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Despite the high prevalence of alcoholic liver disease, its identification and characterization remain poor, especially in early stages such as alcoholic fatty liver disease and alcoholic steatohepatitis. This latter implies diagnostic difficulties, few therapeutic options and unclear mechanisms of action. To elucidate the metabolic alterations and pinpoint affected biochemical pathways, alcoholic steatohepatitis was simulated in vitro by exposing HepaRG cells to ethanol (IC10, 368 mM) and tumor necrosis factor alpha (TNF-α, 50 ng/mL) for 24 h. This combined exposure was compared to solely ethanol-exposed as well as -nonexposed cells. Four different metabolomics platforms were used combining liquid chromatography, high-resolution mass spectrometry and drift tube ion mobility to elucidate both intracellular and extracellular metabolic alterations. Some of the key findings include the influence of TNF-α in the upregulation of hepatic triglycerides and the downregulation of hepatic phosphatidylethanolamines and phosphatidylcholines. S-Adenosylmethionine showed to play a central role in the progression of alcoholic steatohepatitis. In addition, fatty acyl esters of hydroxy fatty acid (FAHFA)-containing triglycerides were detected for the first time in human hepatocytes and their alterations showed a potentially important role during the progression of alcoholic steatohepatitis. Ethoxylated phosphorylcholine was identified as a potential new biomarker of ethanol exposure.

BACKGROUND: It is unclear whether the patatin-like phospholipase domain-containing protein 3 (PNPLA3) rs738409 C-to-G single nucleotide polymorphism, resulting in the substitution of isoleucine to methionine at position 148 (I148M), impedes regression of hepatic steatosis when treating non-alcoholic fatty liver disease (NAFLD). OBJECTIVES: Investigate if carriage of the PNPLA3 148M allele affects the anti-steatotic efficacy of all possible anti-NAFLD interventions, identify gaps in current knowledge and provide guidance for individual treatment. METHODS: Research available in public databases was searched up to 13 November 2022. Studies were included if a treatment in NAFLD patients decreased hepatic steatosis in the pooled patient group or a PNPLA3 I148M polymorphism subgroup (II/IM/MM). The risk of bias was assessed using the Cochrane Risk-Of-Bias 2 Tool and the Newcastle-Ottawa Scale. RESULTS: Moderate evidence indicates that NAFLD patients homozygous for the PNPLA3 148M allele benefit less or not at all from omega-3 carboxylic acids to decrease liver fat, while the PNPLA3 148I allele shows moderate benefit. Low evidence suggests that interventions employing lifestyle changes are more effective to reduce liver fat in NAFLD patients homozygous for the PNPLA3 148M allele compared to patients with wild-type PNPLA3. CONCLUSIONS: NAFLD patients homozygous for the PNPLA3 148M allele might not benefit from omega-3 carboxylic acids to reduce hepatic steatosis in contrast to patients with wild-type PNPLA3. Instead, patients with two PNPLA3 148M alleles should be especially advised to adopt lifestyle changes. Genotyping for PNPLA3 I148M should be encouraged in therapeutic studies for NAFLD. REGISTRATION NUMBER (PROSPERO): CRD42022375028.

Nonalcoholic fatty liver disease (NAFLD) is the leading cause of chronic liver disease worldwide. Despite extensive research and multiple clinical trials, there are still no FDA-approved therapies to treat the most severe forms of NAFLD. This is largely due to its complicated etiology and pathogenesis, which involves visceral obesity, insulin resistance, gut dysbiosis, etc. Although inflammation is generally believed to be one of the critical factors that drive the progression of simple steatosis to nonalcoholic steatohepatitis (NASH), the exact type of inflammation and how it contributes to NASH pathogenesis remain largely unknown. Liver inflammation is accompanied by the elevation of inflammatory mediators, including cytokines and chemokines and consequently intrahepatic infiltration of multiple types of immune cells. Recent studies revealed that extracellular vesicles (EVs) derived from inflammatory cells and hepatocytes play an important role in controlling liver inflammation during NASH. In this review, we highlight the roles of innate and adaptive immune cells and their microRNA-enriched EVs during NAFLD development and discuss potential drugs that target inflammatory pathways for the treatment of NAFLD.

