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- M.L.J. Coonen (1)
- D.A.M. Dartel van (1)
- J. Delft van (1)
- J.H.M. Delft van (1)
- Y.E.M. Dommels (1)
- S. Gaj (1)
- R.W.L. Godschalk (1)
- W.H. Gottschalk (1)
- R.M.M. Haenen (1)
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- M. Herwijnen van (1)
- W.F.P.M. Hof (1)
- P.C.H. Hollman (1)
- D.G.J. Jennen (1)
- D. Jennen (1)
- M.J.A. Jetten (1)
- H.B. Ketelslegers (1)
- J.C.S. Kleinjans (8)
- T.M.C.M. Kok de (1)
- T.M. Kok de (1)
- S. Kol (1)
- A. Lommen (3)
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- L.M. Maas (1)
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- H.J.J. Moonen (1)
- A.M. Osman (1)
- M.F.A. Pachen (1)
- A.A.C.M. Peijnenburg (3)
- J.L.A. Pennings (2)
- A.H. Piersma (1)
- J.F. Robinson (1)
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- A. Ruiz-Aracama (1)
- F.J. Schooten (1)
- E.P. Someren van (1)
- R. Stierum (1)
- A. Summeren van (1)
- P.T. Theunissen (1)
- L.C. Wilms (2)
- W.K.W.H. Wodzig (1)
- M. Zwam (1)
Integrative cross-omics analysis in primary mouse hepatocytes unravels mechanisms of cyclosporin A-induced hepatotoxicity
Hof, W.F.P.M. ; Summeren, A. van; Lommen, A. ; Coonen, M.L.J. ; Brauers, K. ; Herwijnen, M. van; Wodzig, W.K.W.H. ; Kleinjans, J.C.S. - \ 2014
Toxicology 324 (2014). - ISSN 0300-483X - p. 18 - 26.
drug-induced hepatotoxicity - gene-expression - micrornas - repression - normalization - accumulation - metabonomics - translation - activation - biomarkers
The liver is responsible for drug metabolism and drug-induced hepatotoxicity is the most frequent reason for drug withdrawal, indicating that better pre-clinical toxicity tests are needed. In order to bypass animal models for toxicity screening, we exposed primary mouse hepatocytes for exploring the prototypical hepatotoxicant cyclosporin A. To elucidate the mechanisms underlying cyclosporin A-induced hepatotoxicity, we analyzed expression levels of proteins, mRNAs, microRNAs and metabolites.
Complementary Detection of Embryotoxic Properties of Substances in the Neural and Cardiac Embryonic Stem Cell Tests
Theunissen, P.T. ; Pennings, J.L.A. ; Dartel, D.A.M. van; Robinson, J.F. ; Kleinjans, J.C.S. ; Piersma, A.H. - \ 2013
Toxicological sciences 132 (2013)1. - ISSN 1096-6080 - p. 118 - 130.
retinoic acid - in-vitro - gene-expression - developmental toxicity - reproductive toxicants - response evaluation - vitamin-a - test estn - differentiation - models
In developmental toxicity testing, in vitro screening assays are highly needed to increase efficiency and to reduce animal use. A promising in vitro assay is the cardiac embryonic stem cell test (ESTc), in which the effect of developmental toxicants on cardiomyocyte differentiation is assessed. Recently, we developed a neural differentiation variant of the stem cell test (neural embryonic stem cell test [ESTn]). In both of these models, we have previously performed a series of transcriptomic studies to characterize gene expression changes (1) across time during normal differentiation and (2) in response to a series of developmental toxicants in the ESTn and ESTc. Here, using the cumulative of these studies, we compared gene expression profiles of ESTn and ESTc over time as well as model-specific changes induced by seven compounds, comprising six known in vivo developmental toxicants and one negative control. Time-related gene expression profiles showed similarities between the two EST systems. However, specific genes could be identified changing over time differently in each model related to the two different lineages of differentiation. Interestingly, compound-induced gene expression changes were generally model specific, especially for methylmercury and flusilazole, which were predicted better in ESTn and ESTc, respectively. Valproic acidinduced gene expression changes were most comparable out of the six developmental toxicants between the ESTn and ESTc. Direct transcriptomic comparisons between the ESTn and ESTc indicate that combined transcriptomic analyses support and complement each other. Therefore, a combined approach incorporating ESTc and ESTn may improve developmental toxicant detection over individual assays.
