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Article ; Online: Genotype Combinations Drive Variability in the Microbiome Configuration of the Rhizosphere of Maize/Bean Intercropping System.

Lanzavecchia, Giovanna / Frascarelli, Giulia / Rocchetti, Lorenzo / Bellucci, Elisa / Bitocchi, Elena / Di Vittori, Valerio / Sillo, Fabiano / Ferraris, Irene / Carta, Giada / Delledonne, Massimo / Nanni, Laura / Papa, Roberto

International journal of molecular sciences

2024 Volume 25, Issue 2

Abstract: In an intercropping system, the interplay between cereals and legumes, which is strongly driven by the complementarity of below-ground structures and their interactions with the soil microbiome, raises a fundamental query: Can different genotypes alter ... ...

Abstract	In an intercropping system, the interplay between cereals and legumes, which is strongly driven by the complementarity of below-ground structures and their interactions with the soil microbiome, raises a fundamental query: Can different genotypes alter the configuration of the rhizosphere microbial communities? To address this issue, we conducted a field study, probing the effects of intercropping and diverse maize (
MeSH term(s)	Agriculture/methods ; Zea mays/genetics ; Plant Roots ; Rhizosphere ; RNA, Ribosomal, 16S/genetics ; Fabaceae/genetics ; Soil ; Bacteria/genetics ; Genotype ; Soil Microbiology
Chemical Substances	RNA, Ribosomal, 16S ; Soil
Language	English
Publishing date	2024-01-20
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2019364-6
ISSN	1422-0067 ; 1422-0067 ; 1661-6596
ISSN (online)	1422-0067
ISSN	1422-0067 ; 1661-6596
DOI	10.3390/ijms25021288
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Article ; Online: Capturing time-dependent activation of genes and stress-response pathways using transcriptomics in iPSC-derived renal proximal tubule cells.

Jennings, Paul / Carta, Giada / Singh, Pranika / da Costa Pereira, Daniel / Feher, Anita / Dinnyes, Andras / Exner, Thomas E / Wilmes, Anja

Cell biology and toxicology

2022 Volume 39, Issue 4, Page(s) 1773–1793

Abstract: Transcriptomic analysis is a powerful method in the utilization of New Approach Methods (NAMs) for identifying mechanisms of toxicity and application to hazard characterization. With this regard, mapping toxicological events to time of exposure would be ... ...

Abstract	Transcriptomic analysis is a powerful method in the utilization of New Approach Methods (NAMs) for identifying mechanisms of toxicity and application to hazard characterization. With this regard, mapping toxicological events to time of exposure would be helpful to characterize early events. Here, we investigated time-dependent changes in gene expression levels in iPSC-derived renal proximal tubular-like cells (PTL) treated with five diverse compounds using TempO-Seq transcriptomics with the aims to evaluate the application of PTL for toxicity prediction and to report on temporal effects for the activation of cellular stress response pathways. PTL were treated with either 50 μM amiodarone, 10 μM sodium arsenate, 5 nM rotenone, or 300 nM tunicamycin over a temporal time course between 1 and 24 h. The TGFβ-type I receptor kinase inhibitor GW788388 (1 μM) was used as a negative control. Pathway analysis revealed the induction of key stress-response pathways, including Nrf2 oxidative stress response, unfolding protein response, and metal stress response. Early response genes per pathway were identified much earlier than 24 h and included HMOX1, ATF3, DDIT3, and several MT1 isotypes. GW788388 did not induce any genes within the stress response pathways above, but showed deregulation of genes involved in TGFβ inhibition, including downregulation of CYP24A1 and SERPINE1 and upregulation of WT1. This study highlights the application of iPSC-derived renal cells for prediction of cellular toxicity and sheds new light on the temporal and early effects of key genes that are involved in cellular stress response pathways.
MeSH term(s)	Transcriptome ; Induced Pluripotent Stem Cells ; Gene Expression Profiling ; Kidney
Chemical Substances	4-(4-(3-(pyridin-2-yl)-1H-pyrazol-4-yl)pyridin-2-yl)-N-(tetrahydro-2H-pyran-4-yl)benzamide
Language	English
Publishing date	2022-12-31
Publishing country	Switzerland
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	48824-0
ISSN	1573-6822 ; 0742-2091
ISSN (online)	1573-6822
ISSN	0742-2091
DOI	10.1007/s10565-022-09783-5
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Article ; Online: Transcriptional landscape of mitochondrial electron transport chain inhibition in renal cells.

