Metabolomics profiling of steatosis progression in HepaRG ® cells using sodium valproate

Abstract

Non-alcoholic Fatty Liver Disease (NAFLD) is a frequently encountered Drug-Induced Liver Injury (DILI). Although this stage of the disease is reversible, it can lead to irreversible damage provoked by non-alcoholic steatohepatitis (NASH), fibrosis and cirrhosis. Therefore, the assessment of NAFLD is a paramount objective in toxicological screenings of new drug candidates. In this study, a metabolomic fingerprint of NAFLD induced in HepaRG cells at four dosing schemes by a reference toxicant, sodium valproate (NaVPA), was obtained using liquid-liquid extraction followed by liquid chromatography and accurate mass-mass spectrometry (LC-AM/MS). The combination of a strict design of experiment with a robust detection method, applied on sodium valproate, validated the possibilities of untargeted metabolomics in hepatic toxicological research. Distinctive patterns between exposed and control cells were consistently observed, multivariate analyses selected up to 200 features of interest, revealing hallmark NAFLD-biomarkers, such as diacylglycerol and triglyceride accumulation and carnitine deficiency. Initial toxic responses show increased levels of S-adenosylmethionine and mono-acetylspermidine in combination with only a moderate increase in triglycerides. New specific markers of toxicity have been observed, such as spermidines, creatine, and acetylcholine. The described design of experiment provides a valuable metabolomics platform for mechanistic research of toxicological hazards and identified new markers for steatotic progression.

Drug-induced hepatic steatosis in absence of severe mitochondrial dysfunction in HepaRG cells: proof of multiple mechanism-based toxicity

“…Among the 12 drugs, AMIO, ALLO, 5FU, INDI, INDO, METHO, SUL, RIF, and TRO induced steatosis in HepaRG cells, thus confirming that the HepaRG cell line represents a suitable model in order to study druginduced steatosis (Anthérieu et al 2011;Cuykx et al 2018;Tolosa et al 2016). In addition, we found that AMIO, ALLO, 5FU, INDI, INDO, RIF, and TRO also induced steatosis in at least one out of six different PHH batches.…”

Managing the challenge of drug-induced liver injury: a roadmap for the development and deployment of preclinical predictive models

“…Furthermore, the ability of this model to detect delayed mitochondrial toxicity has been shown for a panel of mitotoxicants with varied mitochondrial targets, including mitochondrial DNA, mitochondrial protein synthesis and fatty acid oxidation 61,62 . Importantly, this model enabled the pathway of mitochondrial toxicity to be delineated at concentrations in the region of drug exposure (Cmax, Css) in clinical practice.…”

Early detection of metabolic changes in drug-induced steatosis using metabolomics approaches

“…The use of drugs is a frequent cause of steatosis, [17][18][19] ranging across multiple therapeutic classes including antiarrhythmic agents, chemotherapeutics, and tetracyclines. 6,[20][21][22] High doses and prolonged use of tetracycline, for treatment of a range of bacterial infections, 23 has been shown to cause steatosis and liver damage in animals [23][24][25][26] clinically similar to the human pathology of liver steatosis assessed by histological staining. [27][28][29] Animal studies play an important role in studying and understanding the pathogenic processes of the human disease.…”

“…6,20,21,25,31,32 Researchers have investigated altered chemical pathways 27,[33][34][35][36] drug-induced steatosis through metabolite changes in serum, urine and hepatic tissues from animals and patients. 19,22,24,25,[37][38][39][40][41] These studies have identied several metabolites associated with various pathways related to the pathogenesis of drug induced steatosis. However, these studies have limitations identifying changes in biomarkers in the early development and progression of the disease.…”