'Omics analysis of low dose acetaminophen intake demonstrates novel response pathways in humans
Jetten, M.J.A. ; Gaj, S. ; Ruiz Aracama, A. ; Kok, T.M. de; Delft, J.H.M. van; Lommen, A. ; Someren, E.P. van; Jennen, D. ; Claessen, S.M. ; Peijnenburg, A.A.C.M. ; Stierum, R. ; Kleinjans, J.C.S. - \ 2012
Toxicology and Applied Pharmacology 259 (2012)3. - ISSN 0041-008X - p. 320 - 328.
induced liver-injury - gene-expression - induced hepatotoxicity - circulating micrornas - liquid-chromatography - potential biomarkers - toxicity - metabolomics - metabolites - paracetamol
Acetaminophen is the primary cause of acute liver toxicity in Europe/USA, which led the FDA to reconsider recommendations concerning safe acetaminophen dosage/use. Unfortunately, the current tests for liver toxicity are no ideal predictive markers for liver injury, i.e. they only measure acetaminophen exposure after profound liver toxicity has already occurred. Furthermore, these tests do not provide mechanistic information. Here, 'omics techniques (global analysis of metabolomic/gene-expression responses) may provide additional insight. To better understand acetaminophen-induced responses at low doses, we evaluated the effects of (sub-)therapeutic acetaminophen doses on metabolite formation and global gene-expression changes (including, for the first time, full-genome human miRNA expression changes) in blood/urine samples from healthy human volunteers. Many known and several new acetaminophen-metabolites were detected, in particular in relation to hepatotoxicity-linked, oxidative metabolism of acetaminophen. Transcriptomic changes indicated immune-modulating effects (2 g dose) and oxidative stress responses (4 g dose). For the first time, effects of acetaminophen on full-genome human miRNA expression have been considered and confirmed the findings on mRNA level. 'Omics techniques outperformed clinical chemistry tests and revealed novel response pathways to acetaminophen in humans. Although no definitive conclusion about potential immunotoxic effects of acetaminophen can be drawn from this study, there are clear indications that the immune system is triggered even after intake of low doses of acetaminophen. Also, oxidative stress-related gene responses, similar to those seen after high dose acetaminophen exposure, suggest the occurrence of possible pre-toxic effects of therapeutic acetaminophen doses. Possibly, these effects are related to dose-dependent increases in levels of hepatotoxicity-related metabolites.
Integrating transcriptomics and metabonomics to unravel modes-of-action of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in HepG2 cells
Jennen, D.G.J. ; Ruiz-Aracama, A. ; Magkoufopoulou, C. ; Peijnenburg, A.A.C.M. ; Lommen, A. ; Delft, J. van; Kleinjans, J.C.S. - \ 2011
BMC Systems Biology 5 (2011). - ISSN 1752-0509 - 14 p.
reactive oxygen production - primary human hepatocytes - elicited gene-expression - nongenotoxic carcinogens - oxidative stress - human liver - hepatocellular-carcinoma - induced hepatotoxicity - hydrocarbon receptor - n-acetylaspartate
Background The integration of different 'omics' technologies has already been shown in several in vivo studies to offer a complementary insight into cellular responses to toxic challenges. Being interested in developing in vitro cellular models as alternative to animal-based toxicity assays, we hypothesize that combining transcriptomics and metabonomics data improves the understanding of molecular mechanisms underlying the effects caused by a toxic compound also in vitro in human cells. To test this hypothesis, and with the focus on non-genotoxic carcinogenesis as an endpoint of toxicity, in the present study, the human hepatocarcinoma cell line HepG2 was exposed to the well-known environmental carcinogen 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Results Transcriptomics as well as metabonomics analyses demonstrated changes in TCDD-exposed HepG2 in common metabolic processes, e.g. amino acid metabolism, of which some of the changes only being confirmed if both 'omics' were integrated. In particular, this integrated analysis identified unique pathway maps involved in receptor-mediated mechanisms, such as the G-protein coupled receptor protein (GPCR) signaling pathway maps, in which the significantly up-regulated gene son of sevenless 1 (SOS1) seems to play an important role. SOS1 is an activator of several members of the RAS superfamily, a group of small GTPases known for their role in carcinogenesis. Conclusions The results presented here were not only comparable with other in vitro studies but also with in vivo studies. Moreover, new insights on the molecular responses caused by TCDD exposure were gained by the cross-omics analysis.
Proteomic analysis of mouse thymoma EL4 cells treated with bis (tri-n-butyltin)oxide (TBTO)
Osman, A.M. ; Kol, S. ; Peijnenburg, A.A.C.M. ; Blokland, M.H. ; Pennings, J.L.A. ; Kleinjans, J.C.S. ; Loveren, H. van - \ 2009
Journal of Immunotoxicology 6 (2009)3. - ISSN 1547-691X - p. 174 - 183.
