30 results on '"Kuil T"'
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2. Insights into PPi metabolism of Clostridium thermocellum for improved cellulosic ethanol production
- Author
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Kuil, T., primary, Yayo, J., additional, and van Maris, A. J. A., additional
- Published
- 2022
- Full Text
- View/download PDF
3. Vascular responses of isolated mesenteric resistance and basilar arteries from short- and long-term diabetic rats
- Author
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Van Buren, T., Vleeming, W., Krutzen, M. M., Van de Kuil, T., Gispen, Willem H., and De Wildt, D. J.
- Published
- 1998
- Full Text
- View/download PDF
4. Michiel de Ruyter, Boegbeeld van de Natie?: Onderzoek naar publieksparticipatie en identiteitsvorming in hedendaagse manifestaties rond het fenomeen De Ruyter
- Author
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Kuil, T. van der, Henrichs, H. (Thesis Advisor), Kuil, T. van der, and Henrichs, H. (Thesis Advisor)
- Abstract
In dit onderzoek wordt de De Ruyter-vering in het licht van de huidige debatten over publieksparticipatie en autoriteit in geschiedschrijving geplaatst. De hoofdvraag die in dit onderzoek wordt beantwoord is: wat kan in termen van identiteitsvorming en participatieve historische cultuur worden gezegd over de hedendaagse manifestaties rond het fenomeen Michiel de Ruyter?
- Published
- 2015
5. Beschrijvingen van de Bokkenrijdersbende Een discoursanalyse van geschriften over de Bokkenrijders uit de 18e, 19e en 20e eeuw
- Author
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Kuil, T. van der, Ruberg, W.G. (Thesis Advisor), Kuil, T. van der, and Ruberg, W.G. (Thesis Advisor)
- Abstract
Dit onderzoek gaat over de beeldvorming van de Bokkenrijdersbende. Er is onderzocht hoe er vanaf de 18e tot en met de 20e eeuw over deze bende is geschreven en in welk opzicht dit is veranderd.
- Published
- 2014
6. Endocrine effects of hexabromocyclododecane (HBCD) in a one-generation reproduction study in Wistar rats.
- Author
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Risk Assessment of Toxic and Immunomodulatory Agents, Dep IRAS, van der Ven, L.T., van de Kuil, T., Leonards, P.E., Slob, W., Lilienthal, H., Litens, S., Herlin, M., Hakansson, H., Canton, R.F., van den Berg, M., Visser, T.J., van Loveren, H., Vos, J.G., Piersma, A.H., Risk Assessment of Toxic and Immunomodulatory Agents, Dep IRAS, van der Ven, L.T., van de Kuil, T., Leonards, P.E., Slob, W., Lilienthal, H., Litens, S., Herlin, M., Hakansson, H., Canton, R.F., van den Berg, M., Visser, T.J., van Loveren, H., Vos, J.G., and Piersma, A.H.
- Published
- 2009
7. A 28-day oral dose toxicity study enhanced to detect endocrine effects of hexabromocyclododecane in wistar rats
- Author
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van der Ven, L.T.M., Verhoef, A., van de Kuil, T., Slob, W., Leonards, P.E.G., Visser, T.J., Hamers, T., Herlin, M., Hakansson, H., Olausson, H., Piersma, A.H., Vos, J.G., van der Ven, L.T.M., Verhoef, A., van de Kuil, T., Slob, W., Leonards, P.E.G., Visser, T.J., Hamers, T., Herlin, M., Hakansson, H., Olausson, H., Piersma, A.H., and Vos, J.G.
- Abstract
A 28-day repeated dose study in rats (OECD407) enhanced for endocrine and immune parameters was performed with hexabromocyclododecane (HBCD). Rats were exposed by daily gavage to HBCD dissolved in corn oil in 8 dose groups with doses ranging between 0 and 200 mg/kg bw per day (mkd). Evaluation consisted of dose-response analysis with calculation of a benchmark dose at the lower 95% one-sided confidence bound (BMDL) at predefined critical effect sizes (CESs) of 10-20%. The most remarkable findings were dose-related effects on the thyroid hormone axis, that is, decreased total thyroxin (TT4, BMDL 55.5 mkd at CES - 10%), increased pituitary weight (29 mkd at 10%) and increased immunostaining of TSH in the pituitary, increased thyroid weight (1.6 mkd at 10%), and thyroid follicle cell activation. These effects were restricted to females. Female rats also showed increased absolute liver weights (22.9 mkd at 20%) and induction of T4-glucuronyl transferase (4.1 mkd at 10%), suggesting that aberrant metabolization of T4 triggers feedback activation of the thyroid hormone system. These effects were accompanied by possibly secondary effects, including increased cholesterol (7.4 mkd at 10%), increased tibial bone mineral density (> 49 mkd at 10%), both in females, and decreased splenocyte counts (0.3-6.3 mkd at 20%; only evaluated in males). Overall, female rats appeared to be more sensitive to HBCD than male rats, and an overall BMDL is proposed at 1.6 mkd, based on a 10% increase of the thyroid weight, which was the most sensitive parameter in the sequence of events. © 2006 Oxford University Press.
- Published
- 2006
- Full Text
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8. Diet during pregnancy and weaning and adult disease in the offspring in the rat
- Author
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LEO, LPI, Siemelink M, Dormans JAMA, Loveren H van, Klerk A de, Vleeming W, Schenk E, Kuil T van de, Biesebeek JD te, Verharen H, Verhoef A, Piersma AH, Opperhuizen A, LEO, LPI, Siemelink M, Dormans JAMA, Loveren H van, Klerk A de, Vleeming W, Schenk E, Kuil T van de, Biesebeek JD te, Verharen H, Verhoef A, Piersma AH, and Opperhuizen A
- Abstract
RIVM rapport:Several epidemiological as well as experimental studies suggest that the composition of the maternal diet during pregnancy may have a programming effect on the fetus leading to epigenetic predisposition of chronic diseases in adulthood. Here, effects of various diets given during pregnancy and weaning on parameters related to chronic diseases during adulthood were studied in the rat. Seven groups of sixteen animals were put on an isocaloric semi-synthetic high-fat diet two weeks before pregnancy, and continuing throughout pregnancy and weaning. Diets differed in macronutrient - or fatty acid composition. Fetal development was studied on gestational day 21. After birth, litter size in parallel groups was reduced to eight pups. A series of physiological parameters were studied in the pups at age 12 weeks. The offspring of dams put on a low protein diet or a fish oil diet in utero and during weaning had a significantly reduced body weight gain from birth up to age 12 weeks in comparison to the control group. Male offspring aged 12 weeks fed the coconut oil diet in utero and during weaning appeared slightly insulin-resistant. No difference in the volume of the islets of Langerhans could be demonstrated in this group. Despite significantly smaller islets of Langerhans and a reduced number of large islets in the pancreas of 12-week-old male rats given the low protein diet in utero and during weaning, no differences in glucose or insulin concentration were found in comparison to the control group. Maternal diet during pregnancy and weaning did not influence immune function or blood pressure in the offspring at the age of 12 weeks. In conclusion, prenatal diet influences postnatal development, as determined by anatomical and physiological parameters., Uit zowel epidemiologische als dierexperimentele studies komen steeds meer aanwijzingen dat de samenstelling van de voeding tijdens de zwangerschap een belangrijke invloed heeft op de vatbaarheid van het nageslacht voor chronische ziekten op latere leeftijd. In deze dierexperimentele studie zijn de effecten van diervoeders met een verschillende samenstelling onderzocht op parameters voor chronische ziekten op volwassen leeftijd. Aan zeven groepen van elk zestien dieren werd een isocalorisch hoog-vet voer gegeven met een verschillende macronutrient- en vetzuursamenstelling vanaf twee weken voor de dracht tot aan het einde van de speenperiode. Op dag 21 van de dracht werd de foetale ontwikkeling onderzocht. Op de leeftijd van 12 weken is een aantal fysiologische en morfologische parameters bepaald. Nageslacht dat in utero en tijdens de speenperiode was blootgesteld aan een laag eiwit voer of een voer rijk in visvetzuren had een significant lager lichaamsgewicht vanaf de geboorte tot aan de leeftijd van 12 weken, ten opzichte van de controlegroep. Het mannelijk nageslacht van dieren van de cocosnootgroep (rijk aan verzadigde vetten) lijkt licht insuline resistent te zijn. Er was echter geen verschil in volume van de eilandjes van Langerhans ten opzichte van de controlegroep. Ondanks significant kleinere eilandjes van Langerhans en de aanwezigheid van minder grotere eilandjes in de pancreas bij 12-weken oude dieren uit de laag eiwit groep verschilde deze groep niet van de controlegroep in glucose- of insuline respons na een orale glucose load. De immuunfunctie en systolische en diastolische bloeddruk in het nageslacht op volwassen leeftijd leek niet te worden beinvloed door de samenstelling van de maternale voeding in utero en tijdens spenen Concluderend kan worden gezegd dat de samenstelling van de voeding tijdens de dracht en de vroege postnatale periode de postnatale ontwikkeling van het nageslacht beinvloedde, hetgeen bleek uit veranderingen in zowel morfologische a
