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The present study predicts in vivo human and rat red blood cell (RBC) acetylcholinesterase (AChE) inhibition upon diazinon (DZN) exposure using physiological based kinetic (PBK) modelling-facilitated reverse dosimetry. Due to the fact that both DZN and its oxon metabolite diazoxon (DZO) can inhibit AChE, a toxic equivalency factor (TEF) was included in the PBK model to combine the effect of DZN and DZO when predicting in vivo AChE inhibition. The PBK models were defined based on kinetic constants derived from in vitro incubations with liver fractions or plasma of rat and human, and were used to translate in vitro concentration–response curves for AChE inhibition obtained in the current study to predicted in vivo dose–response curves. The predicted dose–response curves for rat matched available in vivo data on AChE inhibition, and the benchmark dose lower confidence limits for 10% inhibition (BMDL10 values) were in line with the reported BMDL10 values. Humans were predicted to be 6-fold more sensitive than rats in terms of AChE inhibition, mainly because of inter-species differences in toxicokinetics. It is concluded that the TEF-coded DZN PBK model combined with quantitative in vitro to in vivo extrapolation (QIVIVE) provides an adequate approach to predict RBC AChE inhibition upon acute oral DZN exposure, and can provide an alternative testing strategy for derivation of a point of departure (POD) in risk assessment. Supplementary Information The online version contains supplementary material available at 10.1007/s00204-021-03015-1.

The aim of the present study was to develop a generic rat physiologically based kinetic (PBK) model that includes a novel testing strategy where active biliary excretion is incorporated using estradiol-17β glucuronide (E217βG) as the model substance. A major challenge was the definition of the scaling factor for the in vitro to in vivo conversion of the PBK-model parameter Vmax. In vitro values for the Vmax and Km for transport of E217βG were found in the literature in four different studies based on experiments with primary rat hepatocytes. The required scaling factor was defined based on fitting the PBK model-based predicted values to reported experimental data on E217βG blood levels and cumulative biliary E217βG excretion. This resulted in a scaling factor of 129 mg protein/g liver. With this scaling factor the PBK model predicted the in vivo data for blood and cumulative biliary E217βG levels with on average of less than 1.8-fold deviation. The study provides a proof of principle on how biliary excretion can be included in a generic PBK model using primary hepatocytes to define the kinetic parameters that describe the biliary excretion.

Human intestinal organoids (HIOs) are a promising in vitro model consisting of different intestinal cell types with a 3D microarchitecture resembling native tissue. In the current study, we aimed to assess the expression of the most common intestinal CYP enzymes in a human induced pluripotent stem cell (hiPSC)-derived HIO model, and the suitability of that model to study chemical-induced changes in CYP expression and activity. We compared this model with the commonly used human colonic adenocarcinoma cell line Caco-2 and with a human primary intestinal epithelial cell (IEC)-based model, closely resembling in vivo tissue. We optimized an existing protocol to differentiate hiPSCs into HIOs and demonstrated that obtained HIOs contain a polarized epithelium with tight junctions consisting of enterocytes, goblet cells, enteroendocrine cells and Paneth cells. We extensively characterized the gene expression of CYPs and activity of CYP3A4/5, indicating relatively high gene expression levels of the most important intestinal CYP enzymes in HIOs compared to the other models. Furthermore, we showed that CYP1A1 and CYP1B1 were induced by β-naphtoflavone in all three models, whereas CYP3A4 was induced by phenobarbital and rifampicin in HIOs, in the IEC-based model (although not statistically significant), but not in Caco-2 cells. Interestingly, CYP2B6 expression was not induced in any of the models by the well-known liver CYP2B6 inducer phenobarbital. In conclusion, our study indicates that hiPSC-based HIOs are a useful in vitro intestinal model to study biotransformation of chemicals in the intestine. Electronic supplementary material The online version of this article (10.1007/s00204-020-02953-6) contains supplementary material, which is available to authorized users.

