Use of the ES-D3 cell differentiation assay, combined with the BeWo transport model, to predict relative in vivo developmenatl toxicity of antifungal compounds
Li, H. ; Rietjens, I. ; Louisse, J. ; Blok, M. ; Wang, X. ; Snijders, L. ; Ravenzwaay, B. van - \ 2015
Toxicology in Vitro 29 (2015)2. - ISSN 0887-2333 - p. 320 - 328.
dose-response curves - placental perfusion - test est - vitro - toxicology - potency - risk - rat
We investigated the applicability of the ES-D3 cell differentiation assay combined with the in vitro BeWo transport model to predict the relative in vivo developmental toxicity potencies. To this purpose, the in vitro developmental toxicity of five antifungal compounds was investigated by characterizing their inhibitory effect on the differentiation of ES-D3 cells into cardiomyocytes. The BeWo transport model, consisting of BeWo b30 cells grown on transwell inserts and mimicking the placental barrier, was used to determine the relative placental transport velocity. The ES-D3 cell differentiation data were first compared to benchmark doses (BMDs) for in vivo developmental toxicity as derived from data reported in the literature. Correlation between the benchmark concentration for 50% effect (BMCd50) values, obtained in the ES-D3 cell differentiation assay, with in vivo BMD10 values showed a reasonable correlation (R2 = 0.57). When the ES-D3 cell differentiation data were combined with the relative transport rates obtained from the BeWo model, the correlation with the in vivo data increased (R2 = 0.95). In conclusion, we show that the ES-D3 cell differentiation assay is able to better predict the in vivo developmental toxicity ranking of antifungal compounds when combined with the BeWo transport model, than as a stand-alone assay.
Relationship between chemical composition and in situ rumen degradation characteristics of maize silages in dairy cows
Ali, M. ; Duinkerken, G. van; Cone, J.W. ; Klop, A. ; Blok, M.C. ; Spek, J.W. ; Bruinenberg, M.H. ; Hendriks, W.H. - \ 2014
Animal 8 (2014)11. - ISSN 1751-7311 - p. 1832 - 1838.
neutral detergent fiber - crude protein - starch degradation - dry-matter - degradability - maturity - digestibility - vitro - digestion - extent
Several in situ studies have been conducted on maize silages to determine the effect of individual factors such as maturity stage, chop length and ensiling of maize crop on the rumen degradation but the information on the relationship between chemical composition and in situ rumen degradation characteristics remains scarce. The objectives of this study were to determine and describe relationships between the chemical composition and the rumen degradation characteristics of dry matter (DM), organic matter (OM), CP, starch and aNDFom (NDF assayed with a heat stable amylase and expressed exclusive of residual ash) of maize silages. In all, 75 maize silage samples were selected, with a broad range in chemical composition and quality parameters. The samples were incubated in the rumen for 2, 4, 8, 16, 32, 72 and 336 h, using the nylon bag technique. Large range was found in the rumen degradable fractions of DM, OM, CP, starch and aNDFom because of the broad range in chemical composition and quality parameters. The new database with in situ rumen degradation characteristics of DM, OM, CP, starch and aNDFom of the maize silages was obtained under uniform experimental conditions; same cows, same incubation protocol and same chemical analysis procedures. Regression equations were developed with significant predictors (P
Relative embryotoxic potency of p-substituted phenols in the embryonic stem cell test (EST) and comparison to their toxic potency in vivo and in the whole embryo culture (WEC) assay
Strikwold, M. ; Woutersen, R.A. ; Spenkelink, A. ; Punt, A. ; Rietjens, I.M.C.M. - \ 2012
Toxicology Letters 213 (2012)2. - ISSN 0378-4274 - p. 235 - 242.
developmental toxicity - rat embryos - vitro - metabolism - workshop - benzene - invivo - glucuronidation - recommendations - cytotoxicity
The applicability of the embryonic stem cell test (EST) as an alternative for in vivo embryotoxicity testing was evaluated for a series of five p-substituted phenols. To this purpose, the potency ranking for this class of compounds derived from the inhibition of cardiomyocyte differentiation in the EST was compared to in vivo embryotoxic potency data obtained from literature and to the potency ranking defined in the in vitro whole embryo culture (WEC) assay. From the results obtained it appears that the EST was able to identify the embryotoxic potential for p-substituted phenols, providing an identical potency ranking compared to the WEC assay. However, the EST was not able to predict an accurate ranking for the phenols compared to their potency observed in vivo. Only phenol, the least potent compound within this series, was correctly ranked. Furthermore, p-mercaptophenol was correctly identified as a relative potent congener of the phenols tested, but its ranking was distorted by p-heptyloxyphenol, of which the toxicity was overestimated in the EST. It is concluded that when attempting to explain the observed disparity in potency rankings between in vitro and in vivo embryotoxicity, the in vitro models should be combined with a kinetic model describing in vivo absorption, distribution, metabolism and excretion processes of the compounds.