- Published
- 2002
9. Diet during pregnancy and weaning and adult disease inthe offspring in the rat
- Author
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LEO, LPI, Siemelink, M, Dormans, JAMA, van Loveren, H, de Klerk, A, Vleeming, W, Schenk, E, van de Kuil, T, te Biesebeek, JD, Verharen, H, Verhoef, A, Piersma, AH, Opperhuizen, A, LEO, LPI, Siemelink, M, Dormans, JAMA, van Loveren, H, de Klerk, A, Vleeming, W, Schenk, E, van de Kuil, T, te Biesebeek, JD, Verharen, H, Verhoef, A, Piersma, AH, and Opperhuizen, A
- Abstract
RIVM rapport:Several epidemiological as well as experimental studies suggest that the composition of the maternal diet during pregnancy may have a programming effect on the fetus leading to epigenetic predisposition of chronic diseases in adulthood. Here, effects of various diets given during pregnancy and weaning on parameters related to chronic diseases during adulthood were studied in the rat. Seven groups of sixteen animals were put on an isocaloric semi-synthetic high-fat diet two weeks before pregnancy, and continuing throughout pregnancy and weaning. Diets differed in macronutrient - or fatty acid composition. Fetal development was studied on gestational day 21. After birth, litter size in parallel groups was reduced to eight pups. A series of physiological parameters were studied in the pups at age 12 weeks. The offspring of dams put on a low protein diet or a fish oil diet in utero and during weaning had a significantly reduced body weight gain from birth up to age 12 weeks in comparison to the control group. Male offspring aged 12 weeks fed the coconut oil diet in utero and during weaning appeared slightly insulin-resistant. No difference in the volume of the islets of Langerhans could be demonstrated in this group. Despite significantly smaller islets of Langerhans and a reduced number of large islets in the pancreas of 12-week-old male rats given the low protein diet in utero and during weaning, no differences in glucose or insulin concentration were found in comparison to the control group. Maternal diet during pregnancy and weaning did not influence immune function or blood pressure in the offspring at the age of 12 weeks. In conclusion, prenatal diet influences postnatal development, as determined by anatomical and physiological parameters., Uit zowel epidemiologische als dierexperimentele studies komen steeds meer aanwijzingen dat de samenstelling van de voeding tijdens de zwangerschap een belangrijke invloed heeft op de vatbaarheid van het nageslacht voor chronische ziekten op latere leeftijd. In deze dierexperimentele studie zijn de effecten van diervoeders met een verschillende samenstelling onderzocht op parameters voor chronische ziekten op volwassen leeftijd. Aan zeven groepen van elk zestien dieren werd een isocalorisch hoog-vet voer gegeven met een verschillende macronutrient- en vetzuursamenstelling vanaf twee weken voor de dracht tot aan het einde van de speenperiode. Op dag 21 van de dracht werd de foetale ontwikkeling onderzocht. Op de leeftijd van 12 weken is een aantal fysiologische en morfologische parameters bepaald. Nageslacht dat in utero en tijdens de speenperiode was blootgesteld aan een laag eiwit voer of een voer rijk in visvetzuren had een significant lager lichaamsgewicht vanaf de geboorte tot aan de leeftijd van 12 weken, ten opzichte van de controlegroep. Het mannelijk nageslacht van dieren van de cocosnootgroep (rijk aan verzadigde vetten) lijkt licht insuline resistent te zijn. Er was echter geen verschil in volume van de eilandjes van Langerhans ten opzichte van de controlegroep. Ondanks significant kleinere eilandjes van Langerhans en de aanwezigheid van minder grotere eilandjes in de pancreas bij 12-weken oude dieren uit de laag eiwit groep verschilde deze groep niet van de controlegroep in glucose- of insuline respons na een orale glucose load. De immuunfunctie en systolische en diastolische bloeddruk in het nageslacht op volwassen leeftijd leek niet te worden beinvloed door de samenstelling van de maternale voeding in utero en tijdens spenen Concluderend kan worden gezegd dat de samenstelling van de voeding tijdens de dracht en de vroege postnatale periode de postnatale ontwikkeling van het nageslacht beinvloedde, hetgeen bleek uit veranderingen in zowel morfologische a
- Published
- 2002
10. Vascular responses of isolated mesenteric resistance and basilar arteries from short- and long-term diabetic rats
- Author
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Gispen, W.H., Buren, T. van, Vleeming, W., Krutzen, M.M., Kuil, T. van de, Wildt, D.J. de, Gispen, W.H., Buren, T. van, Vleeming, W., Krutzen, M.M., Kuil, T. van de, and Wildt, D.J. de
- Published
- 1998
11. Increased frequencies of diploid sperm detected by multicolour FISH after treatment of rats with carbendazim without micronucleus induction in peripheral blood erythrocytes
- Author
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de Stoppelaar, J. M., primary, van de Kuil, T., additional, Bedaf, M., additional, Verharen, H. W., additional, Slob, W., additional, Mohn, G. R., additional, Hoebee, B., additional, and van Benthem, J., additional
- Published
- 1999
- Full Text
- View/download PDF
12. The 6-phosphofructokinase reaction in Acetivibrio thermocellus is both ATP- and pyrophosphate-dependent.
- Author
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Koendjbiharie JG, Kuil T, Nurminen CMK, and van Maris AJA
- Abstract
Acetivibrio thermocellus (formerly Clostridium thermocellum) is a potential platform for lignocellulosic ethanol production. Its industrial application is hampered by low product titres, resulting from a low thermodynamic driving force of its central metabolism. It possesses both a functional ATP- and a functional PP
i -dependent 6-phosphofructokinase (PPi -Pfk), of which only the latter is held responsible for the low driving force. Here we show that, following the replacement of PPi -Pfk by cytosolic pyrophosphatase and transaldolase, the native ATP-Pfk is able to carry the full glycolytic flux. Interestingly, the barely-detectable in vitro ATP-Pfk activities are only a fraction of what would be required, indicating its contribution to glycolysis has consistently been underestimated. A kinetic model demonstrated that the strong inhibition of ATP-Pfk by PPi can prevent futile cycling that would arise when both enzymes are active simultaneously. As such, there seems to be no need for a long-sought-after PPi -generating mechanism to drive glycolysis, as PPi -Pfk can simply use whatever PPi is available, and ATP-Pfk complements the rest of the PFK-flux. Laboratory evolution of the ΔPPi -Pfk strain, unable to valorize PPi , resulted in a mutation in the GreA transcription elongation factor. This mutation likely results in reduced RNA-turnover, hinting at transcription as a significant (and underestimated) source of anabolic PPi . Together with other mutations, this resulted in an A. thermocellus strain with the hitherto highest biomass-specific cellobiose uptake rate of 2.2 g/gx /h. These findings are both relevant for fundamental insight into dual ATP/PPi Pfk-nodes, which are not uncommon in other microorganisms, as well as for further engineering of A. thermocellus for consolidated bioprocessing., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)- Published
- 2024
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13. Arabinoxylan source and xylanase specificity influence the production of oligosaccharides with prebiotic potential.
