Chlorpromazine toxicity is associated with disruption of cell membrane integrity and initiation of a pro-inflammatory response in the HepaRG hepatic cell line

Morgan, Katie; Martucci, Nicole J.; Kozlowska, Ada; Gamal, Wesam; Brzeszczyński, Filip; Treskes, Philipp; Samuel, Kay; Hayes, Peter; Nelson, Leonard J.; Bagnaninchi, Pierre O.; Brzeszczyńska, Joanna; Plevris, John

doi:10.1016/j.biopha.2019.01.020

Cited by 34 publications

(32 citation statements)

References 25 publications

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“…The cell culture was carried out according to previously published protocol [9,11,17,19]. HepaRG fully differentiated cells (HPR116-TA08; Cryopreserved HepaRG™ cells; Biopredic Int., Rennes, France) were thawed from liquid nitrogen and plated in Biopredic's proprietary General Purpose Medium (GPS) without dimethyl sulfoxide (DMSO) for up to 2 days.…”

Section: Cell Culturementioning

confidence: 99%

“…Throughout the experimental process, differentiated HepaRG cells were regularly assessed for viability according to previously published procedure [17,19]. The cells were seeded on a 96 well standard tissue culture plate and grown to confluency (day 8).…”

Section: Viability Assaysmentioning

confidence: 99%

“…Both CPZ and APAP did not compromise HepaRG cell viability at sub toxic concentration. Using total ATP and PrestoBlue assays, the effect of CPZ and APAP on HepaRG cells have already been published from our group [17,19].…”

Section: Cell Viability: Total Adenosine Triphosphate (Atp) and Prestmentioning

confidence: 99%

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Validation of Reference Genes for Gene Expression Studies by RT-qPCR in HepaRG Cells during Toxicity Testing and Disease Modelling

Brzeszczyńska

Brzeszczyński

Samuel

et al. 2020

Cells

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Gene expression analysis by quantitative real-time polymerase chain reaction (RT-qPCR) is routinely used in biomedical studies. The reproducibility and reliability of the data fundamentally depends on experimental design and data interpretation. Despite the wide application of this assay, there is significant variation in the validation process of gene expression data from research laboratories. Since the validity of results depends on appropriate normalisation, it is crucial to select appropriate reference gene(s), where transcription of the selected gene is unaffected by experimental setting. In this study we have applied geNorm technology to investigate the transcription of 12 'housekeeping' genes for use in the normalisation of RT-qPCR data acquired using a widely accepted HepaRG hepatic cell line in studies examining models of pre-clinical drug testing. geNorm data identified a number of genes unaffected by specific drug treatments and showed that different genes remained invariant in response to different drug treatments, whereas the transcription of 'classical' reference genes such as GAPDH (glyceralde-hyde-3-phosphate dehydrogenase) was altered by drug treatment. Comparing data normalised using the reference genes identified by geNorm with normalisation using classical housekeeping genes demonstrated substantial differences in the final results. In light of cell therapy application, RT-qPCR analyses has to be carefully evaluated to accurately interpret data obtained from dynamic cellular models undergoing sequential stages of phenotypic change.

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Section: Cell Culturementioning

confidence: 99%

Section: Viability Assaysmentioning

confidence: 99%

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Validation of Reference Genes for Gene Expression Studies by RT-qPCR in HepaRG Cells during Toxicity Testing and Disease Modelling

Brzeszczyńska

Brzeszczyński

Samuel

et al. 2020

Cells

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“…Using standard assays and comparing with LC50 cytotoxicity curves generated from xCELLigence TM data, they showed that all cell lines were sensitive to hepatotoxins, though none at a high enough level to use in predictive experiments [38]. It is important to note here that a loss of impedance does not necessarily mean a loss of cell viability [3,40,41], but detects a change in the membrane.…”