Genotoxic effects in the Eastern mudminnow (Umbra pygmaea) after prolonged exposure to River Rhine water, as assessed by use of the in vivo SCE and Comet assays
Penders, E.J.M. ; Spenkelink, A. ; Hoogenboezem, W. ; Rotteveel, S.G.P. ; Maas, J.L. ; Alink, G.M. - \ 2012
Environmental and Molecular Mutagenesis 53 (2012)4. - ISSN 0893-6692 - p. 304 - 310.
surface waters - fish - exchanges - vitro
The production of drinking water from river water requires a certain minimal river water quality. The Association of River Rhine Water Works (RIWA), therefore, operates a monitoring network. In vitro mutagenicity studies have shown that the genotoxicity of the River Rhine water steadily decreased from 1981 until 2001. Compared to a study in 1978, a decrease in genotoxicity was also observed in an in vivo genotoxicity study in 2005, in which Eastern mudminnows (Umbra pygmaea) were exposed to River Rhine water, and gill cells were used for the Sister Chromatid Exchange (SCE) test and the Comet assay. In this 2005 study, the in vivo genotoxicity increased upon extending exposure of the fish from 3 to 11 days. Therefore, the objectives of this study were to investigate (i) whether new data corroborate that in vivo genotoxicity of River Rhine water is at present lower than in 1978, (ii) whether the Comet assay is a suitable alternative to the SCE assay, and (iii) whether further prolonged exposure results in a further increase in in vivo genotoxicity. The new data corroborate that in vivo genotoxicity of River Rhine water is at present lower than in 1978. The Comet assay is a useful addition but does not provide a substitute for the SCE endpoint in these in vivo genotoxicity studies. Prolonging the exposure time of Eastern mudminnows to River Rhine water from 11 to 42 days did not give a significant increase in SCEs and DNA damage (Comet assay) in gill cells. Mol. Mutagen. 2012. (c) 2012 Wiley Periodicals, Inc.
Affecting osteoblastic responses with in vivo engineered potato pectin fragments
Kokkonen, H. ; Verhoef, R.P. ; Kauppinen, K. ; Muhonen, V. ; Jorgensen, B. ; Damager, I. ; Schols, H.A. ; Morra, M. ; Ulvskov, P. ; Tuukkanen, J. - \ 2012
Journal of Biomedical Materials Research Part A 100A (2012)1. - ISSN 1549-3296 - p. 111 - 119.
enzymatically-tailored pectins - hairy ramified regions - plant-cell wall - rhamnogalacturonan-i - citrus pectin - differentiation - vitro - mineralization - proliferation - architecture
Pectins, complex plant-derived polysaccharides, are novel candidates for biomaterial nanocoatings. Pectic rhamnogalacturonan-I regions (RG-I) can be enzymatically treated to so-called modified hairy regions (MHR). We surveyed the growth and differentiation of murine preosteoblastic MC3T3-E1 cells on Petri dishes coated with RG-Is from native or genetically engineered potato tubers. Uncoated tissue culture polystyrene (TCPS) and aminated (AMI) dishes served as controls. MHRPTR_GAL sample was depleted of galactose (9 mol % galactose; 23 mol % arabinose) and MHRPTR_ARA of arabinose (61 mol % galactose; 6 mol % arabinose). Wild-type (modified hairy region from potato pectin (MHRP)_WT) fragment contained default amounts (58 mol % galactose; 13 mol % arabinose) of both sugars. Focal adhesions (FAs) indicating cellular attachment were quantified. Reverse transcriptase polymerase chain reaction (RT-PCR) of alkaline phosphatase and osteocalcin genes indicating osteoblastic differentiation was performed along with staining the produced calcium with tetracycline as an indicator of osteoblastic differentiation. Osteoblasts proliferated on all the samples to some extent. The control surfaces performed better than any of the pectin samples, of which the MHRP_WT seemed to function best. FA length was greater on MHRPTR_GAL than on other pectin samples, otherwise the mutants did not significantly deviate. RT-PCR results indicate that differences between the samples at the gene expression level might be even subtler. However, tetracycline-stained calcium-containing mineral was detected merely only on uncoated TCPS. These results indicate the possibility to affect bone cell growth with in vivo-modified pectin fragments, consecutively providing information on the significance of certain monosaccharides on the biocompatibility of these polysaccharides.