- Author
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Rudjito RC, Jiménez-Quero A, Muñoz MDCC, Kuil T, Olsson L, Stringer MA, Krogh KBRM, Eklöf J, and Vilaplana F
- Abstract
Cereal arabinoxylans (AXs) are complex polysaccharides in terms of their pattern of arabinose and ferulic acid substitutions, which influence their properties in structural and nutritional applications. We have evaluated the influence of the molecular structure of three AXs from wheat and rye with distinct substitutions on the activity of β-xylanases from different glycosyl hydrolase families (GH 5_34, 8, 10 and 11). The arabinose and ferulic acid substitutions influence the accessibility of the xylanases, resulting in specific profiles of arabinoxylan-oligosaccharides (AXOS). The GH10 xylanase from Aspergillus aculeatus (AcXyn10A) and GH11 from Thermomyces lanuginosus (TlXyn11) showed the highest activity, producing larger amounts of small oligosaccharides in shorter time. The GH8 xylanase from Bacillus sp. (BXyn8) produced linear xylooligosaccharides and was most restricted by arabinose substitution, whereas GH5_34 from Gonapodya prolifera (GpXyn5_34) required arabinose substitution and produced longer (A)XOS substituted on the reducing end. The complementary substrate specificity of BXyn8 and GpXyn5_34 revealed how arabinoses were distributed along the xylan backbones. This study demonstrates that AX source and xylanase specificity influence the production of oligosaccharides with specific structures, which in turn impacts the growth of specific bacteria (Bacteroides ovatus and Bifidobacterium adolescentis) and the production of beneficial metabolites (short-chain fatty acids)., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Francisco Vilaplana reports financial support was provided by Swedish Research Council Formas. Francisco Vilaplana reports financial support was provided by Lantmännen Research Foundation. Francisco Vilaplana reports a relationship with Oatly AB that includes: employment. Mary Ann STRINGER reports a relationship with Novozymes Inc. that includes: employment. Kristian Bertel Romer Morkeberg KROGH reports a relationship with Novozymes Inc. that includes: employment. Jens EKLOF reports a relationship with Novozymes Inc. that includes: employment. None., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)
- Published
- 2023
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14. Pyrophosphate as allosteric regulator of ATP-phosphofructokinase in Clostridium thermocellum and other bacteria with ATP- and PP i -phosphofructokinases.
- Author
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Kuil T, Nurminen CMK, and van Maris AJA
- Subjects
- Diphosphates, Amino Acid Sequence, Phosphofructokinase-1 genetics, Phosphofructokinase-1 metabolism, Bacteria metabolism, Adenosine Triphosphate, Guanosine Triphosphate, Kinetics, Phosphofructokinases metabolism, Clostridium thermocellum metabolism
- Abstract
The phosphofructokinase (Pfk) reaction represents one of the key regulatory points in glycolysis. While most organisms encode for Pfks that use ATP as phosphoryl donor, some organisms also encode for PP
i -dependent Pfks. Despite this central role, the biochemical characteristics as well as the physiological role of both Pfks is often not known. Clostridium thermocellum is an example of a microorganism that encodes for both Pfks, however, only PPi -Pfk activity has been detected in cell-free extracts and little is known about the regulation and function of both enzymes. In this study, the ATP- and PPi -Pfk of C. thermocellum were purified and biochemically characterized. No allosteric regulators were found for PPi -Pfk amongst common effectors. With fructose-6-P, PPi , fructose-1,6-bisP, and Pi PPi -Pfk showed high specificity (KM < 0.62 mM) and maximum activity (Vmax > 156 U mg-1 ). In contrast, ATP-Pfk showed much lower affinity (K0.5 of 9.26 mM) and maximum activity (14.5 U mg-1 ) with fructose-6-P. In addition to ATP, also GTP, UTP and ITP could be used as phosphoryl donors. The catalytic efficiency with GTP was 7-fold higher than with ATP, suggesting that GTP is the preferred substrate. The enzyme was activated by NH4 + , and pronounced inhibition was observed with GDP, FBP, PEP, and especially with PPi (Ki of 0.007 mM). Characterization of purified ATP-Pfks originating from eleven different bacteria, encoding for only ATP-Pfk or for both ATP- and PPi -Pfk, identified that PPi inhibition of ATP-Pfks could be a common phenomenon for organisms with a PPi -dependent glycolysis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)- Published
- 2023
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15. A detailed genome-scale metabolic model of Clostridium thermocellum investigates sources of pyrophosphate for driving glycolysis.
- Author
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Schroeder WL, Kuil T, van Maris AJA, Olson DG, Lynd LR, and Maranas CD
- Subjects
- Diphosphates metabolism, Glycolysis genetics, Fermentation, Adenosine Triphosphate metabolism, Clostridium thermocellum genetics, Clostridium thermocellum metabolism
- Abstract
Lignocellulosic biomass is an abundant and renewable source of carbon for chemical manufacturing, yet it is cumbersome in conventional processes. A promising, and increasingly studied, candidate for lignocellulose bioprocessing is the thermophilic anaerobe Clostridium thermocellum given its potential to produce ethanol, organic acids, and hydrogen gas from lignocellulosic biomass under high substrate loading. Possessing an atypical glycolytic pathway which substitutes GTP or pyrophosphate (PP
i ) for ATP in some steps, including in the energy-investment phase, identification, and manipulation of PPi sources are key to engineering its metabolism. Previous efforts to identify the primary pyrophosphate have been unsuccessful. Here, we explore pyrophosphate metabolism through reconstructing, updating, and analyzing a new genome-scale stoichiometric model for C. thermocellum, iCTH669. Hundreds of changes to the former GEM, iCBI655, including correcting cofactor usages, addressing charge and elemental balance, standardizing biomass composition, and incorporating the latest experimental evidence led to a MEMOTE score improvement to 94%. We found agreement of iCTH669 model predictions across all available fermentation and biomass yield datasets. The feasibility of hundreds of PPi synthesis routes, newly identified and previously proposed, were assessed through the lens of the iCTH669 model including biomass synthesis, tRNA synthesis, newly identified sources, and previously proposed PPi -generating cycles. In all cases, the metabolic cost of PPi synthesis is at best equivalent to investment of one ATP suggesting no direct energetic advantage for the cofactor substitution in C. thermocellum. Even though no unique source of PPi could be gleaned by the model, by combining with gene expression data two most likely scenarios emerge. First, previously investigated PPi sources likely account for most PPi production in wild-type strains. Second, alternate metabolic routes as encoded by iCTH669 can collectively maintain PPi levels even when previously investigated synthesis cycles are disrupted. Model iCTH669 is available at github.com/maranasgroup/iCTH669., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)- Published
- 2023
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16. The Roles of Nicotinamide Adenine Dinucleotide Phosphate Reoxidation and Ammonium Assimilation in the Secretion of Amino Acids as Byproducts of Clostridium thermocellum.
