Host: The Japanese Society of Toxicology
Name : The 47th Annual Meeting of the Japanese Society of Toxicology
Date : 2020 -
【Purpose】Mitochondrial toxicity is paid attention as a factor of drug-induced liver injury. For the assessment of mitochondrial toxicity induced cell death in vitro, sugar resource substitution from glucose in the culture medium to galactose helps maintain higher mitochondrial activity in cultured hepatocytes. However, mitochondrial permeability transition (MPT)-induced toxicity can’t be detected by only substituting sugar resource. Recently, we reported that lipopolysaccharide-induced mitochondrial reactive oxygen species (mtROS) sensitized MPT-induced cell death by diclofenac in vivo. Based on this, we aim to build a comprehensive cell-based assay system for MPT-inducers by loading mtROS in vitro.
【Methods】To load mtROS, primary mouse hepatocytes cultured in either glucose or galactose media were incubated in 1% O2 for 4 hr, then 40% O2 for 20 hr (designated H/R). During this time, the drug was exposed for total 24 hr. Cell death was evaluated by lactate dehydrogenase leakage.
【Result・Discussion】Three well-known MPT-inducers (benzbromarone, flutamide, troglitazone) significantly induced cell death only in combination with H/R and galactose. Such enhancement of cell death was not observed in hepatocytes lacking cyclophilin D (regulator of MPT pore opening). Drugs which have other mitochondrial toxicity mechanisms caused highest cell death in H/R and galactose. In conclusion, MPT could be evaluated with a new cell-based assay combining galactose and H/R. This cell-based system is useful for comprehensive assessment of the MPT inducibility of parent drugs and drug metabolites.
