Honey is known for its wound healing potential. In this study we investigated free radical format... more Honey is known for its wound healing potential. In this study we investigated free radical formation and inhibition in different honeys and the influence on bacterial growth inhibition. α-Oxaldehydes and free amino acids were shown to be present in manuka and non-manuka honeys. With electron spin resonance we measured that methylglyoxal (MG) and glyoxal (GO) form different carbon centered radicals under the influence of either hydrogen peroxide or arginine and lysine. These radicals were also formed in a combination of α-oxaldehydes and amino acids in artificial honey, which was quantified by the cytochrome C assay. Also, a strong bactericidal activity for MG and GO (minimum concentration 0.6 and 1.25 mM) was shown against different bacterial strains. Interestingly, honeys with a high MG content were shown to be better free radical scavengers for hydroxyl and superoxide radicals, implying a complex mechanism for free radical donation and scavenging in honey.
The Journal of Maternal-Fetal & Neonatal Medicine, 2016
Aims: Oxidative stress is crucial in the pathophysiology of preeclampsia. Reactive oxygen species... more Aims: Oxidative stress is crucial in the pathophysiology of preeclampsia. Reactive oxygen species (ROS) like the superoxide radical O2-, are produced by the ischemic placenta, causing systemic inflammation and endothelial dysfunction. EPR uses microwave radiation and a high magnetic field and is the most direct and reliable method to detect free radicals. We explored the feasibility of determining O2-concentration in maternal serum and compared pre-eclamptic and uncomplicated pregnancies. Methods: In this study a CMH (1-hydroxyl-3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine) spin trap stabilizing free O2-radicals for EPR detection, has been used. Twenty-four patients were included of which 12 pre-eclamptic pregnancies and 12 uncomplicated pregnancies. Results: EPR successfully detects O2-concentration in maternal serum. Results are presented in Figure 1. Since there was an overt difference (p ¼ 0.007) between patients with HELLP syndrome versus pre-eclamptic patients with only hypertension and proteinuria, these patients were observed as a distinct group. A significant difference between uncomplicated and pre-eclamptic pregnancies (p ¼ 0.005) was observed. Discussion: We hypothesize that an increased protective scavenger system for ROS is activated in pre-eclampsia. In case of further decompensation as in HELLP syndrome, this system collapses resulting in increased vascular endothelial damage.
Background During normal pregnancy, placental oxidative stress (OS) is present during all three t... more Background During normal pregnancy, placental oxidative stress (OS) is present during all three trimesters and is necessary to obtain normal cell function. However, if OS reaches a certain level, pregnancy complications might arise. In preeclampsia (PE), a dangerous pregnancy specific hypertensive disorder, OS induced in the ischemic placenta causes a systemic inflammatory response and activates maternal endothelial cells. In this study, we aimed to quantify superoxide concentrations (as a measure of systemic OS) using electron paramagnetic resonance (EPR) and correlate them to markers of systemic inflammation, iron status and vascular function. Methods Fifty-nine women with a healthy pregnancy (HP), 10 non-pregnant controls (NP) and 28 PE patients (32±3.3weeks) were included. During HP, blood samples for superoxide, neutrophil to lymphocyte ratio (NLR), mean platelet volume (MPV) and iron status were taken at 10, 25 and 39 weeks. Vascular measurements for arterial stiffness (carotid-femoral pulse wave velocity (CF-PWV), augmentation index (AIx), augmentation Pressure (AP)) and microvascular endothelial function (reactive hyperemia index (RHI)) were performed at 35 weeks. In PE, all measurements were performed at diagnosis. CMH (1-hydroxy-3-methoxycarbonyl
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
Hereditary spherocytosis (HS) is a congenital haemolytic anaemia which is characterized by a grea... more Hereditary spherocytosis (HS) is a congenital haemolytic anaemia which is characterized by a great variety of structural defects in the red cell's membrane skeleton and/or deficiencies in particular membrane (skeletal) proteins. Enhanced (Mg2+)‐dependent adenosine triphosphatase (Mg2+‐ATPase) activities, varying from 115% to 160%, were invariably found in erythrocyte ghosts derived from 13 HS patients. Similarly, an enhancement of Mg2+‐ATPase activity by 30% is observed in normal red cell ghosts that have been stripped of the greater part of their membrane skeletal proteins by treatment with a low ionic strength buffer. Reassociation of those stripped ghosts with spectrin reduces the enhanced Mg2+‐ATPase activity to its original level. Since in both cases, HS ghosts and stripped normal ghosts, the stabilizing effects that the membrane skeleton exerts on the maintenance of an endofacial localization of the aminophospholipids are impaired, the enhanced Mg2+‐ATPase activity is interpreted to reflect an increased activity of the aminophospholipid translocase. The present observations therefore support a role of the membrane skeleton in the stabilization of phospholipid asymmetry in the red cell membrane and consequently in reducing the energy consumption of the translocase.