Carta, Giada / van der Stel, Wanda / Scuric, Emma W J / Capinha, Liliana / Delp, Johannes / Bennekou, Susanne Hougaard / Forsby, Anna / Walker, Paul / Leist, Marcel / van de Water, Bob / Jennings, Paul

Cell biology and toxicology

2023 Volume 39, Issue 6, Page(s) 3031–3059

Abstract: Analysis of the transcriptomic alterations upon chemical challenge, provides in depth mechanistic information on the compound's toxic mode of action, by revealing specific pathway activation and other transcriptional modulations. Mapping changes in ... ...

Abstract	Analysis of the transcriptomic alterations upon chemical challenge, provides in depth mechanistic information on the compound's toxic mode of action, by revealing specific pathway activation and other transcriptional modulations. Mapping changes in cellular behaviour to chemical insult, facilitates the characterisation of chemical hazard. In this study, we assessed the transcriptional landscape of mitochondrial impairment through the inhibition of the electron transport chain (ETC) in a human renal proximal tubular cell line (RPTEC/TERT1). We identified the unfolded protein response pathway (UPR), particularly the PERK/ATF4 branch as a common cellular response across ETC I, II and III inhibitions. This finding and the specific genes elaborated may aid the identification of mitochondrial liabilities of chemicals in both legacy data and prospective transcriptomic studies.
MeSH term(s)	Humans ; Electron Transport/genetics ; Prospective Studies ; Kidney/metabolism ; Cell Line ; Epithelial Cells/metabolism
Language	English
Publishing date	2023-06-23
Publishing country	Switzerland
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	48824-0
ISSN	1573-6822 ; 0742-2091
ISSN (online)	1573-6822
ISSN	0742-2091
DOI	10.1007/s10565-023-09816-7
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Article ; Online: Transcriptomic-based evaluation of trichloroethylene glutathione and cysteine conjugates demonstrate phenotype-dependent stress responses in a panel of human in vitro models

Capinha, Liliana / Zhang, Yaran / Holzer, Anna-Katharina / Ückert, Anna-Katharina / Zana, Melinda / Carta, Giada / Murphy, Cormac / Baldovini, Jenna / Mazidi, Zahra / Grillari, Johannes / Dinnyes, Andras / van de Water, Bob / Leist, Marcel / Commandeur, Jan N. M. / Jennings, Paul

Arch Toxicol. 2023 Feb., v. 97, no. 2, p. 523-545

2023 , Page(s) 523–545

Abstract: Environmental or occupational exposure of humans to trichloroethylene (TCE) has been associated with different extrahepatic toxic effects, including nephrotoxicity and neurotoxicity. Bioactivation of TCE via the glutathione (GSH) conjugation pathway has ... ...