- Author
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Yayo J, Rydzak T, Kuil T, Karlsson A, Harding DJ, Guss AM, and van Maris AJA
- Subjects
- Ethanol metabolism, Ferredoxins metabolism, Malates metabolism, NAD metabolism, Pyruvates metabolism, Oxidation-Reduction, Amino Acids biosynthesis, Amino Acids metabolism, Ammonium Compounds metabolism, Clostridium thermocellum genetics, Clostridium thermocellum metabolism, NADP metabolism
- Abstract
Clostridium thermocellum is a cellulolytic thermophile that is considered for the consolidated bioprocessing of lignocellulose to ethanol. Improvements in ethanol yield are required for industrial implementation, but the incompletely understood causes of amino acid secretion impede progress. In this study, amino acid secretion was investigated via gene deletions in ammonium-regulated, nicotinamide adenine dinucleotide phosphate (NADPH)-supplying and NADPH-consuming pathways as well as via physiological characterization in cellobiose-limited or ammonium-limited chemostats. First, the contribution of the NADPH-supplying malate shunt was studied with strains using either the NADPH-yielding malate shunt (Δ ppdk ) or a redox-independent conversion of PEP to pyruvate (Δ ppdk Δ malE::P
eno -pyk ). In the latter, branched-chain amino acids, especially valine, were significantly reduced, whereas the ethanol yield increased from 46 to 60%, suggesting that the secretion of these amino acids balances the NADPH surplus from the malate shunt. The unchanged amino acid secretion in Δ ppdk falsified a previous hypothesis on an ammonium-regulated PEP-to-pyruvate flux redistribution. The possible involvement of another NADPH-supplier, namely, NADH-dependent reduced ferredoxin:NADP+ oxidoreductase ( nfnAB ), was also excluded. Finally, the deletion of glutamate synthase ( gogat ) in ammonium assimilation resulted in the upregulation of NADPH-linked glutamate dehydrogenase activity and decreased amino acid yields. Since gogat in C. thermocellum is putatively annotated as ferredoxin-linked, a claim which is supported by the product redistribution observed in this study, this deletion likely replaced ferredoxin with NADPH in ammonium assimilation. Overall, these findings indicate that a need to reoxidize NADPH is driving the observed amino acid secretion, likely at the expense of the NADH needed for ethanol formation. This suggests that metabolic engineering strategies that simplify the redox metabolism and ammonium assimilation can contribute to increased ethanol yields. IMPORTANCE Improving the ethanol yield of C. thermocellum is important for the industrial implementation of this microorganism in consolidated bioprocessing. A central role of NADPH in driving amino acid byproduct formation was demonstrated by eliminating the NADPH-supplying malate shunt and separately by changing the cofactor specificity in ammonium assimilation. With amino acid secretion diverting carbon and electrons away from ethanol, these insights are important for further metabolic engineering to reach industrial requirements on ethanol yield. This study also provides chemostat data that are relevant for training genome-scale metabolic models and for improving the validity of their predictions, especially considering the reduced degree-of-freedom in the redox metabolism of the strains generated here. In addition, this study advances the fundamental understanding on the mechanisms underlying amino acid secretion in cellulolytic Clostridia as well as on the regulation and cofactor specificity in ammonium assimilation. Together, these efforts aid in the development of C. thermocellum for the sustainable consolidated bioprocessing of lignocellulose to ethanol with minimal pretreatment.- Published
- 2023
- Full Text
- View/download PDF
17. Ethanol tolerance of Clostridium thermocellum: the role of chaotropicity, temperature and pathway thermodynamics on growth and fermentative capacity.
- Author
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Kuil T, Yayo J, Pechan J, Küchler J, and van Maris AJA
- Subjects
- Fermentation, Temperature, Glycolysis, Thermodynamics, Clostridium thermocellum genetics, Clostridium thermocellum metabolism
- Abstract
Background: Clostridium thermocellum is a promising candidate for consolidated bioprocessing of lignocellulosic biomass to ethanol. The low ethanol tolerance of this microorganism is one of the remaining obstacles to industrial implementation. Ethanol inhibition can be caused by end-product inhibition and/or chaotropic-induced stress resulting in increased membrane fluidization and disruption of macromolecules. The highly reversible glycolysis of C. thermocellum might be especially sensitive to end-product inhibition. The chaotropic effect of ethanol is known to increase with temperature. This study explores the relative contributions of these two aspects to investigate and possibly mitigate ethanol-induced stress in growing and non-growing C. thermocellum cultures., Results: To separate chaotropic from thermodynamic effects of ethanol toxicity, a non-ethanol producing strain AVM062 (P
clo1313_2638 ::ldh* ∆adhE) was constructed by deleting the bifunctional acetaldehyde/alcohol dehydrogenase gene, adhE, in a lactate-overproducing strain. Exogenously added ethanol lowered the growth rate of both wild-type and the non-ethanol producing mutant. The mutant strain grew quicker than the wild-type at 50 and 55 °C for ethanol concentrations ≥ 10 g L-1 and was able to reach higher maximum OD600 at all ethanol concentrations and temperatures. For the wild-type, the maximum OD600 and relative growth rates were higher at 45 and 50 °C, compared to 55 °C, for ethanol concentrations ≥ 15 g L-1 . For the mutant strain, no positive effect on growth was observed at lower temperatures. Growth-arrested cells of the wild-type demonstrated improved fermentative capacity over time in the presence of ethanol concentrations up to 40 g L-1 at 45 and 50 °C compared to 55 °C., Conclusion: Positive effects of temperature on ethanol tolerance were limited to wild-type C. thermocellum and are likely related to mechanisms involved in the ethanol-formation pathway and redox cofactor balancing. Lowering the cultivation temperature provides an attractive strategy to improve growth and fermentative capacity at high ethanol titres in high-cellulose loading batch cultivations. Finally, non-ethanol producing strains are useful platform strains to study the effects of chaotropicity and thermodynamics related to ethanol toxicity and allow for deeper understanding of growth and/or fermentation cessation under industrially relevant conditions., (© 2022. The Author(s).)- Published
- 2022
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18. Functional Analysis of H + -Pumping Membrane-Bound Pyrophosphatase, ADP-Glucose Synthase, and Pyruvate Phosphate Dikinase as Pyrophosphate Sources in Clostridium thermocellum.
- Author
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Kuil T, Hon S, Yayo J, Foster C, Ravagnan G, Maranas CD, Lynd LR, Olson DG, and van Maris AJA
- Subjects
- Diphosphates metabolism, Glucose-1-Phosphate Adenylyltransferase metabolism, Inorganic Pyrophosphatase metabolism, Phosphates metabolism, Pyruvate, Orthophosphate Dikinase genetics, Pyruvate, Orthophosphate Dikinase metabolism, Pyruvic Acid metabolism, Clostridium thermocellum metabolism
- Abstract
The atypical glycolysis of Clostridium thermocellum is characterized by the use of pyrophosphate (PP
i ) as a phosphoryl donor for phosphofructokinase (Pfk) and pyruvate phosphate dikinase (Ppdk) reactions. Previously, biosynthetic PPi was calculated to be stoichiometrically insufficient to drive glycolysis. This study investigates the role of a H+ -pumping membrane-bound pyrophosphatase, glycogen cycling, a predicted Ppdk-malate shunt cycle, and acetate cycling in generating PPi . Knockout studies and enzyme assays confirmed that clo1313_0823 encodes a membrane-bound pyrophosphatase. Additionally, clo1313_0717-0718 was confirmed to encode ADP-glucose synthase by knockouts, glycogen measurements in C. thermocellum , and heterologous expression in Escherichia coli. Unexpectedly, individually targeted gene deletions of the four putative PPi sources did not have a significant phenotypic effect. Although combinatorial deletion of all four putative PPi sources reduced the growth rate by 22% (0.30 ± 0.01 h-1 ) and the biomass yield by 38% (0.18 ± 0.00 gbiomass gsubstrate -1 ), this change was much smaller than what would be expected for stoichiometrically essential PPi -supplying mechanisms. Growth-arrested cells of the quadruple knockout readily fermented cellobiose, indicating that the unknown PPi -supplying mechanisms are independent of biosynthesis. An alternative hypothesis that ATP-dependent Pfk activity circumvents a need for PPi altogether was falsified by enzyme assays, heterologous expression of candidate genes, and whole-genome sequencing. As a secondary outcome, enzymatic assays confirmed functional annotation of clo1313_1832 as ATP- and GTP-dependent fructokinase. These results indicate that the four investigated PPi sources individually and combined play no significant PPi -supplying role, and the true source(s) of PPi , or alternative phosphorylating mechanisms, that drive(s) glycolysis in C. thermocellum remain(s) elusive. IMPORTANCE Increased understanding of the central metabolism of C. thermocellum is important from a fundamental as well as from a sustainability and industrial perspective. In addition to showing that H+ -pumping membrane-bound PPase, glycogen cycling, a Ppdk-malate shunt cycle, and acetate cycling are not significant sources of PPi supply, this study adds functional annotation of four genes and availability of an updated PPi stoichiometry from biosynthesis to the scientific domain. Together, this aids future metabolic engineering attempts aimed to improve C. thermocellum as a cell factory for sustainable and efficient production of ethanol from lignocellulosic material through consolidated bioprocessing with minimal pretreatment. Getting closer to elucidating the elusive source of PPi , or alternative phosphorylating mechanisms, for the atypical glycolysis is itself of fundamental importance. Additionally, the findings of this study directly contribute to investigations into trade-offs between thermodynamic driving force versus energy yield of PPi - and ATP-dependent glycolysis.- Published
- 2022
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19. Laboratory Evolution and Reverse Engineering of Clostridium thermocellum for Growth on Glucose and Fructose.