Inhibition of LPS-induced proinflammatory responses of J774.2 macrophages by immobilized enzymatically tailored pectins
Gallet, M. ; Vayssade, M. ; Morra, M. ; Verhoef, R.P. ; Perrone, S. ; Cascardo, G. ; Vigneron, P. ; Schols, H.A. ; Nagel, M.D. - \ 2009
Acta Biomaterialia 5 (2009). - ISSN 1742-7061 - p. 2618 - 2622.
hairy ramified regions - glinus-oppositifolius - rhamnogalacturonan-i - cell - polysaccharides - adhesion - activation - polymer - plant - vitro
The surface of an implant device can be modified by immobilizing biological molecules on it to improve its integration into the host tissue. We have previously demonstrated that enzymatically tailored plant pectins are promising nanocoatings for biomaterials. This study investigates whether a coating of modified hairy region (rhamnogalacturonan-I) from apple pectin (MHR-a) which has anti-adhesive properties can inhibit the generation of inflammatory mediators by lipopolysaccharide (LPS)-activated macrophages. For that purpose, J774.2 murine macrophages were cultured for 24 h on MHR-a-coated Petri dishes and tissue culture polystyrene controls, with and without LPS. Cell morphology, cell growth, nitrite and TNF-a secretion were studied. The results indicate that MHR-a coating inhibits the LPS-induced activation of macrophages.
In Vivo Cell Wall Loosening by Hydroxyl Radicals during Cress Seed Germination and Elongation Growth
Muller, K. ; Linkies, A. ; Vreeburg, R.A.M. ; Fry, S.C. ; Krieger-Liszkay, A. ; Leubner-Metzger, G. - \ 2009
Plant Physiology 150 (2009)4. - ISSN 0032-0889 - p. 1855 - 1865.
endo-beta-mannanase - pollen-tube growth - nadph oxidase - endosperm cap - abscisic-acid - dormancy alleviation - hydrogen-peroxide - arabidopsis - mechanisms - vitro
Loosening of cell walls is an important developmental process in key stages of the plant life cycle, including seed germination, elongation growth, and fruit ripening. Here, we report direct in vivo evidence for hydroxyl radical (·OH)-mediated cell wall loosening during plant seed germination and seedling growth. We used electron paramagnetic resonance spectroscopy to show that ·OH is generated in the cell wall during radicle elongation and weakening of the endosperm of cress (Lepidium sativum; Brassicaceae) seeds. Endosperm weakening precedes radicle emergence, as demonstrated by direct biomechanical measurements. By 3H fingerprinting, we showed that wall polysaccharides are oxidized in vivo by the developmentally regulated action of apoplastic ·OH in radicles and endosperm caps: the production and action of ·OH increased during endosperm weakening and radicle elongation and were inhibited by the germination-inhibiting hormone abscisic acid. Both effects were reversed by gibberellin. Distinct and tissue-specific target sites of ·OH attack on polysaccharides were evident. In vivo ·OH attack on cell wall polysaccharides were evident not only in germinating seeds but also in elongating maize (Zea mays; Poaceae) seedling coleoptiles. We conclude that plant cell wall loosening by ·OH is a controlled action of this type of reactive oxygen species.
Food-associated estrogenic compounds induce estrogen receptor-mediated luciferase gene expression in transgenic male mice
Veld, M.G.R. ter; Zawadzka, E. ; Berg, J.H.J. van den; Saag, P.T. van der; Rietjens, I.M.C.M. ; Murk, A.J. - \ 2008
Chemico-Biological Interactions 174 (2008)2. - ISSN 0009-2797 - p. 126 - 133.