colorectal tumor patients - patients in-vitro - prothymosin-alpha - organotin compounds - endoplasmic-reticulum - induced apoptosis - oxidative stress - rat thymocytes - nonspecific resistance - antitumor-activity
Here, we report the results of proteomic analysis of the mouse thymoma EL4 cell line exposed to bis(tri-n-butylin)oxide (TBTO), an immunotoxic organotin compound. The objective of the work was to examine whether TBTO affects the expression of proteins in this cell line and to compare the differentially expressed proteins with the corresponding mRNA expression data. The identified proteins were quantified using a label-free quantitative method based on counting the observed peptides as an index of protein abundance. The calculation of the ratio of peptides obtained from exposed and control samples allowed us to evaluate the effect of TBTO on protein expression and to compare these results to those obtained in gene expression profiling studies. Correlation of some of the differentially expressed proteins and their corresponding mRNAs was observed. The analysis of the protein ratios revealed that 12 proteins were significantly affected. These proteins included cytoskeleton proteins myosin-9, spectrin beta 2 and plectin 8. The first two proteins were down-regulated 3-fold, whereas the third was up-regulated 2-fold. Ras-related Rab1, a GTP binding protein and T-complex protein-1 subunit alpha, a chaperonin, were decreased 2- and 3.6-fold, respectively. The ribosomal S10 and eukaryotic translation factor (eIf4G1), which are involved in protein synthesis, were down-regulated 2.6- and 3.7-fold, respectively. Also, proteins involved in splicing of pre-mRNA and in transcription, splicing factor arginine/serine-rich 2 and chromodomain-helicase-DNA binding protein 4 (Chd4), were decreased 2.6- and 4.5 times, respectively. Nuclear RNA helicase II was reduced 2.8-fold. Finally, prothymosin-alpha (ProT¿), an essential protein for cell proliferation, and a protein similar to ProT¿, (with a molecular weight and a pI (3.54) comparable to that of ProT¿) were also down-regulated 6-and 8-fold, respectively. We propose that the observed down-regulation of the expression level of ProT¿ in the TBTO-exposed cells could account for the previously reported anti-proliferative effect of TBTO
Impact of multiple genetic polymorphisms on effects of a 4-week blueberry juice intervention on ex vivo induced lymphocytic DNA damage in human volunteers
Wilms, L.C. ; Boots, A.W. ; Boer, V.C.J. de; Maas, L.M. ; Pachen, M.F.A. ; Gottschalk, W.H. ; Ketelslegers, H.B. ; Godschalk, R.W.L. ; Haenen, R.M.M. ; Schooten, F.J. ; Kleinjans, J.C.S. - \ 2007
Carcinogenesis 28 (2007)8. - ISSN 0143-3334 - p. 1800 - 1806.
catechol-o-methyltransferase - epidemiologic evidence - vegetable consumption - antioxidant capacity - adduct levels - comet assay - fruit juice - vitamin-c - in-vitro - flavonoids
Consumption of fruits and vegetables has been associated with a decrease in cancer incidence and cardiovascular disease, presumably caused by antioxidants. We designed a human intervention study to assess antioxidative and possible anti-genotoxic properties of fruit-borne antioxidants. We hypothesized that individuals bearing genetic polymorphisms for genes related to quercetin metabolism, benzo[a]pyrene metabolism, oxidative stress and DNA repair differ in their response to DNA protective effects of increased antioxidant intake. In the present study, 168 healthy volunteers consumed a blueberry/apple juice that provided 97 mg quercetin and 16 mg ascorbic acid a day. After a 4-week intervention period, plasma concentrations of quercetin and ascorbic acid and trolox equivalent antioxidant capacity (TEAC) were significantly increased. Further, we found 20% protection (P <0.01) against ex vivo H2O2-provoked oxidative DNA damage, measured by comet assay. However, the level of ex vivo induced benzo[a]pyrene-diol-epoxide (BPDE)-DNA adducts was 28% increased upon intervention (P <0.01). Statistical analysis of 34 biologically relevant genetic polymorphisms revealed that six significantly influenced the outcome of the intervention. Lymphocytes from individuals bearing variant genotype for Cyp1B1*5 seemed to benefit more than wild-types from DNA damage-protecting effects upon intervention. Variants for COMT tended to benefit less or even experienced detrimental effects from intervention. With respect to GSTT1, the effect is ambiguous; variants respond better in terms of intervention-related increase in TEAC, but wild-types benefit more from its protecting effects against oxidative DNA damage. We conclude that genotyping for relevant polymorphisms enables selecting subgroups among the general population that benefit more of DNA damage-modulating effects of micronutrients.