Abstract	Environmental or occupational exposure of humans to trichloroethylene (TCE) has been associated with different extrahepatic toxic effects, including nephrotoxicity and neurotoxicity. Bioactivation of TCE via the glutathione (GSH) conjugation pathway has been proposed as underlying mechanism, although only few mechanistic studies have used cell models of human origin. In this study, six human derived cell models were evaluated as in vitro models representing potential target tissues of TCE-conjugates: RPTEC/TERT1 (kidney), HepaRG (liver), HUVEC/TERT2 (vascular endothelial), LUHMES (neuronal, dopaminergic), human induced pluripotent stem cells (hiPSC) derived peripheral neurons (UKN5) and hiPSC-derived differentiated brain cortical cultures containing all subtypes of neurons and astrocytes (BCC42). A high throughput transcriptomic screening, utilizing mRNA templated oligo-sequencing (TempO-Seq), was used to study transcriptomic effects after exposure to TCE-conjugates. Cells were exposed to a wide range of concentrations of S-(1,2-trans-dichlorovinyl)glutathione (1,2-DCVG), S-(1,2-trans-dichlorovinyl)-L-cysteine (1,2-DCVC), S-(2,2-dichlorovinyl)glutathione (2,2-DCVG), and S-(2,2-dichlorovinyl)-L-cysteine (2,2-DCVC). 1,2-DCVC caused stress responses belonging to the Nrf2 pathway and Unfolded protein response in all the tested models but to different extents. The renal model was the most sensitive model to both 1,2-DCVC and 1,2-DCVG, with an early Nrf2-response at 3 µM and hundreds of differentially expressed genes at higher concentrations. Exposure to 2,2-DCVG and 2,2-DCVC also resulted in the upregulation of Nrf2 pathway genes in RPTEC/TERT1 although at higher concentrations. Of the three neuronal models, both the LUHMES and BCC42 showed significant Nrf2-responses and at higher concentration UPR-responses, supporting recent hypotheses that 1,2-DCVC may be involved in neurotoxic effects of TCE. The cell models with the highest expression of γ-glutamyltransferase (GGT) enzymes, showed cellular responses to both 1,2-DCVG and 1,2-DCVC. Little to no effects were found in the neuronal models from 1,2-DCVG exposure due to their low GGT-expression. This study expands our knowledge on tissue specificity of TCE S-conjugates and emphasizes the value of human cell models together with transcriptomics for such mechanistic studies.
Keywords	astrocytes ; brain ; cysteine ; gene expression regulation ; glutathione ; humans ; kidneys ; liver ; models ; nephrotoxicity ; neurons ; neurotoxicity ; occupational exposure ; transcriptomics ; trichloroethylene ; unfolded protein response
Language	English
Dates of publication	2023-02
Size	p. 523-545
Publishing place	Springer Berlin Heidelberg
Document type	Article ; Online
ZDB-ID	124992-7
ISSN	1432-0738 ; 0340-5761
ISSN (online)	1432-0738
ISSN	0340-5761
DOI	10.1007/s00204-022-03436-6
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Article ; Online: A quantitative AOP of mitochondrial toxicity based on data from three cell lines.

Tebby, Cleo / Gao, Wang / Delp, Johannes / Carta, Giada / van der Stel, Wanda / Leist, Marcel / Jennings, Paul / van de Water, Bob / Bois, Frederic Y

Toxicology in vitro : an international journal published in association with BIBRA

2022 Volume 81, Page(s) 105345

Abstract: Adverse Outcome Pathways (AOPs) are increasingly used to support the integration of in vitro data in hazard assessment for chemicals. Quantitative AOPs (qAOPs) use mathematical models to describe the relationship between key events (KEs). In this paper, ... ...

Abstract	Adverse Outcome Pathways (AOPs) are increasingly used to support the integration of in vitro data in hazard assessment for chemicals. Quantitative AOPs (qAOPs) use mathematical models to describe the relationship between key events (KEs). In this paper, data obtained in three cell lines, LHUMES, HepG2 and RPTEC/TERT1, using similar experimental protocols, was used to calibrate a qAOP of mitochondrial toxicity for two chemicals, rotenone and deguelin. The objectives were to determine whether the same qAOP could be used for the three cell types, and to test chemical-independence by cross-validation with a dataset obtained on eight other chemicals in LHUMES cells. Repeating the calibration approach for both chemicals in three cell lines highlighted various practical difficulties. Even when the same readouts of KEs are measured, the mathematical functions used to describe the key event relationships may not be the same. Cross-validation in LHUMES cells was attempted by estimating chemical-specific potency at the molecular initiating events and using the rest of the calibrated qAOP to predict downstream KEs: toxicity of azoxystrobin, carboxine, mepronil and thifluzamide was underestimated. Selection of most relevant readouts and accurate characterization of the molecular initiating event for cross-validation are critical when designing in vitro experiments targeted at calibrating qAOPs.
MeSH term(s)	Adverse Outcome Pathways ; Cell Line ; Models, Theoretical ; Risk Assessment ; Toxicity Tests
Language	English
Publishing date	2022-03-10
Publishing country	England
Document type	Journal Article
ZDB-ID	639064-x
ISSN	1879-3177 ; 0887-2333
ISSN (online)	1879-3177
ISSN	0887-2333
DOI	10.1016/j.tiv.2022.105345
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Article ; Online: Transcriptomic-based evaluation of trichloroethylene glutathione and cysteine conjugates demonstrate phenotype-dependent stress responses in a panel of human in vitro models.

