lnc RNA MAFG-AS1调控miR-532-3p表达对肺癌A549细胞糖酵解的影响

Effect of lncRNA MAFG-AS1 regulating miR-532-3p expression on glycolysis of lung cancer A549 cells

目的:探讨lncRNA MAFG反义RNA1(MAFG-AS1)调控miR-532-3p表达对肺癌A549细胞糖酵解的影响。方法:用qPCR法检测人肺癌A549细胞和正常肺上皮细胞BEAS-2B中MAFG-AS1和miR-532-3p的表达水平。利用脂质体介导转染技术,分别构建MAFG-AS1表达下调和miR-532-3p过表达的A549细胞,用分光光度法检测细胞培养上清中葡萄糖消耗量与乳酸含量,qPCR和WB法分别检测细胞中M2型丙酮酸激酶(pyruvate kinase M2,PKM2)、己糖激酶2(hexokinase 2,HK2)mRNA和蛋白的表达水平。用双荧光素酶报告基因实验验证MAFG-AS1与miR-532-3p的靶向关系,观察MAFG-AS1和miR-532-3p同时低表达对A549细胞葡萄糖及乳酸含量和PKM2、HK2表达的影响。结果:与BEAS-2B细胞比较,A549细胞中MAFG-AS1表达上调而miR-532-3p表达下调(均P<0.01)。下调MAFG-AS1或miR-532-3p过表达均可降低A549细胞葡萄糖消耗量及乳酸含量,并抑制PKM2、HK2 mRNA和蛋白的表达(均P<0.01)。miR-532-3p可与MAFG-AS1靶向结合抑制野生型MAFG-AS1细胞的荧光素酶活性(P<0.01),下调MAFG-AS1可使A549细胞中miR-532-3p表达升高(P<0.01)。miR-532-3p低表达可逆转MAFG-AS1表达下调对细胞葡萄糖消耗量及乳酸含量和PKM2、HK2表达的影响(均P<0.01)。结论:下调MAFG-AS可通过促进miR-532-3p表达而抑制肺癌A549细胞糖酵解。

Objective:To investigate the effect of lncRNA MAFG antisense RNA1(lncRNA MAFG-AS1)regulating miR-532-3 p on the glycolysis of lung cancer A549 cells.Methods:The expression levels of MAFG-AS1 and miR-532-3 p in human lung cancer A549 cells and normal lung epithelial BEAS-2 B cells were detected by q PCR.A549 cells with MAFG-AS1 downregulation or miR-532-3 p overexpression were constructed by liposome transfection technique,respectively.The glucose consumption and lactate secretion in cell culture supernatant of A549 cells were detected by visible spectrophotometry,and the mRNA and protein expression levels of pyruvate kinase M2(PKM2)and hexokinase 2(HK2)in A549 cells were detected using qPCR and WB,respectively.The targeting relationship between MAFG-AS1 and miR-532-3 p was verified by Dual luciferase reporter gene assay,and the effects of co-downregulation of MAFG-AS1 and miR-532-3 p on glucose uptake,lactate secretion and expression of PKM2 and HK2 in A549 cells were observed.Results:Compared with BEAS-2 B cells,the expression of MAFG-AS1 was upregulated while miR-532-3 p expression was downregulated in A549 cells(all P<0.01).Glucose consumption,lactate secretion and the protein and m RNA expressions of PKM2 and HK2 were inhibited by MAFG-AS1 downregulation or miR-532-3 p overexpression(all P<0.01).miR-532-3 p could bind to MAFG-AS1 and inhibit the luciferase activity of wild-type MAFG-AS1 cells(P<0.01).Downregulation of MAFG-AS1 could increase the expression of miR-532-3 p(P<0.01),while the co-downregulation of miR-532-3 p could reverse the effect of MAFG-AS1 downregulation on glucose uptake,lactate secretion and the expression of PKM2 and HK2 in A549 cells(all P<0.01).Conclusion:Downregulation of MAFG-AS1 can inhibit the glycolysis of A549 cells by promoting the expression of miR-532-3 p.