Background: Healthcare-associated SARS-CoV-2 infections need to be explored further. Our study is an analysis of hospital-acquired infections (HAIs) and ambulatory healthcare workers (aHCWs) with SARS-CoV-2 across the pandemic in a Belgian university hospital. Methods: We compared HAIs with community-associated infections (CAIs) to identify the factors associated with having an HAI. We then performed a genomic cluster analysis of HAIs and aHCWs. We used this alongside the European Centre for Disease Control (ECDC) case source classifications of an HAI. Results: Between March 2020 and March 2022, 269 patients had an HAI. A lower BMI, a worse frailty index, lower C-reactive protein (CRP), and a higher thrombocyte count as well as death and length of stay were significantly associated with having an HAI. Using those variables to predict HAIs versus CAIs, we obtained a positive predictive value (PPV) of 83.6% and a negative predictive value (NPV) of 82.2%; the area under the ROC was 0.89. Genomic cluster analyses and representations on epicurves and minimal spanning trees delivered further insights into HAI dynamics across different pandemic waves. The genomic data were also compared with the clinical ECDC definitions for HAIs; we found that 90.0% of the ‘definite’, 87.8% of the ‘probable’, and 70.3% of the ‘indeterminate’ HAIs belonged to one of the twenty-two COVID-19 genomic clusters we identified. Conclusions: We propose a novel prediction model for HAIs. In addition, we show that the management of nosocomial outbreaks will benefit from genome sequencing analyses.

Non-alcoholic steatohepatitis (NASH) is a highly prevalent, chronic liver disease characterized by hepatic lipid accumulation, inflammation, and concomitant fibrosis. Up to date, no anti-NASH drugs have been approved. In this study, we reproduced key NASH characteristics in vitro by exposing primary human hepatocytes (PHH), human skin stem cell-derived hepatic cells (hSKP-HPC), HepaRG and HepG2 cell lines, as well as LX-2 cells to multiple factors that play a role in the onset of NASH. The obtained in vitro disease models showed intracellular lipid accumulation, secretion of inflammatory chemokines, induced ATP content, apoptosis, and increased pro-fibrotic gene expression. These cell systems were then used to evaluate the anti-NASH properties of eight peroxisome proliferator-activated receptor (PPAR) agonists (bezafibrate, elafibranor, fenofibrate, lanifibranor, pemafibrate, pioglitazone, rosiglitazone, and saroglitazar). PPAR agonists differently attenuated lipid accumulation, inflammatory chemokine secretion, and pro-fibrotic gene expression.Based on the obtained readouts, a scoring system was developed to grade the anti-NASH potencies. The in vitro scoring system, based on a battery of the most performant models, namely PHH, hSKP-HPC, and LX-2 cultures, showed that elafibranor, followed by saroglitazar and pioglitazone, induced the strongest anti-NASH effects. These data corroborate available clinical data and show the relevance of these in vitro models for the preclinical investigation of anti-NASH compounds.

BackgroundWorldwide, healthcare-associated SARS-CoV-2 infections are a major problem: they are associated with increased morbidity, mortality, and hospitalization costs. In-depth studies across the pandemic are crucial to understand and prevent transmission in hospital settings. The principal aims of this study were to characterise patients and validate ECDC definitions of healthcare-associated COVID-19 infections.MethodsWe set up a retrospective observational study spanning the first three waves of the COVID-19 pandemic in a Belgian university hospital: it describes the characteristics of COVID-19 patients admitted, with either healthcare- or community-associated infections. We performed a cluster analysis through epidemiological and viral genome analyses of the healthcare-associated infections, in order to validate the ECDC definitions of healthcare-associated COVID-19 infections.ResultsBetween week 10 of 2020 and week 22 of 2021, 168 patients were hospitalized with healthcare-associated COVID-19. The following factors were found more often in symptomatic healthcare- than in community-associated hospitalized patients: older age, increased frailty, smoking habits, and comorbidities. The genome-based cluster analyses showed that different viral lineages predominated in different timeframes. We observed a good correlation of epidemiological data with genome sequencing results in at least 12 different outbreaks in our hospital, thus validating the ECDC definitions. ConclusionsThis in-depth characterization sheds new light on the problem of healthcare-associated COVID-19 infections, in particular on patients’ characteristics, epidemiology, and cluster dynamics. Even though epidemiological evaluation of nosocomial infections is vital, management of nosocomial outbreaks can undoubtedly benefit from genome sequencing analyses to reinforce their strategy.