Archives of toxicology

2022 Volume 97, Issue 2, Page(s) 523–545

Abstract	Environmental or occupational exposure of humans to trichloroethylene (TCE) has been associated with different extrahepatic toxic effects, including nephrotoxicity and neurotoxicity. Bioactivation of TCE via the glutathione (GSH) conjugation pathway has been proposed as underlying mechanism, although only few mechanistic studies have used cell models of human origin. In this study, six human derived cell models were evaluated as in vitro models representing potential target tissues of TCE-conjugates: RPTEC/TERT1 (kidney), HepaRG (liver), HUVEC/TERT2 (vascular endothelial), LUHMES (neuronal, dopaminergic), human induced pluripotent stem cells (hiPSC) derived peripheral neurons (UKN5) and hiPSC-derived differentiated brain cortical cultures containing all subtypes of neurons and astrocytes (BCC42). A high throughput transcriptomic screening, utilizing mRNA templated oligo-sequencing (TempO-Seq), was used to study transcriptomic effects after exposure to TCE-conjugates. Cells were exposed to a wide range of concentrations of S-(1,2-trans-dichlorovinyl)glutathione (1,2-DCVG), S-(1,2-trans-dichlorovinyl)-L-cysteine (1,2-DCVC), S-(2,2-dichlorovinyl)glutathione (2,2-DCVG), and S-(2,2-dichlorovinyl)-L-cysteine (2,2-DCVC). 1,2-DCVC caused stress responses belonging to the Nrf2 pathway and Unfolded protein response in all the tested models but to different extents. The renal model was the most sensitive model to both 1,2-DCVC and 1,2-DCVG, with an early Nrf2-response at 3 µM and hundreds of differentially expressed genes at higher concentrations. Exposure to 2,2-DCVG and 2,2-DCVC also resulted in the upregulation of Nrf2 pathway genes in RPTEC/TERT1 although at higher concentrations. Of the three neuronal models, both the LUHMES and BCC42 showed significant Nrf2-responses and at higher concentration UPR-responses, supporting recent hypotheses that 1,2-DCVC may be involved in neurotoxic effects of TCE. The cell models with the highest expression of γ-glutamyltransferase (GGT) enzymes, showed cellular responses to both 1,2-DCVG and 1,2-DCVC. Little to no effects were found in the neuronal models from 1,2-DCVG exposure due to their low GGT-expression. This study expands our knowledge on tissue specificity of TCE S-conjugates and emphasizes the value of human cell models together with transcriptomics for such mechanistic studies.
MeSH term(s)	Humans ; Cysteine/toxicity ; Cysteine/metabolism ; Trichloroethylene/toxicity ; Trichloroethylene/metabolism ; Transcriptome ; NF-E2-Related Factor 2/metabolism ; Induced Pluripotent Stem Cells/metabolism ; Glutathione/metabolism ; Phenotype
Chemical Substances	S-(1,2-dichlorovinyl)cysteine (627-72-5) ; Cysteine (K848JZ4886) ; Trichloroethylene (290YE8AR51) ; NF-E2-Related Factor 2 ; Glutathione (GAN16C9B8O)
Language	English
Publishing date	2022-12-28
Publishing country	Germany
Document type	Journal Article
ZDB-ID	124992-7
ISSN	1432-0738 ; 0340-5761
ISSN (online)	1432-0738
ISSN	0340-5761
DOI	10.1007/s00204-022-03436-6
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Article ; Online: Expression of xenobiotic transporters in the human renal proximal tubule cell line RPTEC/TERT1.

