五氯联苯PCB126对人肝癌细胞SMMC-7721有氧糖酵解的影响

Effect of PCB126 on aerobic glycolysis of human hepatocellular carcinoma cell SMMC-7721

目的探讨五氯联苯PCB126对人肝癌细胞有氧糖酵解的影响和机制。方法取人肝癌细胞SMMC-7721,用浓度分别为10^(-11)、10^(-10)、10^(-9)、10^(-8)、10^(-7) mol/L PCB126分别处理24、48 h,并以0.1%DMSO作为对照组。用试剂盒检测培养基中葡萄糖含量和乳酸生成量,以实时荧光定量PCR法检测丙酮酸激酶M2(PKM2)和有氧糖酵解通路中关键因子葡萄糖转运蛋白1(GLUT1)、乳酸脱氢酶(LDHA)、丙酮酸脱氢酶激酶(PDK)的表达。用RNA干扰技术敲减PKM2,检测其对培养基中葡萄糖含量、乳酸生成量和有氧糖酵解关键因子表达的影响。结果与对照组相比,人肝癌细胞SMMC-7721经10^(-10)、10^(-9)、10^(-8)mol/L PCB126处理48 h后,培养基中葡萄糖含量明显下降,乳酸生成量明显增加,差异有统计学意义(P<0.05);与对照组相比,处理48 h后对细胞存活率无明显影响,72 h时细胞存活率明显升高。PCB126暴露可明显升高PKM2、GLUT1、LDHA和PDK mRNA水平,差异有统计学意义(P<0.05)。用PKM2 shRNA敲减PKM2后,与10^(-9) mol/L PCB126处理组相比,培养基中葡萄糖含量升高,乳酸生成量下降,GLUT1、LDHA和PDK mRNA水平明显降低,差异均有统计学意义(P<0.01)。结论 PCB126可通过PKM2上调肝癌细胞GLUT1、LDHA和PDK的表达,从而促进有氧糖酵解,这可能与PCB126促进肝癌的发展有关。

Objective To investigate the effects and mechanism of PCB126 on the aerobic glycolysis of human hepatocellular carcinoma cell SMMC-7721. Methods The cells were treated with PCB126 at the doses of 10^-11, 10^-10, 10^-9, 10^-8, 10^-7mol/L and 0.1% DMSO treatment as the control for 24 or 48 h, respectively. Then, glucose residues and lactate production in the medium were measured by using glucose and lactate assay kits. Quantitative real-time PCR assay was employed for assessing the expression of PKM2 and key cytokines of aerobic glycolysis including GLUT1, LDHA and PDK. PKM2 sh RNA were used to detect the influence of PKM2 on PCB126-induced glucose consumption, lactate production and the expressions of GLUT1,LDHA and PDK. Results Glucose content was significantly reduced（P〈0.05） after human hepatocellular carcinoma cell BEL-7402 was treated with PCB126 10^-10, 10^-9, 10^-8mol/L for 48 h. And lactate acid production was significantly increased（P〈0.05）.Exposure to PCB126 at the doses of 10^-10, 10^-9, 10^-8mol/L for 48 hours induced the up-regulation of PKM2, GLUT1, LDHA and PDK. After knockdown of PKM2, the glucose content was increased and lactate acid production was reduced, compared with the PCB126 treatment group. Meanwhile, the expression of GLUT1, LDHA and PDK were significantly up-regulated（P〈0.01）. Conclusion PCB126 exposure may increase the expression of GLUT1, LDHA and PDK via PKM2, which may be involved in the development of hepatocellular carcinoma.