Oxidative-stress and long-term hepatotoxicity: comparative study in Upcyte human hepatocytes and hepaRG cells

Donato, M. Teresa; Jiménez, Núria; Pelechá, María; Tolosa, Laia

doi:10.1007/s00204-022-03236-y

Cited by 9 publications

(7 citation statements)

References 56 publications

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“…on the order of one to two weeks may resolve false negative results. Encouragingly, in previous studies hepatotoxicity was effectively characterized in HepaRG cells after up to 14 days of chemical exposure (Donato et al, 2022;Dietrich et al, 2020;Anthérieu et al, 2011).…”

Section: Discussionmentioning

confidence: 99%

Chemical screening of food-related chemicals for human fatty liver risk: Combining high content imaging of cellular responses with in vitro to in vivo extrapolation

Müller

Stamou

Englert

et al. 2022

Preprint

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Nonalcoholic fatty liver disease (NAFLD) is an increasingly prevalent human disease with accumulating evidence linking its pathophysiology and co-morbidities to chemical exposures. The complex pathophysiology of NAFLD has limited the elucidation of potential chemical etiologies. In this study we generated a high-content imaging analysis method for the simultaneous quantification of sentinel steatosis cellular markers in chemically exposed human liver cells in vitro combined with a computational model for the extrapolation of human oral equivalent doses (OED). First, the in vitro test method was generated using 14 reference chemicals with known capacities to induce cellular alterations in nuclear morphology, lipid accumulation, mitochondrial membrane potential and oxidative stress. These effects were quantified on a single cell- and population-level, and then, using physiologically based pharmacokinetic modelling and reverse dosimetry, OEDs were extrapolated from these in vitro data. The extrapolated OEDs were confirmed to be within biologically relevant exposure ranges for the reference chemicals. Next, we tested 14 chemicals found in food, selected from thousands of putative chemicals on the basis of structure-based prediction for nuclear receptor activation. Amongst these, orotic acid had an extrapolated OED overlapping with realistic exposure ranges. By the strategy developed in this study, we were able to characterize known NAFLD-inducing chemicals and translate to data scarce food-related chemicals, amongst which we identified orotic acid to induce steatosis. This strategy addresses needs of next generation risk assessment, and can be used as a first chemical prioritization hazard screening step in a tiered approach to identify chemical risk factors for NAFLD.

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Section: Discussionmentioning

confidence: 99%

Chemical screening of food-related chemicals for human fatty liver risk: Combining high content imaging of cellular responses with in vitro to in vivo extrapolation

Müller

Stamou

Englert

et al. 2022

Preprint

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“…Several authors have used differentiated HepaRG for long-term toxicity evaluations for a maximum of 14 days after differentiation at clinically relevant concentrations [ 91 , 92 , 93 , 94 ]. The use of HepaRG cells following repeated-dose regimes allow using lower concentrations more relevant to the clinics and obtaining better predictions.…”

Section: In Vitro Cell Models For Chronic Hepatotoxicitymentioning

confidence: 99%

“…The study was based on the use of high-content screening (HCS) technology that allows the detection of the underlying mechanisms of drug-induced toxicity before cytotoxicity occurs [ 95 ], and revealed the suitability of UHHs for long-term repeated-dose studies that can detect the potential hepatotoxicity in preclinical development. Recently, we have also demonstrated that UHHs maintained the activity of important antioxidant enzymes and allow studying different mechanisms implicated in DILI, including oxidative stress, at sub-lethal concentrations [ 92 ]. In this case, UHH donors seemed to show a higher sensitivity than HepaRG detecting oxidative stress induced by test compounds and also provided information about differential responses in the distinct donors [ 92 ].…”

Section: In Vitro Cell Models For Chronic Hepatotoxicitymentioning

confidence: 99%

“…Recently, we have also demonstrated that UHHs maintained the activity of important antioxidant enzymes and allow studying different mechanisms implicated in DILI, including oxidative stress, at sub-lethal concentrations [ 92 ]. In this case, UHH donors seemed to show a higher sensitivity than HepaRG detecting oxidative stress induced by test compounds and also provided information about differential responses in the distinct donors [ 92 ]. The importance of this study is that cells were exposed to concentrations around the therapeutic peak plasmatic concentration (C max ), which is close to what happens in vivo.…”

Section: In Vitro Cell Models For Chronic Hepatotoxicitymentioning

confidence: 99%

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In Vitro Models for Studying Chronic Drug-Induced Liver Injury