sprague-dawley rats - reporter male-mice - in-vivo - bisphenol-a - phthalate-esters - p-nonylphenol - er-alpha - beta - vitro - assays
The present paper aims at clarifying to what extent seven food-associated compounds, shown before to be estrogenic in vitro, can induce estrogenic effects in male mice with an estrogen receptor (ER)-mediated luciferase (luc) reporter gene system. The luc induction was determined in different tissues 8 h after dosing the ER-luc male mice intraperitoneally (IP) or 14 h after oral dosing. Estradiol-propionate (EP) was used as a positive control at 0.3 and 1 mg/kg bodyweight (bw), DMSO as solvent control. The food-associated estrogenic compounds tested at non-toxic doses were bisphenol A (BPA) and nonylphenol (NP) (both at 10 and 50 mg/kg bw), dichlorodiphenyldichloroethylene (p,p¿-DDE; at 5 and 25 mg/kg bw), quercetin (at 1.66 and 16.6 mg/kg bw), di-isoheptyl phthalate (DIHP), di-(2-ethylhexyl) phthalate (DEHP) and di-(2-ethylhexyl) adipate (DEHA) all at 30 and 100 mg/kg bw. In general IP dosing resulted in higher luc inductions than oral dosing. EP induced luc activity in the liver in a statistically significant dose-related way with the highest induction of all compounds tested which was 20,000 times higher than the induction by the DMSO-control. NP, DDE, DEHA and DIHP did not induce luc activity in any of the tissues tested. BPA induced luc in the liver up to 420 times via both exposure routes. BPA, DEHP and quercetin induced luc activity in the liver after oral exposure. BPA (50 mg/kg bw IP) also induced luc activity in the testis, kidneys and tibia. The current study reveals that biomarker-responses in ER-luc male mice occur after a single oral exposure to food-associated estrogenic model compounds at exposure levels 10 to 104 times higher than the established TDI's for some of these compounds. Given the facts that (i) the present study did not include chronic exposure and that (ii) simultaneous exposure to multiple estrogenic compounds may be a realistic exposure scenario, it remains to be seen whether this margin is sufficiently high
Self-assembling protein arrays on DNA chips by auto-labeling fusion proteins with a single DNA address
Jongsma, M.A. ; Litjens, R.H.G.M. - \ 2006
Proteomics 6 (2006)9. - ISSN 1615-9853 - p. 2650 - 2655.
molecules in-vivo - o-6-alkylguanine-dna alkyltransferase - directed immobilization - microarrays - technology - efficient - ligation - vitro - hagt
The high-throughput deposition of recombinant proteins on chips, beads or biosensor devices would be greatly facilitated by the implementation of self-assembly concepts. DNA-directed immobilization via conjugation of proteins to an oligonucleotide would be preeminently suited for this purpose. Here, we present a unique method to attach a single DNA address to proteins in one step during the purification from the E. coli lysate by fusion to human O6-alkylguanine-DNA-alkyltransferase (SNAP-tagTM) and the Avitag. Use of the conjugates in converting a DNA chip into a protein chip by self assembly is demonstrated.
Estrogenic endpoints in fish early life-stage tests: luciferase and vitellogenin induction in estrogen-responsive transgenic zebrafish
Bogers, R. ; Mutsaerds, E. ; Druke, J. ; Roode, D.F. de; Murk, A.J. ; Burg, B. van der; Legler, J. - \ 2006
Environmental Toxicology and Chemistry 25 (2006)1. - ISSN 0730-7268 - p. 241 - 247.
in-vivo - pimephales-promelas - fathead minnow - danio-rerio - sewage effluent - rainbow-trout - stw effluent - chemicals - exposure - vitro
This study incorporated specific endpoints for estrogenic activity in the early life-stage (ELS) test, as described in Guideline 210 of the Organization for Economic Cooperation and Development and traditionally used for toxicity screening of chemicals. A transgenic zebrafish model expressing an estrogen receptor-mediated luciferase reporter gene was exposed to ethinylestradiol (EE2), and luciferase activity as well as vitellogenin (VTG) was measured. Concentrations of EE2 were tested at 1, 3, or 10 ng/L for 30 d from fertilization or during only the last 4 d with dimethylsulfoxide (DMSO) as presolvent (0.01%). Exposure to EE2 induced no toxic effects. Mean body weights were significantly higher in groups exposed for 30 d in the presence of DMSO, but condition factors were not affected. Significant luciferase and VTG induction occurred following 30-d exposure (3 and 10 ng EE2/L), while only VTG levels were affected in the 4-d exposure (10 ng EE2/L). This study demonstrated the usefulness of incorporating estrogenic endpoints in the OECD ELS test, fitting the requirements for screening estrogenic activity of chemicals. Quantitative measurement of both VTG and luciferase activity proved to be rapid and sensitive. Additional value of using transgenic zebrafish lies in combining VTG measurement with the more mechanistic approach of luciferase induction in one experiment.