- Author
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Yayo J, Kuil T, Olson DG, Lynd LR, Holwerda EK, and van Maris AJA
- Subjects
- Clostridium thermocellum genetics, Clostridium thermocellum growth & development, Genome, Bacterial, Laboratories, Metabolic Engineering, Mutation, Whole Genome Sequencing, Clostridium thermocellum metabolism, Fructose metabolism, Glucose metabolism
- Abstract
The native ability of Clostridium thermocellum to efficiently solubilize cellulose makes it an interesting platform for sustainable biofuel production through consolidated bioprocessing. Together with other improvements, industrial implementation of C. thermocellum , as well as fundamental studies into its metabolism, would benefit from improved and reproducible consumption of hexose sugars. To investigate growth of C. thermocellum on glucose or fructose, as well as the underlying molecular mechanisms, laboratory evolution was performed in carbon-limited chemostats with increasing concentrations of glucose or fructose and decreasing cellobiose concentrations. Growth on both glucose and fructose was achieved with biomass yields of 0.09 ± 0.00 and 0.18 ± 0.00 g
biomass gsubstrate -1 , respectively, compared to 0.15 ± 0.01 gbiomass gsubstrate -1 for wild type on cellobiose. Single-colony isolates had no or short lag times on the monosaccharides, while wild type showed 42 ± 4 h on glucose and >80 h on fructose. With good growth on glucose, fructose, and cellobiose, the fructose isolates were chosen for genome sequence-based reverse metabolic engineering. Deletion of a putative transcriptional regulator (Clo1313_1831), which upregulated fructokinase activity, reduced lag time on fructose to 12 h with a growth rate of 0.11 ± 0.01 h-1 and resulted in immediate growth on glucose at 0.24 ± 0.01 h-1 Additional introduction of a G-to-V mutation at position 148 in cbpA resulted in immediate growth on fructose at 0.32 ± 0.03 h-1 These insights can guide engineering of strains for fundamental studies into transport and the upper glycolysis, as well as maximizing product yields in industrial settings. IMPORTANCE C. thermocellum is an important candidate for sustainable and cost-effective production of bioethanol through consolidated bioprocessing. In addition to unsurpassed cellulose deconstruction, industrial application and fundamental studies would benefit from improvement of glucose and fructose consumption. This study demonstrated that C. thermocellum can be evolved for reproducible constitutive growth on glucose or fructose. Subsequent genome sequencing, gene editing, and physiological characterization identified two underlying mutations with a role in (regulation of) transport or metabolism of the hexose sugars. In light of these findings, such mutations have likely (and unknowingly) also occurred in previous studies with C. thermocellum using hexose-based media with possible broad regulatory consequences. By targeted modification of these genes, industrial and research strains of C. thermocellum can be engineered to (i) reduce glucose accumulation, (ii) study cellodextrin transport systems in vivo , (iii) allow experiments at >120 g liter-1 soluble substrate concentration, or (iv) reduce costs for labeling studies., (Copyright © 2021 Yayo et al.)- Published
- 2021
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20. Endocrine effects of hexabromocyclododecane (HBCD) in a one-generation reproduction study in Wistar rats.
- Author
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van der Ven LT, van de Kuil T, Leonards PE, Slob W, Lilienthal H, Litens S, Herlin M, Håkansson H, Cantón RF, van den Berg M, Visser TJ, van Loveren H, Vos JG, and Piersma AH
- Subjects
- Animals, Body Burden, Body Weight drug effects, Dose-Response Relationship, Drug, Eating drug effects, Female, Gonadal Steroid Hormones physiology, Immune System drug effects, Liver drug effects, Liver metabolism, Male, Organ Size drug effects, Pregnancy, Rats, Retinoids metabolism, Spermatozoa drug effects, Endocrine Disruptors toxicity, Fetus drug effects, Flame Retardants toxicity, Hydrocarbons, Brominated toxicity, Reproduction drug effects
- Abstract
The brominated flame retardant (BFR) hexabromocyclododecane was tested in a one-generation reproduction assay in Wistar rats, enhanced for endocrine parameters. A solution of the compound in corn oil was mixed in the feed, targeting at dietary exposure of 0-0.1-0.3-1-3-10-30-100 mg/kg body weight/day (mkd) in parental rats during 10 (males) or 2 (females) weeks premating, during gestation and lactation, and in their F1 offspring from weaning until final necropsy. Effects were assessed in F1 animals. Livers of these animals showed increased HBCD concentrations, in a dose-dependent way. The trabecular bone mineral density of the tibia was dose-dependently decreased in females (BenchMark Dose Lower confidence bound, BMDL=0.056 mkd). The IgG response after immunization with sheep red blood cells (SRBC) was increased in males (BMDL=0.46 mkd). Further sensitive effects were decreased weight of the testis (BMDL=1.5 mkd), increased fraction of neutrophilic granulocytes (BMDL=7.7 mkd), decreased concentration of apolar retinoids in female livers (BMDL=1.3 mkd), and decreased plasma alkaline phosphatase in females (BMDL=8.6 mkd). CYP19/aromatase activity in the ovary was correlated to the concentration of gamma-HBCD in the liver. A developmental origin of these effects is considered, and this is also true for sensitive effects observed in neurobehavioural testing in littermates from the same experiment, i.e. in the brainstem auditory evoked potentials and in a catalepsy test [Lilienthal, H., Van der Ven, L.T.M., Piersma, A.H., Vos, J.G. Neurobehavioral effects of the brominated flame retardant hexabromocyclododecane (HBCD) in rats after pre- and postnatal exposure, in press]. The low BMDLs of these effects may raise concern for human health, particularly when based on body burdens of HBCD, which leads to critical margins of exposure particularly for the occupational setting.
- Published
- 2009
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21. A 28-day oral dose toxicity study in Wistar rats enhanced to detect endocrine effects of decabromodiphenyl ether (decaBDE).
