抑制线粒体氧化磷酸化系统各复合物功能对糖酵解的影响

摘要

目的 探讨抑制线粒体氧化磷酸化系统各复合物对细胞糖酵解的影响.方法 培养HepG2细胞,铺于96孔板,用不同浓度的线粒体氧化磷酸化系统各复合物抑制剂处理24h后,通过检测乳酸脱氢酶(LDH)释放量确定其最大无毒剂量;用小于该最大无毒剂量的各复合物抑制剂处理细胞后,通过检测培养基的葡萄糖和乳酸浓度,判断各复合物抑制剂对细胞糖酵解的影响.结果 线粒体复合物Ⅰ、Ⅲ、Ⅴ抑制剂二甲双胍(metformin,Met)、黏噻唑(myxothiazol,Myxo)和寡霉素A(oligomycin A,Olig)可促进HepG2细胞葡萄糖消耗和乳酸生成,且其中二甲双胍的作用最强,黏噻唑其次,寡霉素A的作用最弱;线粒体复合物Ⅱ抑制剂2-噻吩甲酰三氟丙酮(2-thenoyltrifluoroacetone,TTFA)则可减少HepG2细胞葡萄糖消耗和乳酸生成;线粒体复合物Ⅳ抑制剂氰化钾(KCN)对HepG2细胞葡萄糖消耗和乳酸生成无影响.结论 抑制线粒体复合物Ⅰ、Ⅲ、Ⅴ对细胞糖代谢是有益处的;抑制线粒体复合物Ⅳ对细胞糖代谢无作用;而抑制线粒体复合物Ⅱ阻碍了细胞的糖代谢.%Objective To investigate the effect of inhibiting each complexes in mitochondrial oxidative phosphorylation (OXPHOS) system on glycolysis.Methods After human liver hepatocellular cell line HepG2 were treated with different concentrations of each inhibitor of the complexes in OXPHOS system about 24 hours,lactic dehydrogenase(LDH) release test was performed to investigate the highest nontoxic dose of each inhibitor.Then,after the cells were treated with lower concentrations than the highest nontoxic dose about 24 hours,glucose consumption and lactate production were tested to evaluate the anaerobic glycolysis effect of these inhibitors.Results The inhibition of complex Ⅰ,Ⅲ and ATP synthase(complex Ⅴ) increased the glucose consumption and lactic release in HepG2,among which the effect of complex Ⅰ inhibitor metformin was the most significant,and the effect of ATP synthase inhibitor Oligomycin A was the weakest.However,The inhibitor of complex Ⅱ 2-Thenoyltrifluoroacetone (TTFA) reduced the glucose consumption and lactic release in HepG2.Last but not least,The inhibitor of complex Ⅳ KCN had no significant effect on glycolysis in HepG2.Conclusion The inhibition of Ⅰ,Ⅲ and Ⅴ can be beneficial to cell glucose metabolism.However,inhibition of complex Ⅱ can reduce cell glycolysis,which is not ideal for promoting glucose consumption,and inhibition of complex Ⅳ have no effect on cell glycolysis.