Donato

Ferrer

Tolosa

2022

IJMS

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Drug-induced liver injury (DILI) is a major clinical problem in terms of patient morbidity and mortality, cost to healthcare systems and failure of the development of new drugs. The need for consistent safety strategies capable of identifying a potential toxicity risk early in the drug discovery pipeline is key. Human DILI is poorly predicted in animals, probably due to the well-known interspecies differences in drug metabolism, pharmacokinetics, and toxicity targets. For this reason, distinct cellular models from primary human hepatocytes or hepatoma cell lines cultured as 2D monolayers to emerging 3D culture systems or the use of multi-cellular systems have been proposed for hepatotoxicity studies. In order to mimic long-term hepatotoxicity in vitro, cell models, which maintain hepatic phenotype for a suitably long period, should be used. On the other hand, repeated-dose administration is a more relevant scenario for therapeutics, providing information not only about toxicity, but also about cumulative effects and/or delayed responses. In this review, we evaluate the existing cell models for DILI prediction focusing on chronic hepatotoxicity, highlighting how better characterization and mechanistic studies could lead to advance DILI prediction.

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“…Nonetheless these limitations, upcyte ® hepatocytes have been shown to be a valuable in vitro tool for drug-interaction studies ( Ramachandran et al, 2015 ) and for assessing acute and long-term hepatotoxicity ( Tolosa et al, 2016 ; Tolosa et al, 2019 ). Regarding the progressive loss of specific hepatic features exhibited by PHH after their isolation, phenomenon called dedifferentiation ( Heslop et al, 2017 ; Kiamehr et al, 2019 ), it has been reported that upcyte ® hepatocytes are characterized by a stable or even increased activity of some phase-I (CYP2B6, 2C9, 2E1, and 3A4) and phase-II (UGT1A1 and 2B7) enzymes over time up to 21 days ( Donato et al, 2022 ). Analogously, the Albumin production was almost constant over time (10 days) in differentiated cells ( Levy et al, 2015 ), thus indicating phenotypic stability.…”

Section: Introductionmentioning

confidence: 99%

Sub-Lethal Concentrations of Graphene Oxide Trigger Acute-Phase Response and Impairment of Phase-I Xenobiotic Metabolism in Upcyte® Hepatocytes

Romaldini

Spanò

Catalano

et al. 2022

Front. Bioeng. Biotechnol.

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The impact of graphene oxide on hepatic functional cells represents a crucial evaluation step for its potential application in nanomedicine. Primary human hepatocytes are the gold standard for studying drug toxicity and metabolism; however, current technical limitations may slow down the large-scale diffusion of this cellular tool for in vitro investigations. To assess the potential hepatotoxicity of graphene oxide, we propose an alternative cell model, the second-generation upcyte® hepatocytes, which show metabolic and functional profiles akin to primary human hepatocytes. Cells were acutely exposed to sub-lethal concentrations of graphene oxide (≤80 μg/ml) for 24 h and stress-related cell responses (such as apoptosis, oxidative stress, and inflammatory response) were evaluated, along with a broad investigation of graphene oxide impact on specialized hepatic functions. Results show a mild activation of early apoptosis but not oxidative stress or inflammatory response in our cell model. Notably, while graphene oxide clearly impacted phase-I drug-metabolism enzymes (e.g., CYP3A4, CYP2C9) through the inhibition of gene expression and metabolic activity, conversely, no effect was observed for phase-II enzyme GST and phase-III efflux transporter ABCG2. The GO-induced impairment of CYP3A4 occurs concomitantly with the activation of an early acute-phase response, characterized by altered levels of gene expression and protein production of relevant acute-phase proteins (i.e., CRP, Albumin, TFR, TTR). These data suggest that graphene oxide induces an acute phase response, which is in line with recent in vivo findings. In conclusion, upcyte® hepatocytes appear a reliable in vitro model for assessing nanomaterial-induced hepatotoxicity, specifically showing that sub-lethal doses of graphene oxide have a negative impact on the specialized hepatic functions of these cells. The impairment of the cytochrome P450 system, along with the activation of an acute-phase response, may suggest potential detrimental consequences for human health, as altered detoxification from xenobiotics and drugs.

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Oxidative-stress and long-term hepatotoxicity: comparative study in Upcyte human hepatocytes and hepaRG cells

Cited by 9 publications

References 56 publications

Chemical screening of food-related chemicals for human fatty liver risk: Combining high content imaging of cellular responses with in vitro to in vivo extrapolation

Chemical screening of food-related chemicals for human fatty liver risk: Combining high content imaging of cellular responses with in vitro to in vivo extrapolation

In Vitro Models for Studying Chronic Drug-Induced Liver Injury

Sub-Lethal Concentrations of Graphene Oxide Trigger Acute-Phase Response and Impairment of Phase-I Xenobiotic Metabolism in Upcyte® Hepatocytes

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