title: High-content imaging technology for the evaluation of drug-induced steatosis using a multiparametric cell-based assay.
abstract: In the present study, we developed a cell-based protocol for the identification of drugs able to induce steatosis. The assay measures multiple markers of toxicity in a 96-well plate format using high-content screening (HCS) technology. After treating HepG2 cells with increasing concentrations of the tested compounds, toxicity parameters were analyzed using fluorescent probes: BODIPY493/503 (lipid content), 2',7'-dihydrodichlorofluorescein diacetate (reactive oxygen species [ROS] generation), tetramethyl rhodamine methyl ester (mitochondrial membrane potential), propidium iodide (cell viability), and Hoechst 33342 (nuclei staining). A total of 16 drugs previously reported to induce liver steatosis through different mechanisms (positive controls) and six nonsteatotic compounds (negative controls) were included in the study. All the steatosis-positive compounds significantly increased BODIPY493/503 fluorescence in HepG2 cells, whereas none of the negative controls induced lipid accumulation. In addition to effects on fat levels, increased ROS generation was produced by certain compounds, which could be indicative of increased risk of liver damage. Our results suggest that this in vitro approach is a simple, rapid, and sensitive screening tool for steatosis-inducing drugs. This conclusion should be confirmed by testing a larger number of steatosis-positive and -negative inducers.
authors: Unidad de Hepatología Experimental, Centro de Investigación, Hospital La Fe, Valencia, Spain. donato_mte@gva.es
authors: M Teresa
authors: MT
authors: Donato
authors: Laia
authors: L
authors: Tolosa
authors: Nuria
authors: N
authors: Jiménez
authors: José V
authors: JV
authors: Castell
authors: M José
authors: MJ
authors: Gómez-Lechón
mesh_terms: D015415:Biomarkers / Q000032:analysis; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D005234:Fatty Liver / Q000139:chemically induced*; D005456:Fluorescent Dyes; D056945:Hep G2 Cells; D006801:Humans; D008099:Liver / Q000473:pathology; D008856:Microscopy, Fluorescence; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D015415:Biomarkers; D005456:Fluorescent Dyes; D017382:Reactive Oxygen Species