同时降低E2F1的表达，则可加强miR320对葡萄糖（t=5.128，P=0.036；t=5.089，P=0.037）和乳酸（t=8.573，P=0.013；t=13.364，P=0.006）含量的抑制作用。结论E2F1是miR320的靶基因，miR320通过靶向E2F1基因调节结直肠癌细胞的肿瘤糖代谢。

ObjectiveTo study the effect of microRNA320 （miR320） targeting E2F1 gene on tumor glycometabolism in colorectal cancer. MethodsThe miR320 expression level in colorectal cancer cell lines and cancer tissues was detected using quantitative realtime polymerase chain reaction （qRTPCR）. The binding sites of miR320 and E2F1 were predicted by bioinformatics. Luciferase assay was used to detect the targeting regulation of miR320 on E2F1. The relationship between E2F1 and miR320 was verified in mRNA level and protein level. When the miR320 in SW480 and LOVO cells was upregulated and the E2F1 was downregulated, the changes of glycometabolism in tumor cells were analyzed using glucose/glucose oxidase kit and lactate test kit. ResultsThe qRTPCR results showed low expressions of miR320 in colorectal cancer cell lines and cancer tissues （F=42.327, P〈0.001; t=4.345, P=0.023）. Luciferase assay showed that miR320 could negatively regulate the expression of E2F1 （t=4.716, P=0.042）. The expression levels of E2F1 protein and mRNA （t=4.780, P=0.041; t=5.506, P=0.031） confirmed that miR320 could interact with E2F1 in LOVO and SW480 cells. Overexpression of miR320 could reduce the contents of glucose （t=5.262, P=0.034; t=21.079, P=0.002） and lactic acid （t=9.609, P=0.011; t=18.582, P=0.003） in the cellular supernatant in SW480 and LOVO cells. Downregulating the expression of E2F1 at the same time could enhance the inhibitory effect of miR320 on glucose （t=5.128, P=0.036; t=5.089, P=0.037） and lactic acid （t=8.573, P=0.013; t=13.364, P=0.006）. ConclusionE2F1 is the target gene of miR320, and miR320 can regulate the glycometabolism of colorectal cancer cells by targeting E2F1 gene.