- Author
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Van der Ven LT, van de Kuil T, Leonards PE, Slob W, Cantón RF, Germer S, Visser TJ, Litens S, Håkansson H, Schrenk D, van den Berg M, Piersma AH, Vos JG, and Opperhuizen A
- Subjects
- Administration, Oral, Animals, Bone Density drug effects, Bone and Bones diagnostic imaging, Bone and Bones drug effects, Bone and Bones metabolism, Cytochrome P-450 Enzyme System metabolism, Dose-Response Relationship, Drug, Endocrine Glands metabolism, Endocrine Glands pathology, Female, Flame Retardants pharmacokinetics, Halogenated Diphenyl Ethers, Immune System drug effects, Immune System pathology, Liver drug effects, Liver enzymology, Male, Organ Size drug effects, Phenyl Ethers pharmacokinetics, Polybrominated Biphenyls pharmacokinetics, Prostate drug effects, Prostate pathology, Radiography, Rats, Rats, Wistar, Reproduction drug effects, Risk Assessment, Seminal Vesicles drug effects, Seminal Vesicles pathology, Toxicity Tests, Endocrine Glands drug effects, Flame Retardants toxicity, Phenyl Ethers toxicity, Polybrominated Biphenyls toxicity
- Abstract
Decabromodiphenyl ether (decaBDE) is a widely used brominated flame retardant, considered to be of low toxicity. However, previous toxicity studies applied exposure methods with low bioavailability of this compound, and the actual hazard of decaBDE for humans, which are environmentally exposed to decaBDE, may thus be underestimated in current risk assessments. The present 28 days oral toxicity study in Wistar rats was designed to facilitate detection of endocrine and immune modulating effects of decaBDE using an exposure protocol with improved bioavailability. A technical preparation of high purity decaBDE was thus tested by daily exposure through gavage with an emulsion of soy phospholipon/lutrol as a carrier. Most sensitive effect in males were increased weight of seminal vesicle/coagulation gland with BMDL of 0.2mg/kg bw/day and increased expression of hepatic CYP1A and CYP2B (BMDLs 0.5-0.7 mg/kg bw/day). In females the most sensitive effect was decreased activity of P450c17 (CYP17), which is a key enzyme in the androgen synthesis pathway, in adrenals (BMDL 0.18 mg/kg bw/day). These results suggest that decaBDE may represent an as yet unreported hazard for reproductive health.
- Published
- 2008
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22. A 28-day oral dose toxicity study enhanced to detect endocrine effects of a purified technical pentabromodiphenyl ether (pentaBDE) mixture in Wistar rats.
- Author
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van der Ven LT, van de Kuil T, Verhoef A, Leonards PE, Slob W, Cantón RF, Germer S, Hamers T, Visser TJ, Litens S, Håkansson H, Fery Y, Schrenk D, van den Berg M, Piersma AH, and Vos JG
- Subjects
- Animals, Body Weight drug effects, Dose-Response Relationship, Drug, Endocrine Disruptors chemistry, Endocrine Disruptors pharmacokinetics, Female, Flame Retardants pharmacokinetics, Halogenated Diphenyl Ethers, Liver drug effects, Liver metabolism, Liver pathology, Male, Organ Size drug effects, Phenyl Ethers chemistry, Phenyl Ethers pharmacokinetics, Polybrominated Biphenyls chemistry, Polybrominated Biphenyls pharmacokinetics, Rats, Rats, Wistar, Spermatozoa drug effects, Spermatozoa pathology, Thyroid Hormones blood, Toxicity Tests, Chronic methods, Endocrine Disruptors toxicity, Flame Retardants toxicity, Phenyl Ethers toxicity, Polybrominated Biphenyls toxicity
- Abstract
A 28-day subacute oral toxicity study was performed in Wistar rats with a purified preparation of the commercial pentabromodiphenyl ether (pentaBDE), DE-71. The applied OECD407 protocol was enhanced for endocrine and immune parameters, and to enable benchmark dose analysis. A vehicle control group and 7 dose groups were included, which received 0.27, 0.82, 2.47, 7.4, 22.2, 66.7 or 200 mg pentaBDE/kg bw/d (mkd). The liver appeared to be a key target organ, showing a marked increase of weight and centrilobular hepatocellular hypertrophy, probably due to the observed induction of P450 enzymes, notably CYP1A and CYP2B. A marked decrease of circulating total thyroxine (TT4) and an increase of plasma cholesterol were probably secondary to the liver effects. Furthermore, dose-dependently decreased weight of epididymis, seminal vesicles, and prostate, as well as sperm head deformities in males, and induction of CYP17 activity in adrenals in females were observed, all possibly related to anti-androgenic activity. Finally, we observed a substantial increase of large unstained cells in the blood and a decrease of apolar retinoids in the liver. All these effects had benchmark doses at the lower confidence bound (BMDL) in the low- or mid-dose range, but particular sensitive, potentially adverse effects were TT4 decrease (BMDLs 1.1 in males and 1.8 mkd in females), and decrease of hepatic apolar retinoids (BMDLs 0.5 mkd in males and 2.3 mkd in females). These results contribute to refinement of the hazard identification of pentaBDE and improved risk assessment of human exposure to this industrial chemical and environmental pollutant.
- Published
- 2008
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23. Endocrine effects of tetrabromobisphenol-A (TBBPA) in Wistar rats as tested in a one-generation reproduction study and a subacute toxicity study.
- Author
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Van der Ven LT, Van de Kuil T, Verhoef A, Verwer CM, Lilienthal H, Leonards PE, Schauer UM, Cantón RF, Litens S, De Jong FH, Visser TJ, Dekant W, Stern N, Håkansson H, Slob W, Van den Berg M, Vos JG, and Piersma AH
- Subjects
- Administration, Oral, Animals, Body Weight drug effects, Bone Development drug effects, Bone and Bones drug effects, Dose-Response Relationship, Drug, Endocrine Disruptors pharmacokinetics, Female, Male, Organ Size drug effects, Polybrominated Biphenyls pharmacokinetics, Rats, Rats, Wistar, Thyroid Hormones blood, Tissue Distribution, Toxicity Tests methods, Endocrine Disruptors toxicity, Polybrominated Biphenyls toxicity, Reproduction drug effects
- Abstract
Endocrine effects of the brominated flame retardant tetrabromobisphenol-A (TBBPA) were studied in a one-generation reproduction assay in Wistar rats via repeated dietary exposure, applying eight dose groups at 0-3-10-30-100-300-1,000-3,000 mg/kg body weight/day (mkd). This design enables dose-response analysis and calculation of benchmark doses (BMDL). This reproduction study was preceded by a 28-day repeat dose subacute toxicity study, at 0-30-100-300 mkd. Major effects in the reproduction study included decreased circulating thyroxine (T4) with BMDLs of 31 (m) and 16 (f) mkd, and increased weight of testis and male pituitary (BMDLs of 0.5 and 0.6 mkd). The hypothyroxinemia correlated to a cluster of developmental parameters including delayed sexual development in females, decreased pup mortality, and effects on brainstem auditory evoked potentials [Lilienthal, H., Verwer, C.M., Van der Ven, L.T.M., Piersma, A.H., Vos, J.G., 2008. Neurobehavioral effects of tetrabromobisphenol A (TBBPA) in rats after pre- and postnatal exposure. Toxicology]. A second cluster of parameters in F1 animals was correlated to increased testis weight, and included female gonad weight, endometrium height, CYP19/aromatase activity in the ovary, and plasma testosterone levels in males. These two correlation clusters suggest a dual action of TBBPA. The only effects in the subacute study were decreased circulating T4 and increased T3 levels in males (BMDLs 48 and 124mkd), and non-significant trends for these parameters in females, suggesting that the other effects in the reproduction study were induced during development. Combined with data of human exposure to environmental TBBPA, the margin of exposure for highly exposed populations can be calculated at 2.6, and current use of TBBPA may therefore be a matter of concern for human health.
- Published
- 2008
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24. A 28-day oral dose toxicity study enhanced to detect endocrine effects of hexabromocyclododecane in Wistar rats.