Examining fatal poisonings, chronic exposure may be reflected by the concentration in tissues known for long-term storage of drugs. Δ9-tetrahydrocannabinol (THC) persists in adipose tissue (AT), but sparse data on synthetic cannabinoids (SC) are available. Thus, a controlled pig study evaluating antemortem (AM) disposition and postmortem (PM) concentration changes of the SC 4-ethylnaphthalene-1-yl-(1-pentylindole-3-yl)methanone (JWH-210) and 2-(4-methoxyphenyl)-1-(1-pentyl-indole-3-yl)methanone (RCS-4) as well as THC in AT was performed. The drugs were administered pulmonarily (200 µg/kg body weight) to twelve pigs. Subcutaneous (s.c.) AT specimens were collected after 15 and 30 min and then hourly up to 8 h. At the end, pigs were sacrificed and s.c., perirenal, and dorsal AT specimens were collected. The carcasses were stored at room temperature (RT; n = 6) or 4 °C (n = 6) and specimens were collected after 24, 48, and 72 h. After homogenization in acetonitrile and standard addition, LC–MS/MS was performed. Maximum concentrations were reached 0.5–2 h after administration amounting to 21 ± 13 ng/g (JWH-210), 24 ± 13 ng/g (RCS-4), and 22 ± 20 ng/g (THC) and stayed at a plateau level. Regarding the metabolites, very low concentrations of N-hydroxypentyl-RCS-4 (HO-RCS-4) were detected from 0.5 to 8 h. PM concentrations of parent compounds did not change significantly (p > 0.05) over time under both storage conditions. Concentrations of HO-RCS-4 significantly (p

Understanding intramammary estrogen homeostasis constitutes the basis of understanding the role of lifestyle factors in breast cancer etiology. Thus, the aim of the present study was to identify variables influencing levels of the estrogens present in normal breast glandular and adipose tissues (GLT and ADT, i.e., 17β-estradiol, estrone, estrone-3-sulfate, and 2-methoxy-estrone) by multiple linear regression models. Explanatory variables (exVARs) considered were (a) levels of metabolic precursors as well as levels of transcripts encoding proteins involved in estrogen (biotrans)formation, (b) data on breast cancer risk factors (i.e., body mass index, BMI, intake of estrogen-active drugs, and smoking) collected by questionnaire, and (c) tissue characteristics (i.e., mass percentage of oil, oil%, and lobule type of the GLT). Levels of estrogens in GLT and ADT were influenced by both extramammary production (menopausal status, intake of estrogen-active drugs, and BMI) thus showing that variables known to affect levels of circulating estrogens influence estrogen levels in breast tissues as well for the first time. Moreover, intratissue (biotrans)formation (by aromatase, hydroxysteroid-17beta-dehydrogenase 2, and beta-glucuronidase) influenced intratissue estrogen levels, as well. Distinct differences were observed between the exVARs exhibiting significant influence on (a) levels of specific estrogens and (b) the same dependent variables in GLT and ADT. Since oil% and lobule type of GLT influenced levels of some estrogens, these variables may be included in tissue characterization to prevent sample bias. In conclusion, evidence for the intracrine activity of the human breast supports biotransformation-based strategies for breast cancer prevention. The susceptibility of estrogen homeostasis to systemic and tissue-specific modulation renders both beneficial and adverse effects of further variables associated with lifestyle and the environment possible. Electronic supplementary material The online version of this article (10.1007/s00204-020-02807-1) contains supplementary material, which is available to authorized users.

Paracetamol (acetaminophen, APAP) overdose is a leading cause of acute drug-induced liver failure. APAP hepatotoxicity is mediated by the reactive metabolite N-acetyl-p-benzoquinone imine (NAPQI). NAPQI is inactivated by conjugation with glutathione (GSH) to APAP-GSH, which is further converted into its cysteine derivative APAP-CYS. Before necrosis of hepatocytes occurs, APAP-CYS is measurable in plasma of the affected patient and it has been proposed as an early biomarker of acetaminophen toxicity. APAP-GSH and APAP-CYS can be extruded by hepatocytes, but the transporters involved are unknown. In this study we examined whether ATP-binding cassette (ABC) transporters play a role in the cellular efflux of APAP, APAP-GSH, and APAP-CYS. The ABC transport proteins P-gp/ABCB1, BSEP/ABCB11, BCRP/ABCG2, and MRP/ABCC1-5 were overexpressed in HEK293 cells and membrane vesicles were produced. Whereas P-gp, BSEP, MRP3, MRP5, and BCRP did not transport any of the compounds, uptake of APAP-GSH was found for MRP1, MRP2 and MRP4. APAP-CYS appeared to be a substrate of MRP4 and none of the ABC proteins transported APAP. The results suggest that the NAPQI metabolite APAP-CYS can be excreted into plasma by MRP4, where it could be a useful biomarker for APAP exposure and toxicity. Characterization of the cellular efflux of APAP-CYS is important for its development as a biomarker, because plasma concentrations might be influenced by drug-transporter interactions and upregulation of MRP4. Electronic supplementary material The online version of this article (10.1007/s00204-020-02793-4) contains supplementary material, which is available to authorized users.

