BJA Advance Access originally published online on November 9, 2008
British Journal of Anaesthesia 2009 102(1):80-89; doi:10.1093/bja/aen322
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Apoptotic insults to human HepG2 cells induced by S-(+)-ketamine occurs through activation of a Bax-mitochondria-caspase protease pathway
1 Department of Pediatrics, Cathay General Hospital, Taipei, Taiwan, Republic of China
2 Graduate Institute of Clinical Medicine
3 Graduate Institute of Medical Sciences, College of Medicine
4 Core Laboratories and Department of Anaesthesiology, Wan-Fang Hospital, Taipei Medical University, 250 Wu-Hsing St., Taipei 110, Taiwan, Republic of China
5 Department of Internal Medicine, Taipei County Hospital, Taipei, Taiwan, Republic of China
6 Department of Anaesthesiology, Renai Branch, Taipei City Hospital, Taipei, Taiwan, Republic of China
* Corresponding author. E-mail: rmchen{at}tmu.edu.tw
Background: Ketamine is widely used as an i.v. anaesthetic agent and as a drug of abuse. Hepatocytes contribute to the metabolism of endogenous and exogenous substances. This study evaluated the toxic effects of S-(+)-ketamine and possible mechanisms using human hepatoma HepG2 cells as the experimental model.
Methods: HepG2 cells were exposed to S-(+)-ketamine. Cell viability and the release of lactate dehydrogenase (LDH) and 
-glutamyl transpeptidase (GPT) were measured to determine the toxicity of S-(+)-ketamine to HepG2 cells. Cell morphology, DNA fragmentation, and apoptotic cells were analysed to evaluate the mechanism of S-(+)-ketamine-induced cell death. Amounts of Bax, an apoptotic protein, and cytochrome c in the cytoplasm or mitochondria were quantified by immunoblotting. Cellular adenosine triphosphate levels were analysed using a bioluminescence assay. Caspases-3, -9, and -6 were measured fluorometrically.
Results: Exposure of HepG2 cells to S-(+)-ketamine increased the release of LDH and GPT, but decreased cell viability (all P<0.01). S-(+)-Ketamine time-dependently caused shrinkage of HepG2 cells. Exposure to S-(+)-ketamine led to significant DNA fragmentation and cell apoptosis (P=0.003 and 0.002). S-(+)-Ketamine increased translocation of Bax from the cytoplasm to mitochondria, but decreased the mitochondrial membrane potential and cellular adenosine triphosphate levels (all P<0.01). Sequentially, cytosolic cytochrome c levels and activities of caspases-9, -3, and -6 were augmented after S-(+)-ketamine administration (all P<0.001). Z-VEID-FMK, an inhibitor of caspase-6, alleviated the S-(+)-ketamine-induced augmentation of caspase-6 activity, DNA fragmentation, and cell apoptosis (all P<0.001).
Conclusions: This study shows that S-(+)-ketamine can induce apoptotic insults to human HepG2 cells via a Bax-mitochondria-caspase protease pathway. Thus, we suggest that S-(+)-ketamine at a clinically relevant or an abused concentration may induce liver dysfunction possibly due to its toxicity to hepatocytes.
Keywords: anaesthetics i.v., ketamine; liver, hepatotoxicity; metabolism, ATP, DNA; theories of anaesthetic action, cellular mechanisms