Hereditary spherocytosis (HS) is a congenital haemolytic anaemia which is characterized by a grea... more Hereditary spherocytosis (HS) is a congenital haemolytic anaemia which is characterized by a great variety of structural defects in the red cell's membrane skeleton and/or deficiencies in particular membrane (skeletal) proteins. Enhanced (Mg2+)‐dependent adenosine triphosphatase (Mg2+‐ATPase) activities, varying from 115% to 160%, were invariably found in erythrocyte ghosts derived from 13 HS patients. Similarly, an enhancement of Mg2+‐ATPase activity by 30% is observed in normal red cell ghosts that have been stripped of the greater part of their membrane skeletal proteins by treatment with a low ionic strength buffer. Reassociation of those stripped ghosts with spectrin reduces the enhanced Mg2+‐ATPase activity to its original level. Since in both cases, HS ghosts and stripped normal ghosts, the stabilizing effects that the membrane skeleton exerts on the maintenance of an endofacial localization of the aminophospholipids are impaired, the enhanced Mg2+‐ATPase activity is inte...
Oxidative stress is an important pathophysiological mechanism in the development of numerous card... more Oxidative stress is an important pathophysiological mechanism in the development of numerous cardiovascular disorders, but few studies have examined the levels of oxidative stress in adults with congenital heart disease (CHD). The objective of this study was to investigate oxidative stress levels in adults with CHD and the association with inflammation, exercise capacity and endothelial function. To this end, 36 adults with different types of CHD and 36 age- and gender-matched healthy controls were enrolled. Blood cell counts, hs-CRP, NT-proBNP, fasting glucose, cholesterol levels, iron saturation and folic acid concentrations were determined in venous blood samples. Levels of superoxide anion radical in whole blood were determined using electron paramagnetic resonance spectroscopy in combination with the spin probe CMH. Physical activity was assessed with the IPAQ-SF questionnaire. Vascular function assessment (EndoPAT) and cardiopulmonary exercise testing were performed in the pat...