Aschauer, Lydia / Carta, Giada / Vogelsang, Nadine / Schlatter, Eberhard / Jennings, Paul

Toxicology in vitro : an international journal published in association with BIBRA

2014 Volume 30, Issue 1 Pt A, Page(s) 95–105

Abstract: The kidney is a major target for drug-induced injury, primarily due the fact that it transports a wide variety of chemical entities into and out of the tubular lumen. Here, we investigated the expression of the main xenobiotic transporters in the human ... ...

Abstract	The kidney is a major target for drug-induced injury, primarily due the fact that it transports a wide variety of chemical entities into and out of the tubular lumen. Here, we investigated the expression of the main xenobiotic transporters in the human renal proximal tubule cell line RPTEC/TERT1 at an mRNA and/or protein level. RPTEC/TERT1 cells expressed OCT2, OCT3, OCTN2, MATE1, MATE2, OAT1, OAT3 and OAT4. The functionality of the OCTs was demonstrated by directional transport of the fluorescent dye 4-Di-1-ASP. In addition, P-glycoprotein activity in RPTEC/TERT1 cells was verified by fluorescent dye retention in presence of various P-glycoprotein inhibitors. In comparison to proliferating cells, contact inhibited RPTEC/TERT1 cells expressed increased mRNA levels of several ABC transporter family members and were less sensitive to cyclosporine A. We conclude that differentiated RPTEC/TERT1 cells are well suited for utilisation in xenobiotic transport and pharmacokinetic studies.
MeSH term(s)	Carrier Proteins/genetics ; Carrier Proteins/metabolism ; Cell Line ; Gene Expression Regulation/physiology ; Humans ; Kidney Tubules, Proximal/cytology ; Kidney Tubules, Proximal/metabolism ; RNA, Messenger/genetics ; RNA, Messenger/metabolism
Chemical Substances	Carrier Proteins ; RNA, Messenger
Language	English
Publishing date	2014-12-10
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	639064-x
ISSN	1879-3177 ; 0887-2333
ISSN (online)	1879-3177
ISSN	0887-2333
DOI	10.1016/j.tiv.2014.12.003
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Article ; Online: Correction to: Multiparametric assessment of mitochondrial respiratory inhibition in HepG2 and RPTEC/TERT1 cells using a panel of mitochondrial targeting agrochemicals.

van der Stel, Wanda / Carta, Giada / Eakins, Julie / Darici, Salihanur / Delp, Johannes / Forsby, Anna / Bennekou, Susanne Hougaard / Gardner, Iain / Leist, Marcel / Danen, Erik H J / Walker, Paul / van de Water, Bob / Jennings, Paul

Archives of toxicology

2020 Volume 94, Issue 8, Page(s) 2731–2732

Abstract: In the original publication of the article. ...

Abstract	In the original publication of the article.
Language	English
Publishing date	2020-07-27
Publishing country	Germany
Document type	Journal Article ; Published Erratum
ZDB-ID	124992-7
ISSN	1432-0738 ; 0340-5761
ISSN (online)	1432-0738
ISSN	0340-5761
DOI	10.1007/s00204-020-02849-5
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Article ; Online: Multiparametric assessment of mitochondrial respiratory inhibition in HepG2 and RPTEC/TERT1 cells using a panel of mitochondrial targeting agrochemicals.

Archives of toxicology

2020 Volume 94, Issue 8, Page(s) 2707–2729

Abstract: Evidence is mounting for the central role of mitochondrial dysfunction in several pathologies including metabolic diseases, accelerated ageing, neurodegenerative diseases and in certain xenobiotic-induced organ toxicity. Assessing mitochondrial ... ...

