Inhibition of energy-producing pathways of HepG2 cells by 3-bromopyruvate
Inhibition of energy-producing pathways of HepG2 cells by 3-bromopyruvate
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DOI：
10.1042/bj20080805
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发表时间：
2009-02-01
影响因子：
4.1
通讯作者：
Galina, Antonio
中科院分区：
文献类型：
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作者：
Pereira da Silva, Ana Paula;El-Bacha, Tatiana;Galina, Antonio
3-溴丙酮酸（3-溴丙酮酸）是一种对肝癌具有抗肿瘤活性的烷基化剂。该化合物通过糖酵解和氧化磷酸化抑制细胞ATP的产生；然而，人类肝细胞癌中3-BrPA作用的具体代谢步骤和机制，特别是其对线粒体能量学的影响，尚不清楚。本研究发现，HepG2细胞与低浓度3-BrPA孵育短时间（150 μ M 30 min）显著影响糖酵解和线粒体呼吸功能。150 μ M 3-BrPA对线粒体己糖激酶活性没有抑制作用，但该浓度对GAPDH（甘油醛-3-磷酸脱氢酶）和3-磷酸甘油激酶活性的抑制作用超过70%。此外，3-BrPA处理显著损害了HepG2细胞的乳酸生成，即使从培养培养基中去除葡萄糖也是如此。用3-BrPA在无葡萄糖培养基中预孵育HepG2细胞时，丙酮酸/苹果酸或琥珀酸支持的HepG2细胞耗氧量均受到抑制。另一方面，当细胞在葡萄糖补充的培养基中预孵育时，仅当琥珀酸盐作为可氧化底物时，耗氧量受到影响。3-BrPA处理的HepG2细胞仅在葡萄糖补充培养基中孵育时，低霉素非依赖性呼吸均增加，表明3-BrPA诱导线粒体质子泄漏并阻断电子传递系统。经3-BrPA处理后，琥珀酸脱氢酶活性被抑制70%。这些结果表明，3-BrPA油琥珀酸脱氢酶和糖酵解的联合作用，抑制葡萄糖磷酸化的下游步骤，在HepG2细胞死亡中起重要作用。
3-BrPA (3-bromopyruvate) is an alkylating agent with anti-tumoral activity on hepatocellular carcinoma. This compound inhibits cellular ATP production owing to its action on glycolysis and oxidative phosphorylation; however, the specific metabolic steps and mechanisms of 3-BrPA action in human hepatocellular carcinomas, particularly its effects on mitochondrial energetics, are poorly understood. In the present study it was found that incubation of HepG2 cells with a low concentration of 3-BrPA for a short period (150 mu M for 30 min) significantly affected both glycolysis and mitochondrial respiratory functions. The activity of mitochondrial hexokinase was not inhibited by 150 mu M 3-BrPA, but this concentration caused more than 70% inhibition of GAPDH (glyceraldehyde-3-phosphate dehydrogenase) and 3-phosphoglycerate kinase activities. Additionally, 3-BrPA treatment significantly impaired lactate production by HepG2 cells, even when glucose was withdrawn from the incubation medium. Oxygen consumption of HepG2 cells supported by either pyruvate/malate or succinate was inhibited when cells were pre- incubated with 3-BrPA in glucose-free medium. Oil the other hand, when cells were pre-incubated in glucose-supplemented medium, oxygen consumption was affected only when succinate was used as the oxidizable substrate. Ail increase in oligomycin-independent respiration was observed in HepG2 cells treated with 3-BrPA only when incubated in glucose-supplemented medium, indicating that 3-BrPA induces mitochondrial proton leakage as well as blocking the electron transport system. The activity of succinate dehydrogenase was inhibited by 70% by 3-BrPA treatment. These results suggest that the combined action of 3-BrPA oil succinate dehydrogenase and on glycolysis, inhibiting steps downstream of the phosphorylation of glucose, play an important role in HepG2 cell death.