Although most same-stage non-alcoholic fatty liver disease (NAFLD) patients exhibit similar histologic sequelae, the underlying mechanisms appear to be highly heterogeneous. Therefore, it was recently proposed to redefine NAFLD to metabolic dysfunction-associated fatty liver disease (MAFLD) in which other known causes of liver disease such as alcohol consumption or viral hepatitis do not need to be excluded. Revised nomenclature envisions speeding up and facilitating anti-MAFLD drug development by means of patient stratification whereby each subgroup would benefit from distinct pharmacological interventions. As human-based in vitro research fulfils an irrefutable step in drug development, action should be taken as well in this stadium of the translational path. Indeed, most established in vitro NAFLD models rely on short-term exposure to fatty acids and use lipid accumulation as a phenotypic benchmark. This general approach to a seemingly ambiguous disease such as NAFLD therefore no longer seems applicable. Human-based in vitro models that accurately reflect distinct disease subgroups of MAFLD should thus be adopted in early preclinical disease modeling and drug testing. In this review article, we outline considerations for setting up translational in vitro experiments in the MAFLD era and allude to potential strategies to implement MAFLD heterogeneity into an in vitro setting so as to better align early drug development with future clinical trial designs.

Alcoholic liver disease is highly prevalent but poorly identified and characterized, leading to knowledge gaps, which impairs early diagnosis. Excessive alcohol consumption is known to alter lipid metabolism, followed by progressive intracellular lipid accumulation, resulting in alcoholic fatty liver disease. In this study, HepaRG cells were exposed to ethanol at IC10 and 1/10 IC10 for 24 and 48 h. Metabolic alterations were investigated intra- and extracellularly with liquid chromatography-high-resolution mass spectrometry. Ion mobility was added as an extra separation dimension for untargeted lipidomics to improve annotation confidence. Distinctive patterns between exposed and control cells were consistently observed, with intracellular upregulation of di- and triglycerides, downregulation of phosphatidylcholines and phosphatidylethanolamines, sphingomyelins, and S-adenosylmethionine, among others. Several intracellular metabolic patterns could be related to changes in the extracellular environment, such as increased intracellular hydrolysis of sphingomyelins, leading to increased phosphorylcholine secretion. Carnitines showed alterations depending on the size of their carbon chain, which highlights the interplay between β-oxidation in mitochondria and peroxisomes. Potential new biomarkers of ethanol-induced hepatotoxicity have been observed, such as ceramides with a sphingadienine backbone, octanoylcarnitine, creatine, acetylcholine, and ethoxylated phosphorylcholine. The combination of the metabolic fingerprint and footprint enabled a comprehensive investigation of the pathophysiology behind ethanol-induced hepatotoxicity.

Summary There is an unmet need for functional primary human hepatocytes to support the pharmaceutical and (bio)medical demand. The unique discovery, a decade ago, that somatic cells can be drawn out of their apparent biological lockdown to reacquire a pluripotent state has revealed a completely new avenue of possibilities for generating surrogate human hepatocytes. Since then, the number of papers reporting the direct conversion of somatic cells into induced hepatocytes (iHeps) has burgeoned. A hepatic cell fate can be established via the ectopic expression of native liver-enriched transcription factors in somatic cells, thereby bypassing the need for an intermediate (pluripotent) stem cell state. That said, understanding and eventually controlling the processes that give rise to functional iHeps remains challenging. In this review, we provide an overview of the state-of-the-art reprogramming cocktails and techniques, as well as their corresponding conversion efficiencies. Special attention is paid to the role of liver-enriched transcription factors as hepatogenic reprogramming tools and small molecules as facilitators of hepatic transdifferentiation. To conclude, we formulate recommendations to optimise, standardise and enrich the in vitro production of iHeps to reach clinical standards, and propose minimal criteria for their characterisation.