- Author
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van der Ven LT, Verhoef A, van de Kuil T, Slob W, Leonards PE, Visser TJ, Hamers T, Herlin M, Håkansson H, Olausson H, Piersma AH, and Vos JG
- Subjects
- Administration, Oral, Animals, Bone Density drug effects, Cell Count, Dose-Response Relationship, Drug, Endocrine Disruptors classification, Female, Glucuronosyltransferase biosynthesis, Hydrocarbons, Brominated classification, Liver drug effects, Liver pathology, Male, Organ Size drug effects, Pituitary Gland metabolism, Pituitary Gland pathology, Rats, Rats, Wistar, Risk Assessment, Spleen drug effects, Spleen pathology, Thyroid Gland metabolism, Thyroid Gland pathology, Thyrotropin metabolism, Tibia drug effects, Tibia metabolism, Toxicity Tests, Endocrine Disruptors toxicity, Hydrocarbons, Brominated toxicity, Pituitary Gland drug effects, Thyroid Gland drug effects
- Abstract
A 28-day repeated dose study in rats (OECD407) enhanced for endocrine and immune parameters was performed with hexabromocyclododecane (HBCD). Rats were exposed by daily gavage to HBCD dissolved in corn oil in 8 dose groups with doses ranging between 0 and 200 mg/kg bw per day (mkd). Evaluation consisted of dose-response analysis with calculation of a benchmark dose at the lower 95% one-sided confidence bound (BMDL) at predefined critical effect sizes (CESs) of 10-20%. The most remarkable findings were dose-related effects on the thyroid hormone axis, that is, decreased total thyroxin (TT4, BMDL 55.5 mkd at CES--10%), increased pituitary weight (29 mkd at 10%) and increased immunostaining of TSH in the pituitary, increased thyroid weight (1.6 mkd at 10%), and thyroid follicle cell activation. These effects were restricted to females. Female rats also showed increased absolute liver weights (22.9 mkd at 20%) and induction of T4-glucuronyl transferase (4.1 mkd at 10%), suggesting that aberrant metabolization of T4 triggers feedback activation of the thyroid hormone system. These effects were accompanied by possibly secondary effects, including increased cholesterol (7.4 mkd at 10%), increased tibial bone mineral density (> 49 mkd at 10%), both in females, and decreased splenocyte counts (0.3-6.3 mkd at 20%; only evaluated in males). Overall, female rats appeared to be more sensitive to HBCD than male rats, and an overall BMDL is proposed at 1.6 mkd, based on a 10% increase of the thyroid weight, which was the most sensitive parameter in the sequence of events.
- Published
- 2006
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25. Pertussis toxin relaxes small arteries with no vascular lesions or vascular smooth muscle cell injury.
- Author
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van Meijeren CE, Vleeming W, Dormans JA, van de Kuil T, Opperhuizen A, Hendriksen CF, and de Wildt DJ
- Subjects
- Animals, Bordetella pertussis chemistry, Injections, Intravenous, Male, Mesenteric Arteries ultrastructure, Microscopy, Electron, Transmission, Muscle Contraction drug effects, Muscle, Smooth, Vascular ultrastructure, Pertussis Toxin administration & dosage, Pertussis Toxin isolation & purification, Rats, Rats, Wistar, Specific Pathogen-Free Organisms, Mesenteric Arteries drug effects, Muscle, Smooth, Vascular drug effects, Pertussis Toxin pharmacology, Vasodilation drug effects
- Abstract
Previous studies showed that pertussis toxin (PT) decreased agonist-induced contractions of isolated rat small mesenteric resistance arteries independently from endothelium, nitric oxide-synthase or intracellular calcium concentrations. In this study, it was investigated if the PT-induced decreased contractile properties of small mesenteric resistance arteries could be a consequence of a PT-induced vascular and/or smooth muscle cell injury, leading to loss of contractile functionality. Male Wistar rats were treated with PT (30 microg/kg, intravenously) and sections of isolated small mesenteric resistance arteries were investigated with light- and electron microscopy. Light microscopic investigation of cross-sectioned small mesenteric resistance arteries of control animals clearly showed a contracted phase, while PT-pretreated animals showed a relaxed smooth inner surface of the vessel, indicating a vasodilated state. Electron microscopic investigation showed that PT-pretreatment neither induced vascular lesions nor caused morphological or numerical changes in cell organelles such as contractile elements of vascular smooth muscle cells. In conclusion, the PT-induced decreased contractile properties of isolated rat small resistance arteries are not caused by a PT-induced vascular and/or smooth muscle cell injury.
- Published
- 2004
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26. Pertussis toxin-induced histamine sensitisation: an aspecific phenomenon independent from the nitric oxide system?
- Author
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van Meijeren CE, Vleeming W, van de Kuil T, Gerards AL, Hendriksen CF, and de Wildt DJ
- Subjects
- Animals, Aorta, Thoracic drug effects, Blood Pressure drug effects, Buffers, Calcium metabolism, Dose-Response Relationship, Drug, Histamine pharmacology, Injections, Intravenous, Male, Mesenteric Arteries drug effects, Mesenteric Arteries injuries, Methods, Muscle, Smooth, Vascular drug effects, NG-Nitroarginine Methyl Ester pharmacology, Netherlands, Norepinephrine antagonists & inhibitors, Norepinephrine pharmacology, Pertussis Toxin administration & dosage, Pertussis Vaccine standards, Phenylephrine antagonists & inhibitors, Phenylephrine pharmacology, Potassium Chloride pharmacology, Rats, Rats, Wistar, Serotonin pharmacology, Vasoconstriction drug effects, Nitric Oxide immunology, Nitric Oxide physiology, Pertussis Toxin adverse effects, Pertussis Toxin immunology
- Abstract
Mechanisms were studied initially to develop an in vitro safety test for detecting pertussis toxin toxicity in acellular pertussis vaccines based on the histamine sensitisation test. Maximal contractions and sensitivities to different agonists and adrenoceptor-induced contractions in Ca2+-free medium of isolated rat small mesenteric resistance arteries were significantly reduced by in vivo [30 microg/kg, intravenously (i.v.), day 5] or in vitro (10 microg/ml, 2 h) pertussis toxin pretreatment. Pertussis toxin-induced decrease in sensitivity of small mesenteric resistance arteries to noradrenaline was endothelium-dependent. Nomega-nitro-L-arginine methyl ester (L-NAME) (100 microM, 20 min) did not reestablish the sensitivity to noradrenaline. In vivo L-NAME treatment (0, 1, 10 or 30 mg/kg) of pertussis toxin-pretreated (15 microg/kg) rats did not reduce pertussis toxin-induced enhancement of the histamine-induced decrease in blood pressure and histamine (10, 30, 100 or 300 mg/kg) induced mortality. Finally, in vivo pertussis toxin pretreatment sensitises rats for sodium nitroprusside (50 microg/kg/min). We conclude that pertussis toxin-induced histamine sensitisation is caused by an interference of pertussis toxin with the contractile mechanisms of vascular smooth muscle of resistance arteries which indicates only an indirect role for histamine in the histamine sensitisation test., (Copyright 2004 Elsevier B.V.)
- Published
- 2004
- Full Text
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27. In vivo pertussis toxin treatment reduces contraction of rat resistance arteries but not that of mouse trachea.