Tamoxifen, a standard therapy for breast cancer, is metabolized to compounds with anti-estrogenic as well as estrogen-like action at the estrogen receptor. Little is known about the formation of estrogen-like metabolites and their biological impact. Thus, we characterized the estrogen-like metabolites tamoxifen bisphenol and metabolite E for their metabolic pathway and their influence on cytochrome P450 activity and ADME gene expression. The formation of tamoxifen bisphenol and metabolite E was studied in human liver microsomes and Supersomes™. Cellular metabolism and impact on CYP enzymes was analyzed in upcyte® hepatocytes. The influence of 5 µM of tamoxifen, anti-estrogenic and estrogen-like metabolites on CYP activity was measured by HPLC MS/MS and on ADME gene expression using RT-PCR analyses. Metabolite E was formed from tamoxifen by CYP2C19, 3A and 1A2 and from desmethyltamoxifen by CYP2D6, 1A2 and 3A. Tamoxifen bisphenol was mainly formed from (E)- and (Z)-metabolite E by CYP2B6 and CYP2C19, respectively. Regarding phase II metabolism, UGT2B7, 1A8 and 1A3 showed highest activity in glucuronidation of tamoxifen bisphenol and metabolite E. Anti-estrogenic metabolites (Z)-4-hydroxytamoxifen, (Z)-endoxifen and (Z)-norendoxifen inhibited the activity of CYP2C enzymes while tamoxifen bisphenol consistently induced CYPs similar to rifampicin and phenobarbital. On the transcript level, highest induction up to 5.6-fold was observed for CYP3A4 by tamoxifen, (Z)-4-hydroxytamoxifen, tamoxifen bisphenol and (E)-metabolite E. Estrogen-like tamoxifen metabolites are formed in CYP-dependent reactions and are further metabolized by glucuronidation. The induction of CYP activity by tamoxifen bisphenol and the inhibition of CYP2C enzymes by anti-estrogenic metabolites may lead to drug–drug-interactions. Electronic supplementary material The online version of this article (10.1007/s00204-017-2147-y) contains supplementary material, which is available to authorized users.

The potential complex formation between microsomal epoxide hydrolase (mEH) and cytochrome P450-dependent monooxygenase (CYP) has been a subject of research for many decades. Such an association would enable efficient substrate channeling between CYP and mEH and as such represent an attractive strategy to prevent deleterious accumulation of harmful metabolic by-products such as CYP-generated epoxide intermediates. However, such complex formation is experimentally difficult to prove, because CYP and mEH are membrane-bound proteins that are prone to unspecific aggregation after solubilization. Here, we report the development of a FRET-based procedure to analyze the mEH–CYP interaction in living cells by fluorescence-activated cell sorting. With this non-invasive procedure, we demonstrate that CYP2J5 and mEH associate in the endoplasmic reticulum of recombinant HEK293 cells to the same extent as do CYP2J5 and its indispensible redox partner cytochrome P450 reductase. This presents final proof for a very close proximity of CYP and mEH in the endoplasmic reticulum, compatible with and indicative of their physical interaction. In addition, we provide with FAMPIR a robust and easy-to-implement general method for analyzing the interaction of ER membrane-resident proteins that share a type I topology. Electronic supplementary material The online version of this article (doi:10.1007/s00204-017-2072-0) contains supplementary material, which is available to authorized users.