Stem cells are characterized by their self-renewal capacity and their ability to differentiate into multiple cell types of the human body. Using directed differentiation strategies, stem cells can now be converted into hepatocyte-like cells (HLCs) and therefore, represent a unique cell source for toxicological applications in vitro. However, the acquired hepatic functionality of stem cell-derived HLCs is still significantly inferior to primary human hepatocytes. One of the main reasons for this is that most in vitro models use traditional two-dimensional (2D) setups where the flat substrata cannot properly mimic the physiology of the human liver. Therefore, 2D-setups are progressively being replaced by more advanced culture systems, which attempt to replicate the natural liver microenvironment, in which stem cells can better differentiate towards HLCs. This review highlights the most recent cell culture systems, including scaffold-free and scaffold-based three-dimensional (3D) technologies and microfluidics that can be employed for culture and hepatic differentiation of stem cells intended for hepatotoxicity testing. These methodologies have shown to improve in vitro liver cell functionality according to the in vivo liver physiology and allow to establish stem cell-based hepatic in vitro platforms for the accurate evaluation of xenobiotics.

Detection of SARS-CoV-2 RNA in nasopharyngeal samples using the real-time reverse transcription polymerase chain reaction (rRT-PCR) is the gold standard for diagnosing COVID-19. Determination of SARS-CoV-2 RNA by rRT-PCR sometimes results in an inconclusive test result due to a high cycle threshold-value. We retrospectively analyzed 30,851 SARS-CoV-2 rRT-PCR test results. Borderline positivity was considered as the presence of ≤25 viral copies per milliliter, while no amplification was considered as a negative test result. Of all test results, 204 were answered as borderline, of which 107 were accompanied by a follow-up test within 96 hours. Of the 107 follow-up samples, 10 (9.35%) were found positive for SARS-CoV-2. COVID-19 symptoms were not predictive for testing positive in the follow-up test. The positive SARS-CoV-2 samples in the follow-up group represented 0.92% of all positive test results, highlighting the need for retesting and increased hygienic measures for borderline SARS-CoV-2 patients [NCT04636294].

Metabolic-associated fatty liver disease (MAFLD) is a chronic liver disease that affects about a quarter of the world population. MAFLD encompasses different disease stadia ranging from isolated liver steatosis to non-alcoholic steatohepatitis (NASH), fibrosis, cirrhosis and hepatocellular carcinoma. Although MAFLD is considered as the hepatic manifestation of the metabolic syndrome, multiple concomitant disease-potentiating factors can accelerate disease progression. Among these risk factors are diet, lifestyle, genetic traits, intake of steatogenic drugs, male gender and particular infections. Although infections often outweigh the development of fatty liver disease, pre-existing MAFLD could be triggered to progress towards more severe disease stadia. These combined disease cases might be underreported because of the high prevalence of both MAFLD and infectious diseases that can promote or exacerbate fatty liver disease development. In this review, we portray the molecular and cellular mechanisms by which the most relevant viral, bacterial and parasitic infections influence the progression of fatty liver disease and steatohepatitis. We focus in particular on how infectious diseases, including coronavirus disease-19, hepatitis C, acquired immunodeficiency syndrome, peptic ulcer and periodontitis, exacerbate MAFLD. We specifically underscore the synergistic effects of these infections with other MAFLD-promoting factors.

The analysis of polar metabolites based on liquid chromatography-mass spectrometry (LC-MS) methods should take into consideration the complexity of interactions in LC columns to be able to cover a broad range of metabolites of key biological pathways. Therefore, in this study, different chromatographic columns were tested for polar metabolites including reversed-phase and hydrophilic interaction liquid chromatography (HILIC) columns. Based on a column screening, two new generations of zwitterionic HILIC columns were selected for further evaluation. A tree-based method optimization was applied to investigate the chromatographic factors affecting the retention mechanisms of polar metabolites with zwitterionic stationary phases. The results were evaluated based on a scoring system which was applied for more than 80 polar metabolites with a high coverage of key human metabolic pathways. The final optimized methods showed high complementarity to analyze a wide range of metabolic classes including amino acids, small peptides, sugars, amino sugars, phosphorylated sugars, organic acids, nucleobases, nucleosides, nucleotides and acylcarnitines. Optimized methods were applied to analyze different biological matrices, including human urine, plasma and liver cell extracts using an untargeted approach. The number of high-quality features (< 30% median relative standard deviation) ranged from 3,755 for urine to 5,402 for the intracellular metabolome of liver cells, showing the potential of the methods for untargeted purposes.