Titanium dioxide as a food additive (E171) has been demonstrated to facilitate growth of chemical... more Titanium dioxide as a food additive (E171) has been demonstrated to facilitate growth of chemically induced colorectal tumours in vivo and induce transcriptomic changes suggestive of an immune system impairment and cancer development. The present study aimed to investigate the molecular mechanisms behind the tumour stimulatory effects of E171 in combination with azoxymethane (AOM)/ dextran sodium sulphate (DSS) and compare these results to a recent study performed under the same conditions with E171 only. BALB/c mice underwent exposure to 5 mg/kg bw /day of E171 by gavage for 2, 7, 14, and 21 days. Whole genome mRNA microarray analyses on the distal colon were performed. The results show that E171 induced a downregulation of genes involved in the innate and adaptive immune system, suggesting impairment of this system. In addition, over time, signalling genes involved in colorectal cancer and other types of cancers were modulated. In relation to cancer development, effects potentially associated with oxidative stress were observed through modulation of genes related to antioxidant production. E171 affected genes involved in biotransformation of xenobiotics which can form reactive intermediates resulting in toxicological effects. These transcriptomics data reflect the early biological responses induced by E171 which precede tumour formation in an AOM/DSS mouse model. Titanium dioxide (TiO 2) is the most produced mineral worldwide 1 and is used for its white colouring properties and stability over time 2. One of the main applications of TiO 2 is its use as a food additive in processed food such as cookies, sweets, toppings and coffee creamer 1-3. TiO 2 was accepted as a food colorant and registered under the code of E171 by the European Union (EU) in 1969 4. This approval followed a risk assessment made earlier that year by the joint Food Agriculture Organisation and World Health Organisation (FAO/WHO) that identified no risk concerning the ingestion of TiO 2 4. Therefore, E171 is permitted by the EU in food at quantum satis, which means that there are no maximum intake levels for ingestion. In the USA, the regulation by the food and drug administration limits titanium dioxide to a maximum of 1% of the weight of the food 5. Over the past decade, characteristics have been studied in order to assess possible adverse effects. The shape and size of E171 have been measured by several groups. E171 consist of approximately 40% of nanoparticles (NPs) (<100 nm) and 60% of microparticles (MPs) (>100 nm) 3,6,7. Studies on possible adverse effects have primarily been focused on inhalation of TiO 2. Based on the outcome of these studies, the International Agency for Research in Cancer (IARC) changed in 2010 the classification of TiO 2 from non-carcinogenic to probable carcinogenic to humans (Group 2B) 8. More recently, there are increasing number of studies to detect and quantify E171 in foods for human consumption 9 , pharmaceutical products 10 , to distinguish between NPs and MPs in foods, and to estimate the daily intake in adults and children 11. In addition, a number of both in vivo and in vitro studies report on the adverse effects of exposure to TiO 2 by the oral route and the biological mechanisms involved. In vitro studies have
The application of transcriptome analyses in molecular epidemiology studies has become a promisin... more The application of transcriptome analyses in molecular epidemiology studies has become a promising tool in order to evaluate the impact of environmental exposures. These analyses have a great value in establishing the exposome, the totality of human exposures, both by identifying the chemical nature of the exposures and the induced molecular responses. Transcriptomic signatures can be regarded as biomarker of exposure as well as markers of effect which reflect the interaction between individual genetic background and exposure levels. However, the biological interpretation of modulated gene expression profiles is a challenging task and translating affected molecular pathways into risk assessment, for instance in terms of cancer promoting or disease preventing responses, is a far from standardised process. Here, we describe the in-depth analyses of the gene expression responses in a human dietary intervention in which the interaction between genotype and exposure to a blueberry-apple juice containing a complex mixture of phytochemicals is investigated. We also describe how data on differences in genetic background combined with different effect markers can provide a better understanding of gene-environment interactions. Pathway analyses of differentially expressed genes in combination with gene were used to identify complex but strong changes in several biological processes like immune response, cell adhesion, lipid metabolism and apoptosis. These observed changes may lead to upgraded growth control, induced