Protection by quercetin and quercetin-rich fruit juice against induction of oxidative DNA damage and formation of BPDE-DNA adducts in human lymphocytes
Wilms, L.C. ; Hollman, P.C.H. ; Boots, A.W. ; Kleinjans, J.C.S. - \ 2005
Mutation research. Genetic toxicology and environmental mutagenesis 582 (2005)1-2. - ISSN 1383-5718 - p. 155 - 162.
gene-expression - flavonoids - cells - bioavailability - cancer - glycosides - assay - diet
Flavonoids are claimed to protect against cardiovascular disease, certain forms of cancer and ageing, possibly by preventing initial DNA damage. Therefore, we investigated the protective effects of the flavonoid quercetin against the formation of oxidative DNA damage and bulky DNA adducts in human lymphocytes, both in vitro and ex vivo. First, human lymphocytes were pre-incubated with various concentrations of quercetin, followed by incubation with hydrogen peroxide; protection against oxidative DNA damage was evaluated by use of the single-cell gel electrophoresis (Comet) assay. Second, quercetin-treated human lymphocytes were challenged by treatment with benzo(a)pyrene (B(a)P), and BPDE-DNA adduct formation was measured by 32P-postlabelling. Third, in a pilot study, lymphocytes from female volunteers who consumed a quercetin-rich blueberry/apple juice mixture for four weeks, were treated ex vivo with an effective dose of H2O2 and benzo(a)pyrene, respectively, at three different time points, i.e. before (t = 0 weeks), during (t = 2 weeks) and after (t = 4 weeks) the intervention. Results in vitro: a significant dose-dependent protection by quercetin against both the formation of oxidative DNA damage (p <0.01) and of BPDE-DNA adducts (p <0.05) was observed. Results in vivo: four weeks of juice intervention led to a significant increase in the total antioxidant capacity of plasma, as reflected by the increase of the TEAC value from 773 ¿M trolox equivalent at t = 0 to 855 ¿M at t = 4 weeks (p = 0.04) and an increase in plasma quercetin content from 5.0 to 10.6 nM (p = 0.03). After intervention, the level of oxidative damage upon ex vivo exposure to H2O2 was non-significantly (p = 0.07) decreased by 41%, and the BPDE-DNA adduct level induced ex vivo was non-significantly decreased by 11%. The combination of our findings in vitro and ex vivo provides evidence that quercetin is able to protect against chemically induced DNA damage in human lymphocytes, which may underlie its suggested anticarcinogenic properties
Effects of polyunsaturated fatty acids on prostaglandin synthesis and cyclooxygenase-mediated DNA adduct formation by heterocyclic aromatic amines in human adenocarcinoma colon cells
Moonen, H.J.J. ; Dommels, Y.E.M. ; Zwam, M. ; Herwijnen, M.H.M. van; Kleinjans, J.C.S. ; Alink, G.M. ; Kok, T.M.C.M. de - \ 2004
Molecular Carcinogenesis 40 (2004)3. - ISSN 0899-1987 - p. 180 - 188.
nonsteroidal antiinflammatory drugs - human colorectal-cancer - docosahexaenoic acid - liver-microsomes - 2-amino-3-methylimidazo<4,5-f>quinoline iq - cytochrome-p450 isozymes - arachidonic-acid - linoleic acids - dietary-fat - in-vitro
Dietary heterocyclic aromatic amines (HCA) and polyunsaturated fatty acids (PUFA) are both believed to play a role in colon carcinogenesis, and are both substrate for the enzyme cyclooxygenase (COX). In HCA-7 cells, highly expressing isoform COX-2, we investigated the effects of PUFA on prostaglandin synthesis and DNA adduct formation by the HCA 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-3-methylimidazo[4,5-f]quinoline (IQ). Furthermore, we studied the role of COX, COX-2 in particular, and cytochrome P4501A2 (CYP1A2) by using the enzyme inhibitors indomethacin (IM), NS-398, and phenethyl isothiocyanate (PEITC), respectively. COX-mediated formation of prostaglandin E-2 (PGE(2)) from linoleic acid (LA) showed that HCA-7 cells can convert LA into arachidonic acid (AA). Alternatively, eicosapentaenoic acid (EPA) was found to compete with AA for COX. Strongly decreased PGE2 levels by addition of IM demonstrated involvement of COX in PUFA metabolism. Both IM and NS-398 inhibited adduct formation by HCA to nearly the same extent, indicating involvement of COX-2 rather than COX-1, while CYP1A2 activity in HCA-7 cells was demonstrated by addition of PEITC. Overall, inhibiting effects were stronger for PhIP than for IQ. HCA-DNA adduct formation was stimulated by addition of PUFA, although high PUFA concentrations partly reduced this stimulating effect. Finally, similar effects for n-3 and n-6 fatty acids suggested that adduct formation may not be the crucial mechanism behind the differential effects of PUFA on colon carcinogenesis that have been described. These results show that COX, and COX-2 in particular, can play a substantial role in HCA activation, especially in extrahepatic tissues like the colon. Furthermore, the obvious interactions between PUFA and HCA in COX-2 expressing cancer cells may be important in modulating colorectal cancer risk. (C) 2004 Wiley-Liss, Inc.