Human skin-derived precursors (SKP) represent a group of somatic stem/precursor cells that reside in dermal skin throughout life that harbor clinical potential. SKP have a high self-renewal capacity, the ability to differentiate into multiple cell types and low immunogenicity, rendering them key candidates for allogeneic cell-based, off-the-shelf therapy. However, potential clinical application of allogeneic SKP requires that these cells retain their therapeutic properties under all circumstances and, in particular, in the presence of an inflammation state. Therefore, in this study, we investigated the impact of pro-inflammatory stimulation on the secretome and immunosuppressive properties of SKP. We demonstrated that pro-inflammatory stimulation of SKP significantly changes their expression and the secretion profile of chemo/cytokines and growth factors. Most importantly, we observed that pro-inflammatory stimulated SKP were still able to suppress the graft-versus-host response when cotransplanted with human PBMC in severe-combined immune deficient (SCID) mice, albeit to a much lesser extent than unstimulated SKP. Altogether, this study demonstrates that an inflammatory microenvironment has a significant impact on the immunological properties of SKP. These alterations need to be taken into account when developing allogeneic SKP-based therapies., info:eu-repo/semantics/published

Non-alcoholic steatohepatitis (NASH) is a severe chronic liver disease that affects 3 to 5 percent of the world population. It is characterized by hepatic lipid accumulation and inflammation and can progress towards fibrosis, cirrhosis and hepatocellular carcinoma. Until today, no drug has been approved for the treatment of NASH. This delay relates to the complex pathogenesis of NASH and also to a lack of appropriate predictive preclinical testing systems. Furthermore, the human specificity of the NASH pathology hampers a fortiori clinical translation of animal studies. Therefore, we recently employed human skin-derived precursors (hSKP) differentiated to hepatocyte-like cells (hSKP-HPC) as a human-relevant cell source for modelling NASH in vitro. Using this in vitro NASH model, it was possible to test novel drugs being developed for anti-NASH therapy, such as elafibranor. Since steatosis is an important aspect of NASH and multiple drugs are being developed to decelerate and reduce lipid accumulation in the liver, we optimized a flow cytometric method for quantifying neutral lipids in ‘NASH’-triggered hSKP-HPC. This methodology enables efficient identification of anti-steatotic properties of new medicines. • NASH-triggered hSKP-HPC robustly accumulate lipids intracellularly. • Flow cytometric quantification of neutral lipids in NASH-triggered hSKP-HPC allows for accurate determination of the steatotic response. • This method enables efficient identification of potential anti-steatotic drugs in a human-specific model, Graphical abstract Image, graphical abstract

Non-alcoholic fatty liver disease (NAFLD) affects one-third of the population worldwide, of which a substantial number of patients suffer from non-alcoholic steatohepatitis (NASH). NASH is a severe condition characterized by steatosis and concomitant liver inflammation and fibrosis, for which no drug is yet available. NAFLD is also generally conceived as the hepatic manifestation of the metabolic syndrome. Consequently, well-established drugs that are indicated for the treatment of type 2 diabetes and hyperlipidemia are thought to exert effects that alleviate the pathological features of NASH. One class of these drugs targets peroxisome proliferator-activated receptors (PPARs), which are nuclear receptors that play a regulatory role in lipid metabolism and inflammation. Therefore, PPARs are now also being investigated as potential anti-NASH druggable targets. In this paper, we review the mechanisms of action and physiological functions of PPARs and discuss the position of the different PPAR agonists in the therapeutic landscape of NASH. We particularly focus on the PPAR agonists currently under evaluation in clinical phase II and III trials. Preclinical strategies and how refinement and optimization may improve PPAR-targeted anti-NASH drug testing are also discussed. Finally, potential caveats related to PPAR agonism in anti-NASH therapy are stipulated.