- Author
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van Meijeren CE, Vleeming W, van de Kuil T, Manni J, Kegler D, Hendriksen CF, and de Wildt DJ
- Subjects
- Animals, Histamine pharmacology, In Vitro Techniques, Male, Mesenteric Arteries drug effects, Mice, Mice, Inbred BALB C, Muscle Contraction drug effects, Muscle Relaxation drug effects, Rats, Rats, Wistar, Vascular Resistance drug effects, Vasoconstrictor Agents pharmacology, Vasodilator Agents pharmacology, Muscle, Smooth drug effects, Muscle, Smooth, Vascular drug effects, Pertussis Toxin pharmacology, Trachea drug effects
- Abstract
In order to develop an in vitro method for detecting residual pertussis toxin activity in acellular pertussis vaccines, the effects of in vivo pertussis toxin treatment on contraction and relaxation properties of isolated mouse trachea and of isolated rat small mesenteric resistance arteries were studied. In vivo pertussis toxin treatment (24 or 72 microg/kg, intraperitoneally (i.p.)) did not affect contraction and relaxation properties of isolated BALB/c or NIH mouse trachea. In vivo pertussis toxin treatment (30 microg/kg, intravenously) significantly reduced noradrenaline- or KCl-induced maximal contraction and reduced sensitivity to noradrenaline in isolated male Wistar rat small mesenteric resistance arteries. However, in vivo pertussis toxin treatment did not affect relaxation properties of isolated rat small mesenteric resistance arteries. These results support the hypothesis that vasoconstriction-regulating mechanisms and not airway constriction mechanisms are involved in pertussis toxin-induced histamine sensitisation. The vasoconstriction-regulating mechanisms may provide a lead for further development of an in vitro method for measuring biologically active pertussis toxin in acellular pertussis vaccines based on mechanisms involved in the histamine sensitisation test.
- Published
- 2004
- Full Text
- View/download PDF
28. In vivo cytokinesis blocked micronucleus assay with carbendazim in rat fibroblasts and comparison with in vitro assays.
- Author
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de Stoppelaar JM, van de Kuil T, Verharen HW, Hokse H, Opperhuizen A, Mohn GR, van Benthem J, and Hoebee B
- Subjects
- Animals, Centromere drug effects, Cytochalasin B pharmacology, Dose-Response Relationship, Drug, In Situ Hybridization, Fluorescence, Male, Micronuclei, Chromosome-Defective drug effects, Nondisjunction, Genetic, Ploidies, Rats, Rats, Wistar, Benzimidazoles pharmacology, Carbamates, Cell Division drug effects, Fibroblasts drug effects, Micronucleus Tests methods, Mutagens pharmacology
- Abstract
A successful in vivo application of the cytokinesis blocked micronucleus assay for the detection of aneuploidy induced by carbendazim (CARB) was carried out in the granuloma pouch assay. This was performed in two ways: (i) in vivo exposure of the skin fibroblasts to cytochalasin B (cytB) and CARB, by simultaneous injection of both substances into the pouch; (ii) in vivo exposure to CARB followed by in vitro culturing of the fibroblasts in the presence of cytB. Only the first assay was successful. Injection of cytB (with or without the test compound) into the pouch resulted in the induction of binucleate cells in vivo, up to a maximum of 5% at 1 mg cytB/pouch. After injection of CARB (0-50 or 0-10 mg/pouch) and cytB (1 mg) into the pouch, aneuploidy was determined in the isolated binucleate fibroblasts by fluorescence in situ hybridization with a general centromeric probe and combinations of chromosome-specific probes (19p + 19q, 4q + Yq). With all probes, the induction of chromosome loss and/or non-disjunction by CARB was very pronounced; at 10 mg CARB/pouch the total malsegregation frequency of chromosomes 4, 19 and Y was approximately 300/1000 binucleate cells. In an in vitro cytokinesis block assay with CARB (0-2.5 microg/ml) in primary skin fibroblasts the induced aneuploidy frequencies were as high as observed in the in vivo assay. The use of two probes for chromosome 19, which enabled the scoring of chromosome breaks in addition to aneuploidy, revealed no significant induction of chromosome breaks by CARB. The frequency of polyploid mononucleate and binucleate cells was decreased after CARB treatment, in both the in vivo and in vitro assays. However, in an additional in vitro assay without cytB a major induction of polyploidy from 2.5 microg/ml CARB and above was observed, showing that cytB may interfere with polyploidy induction.
- Published
- 2000
- Full Text
- View/download PDF
29. Repeated administration of whole-cell and acellular pertussis vaccines affects haemodynamics and autonomic responsiveness.
- Author
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van Amsterdam JG, te Biesebeek JD, van de Kuil T, van der Laan JW, de Wildt DJ, and Vleeming W
- Subjects
- Animals, Male, Rats, Rats, Wistar, Autonomic Nervous System physiology, Blood Pressure, Heart Rate, Pertussis Vaccine adverse effects
- Abstract
Rats were treated in a repetitive way one to four times with either pertussis toxin, combined Diphtheria-Tetanus-Poliomyelitis-Pertussis vaccine (DTP-IPV vaccine, which includes inactivated polio virus and whole-cell pertussis), DT-IPV vaccine (lacking the whole-cell pertussis component) or acellular pertussis (aP) vaccine or Haemophilus influenzae type b vaccine. Baseline diastolic blood pressure, baseline heart rate and adrenergic and cholinergic responses were evaluated 4 days after last treatment. Pertussis toxin decreased baseline diastolic blood pressure (28-43%) and increased baseline heart rate (28-40%). Adrenergic and cholinergic response were inhibited by 65-75% and 70-78%. Multiple treatments were grossly as effective as single treatment. Similar results were obtained with DTP-IPV, while DT-IPV did not affect any of the four responses measured. Acellular pertussis vaccine did not affect baseline diastolic blood pressure, but significantly increased baseline heart rate (14%) and inhibited the adrenergic (19-23%) and cholinergic response (39-50%). This indicates that the acellular vaccine tested contains pharmacologically active pertussis toxin. As the effects were less pronounced compared to DTP-IPV, it is concluded that acellular pertussis retains less residual toxicological effects than whole-cell pertussis vaccine and may therefore be a safer vaccine. The observed effects on haemodynamics and autonomic control seem to be specific for pertussis toxin and pertussis-related vaccines as Haemophilus influenzae type b vaccine is in this respect virtually inactive.
- Published
- 1998
- Full Text
- View/download PDF
30. The effect of pertussis toxin and whole-cell pertussis vaccine on haemodynamics and autonomic responsiveness in the rat depends on route of administration and age.
- Author
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van Amsterdam JG, te Biesebeek JD, van de Kuil T, van der Laan JW, Wemer J, de Wildt DJ, and Vleeming W
- Subjects
- Animals, Dose-Response Relationship, Drug, Male, Rats, Rats, Wistar, Aging physiology, Autonomic Nervous System drug effects, Blood Pressure drug effects, Heart Rate drug effects, Pertussis Toxin, Pertussis Vaccine pharmacology, Virulence Factors, Bordetella pharmacology
- Abstract
Vaccination of children with Diphtheria, Tetanus, Poliomyelitis and pertussis vaccine (DTPoP-vaccine) containing the whole-cell pertussis component is known to be associated with manifestation of side-effects such as acute encephalopathy, convulsions and hypotensive-hyporesponsive episodes. In young and adult rats the effects of pertussis toxin and DTPoP-vaccine on haemodynamics and autonomic responsiveness are evaluated following treatment with high dose via different routes of administration (s.c., i.p. and i.v.). The effect of pertussis toxin is dose-dependent (between 1 and 20 micrograms kg-1) and largest responses are observed after i.v. administration. At 20 micrograms kg-1, i.v. pertussis toxin decreases baseline diastolic blood pressure and increases baseline heart rate by 31% and inhibits autonomic responsiveness (salbutamol-induced increase in diastolic blood pressure and arecoline-induced decrease in heart rate). In adult rats DTPoP-vaccine induces generally more prominent effects than in young rats. In adult rats DTPoP-vaccine reduces baseline diastolic blood pressure by 25% while no response is observed in young rats. In adult rats DTPoP inhibits the adrenergic response though less compared to treatment of pertussis toxin. After treatment with DTPoP-vaccine (single or twice) only minor differences are observed between young and adult rats. Present results show that adult rats are more sensitive to pertussis toxin and pertussis vaccine than young rats and that the responses depend on the route of administration.
- Published
- 1998
- Full Text
- View/download PDF
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