UDP-glucuronosyltransferases (UGTs) are involved in phase II conjugation reactions of xenobiotics and differences in their isoform activities result in interindividual kinetic differences of UGT probe substrates. Here, extensive literature searches were performed to identify probe substrates (14) for various UGT isoforms (UGT1A1, UGT1A3, UGT1A4, UGT1A6, UGT1A9, UGT2B7 and UGT2B15) and frequencies of human polymorphisms. Chemical-specific pharmacokinetic data were collected in a database to quantify interindividual differences in markers of acute (Cmax) and chronic (area under the curve, clearance) exposure. Using this database, UGT-related uncertainty factors were derived and compared to the default factor (i.e. 3.16) allowing for interindividual differences in kinetics. Overall, results show that pharmacokinetic data are predominantly available for Caucasian populations and scarce for other populations of different geographical ancestry. Furthermore, the relationships between UGT polymorphisms and pharmacokinetic parameters are rarely addressed in the included studies. The data show that UGT-related uncertainty factors were mostly below the default toxicokinetic uncertainty factor of 3.16, with the exception of five probe substrates (1-OH-midazolam, ezetimibe, raltegravir, SN38 and trifluoperazine), with three of these substrates being metabolised by the polymorphic isoform 1A1. Data gaps and future work to integrate UGT-related variability distributions with in vitro data to develop quantitative in vitro–in vivo extrapolations in chemical risk assessment are discussed. Electronic supplementary material The online version of this article (10.1007/s00204-020-02765-8) contains supplementary material, which is available to authorized users., Article Highlights Extensive literature search of human kinetic parameters for UGT probe substrates. Bayesian meta-analysis quantifying human variability in acute and chronic kinetic parameters. UGT isoform-related uncertainty factors were below the 3.16 kinetic default uncertainty factor for most probe substrates. Quantifying human variability in UGT polymorphisms. Electronic supplementary material The online version of this article (10.1007/s00204-020-02765-8) contains supplementary material, which is available to authorized users.

Physiologically based kinetic (PBK) modelling-based reverse dosimetry is a promising tool for the prediction of in vivo developmental toxicity using in vitro concentration–response data. In the present study, the potential of this approach to predict the dose-dependent increase of uterus weight in rats upon exposure to estrogenic chemicals was assessed. In vitro concentration–response data of 17β-estradiol (E2) and bisphenol A (BPA) obtained in the MCF-7/BOS proliferation assay, the U2OS ER-CALUX assay and the yeast estrogen screen (YES) assay, were translated into in vivo dose–response data in rat, using a PBK model with a minimum number of in vitro and in silico determined parameter values. To evaluate the predictions made, benchmark dose (BMD) analysis was performed on the predicted dose–response data and the obtained BMDL10 values were compared with BMDL10 values derived from data on the effects of E2 and BPA in the uterotrophic assay reported in the literature. The results show that predicted dose–response data of E2 and BPA matched with the data from in vivo studies when predictions were made based on YES assay data. The YES assay-based predictions of the BMDL10 values differed 3.9-fold (E2) and 4.7- to 13.4-fold (BPA) from the BMDL10 values obtained from the in vivo data. The present study provides the proof-of-principle that PBK modelling-based reverse dosimetry of YES assay data using a minimum PBK model can predict dose-dependent in vivo uterus growth caused by estrogenic chemicals. In future studies, the approach should be extended to include other estrogens. Electronic supplementary material The online version of this article (10.1007/s00204-017-2140-5) contains supplementary material, which is available to authorized users.

Human precision-cut liver slices (hPCLS) are a valuable ex vivo model that can be used in acute toxicity studies. However, a rapid decline in metabolic enzyme activity limits their use in studies that require a prolonged xenobiotic exposure. The aim of the study was to extend the viability and function of hPCLS to 5 days of incubation. hPCLS were incubated in two media developed for long-term culture of hepatocytes, RegeneMed(®), and Cellartis(®), and in the standard medium WME. Maintenance of phase I and II metabolism was studied both on gene expression as well as functional level using a mixture of CYP isoform-specific substrates. Albumin synthesis, morphological integrity, and glycogen storage was assessed, and gene expression was studied by transcriptomic analysis using microarrays with a focus on genes involved in drug metabolism, transport and toxicity. The data show that hPCLS retain their viability and functionality during 5 days of incubation in Cellartis(®) medium. Albumin synthesis as well as the activity and gene expression of phase I and II metabolic enzymes did not decline during 120-h incubation in Cellartis(®) medium, with CYP2C9 activity as the only exception. Glycogen storage and morphological integrity were maintained. Moreover, gene expression changes in hPCLS during incubation were limited and mostly related to cytoskeleton remodeling, fibrosis, and moderate oxidative stress. The expression of genes involved in drug transport, which is an important factor in determining the intracellular xenobiotic exposure, was also unchanged. Therefore, we conclude that hPCLS cultured in Cellartis(®) medium are a valuable human ex vivo model for toxicological and pharmacological studies that require prolonged xenobiotic exposure. CC BY 4.0Erratum in: Archives of Toxicology, volume 91, issue 7, page 2711 (2017). doi:10.1007/s00204-016-1895-4Received: 7 July 2016 / Accepted: 28 September 2016 / Published online: 7 October 2016The authors thank Prof. Dr. Robert Porte and all the surgeons of the University Medical Center Groningen for providing the human liver tissue. This study was financially supported by the EU-funded project NanoBio4Trans (Grant No. 304842).