Non-alcoholic steatohepatitis (NASH) is a chronic liver disease characterized by excessive triglyceride accumulation in the liver accompanied by inflammation, cell stress and apoptosis. It is the tipping point to the life-threatening stages of non-alcoholic fatty liver disease (NAFLD). Despite the high prevalence of NASH, up to five percent of the global population, there are currently no approved drugs to treat this disease. Animal models, mostly based on specific diets and genetic modifications, are often employed in anti-NASH drug development. However, due to interspecies differences and artificial pathogenic conditions, they do not represent the human situation accurately and are inadequate for testing the efficacy and safety of potential new drugs. Human-based in vitro models provide a more legitimate representation of the human NASH pathophysiology and can be used to investigate the dysregulation of cellular functions associated with the disease. Also in silico methodologies and pathway-based approaches using human datasets, may contribute to a more accurate representation of NASH, thereby facilitating the quest for new anti-NASH drugs. In this review, we describe the molecular components of NASH and how human-based tools can contribute to unraveling the pathogenesis of this disease and be used in anti-NASH drug development. We also propose a roadmap for the development and application of human-based approaches for future investigation of NASH.

Non-alcoholic steatohepatitis (NASH) is characterized by hepatocellular steatosis with concomitant hepatic inflammation. Despite its pandemic proportions, no anti-NASH drugs have been approved yet. This is partially because drug development is decelerated due to the lack of adequate tools to assess the efficacy of potential new drug candidates. The present study describes the development and application of a new preclinical model for NASH using hepatic cells generated from human skin-derived precursors. Exposure of these cells to lipogenic (insulin, glucose, fatty acids) and pro-inflammatory factors (IL-1β, TNF-α, TGF-β) resulted in a characteristic NASH response, as indicated by intracellular lipid accumulation, modulation of NASH-specific gene expression, increased caspase-3/7 activity and the expression and/or secretion of inflammatory markers, including CCL2, CCL5, CCL7, CCL8, CXCL5, CXCL8, IL1a, IL6 and IL11. The human relevance of the proposed NASH model was verified by transcriptomics analyses that revealed commonly modulated genes and the identification of the same gene classes between the in vitro system and patients suffering from NASH. The application potential of this in vitro model was demonstrated by testing elafibranor, a promising anti-NASH compound currently under clinical phase III trial evaluation. Elafibranor attenuated in vitro key features of NASH, and dramatically lowered lipid load as well as the expression and secretion of inflammatory chemokines, which in vivo are responsible for the recruitment of immune cells. This reduction in inflammatory response was NFκB-mediated. In summary, this human-relevant, in vitro system proved to be a sensitive testing tool for the investigation of novel anti-NASH compounds.

In this study, we suggest optimizing the methodology to determine the Cohesion Index (Icd) in order to avoid mistaken characterizations due to powder bulk density. For this purpose, five different excipients, with different bulk densities and of different chemical nature, were compressed at different heights. Their compression and their tablet characterization enable establishing a powder weight for compression in accordance with its bulk density. Therefore, the resulting tablet will have a height within a defined range of heights where it has no critical effects on its hardness. Then, the impact of this optimization is shown in a formula development, one of the main SeDeM's applications. A mathematical equation was used to calculate the theoretical amount of excipient to formulate the API according to both methodologies. The compression results demonstrate that the characterization with the NM-Icd is more accurate than the previous one while preserving its simplicity.

Phospholipidosis is a metabolic disorder characterized by intracellular accumulation of phospholipids. It can be caused by short-term or chronic exposure to cationic amphiphilic drugs (CADs). These compounds bind to phospholipids, leading to inhibition of their degradation and consequently to their accumulation in lysosomes. Drug-induced phospholipidosis (DIPL) is frequently at the basis of discontinuation of drug development and post-market drug withdrawal. Therefore, reliable human-relevant in vitro models must be developed to speed up the identification of compounds that are potential inducers of phospholipidosis. Here, hepatic cells derived from human skin (hSKP-HPC) were evaluated as an in vitro model for DIPL. These cells were exposed over time to amiodarone, a CAD known to induce phospholipidosis in humans. Transmission electron microscopy revealed the formation of the typical lamellar inclusions in the cell cytoplasm. Increase of phospholipids was already detected after 24 h exposure to amiodarone, whereas a significant increase of neutral lipid vesicles could be observed after 72 h. At the transcriptional level, the modulation of genes involved in DIPL was detected. These results provide a valuable indication of the applicability of hSKP-HPC for the quick assessment of drug-induced phospholipidosis in vitro, early in the drug development process.