Abstract	Evidence is mounting for the central role of mitochondrial dysfunction in several pathologies including metabolic diseases, accelerated ageing, neurodegenerative diseases and in certain xenobiotic-induced organ toxicity. Assessing mitochondrial perturbations is not trivial and the outcomes of such investigations are dependent on the cell types used and assays employed. Here we systematically investigated the effect of electron transport chain (ETC) inhibitors on multiple mitochondrial-related parameters in two human cell types, HepG2 and RPTEC/TERT1. Cells were exposed to a broad range of concentrations of 20 ETC-inhibiting agrochemicals and capsaicin, consisting of inhibitors of NADH dehydrogenase (Complex I, CI), succinate dehydrogenase (Complex II, CII) and cytochrome bc1 complex (Complex III, CIII). A battery of tests was utilised, including viability assays, lactate production, mitochondrial membrane potential (MMP) and the Seahorse bioanalyser, which simultaneously measures extracellular acidification rate [ECAR] and oxygen consumption rate [OCR]. CI inhibitors caused a potent decrease in OCR, decreased mitochondrial membrane potential, increased ECAR and increased lactate production in both cell types. Twenty-fourhour exposure to CI inhibitors decreased viability of RPTEC/TERT1 cells and 3D spheroid-cultured HepG2 cells in the presence of glucose. CI inhibitors decreased 2D HepG2 viability only in the absence of glucose. CII inhibitors had no notable effects in intact cells up to 10 µM. CIII inhibitors had similar effects to the CI inhibitors. Antimycin A was the most potent CIII inhibitor, with activity in the nanomolar range. The proposed CIII inhibitor cyazofamid demonstrated a mitochondrial uncoupling signal in both cell types. The study presents a comprehensive example of a mitochondrial assessment workflow and establishes measurable key events of ETC inhibition.
Language	English
Publishing date	2020-07-18
Publishing country	Germany
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	124992-7
ISSN	1432-0738 ; 0340-5761
ISSN (online)	1432-0738
ISSN	0340-5761
DOI	10.1007/s00204-020-02792-5
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Article ; Online: Generation and characterization of iPSC-derived renal proximal tubule-like cells with extended stability.

Chandrasekaran, Vidya / Carta, Giada / da Costa Pereira, Daniel / Gupta, Rajinder / Murphy, Cormac / Feifel, Elisabeth / Kern, Georg / Lechner, Judith / Cavallo, Anna Lina / Gupta, Shailesh / Caiment, Florian / Kleinjans, Jos C S / Gstraunthaler, Gerhard / Jennings, Paul / Wilmes, Anja

Scientific reports

2021 Volume 11, Issue 1, Page(s) 11575

Abstract: The renal proximal tubule is responsible for re-absorption of the majority of the glomerular filtrate and its proper function is necessary for whole-body homeostasis. Aging, certain diseases and chemical-induced toxicity are factors that contribute to ... ...

Abstract	The renal proximal tubule is responsible for re-absorption of the majority of the glomerular filtrate and its proper function is necessary for whole-body homeostasis. Aging, certain diseases and chemical-induced toxicity are factors that contribute to proximal tubule injury and chronic kidney disease progression. To better understand these processes, it would be advantageous to generate renal tissues from human induced pluripotent stem cells (iPSC). Here, we report the differentiation and characterization of iPSC lines into proximal tubular-like cells (PTL). The protocol is a step wise exposure of small molecules and growth factors, including the GSK3 inhibitor (CHIR99021), the retinoic acid receptor activator (TTNPB), FGF9 and EGF, to drive iPSC to PTL via cell stages representing characteristics of early stages of renal development. Genome-wide RNA sequencing showed that PTL clustered within a kidney phenotype. PTL expressed proximal tubular-specific markers, including megalin (LRP2), showed a polarized phenotype, and were responsive to parathyroid hormone. PTL could take up albumin and exhibited ABCB1 transport activity. The phenotype was stable for up to 7 days and was maintained after passaging. This protocol will form the basis of an optimized strategy for molecular investigations using iPSC derived PTL.
MeSH term(s)	Biomarkers/metabolism ; Cell Differentiation ; Cells, Cultured ; Humans ; Induced Pluripotent Stem Cells/cytology ; Kidney Tubules, Proximal/cytology ; Sequence Analysis, RNA/methods
Chemical Substances	Biomarkers
Language	English
Publishing date	2021-06-02
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2615211-3
ISSN	2045-2322 ; 2045-2322
ISSN (online)	2045-2322
ISSN	2045-2322
DOI	10.1038/s41598-021-89550-4
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