The present study describes physiologically based kinetic (PBK) models for the alkenylbenzene myristicin that were developed by extension of the PBK models for the structurally related alkenylbenzene safrole in rat and human. The newly developed myristicin models revealed that the formation of the proximate carcinogenic metabolite 1′-hydroxymyristicin in liver is at most 1.8 fold higher in rat than in human and limited for the ultimate carcinogenic metabolite 1′-sulfoxymyristicin to (2.8–4.0)-fold higher in human. In addition, a comparison was made between the relative importance of bioactivation for myristicin and safrole. Model predictions indicate that for these related compounds, the formation of the 1′-sulfoxy metabolites in rat and human liver is comparable with a difference of

Urine and plasma have been used to date for the biomonitoring of exposure to pollutants and are still the preferred fluids for this purpose; however, these fluids mainly provide information on the short term and may present a high level of variability regarding pesticide concentrations, especially for nonpersistent compounds. Hair analysis may provide information about chronic exposure that is averaged over several months; therefore, this method has been proposed as an alternative to solely relying on these fluids. Although the possibility of detecting pesticides in hair has been demonstrated over the past few years, the unknown linkage between exposure and pesticides concentration in hair has limited the recognition of this matrix as a relevant tool for assessing human exposure. Based on a rat model in which there was controlled exposure to a mixture of pesticides composed of lindane, β-hexachlorocyclohexane, β-endosulfan, p,p′-DDT, p,p′-DDE, dieldrin, pentachlorophenol, diazinon, chlorpyrifos, cyhalothrin, permethrin, cypermethrin, propiconazole, fipronil, oxadiazon, diflufenican, trifluralin, carbofuran, and propoxur, the current work demonstrates the association between exposure intensity and resulting pesticide concentration in hair. We also compared the results obtained from a hair analysis to urine and plasma collected from the same rats. Hair, blood, and urine were collected from rats submitted to 90-day exposure by gavage to the aforementioned mixture of common pesticides at different levels. We observed a linear relationship between exposure intensity and the concentration of pesticides in the rats’ hair (R Pearson 0.453–0.978, p

Microsomal epoxide hydrolase (mEH) is a detoxifying enzyme for xenobiotic compounds. Enzymatic activity of mEH can be greatly increased by a point mutation, leading to an E404D amino acid exchange in its catalytic triad. Surprisingly, this variant is not found in any vertebrate species, despite the obvious advantage of accelerated detoxification. We hypothesized that this evolutionary avoidance is due to the fact that the mEH plays a dualistic role in detoxification and control of endogenous vascular signaling molecules. To test this, we generated mEH E404D mice and assessed them for detoxification capacity and vascular dynamics. In liver microsomes from these mice, turnover of the xenobiotic compound phenanthrene-9,10-oxide was four times faster compared to WT liver microsomes, confirming accelerated detoxification. mEH E404D animals also showed faster metabolization of a specific class of endogenous eicosanoids, arachidonic acid-derived epoxyeicosatrienoic acids (EETs) to dihydroxyeicosatrienoic acids (DHETs). Significantly higher DHETs/EETs ratios were found in mEH E404D liver, urine, plasma, brain and cerebral endothelial cells compared to WT controls, suggesting a broad impact of the mEH mutant on endogenous EETs metabolism. Because EETs are strong vasodilators in cerebral vasculature, hemodynamics were assessed in mEH E404D and WT cerebral cortex and hippocampus using cerebral blood volume (CBV)-based functional magnetic resonance imaging (fMRI). Basal CBV0 levels were similar between mEH E404D and control mice in both brain areas. But vascular reactivity and vasodilation in response to the vasodilatory drug acetazolamide were reduced in mEH E404D forebrain compared to WT controls by factor 3 and 2.6, respectively. These results demonstrate a critical role for mEH E404D in vasodynamics and suggest that deregulation of endogenous signaling pathways is the undesirable gain of function associated with the E404D variant. Electronic supplementary material The online version of this article (doi:10.1007/s00204-016-1666-2) contains supplementary material, which is available to authorized users.