immunity, reduced platelet aggregation and activation, diminished production of reactive oxidative species by platelets, blood glucose homeostasis, regulation of blood lipid levels and increased apoptosis. Our findings demonstrate that applying transcriptomics to well-controlled human dietary intervention studies can provide insight into mechanistic pathways involved in disease prevention by dietary factors. Analysing 'omics-' responses to dietary and environmental exposures in human population studies With the introduction of genomics-based technologies, providing large scale data on biological responses to dietary or environmental factors, our understanding of the molecular mechanisms behind increased cancer risk as well as the preventive action of specific dietary factors has started to grow. This understanding strengthens the biological plausibility of the observed links between exposures and cancer risk in epidemiological studies, whereas comparison of gene
The biological response to four well-characterized amorphous silica nanoparticles was investigate... more The biological response to four well-characterized amorphous silica nanoparticles was investigated in RAW 264.7 macrophages in view of their potential application as drug carriers to sites of inflammation. All silica nanoparticles-induced cell membrane damage, reduced metabolic activity, generated ROS and released various cytokines, but to different extents. Two silica nanoparticles of 34 nm (A and B) with different zetapotentials were more cytotoxic than (aggregated) 11 and 248 nm nanoparticles, while cytokines were mostly induced by the (aggregated) 11 nm and only one of the 34 nm nanoparticles (34A). The results indicate that specific silica nanoparticles may have counterproductive effects, for example when used as carriers of anti-inflammatory drugs. The physicochemical properties determining the response of nanoparticles vary for different responses, implying that a screening approach for the safe development of nanoparticles needs to consider the role of combinations of (dynamic) physicochemical properties and needs to include multiple toxicity endpoints. Keywords Nanoparticles Á Silica Á Cytotoxicity Á Reactive oxygen species Á Inflammation Á Cytokines Á Environmental and health effects 2010; Roy et al. 2008). An interesting aspect of nanoparticles biodistribution is that they tend to accumulate at sites of inflammation due to increased vascular permeability (Moulari et al. 2008). In
Food-derived heterocyclic aromatic amines (HCAs) have proved to be carcinogenic in both rodents a... more Food-derived heterocyclic aromatic amines (HCAs) have proved to be carcinogenic in both rodents and nonhuman primates. Two different metabolic pathways are suggested for the metabolic activation of HCA. The hepatic pathway proceeds via a two-step process involving N-hydroxylation by cytochrome P4501A2 and subsequent O-acetylation by N-acetyltransferase-2. An alternative pathway may be of particular interest in extrahepatic tissues and proceeds via one-electron oxidation catalyzed by prostaglandin H synthase (PHS), rendering free-radical metabolites. In this study, we investigated the metabolic activation of two HCAs, 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), by two different enzyme systems in vitro, generating different primary and secondary reactive metabolites. Rat liver S9 mix and PHS were used as the activating system and represent the hepatic and extrahepatic pathways, respectively. Electron-spin resonance spectroscopy showed that both IQ and PhIP exerted inhibiting effects on PHS-mediated formation of hydroxyl radicals during the conversion of arachidonic acid to prostaglandins. Evidence for the formation of HCA free radicals was presented in an indirect way by the formation of glutathione-derived thiyl radicals, with purified PHS as the activating system. Activation by S9 mix did not result in the formation of detectable radical metabolites, showing that the two metabolic routes primarily led to the formation of different metabolites. In all electron-spin resonance experiments, IQ appeared to be more effective than PhIP. In contrasts, DNA adduct analysis by means of (32)P-postlabeling showed similar adduct patterns for S9 and PHS in single-stranded and double-stranded salmon testes DNA after incubation with PhIP, indicating the ultimate formation of a common reactive intermediate. For IQ, activation by PHS led to an additional adduct spot that was not present after S9 activation. Furthermore, activation of IQ resulted in higher adduct levels compared with PhIP for both activation pathways. Overall, adduct levels were higher in single-stranded DNA than double-stranded DNA. Our results showed that the hepatic and extrahepatic pathways resulted in different primary metabolites, while the ultimate formation of a similar reactive intermediate for PhIP, possibly an arylnitrenium ion, suggested that both pathways could play an important role in the onset of carcinogenesis.