Section: Drug Induced Liver Injurymentioning

confidence: 99%

“…The cultures remained stable for 4 weeks and beyond, and retained characteristics of differentiation, making HepaRG™ cells a viable model for drug toxicity testing. In contrast to other hepatic cell lines, fully differentiated HepaRGs™ provide a unique co-culture of cholagiocyte-like and hepatocyte cells, making them a useful model for drug transporter studies and for use in cholestatic models [40,43,45,46].…”

Section: Toxicity Assays Using Heparg Cellsmentioning

confidence: 99%

Application of Impedance-Based Techniques in Hepatology Research

Morgan

Gamal

Samuel

et al. 2019

JCM

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There are a variety of end-point assays and techniques available to monitor hepatic cell cultures and study toxicity within in vitro models. These commonly focus on one aspect of cell metabolism and are often destructive to cells. Impedance-based cellular assays (IBCAs) assess biological functions of cell populations in real-time by measuring electrical impedance, which is the resistance to alternating current caused by the dielectric properties of proliferating of cells. While the uses of IBCA have been widely reported for a number of tissues, specific uses in the study of hepatic cell cultures have not been reported to date. IBCA monitors cellular behaviour throughout experimentation non-invasively without labelling or damage to cell cultures. The data extrapolated from IBCA can be correlated to biological events happening within the cell and therefore may inform drug toxicity studies or other applications within hepatic research. Because tight junctions comprise the blood/biliary barrier in hepatocytes, there are major consequences when these junctions are disrupted, as many pathologies centre around the bile canaliculi and flow of bile out of the liver. The application of IBCA in hepatology provides a unique opportunity to assess cellular polarity and patency of tight junctions, vital to maintaining normal hepatic function. Here, we describe how IBCAs have been applied to measuring the effect of viral infection, drug toxicity/IC50, cholangiopathies, cancer metastasis and monitoring of the gut-liver axis. We also highlight key areas of research where IBCAs could be used in future applications within the field of hepatology.

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Multivalent Targeting of the Asialoglycoprotein Receptor by Virus‐Like Particles

Hincapie,

Bhattacharya,

Baksh

et al. 2023

Small

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The asialoglycoprotein receptor (ASGPR) is expressed in high density on hepatocytes. Multivalent variants of galactosyl carbohydrates bind ASGPR with high affinity, enabling hepatic delivery of ligand‐bound cargo. Virus‐like particle (VLP) conjugates of a relatively high‐affinity ligand were efficiently endocytosed by ASGPR‐expressing cells in a manner strongly dependent on the nature and density of ligand display, with the best formulation using a nanomolar‐, but not a picomolar‐level, binder. Optimized particles were taken up by HepG2 cells with greater efficiency than competing small molecules or the natural multigalactosylated ligand, asialoorosomucoid. Upon systemic injection in mice, these VLPs were rapidly cleared to the liver and were found in association with sinusoidal endothelial cells, Kupffer cells, hepatocytes, dendritic cells, and other immune cells. Both ASGPR‐targeted and nontargeted particles were distributed similarly to endothelial and Kupffer cells, but targeted particles were distributed to a greater number and fraction of hepatocytes. Thus, selective cellular trafficking in the liver is difficult to achieve: even with the most potent ASGPR targeting available, barrier cells take up much of the injected particles and hepatocytes are accessed only approximately twice as efficiently in the best case.

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Chlorpromazine toxicity is associated with disruption of cell membrane integrity and initiation of a pro-inflammatory response in the HepaRG hepatic cell line

Cited by 34 publications

References 25 publications

Validation of Reference Genes for Gene Expression Studies by RT-qPCR in HepaRG Cells during Toxicity Testing and Disease Modelling

Validation of Reference Genes for Gene Expression Studies by RT-qPCR in HepaRG Cells during Toxicity Testing and Disease Modelling

Application of Impedance-Based Techniques in Hepatology Research

Multivalent Targeting of the Asialoglycoprotein Receptor by Virus‐Like Particles

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