The present dataset contains the transcriptomic characterization of a novel in vitro model of non-alcoholic steatohepatitis (NASH) as well as its transcriptomics read-outs for the evaluation of elafibranor, a potential anti-NASH compound. We report whole genome microarray data (Affymetrix HG U133 plus 2.0) of human multipotent stem cell-derived hepatic cells (hSKP-HPC) exposed to mediators of NASH. These cells were exposed to lipogenic inducers (insulin, glucose, fatty acids) and pro-inflammatory factors (IL-1β, TNF-α, TGF-β) to trigger hepatocellular responses characteristic of NASH. In addition, to evaluate the anti-NASH features of elafibranor, a dual peroxisome proliferator-activated receptor (PPAR) agonist that currently is under investigation as a potential anti-NASH therapeutic, was tested this in vitro set-up. This paper provides a detailed description of the microarray data as well as an indication of their value for evaluating cell signaling pathways (e.g. NFκB network) during the in vitro evaluation of anti-NASH compounds. Raw microarray data of different testing conditions were deposited as.CEL files in the Gene Expression Omnibus of NCBI with GEO Series accession number {"type":"entrez-geo","attrs":{"text":"GSE126484","term_id":"126484"}}GSE126484. Further interpretation and discussion of these data can be found in the corresponding research article (DOI: 10.1016/j.phrs.2019.04.016) Boeckmans et al., 2019.

Background: Metabolic dysfunction-associated fatty liver disease (MAFLD) distinguishes metabolic-, gene-tic- and environmental disease drivers. In this study, we aim to develop an in vitro model based on human skin-derived precursors (hSKPs) differentiated to hepatic cells (hSKP-HPC) for environmentally driven MAFLD. This model will be used to assess the efficacy of the thyroid hormone receptor beta (THRB)-selective agonist resmetirom (MGL-3196, Madrigal Pharmaceuticals). Methods: hSKP-HPCs are sequentially exposed for 24h to MAFLD-inducing triggers (fatty acids, fructose and ethanol) followed by an additional 24h-exposure to lipopolysaccharide (LPS) and resmetirom. Then, the cells are fixed with paraformaldehyde and stained with DAPI (nuclei) and BODIPY™ (neutral lipids) for fluorescence microscopy quantification of lipids using ImageJ. Gene expression analysis using RT-qPCR is performed for APOB, CD36, FASN, THRB, SCD and SREBF1. Intracellular ATP levels are measured using the CellTiter Glo® Luminescent Cell Viability Assay. Results: After 24h, hSKP-HPCs significantly accumulate intracellular lipids. Subsequent exposure to LPS potentiates this effect, along with reduced intracellular ATP content and expression of THRB, which is in line with clinical data. Expression of APOB potently decreases after 48h, indicating diminished lipid export by very low-density lipoproteins (VLDL). Expression of CD36 (fatty acid import) and SREBF1 and its downstream targets SCD and FASN (de novo lipogenesis) also decreases in this condition. Exposure to resmetirom reduces the increased lipid load and restores THRB expression as well as ATP levels back to baseline. Resmetirom also induces the expression of CD36, FASN, SCD and SREBF1, suggesting increased energy turnover by thyroid signaling. In addition, resmetirom potently induces APOB expression, indicating VLDL-mediated triglyceride clearance. Conclusions: hSKP-HPCs triggered with environmental MAFLD factors exhibit key MAFLD characteristics, including increased lipid load,reduced ATP levels and reduced THRB expression. Resmetirom reduces the lipid load and restores ATP and THRB levels. Therefore, hSKP-HPCs could represent a promising tool for the development of thyromimetic anti-MAFLD drugs.