HepG2 세포에서 에너지 상태를 활용한 화학물질의 미토콘드리아 손상 평가

Evaluation of Chemical-induced Mitochondrial Damage in HepG2 Cells Using Energy Charge

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In the presence study, we evaluated chemical-induced mitochondrial damage in HepG2 cells using energy charge, calculated by (ATP+0.5*ADP)/(ATP+ADP+AMP). ATP and its metabolites, ADP and AMP, were determined using LC-MS/MS. The cellular ATP, ADP and AMP were identified by comparison of mass spectra and retention time with their authentic standards. To establish sampling method, effects of extraction solvent composition on determination of ATP, ADP and AMP were evaluated. The stability of ATP, ADP and AMP was improved by using the sampling solution consisted of acetonitrile:methanol:water (4:4:2), relative to 10% perchloric acid solution. The energy charge of HepG2 cells treated with 0.02 μM rotenone, 25 μM troglitazone or 20 μM flutamide was decreased to 52, 83, or 84%, respectively, relative to control cells. The ratio of AMP to ATP was 13-, 2- or 2-fold higher in HepG2 cells treated with rotenone, troglitazone or flutamide, respectively, compared with control cells. In conclusion, the method established in this study can provide information of cellular energy status to predict chemical-induced mitochondrial damage.