title: Inhibition of bile canalicular network formation in rat sandwich cultured hepatocytes by drugs associated with risk of severe liver injury.
abstract: Idiosyncratic drug-induced liver injury is a clinical concern with serious consequences. Although many preclinical screening methods have been proposed, it remains difficult to identify compounds associated with this rare but potentially fatal liver condition. Here, we propose a novel assay system to assess the risk of liver injury. Rat primary hepatocytes were cultured in a sandwich configuration, which enables the formation of a typical bile canalicular network. From day 2 to 3, test drugs, mostly selected from a list of cholestatic drugs, were administered, and the length of the network was semi-quantitatively measured by immunofluorescence. Liver injury risk information was collected from drug labels and was compared with in vitro measurements. Of 23 test drugs examined, 15 exhibited potent inhibition of bile canalicular network formation (<60% of control). Effects on cell viability were negligible or minimal as confirmed by lactate dehydrogenase leakage and cellular ATP content assays. For the potent 15 drugs, IC50 values were determined. Finally, maximum daily dose divided by the inhibition constant gave good separation of the highest risk of severe liver toxicity drugs such as troglitazone, benzbromarone, flutamide, and amiodarone from lower risk drugs. In conclusion, inhibitory effect on the bile canalicular network formation observed in in vitro sandwich cultured hepatocytes evaluates a new aspect of drug toxicity, particularly associated with aggravation of liver injury.
authors.affiliation: The Laboratory of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Chiba University, Inohana 1-8-1, Chuo-ku, Chiba, 260-8675, Japan.
authors.forename: Akinori
authors.initials: A
authors.lastname: Takemura
authors.affiliation: The Laboratory of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Chiba University, Inohana 1-8-1, Chuo-ku, Chiba, 260-8675, Japan.
authors.forename: Aya
authors.initials: A
authors.lastname: Izaki
authors.affiliation: The Laboratory of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Chiba University, Inohana 1-8-1, Chuo-ku, Chiba, 260-8675, Japan.
authors.forename: Shuichi
authors.initials: S
authors.lastname: Sekine
authors.affiliation: The Laboratory of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Chiba University, Inohana 1-8-1, Chuo-ku, Chiba, 260-8675, Japan. Electronic address: itokousei@chiba-u.jp.
authors.forename: Kousei
authors.initials: K
authors.lastname: Ito
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001648:Bile Canaliculi / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D056486:Chemical and Drug Induced Liver Injury; D022781:Hepatocytes / Q000187:drug effects*; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D000255:Adenosine Triphosphate; D007770:L-Lactate Dehydrogenase
keywords: Bile canaliculus; Cholestasis; Hepatotoxicity; In vitro screening; Jaundice; Sandwich cultured hepatocytes