Studies on the genotoxicity of Alternaria mycotoxins focus primarily on the native compounds. Alternariol (AOH) and its methyl ether (AME) have been reported to represent substrates for cytochrome P450 enzymes, generating hydroxylated metabolites. The impact of these phase I metabolites on genotoxicity remains unknown. In the present study, the synthesis and the toxicological effects of the metabolites 4-hydroxy alternariol (4-OH-AOH) and 4-hydroxy alternariol monomethyl ether (4-OH-AME) are presented and compared to the effects of the parent molecules. Although the two phase I metabolites contain a catecholic structure, which is expected to be involved in redox cycling, only 4-OH-AOH increased reactive oxygen species (ROS) in human esophageal cells (KYSE510), 4 times more pronounced than AOH. No ROS induction was observed for 4-OH-AME, although the parent compound showed some minor impact. Under cell-free conditions, both metabolites inhibited topoisomerase II activity comparable to their parent compounds. In KYSE510 cells, both metabolites were found to enhance the level of transient DNA–topoisomerase complexes in the ICE assay. Although the level of ROS was significantly increased by 4-OH-AOH, neither DNA strand breaks nor enhanced levels of formamidopyrimidine-DNA-glycosylase (FPG)-sensitive sites were observed. In contrast, AOH induced significant DNA damage in KYSE510 cells. Less pronounced or even absent effects of hydroxylated metabolites compared to the parent compounds might at least partly be explained by their poor cellular uptake. Glucuronidation as well as sulfation appear to have only a minor influence. Instead, methylation of 4-OH-AOH seems to be the preferred way of metabolism in KYSE510 cells, whereby the toxicological relevance of the methylation product remains to be clarified. Electronic supplementary material The online version of this article (doi:10.1007/s00204-016-1801-0) contains supplementary material, which is available to authorized users.

5-Hydroxymethylfurfural (HMF) and furfuryl alcohol (FFA) are moderately potent rodent carcinogens that are present in thermally processed foodstuffs. The carcinogenic effects were hypothesized to originate from sulfotransferase (SULT)-mediated bioactivation yielding DNA-reactive and mutagenic sulfate esters, a confirmed metabolic pathway of HMF and FFA in mice. It is known that orthologous SULT forms substantially differ in substrate specificity and tissue distribution. This could influence HMF- and FFA-induced carcinogenic effects. Here, we studied HMF and FFA sulfoconjugation by 30 individual SULT forms of humans, mice and rats. The catalytic efficiencies (kcat/KM) of HMF sulfoconjugation of human SULT1A1 (13.7 s−1 M−1), mouse Sult1a1 (15.8 s−1 M−1) and 1d1 (4.8 s−1 M−1) and rat Sult1a1 (5.3 s−1 M−1) were considerably higher than those of all other SULT forms investigated (≤0.73 s−1 M−1). FFA sulfoconjugation was monitored using adenosine as a nucleophilic scavenger for the reactive 2-sulfoxymethylfuran (t1/2 = 20 s at 37 °C). The resulting adduct N6-((furan-2-yl)methyl)-adenosine (N6-MF-A) was quantified by isotope-dilution UPLC-MS/MS. The rates of N6-MF-A formation showed that hSULT1A1 and its orthologues in mice and rats were also the most important contributors to FFA sulfoconjugation in each of the species. Taken together, the catalytic capacity of hSULT1A1 is comparable to that of mSult1a1 in mice, the species in which carcinogenic effects of HMF and FFA were detected. This is of primary concern due to the expression of hSULT1A1 in many different tissues. Electronic supplementary material The online version of this article (doi:10.1007/s00204-014-1392-6) contains supplementary material, which is available to authorized users.

