Detection of metabolic activation leading to drug-induced phospholipidosis in rat hepatocyte spheroids
| Autor: | Yoko Ejiri, Yaichiro Kotake, Shigeru Ohta, Seigo Sanoh, Masashi Takagi, Masataka Santoh |
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| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
Metabolite Phospholipid Cell Culture Techniques Pharmacology Loratadine Toxicology Rats Sprague-Dawley 03 medical and health sciences chemistry.chemical_compound Surface-Active Agents Cytochrome P-450 Enzyme System In vivo Spheroids Cellular medicine Animals Cells Cultured Phospholipids Fluorescent Dyes Phospholipidosis Desloratadine Chemistry Phosphatidylethanolamines enzymes and coenzymes (carbohydrates) 030104 developmental biology medicine.anatomical_structure Biochemistry Hepatocyte Hepatocytes lipids (amino acids peptides and proteins) Drug metabolism medicine.drug |
| Zdroj: | The Journal of toxicological sciences. 41(1) |
| ISSN: | 1880-3989 |
| Popis: | Drug-induced phospholipidosis (PLD) is one of the adverse reactions to treatment with cationic amphiphilic drugs. Recently, simple and reliable evaluation methods for PLD have been reported. However, the predictive power of these methods for in vivo PLD induction is insufficient in some cases. To accurately predict PLD, we focused on drug metabolism and used three-dimensional cultures of hepatocytes known as spheroids. Here we used the fluorescent phospholipid dye N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-1,2-dihexadecanoyl-sn-glycero-3-phosphoethanolamine (NBD-PE) to detect PLD induction. After 48 hr exposure to 20 µM amiodarone and amitriptyline, PLD inducers, NBD-PE fluorescence in the spheroids was significantly higher than that in the control. In contrast, 1 mM acetaminophen, as a negative control, did not increase fluorescence. Furthermore, the combination of NBD-PE fluorescence and LysoTracker Red fluorescence and the accumulation of intrinsic phospholipids reflected PLD induction in spheroids. To evaluate metabolic activation, we assessed PLD induction by loratadine. NBD-PE fluorescence intensity was significantly increased by 50 µM loratadine treatment. However, the fluorescence was markedly decreased by co-treatment with 500 µM 1-aminobenzotriazole, a broad cytochrome P450 inhibitor. The formation of desloratadine, a metabolite of loratadine, was observed in spheroids after treatment with loratadine alone. These results showed that metabolic activation is the key factor in PLD induction by treatment with loratadine. We demonstrated that rat primary hepatocyte spheroid culture is a useful model for evaluating drug-induced PLD induction mediated by metabolic activation of the drug using the fluorescence probe technique. |
| Databáze: | OpenAIRE |
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