Dan Hu1, Chun-qi Wu1, Ze-jun Li2, Yue Liu1, Xing Fan1, Quan-jun Wang3, Ri-gao Ding4. 1. State Key Laboratory of Toxicology and Medical Countermeasures, Institute of Pharmacology and Toxicology, Academy of Military Medical Sciences, 27 Taiping Road, Beijing 100850, PR China. 2. State Key Laboratory of Toxicology and Medical Countermeasures, Institute of Pharmacology and Toxicology, Academy of Military Medical Sciences, 27 Taiping Road, Beijing 100850, PR China; Guang Dong Pharmaceutical University, Guangzhou 510006, PR China. 3. State Key Laboratory of Toxicology and Medical Countermeasures, Institute of Pharmacology and Toxicology, Academy of Military Medical Sciences, 27 Taiping Road, Beijing 100850, PR China. Electronic address: wangquanjunbeijing@163.com. 4. State Key Laboratory of Toxicology and Medical Countermeasures, Institute of Pharmacology and Toxicology, Academy of Military Medical Sciences, 27 Taiping Road, Beijing 100850, PR China. Electronic address: dingrigao@nic.bmi.ac.cn.
Abstract
OBJECTIVE: To characterize the mechanism of action of thiazolidinedione (TZD)-induced liver mitochondrial toxicity caused by troglitazone, rosiglitazone, and pioglitazone in HepaRG cells. METHODS: Human hepatoma cells (HepaRG) were treated with troglitazone, rosiglitazone, or pioglitazone (12.5, 25, and 50μM) for 48h. The Seahorse Biosciences XF24 Flux Analyzer was used to measure mitochondrial oxygen consumption. The effect of TZDs on reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) were detected by flow cytometry. The mitochondrial ultrastructure of HepaRG cells was observed under a transmission electrical microscope (TEM). mtDNA content was evaluated by real-time PCR, and ATP content and mitochondrial respiratory chain (MRC) complex I, II, III, IV activity were measured via chemiluminescence. Results were considered statistically significant at p<0.05. RESULTS: Among the three drugs, troglitazone exhibited the highest potency, followed by rosiglitazone, and then pioglitazone. The TZDs caused varying degrees of mitochondrial respiratory function disorders including decreases in oxygen consumption, MRC activity, and ATP level, and an elevation in ROS level. TZD treatment resulted in mtDNA content decline, reduction in MMP, and alterations of mitochondrial structure. CONCLUSION: All investigated TZDs show a certain degree of mitochondrial toxicity, with troglitazone exhibiting the highest potency. The underlying mechanism of TZD-induced hepatotoxicity may be associated with alterations in mitochondrial respiratory function disorders, oxidative stress, and changes in membrane permeability. These parameters may be used early in drug development to further optimize risk:benefit profiles.
OBJECTIVE: To characterize the mechanism of action of thiazolidinedione (TZD)-induced liver mitochondrial toxicity caused by troglitazone, rosiglitazone, and pioglitazone in HepaRG cells. METHODS:Humanhepatoma cells (HepaRG) were treated with troglitazone, rosiglitazone, or pioglitazone (12.5, 25, and 50μM) for 48h. The Seahorse Biosciences XF24 Flux Analyzer was used to measure mitochondrial oxygen consumption. The effect of TZDs on reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) were detected by flow cytometry. The mitochondrial ultrastructure of HepaRG cells was observed under a transmission electrical microscope (TEM). mtDNA content was evaluated by real-time PCR, and ATP content and mitochondrial respiratory chain (MRC) complex I, II, III, IV activity were measured via chemiluminescence. Results were considered statistically significant at p<0.05. RESULTS: Among the three drugs, troglitazone exhibited the highest potency, followed by rosiglitazone, and then pioglitazone. The TZDs caused varying degrees of mitochondrial respiratory function disorders including decreases in oxygen consumption, MRC activity, and ATP level, and an elevation in ROS level. TZD treatment resulted in mtDNA content decline, reduction in MMP, and alterations of mitochondrial structure. CONCLUSION: All investigated TZDs show a certain degree of mitochondrial toxicity, with troglitazone exhibiting the highest potency. The underlying mechanism of TZD-induced hepatotoxicity may be associated with alterations in mitochondrial respiratory function disorders, oxidative stress, and changes in membrane permeability. These parameters may be used early in drug development to further optimize risk:benefit profiles.
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