Differentiated human bronchial epithelial cells in air liquid interface cultures (ALI-PBEC) represent a promising alternative for inhalation studies with rodents as these 3D airway epithelial tissue cultures recapitulate the human airway in multiple aspects, including morphology, cell type composition, gene expression and xenobiotic metabolism. We performed a detailed longitudinal gene expression analysis during the differentiation of submerged primary human bronchial epithelial cells into ALI-PBEC to assess the reproducibility and inter-individual variability of changes in transcriptional activity during this process. We generated ALI-PBEC cultures from four donors and focussed our analysis on the expression levels of 362 genes involved in biotransformation, which are of primary importance for toxicological studies. Expression of various of these genes (e.g., GSTA1, ADH1C, ALDH1A1, CYP2B6, CYP2F1, CYP4B1, CYP4X1 and CYP4Z1) was elevated following the mucociliary differentiation of airway epithelial cells into a pseudo-stratified epithelial layer. Although a substantial number of genes were differentially expressed between donors, the differences in fold changes were generally small. Metabolic activity measurements applying a variety of different cytochrome p450 substrates indicated that epithelial cultures at the early stages of differentiation are incapable of biotransformation. In contrast, mature ALI-PBEC cultures were proficient in the metabolic conversion of a variety of substrates albeit with considerable variation between donors. In summary, our data indicate a distinct increase in biotransformation capacity during differentiation of PBECs at the air–liquid interface and that the generation of biotransformation competent ALI-PBEC cultures is a reproducible process with little variability between cultures derived from four different donors. Electronic supplementary material The online version of this article (doi:10.1007/s00204-016-1868-7) contains supplementary material, which is available to authorized users.

Diphenylarsinic acid (DPAA) is an organic arsenic compound used for the synthesis of chemical weapons. We previously found that the residents of Kamisu city in Ibaraki Prefecture, Japan, were exposed to DPAA through contaminated well water in 2003. Although mounting evidence strongly suggests that their neurological symptoms were caused by DPAA, the dynamics of DPAA distribution and metabolism after ingestion by humans remain to be elucidated. To accurately predict the distribution of DPAA in the human body, we administrated DPAA (1.0 mg/kg/day) to cynomolgus monkeys (n = 28) for 28 days. The whole tissues from these monkeys were collected at 5, 29, 170, and 339 days after the last administration. The concentration of DPAA in these tissues was measured by liquid chromatography–mass spectrometry. We found that DPAA accumulated in the central nervous system tissues for a longer period than in other tissues. This finding would extend our knowledge on the distribution dynamics and metabolism of DPAA in primates, including humans. Furthermore, it may be useful for developing a treatment strategy for patients who are exposed to DPAA.

Considering the rapid developments in food safety in the past decade in China, it is of importance to obtain insight into what extent safety and risk assessments of chemicals performed for the Caucasian population apply to the Chinese population. The aim of the present study was to determine physiologically based kinetic (PBK) modeling-based predictions for differences between Chinese and Caucasians in terms of metabolic bioactivation and detoxification of the food-borne genotoxic carcinogen estragole. The PBK models were defined based on kinetic constants for hepatic metabolism derived from in vitro incubations using liver fractions of the two ethnic groups, and used to evaluate the inter-ethnic differences in metabolic activation and detoxification of estragole. The models predicted that at realistic dietary intake levels, only 0.02% of the dose was converted to the ultimate carcinogenic metabolite 1′-sulfooxyestragole in Chinese subjects, whereas this amounted to 0.09% of the dose in Caucasian subjects. Detoxification of 1′-hydroxyestragole, mainly via conversion to 1′-oxoestragole, was similar within the two ethnic groups. The 4.5-fold variation in formation of the ultimate carcinogenic metabolite of estragole accompanied by similar rates of detoxification may indicate a lower risk of estragole for the Chinese population at similar levels of exposure. The study provides a proof of principle for how PBK modeling can identify differences in ethnic sensitivity and provide a more refined risk assessment for a specific ethnic group for a compound of concern. Electronic supplementary material The online version of this article (doi:10.1007/s00204-017-1941-x) contains supplementary material, which is available to authorized users.