Reactive oxygen species-induced oxidative stress in the colon is involved in inflammatory bowel d... more Reactive oxygen species-induced oxidative stress in the colon is involved in inflammatory bowel diseases and suggested to be associated with colorectal cancer risk. However, our insight in molecular responses to different oxygen radicals is still fragmentary. Therefore, we studied global gene expression by an extensive time series (0.08, 0.25, 0.5, 1, 2, 4, 8, 16, or 24 h) analyses in human colon cancer (caco-2) cells after exposure to H 2 O 2 or the superoxide anion donor menadione. Differences in gene expression were investigated by hybridization on two-color microarrays against nonexposed time-matched control cells. Next to gene expression, correlations with related phenotypic markers (8-oxodG levels and cell cycle arrest) were investigated. Gene expression analysis resulted in 1404 differentially expressed genes upon H 2 O 2 challenge and 979 genes after menadione treatment. Further analysis of gene expression data revealed how these oxidant responses can be discriminated. Time-dependent coregulated genes immediately showed a pulse-like response to H 2 O 2 , while the menadione-induced expression is not restored over 24 h. Pathway analyses demonstrated that H 2 O 2 immediately influences pathways involved in the immune function, while menadione constantly regulated cell cycle-related pathways Altogether, this study offers a novel and detailed insight in the similarities and differences of the time-dependent oxidative stress responses induced by the oxidants H 2 O 2 and menadione and show that these can be discriminated regarding their modulation of particular colon carcinogenesis-related mechanisms.
Journal of toxicology and environmental health, Feb 1, 2006
The mechanisms by which particulate matter (PM) produces adverse effects on the respiratory syste... more The mechanisms by which particulate matter (PM) produces adverse effects on the respiratory system, such as pulmonary dysfunction in children, are largely unknown. However, oxidative stress is thought to play an important role. Various chemical compounds in ambient particulate matter, including transition metals and aromatic organic compounds, may contribute to adverse effects through intrinsic generation of reactive oxygen species (ROS). It was hypothesized that ROS generation by PM, as determined through electron spin resonance (ESR) spectroscopy, may be negatively associated with pulmonary function in school children. PM(2.5), PM(10), and total suspended particulates (TSP) were sampled at the playgrounds of six elementary schools in the city of Maastricht, the Netherlands. All children (8-13 yr) from the six schools were asked to undergo spirometry. Multivariate linear regression models were constructed to evaluate associations between oxygen radical formation by PM and lung function. The radical-generating capacity per microgram PM correlated negatively to forced expiratory volume in 1 s (FEV(1)) and forced expiratory flow at 50% (FEF(50%)) of forced vital capacity (FVC). The data indicate that chemical features that contribute to intrinsic generation of ROS may be relevant for PM risk assessment.
Exposure to increased levels of ambient particulate matter (PM) are associated with several healt... more Exposure to increased levels of ambient particulate matter (PM) are associated with several health effects, including cardiopulmonary diseases. The formation of reactive oxygen species (ROS) is thought to play an important role in the induction of these health effects. To quantify the ROS generating capacity of PM, we developed an improved electron spin resonance (ESR) spectrometry-based method. ROS formation was measured directly on PMcontaining filters, thereby avoiding the selective extraction of components and loss of material or reactivity, which is likely to occur during filter extraction. Also, ascorbic acid was added to stimulate ROS formation. This method was applied to PM 10 samples originating from different sources. The radical generating capacity of PM 10 from both gasoline and diesel engine exhaust was significantly higher as compared to that of PM 10 from ambient or indoor air. Furthermore, in urban PM 10 and PM 2.5 , ROS-generating capacity significantly correlated with concentrations of polycyclic aromatic hydrocarbon content and particular transition metals. This indicates that this improved ESR method may be a valuable tool for evaluating the relationship between ROS formation by PM and the adverse health effects associated with this type of air pollution.
People interested in the research are advised to contact the author for the final version of the ... more People interested in the research are advised to contact the author for the final version of the publication, or visit the DOI to the publisher's website. • The final author version and the galley proof are versions of the publication after peer review. • The final published version features the final layout of the paper including the volume, issue and page numbers. Link to publication General rights Copyright and moral rights for the publications made accessible in the public portal are retained by the authors and/or other copyright owners and it is a condition of accessing publications that users recognise and abide by the legal requirements associated with these rights. • Users may download and print one copy of any publication from the public portal for the purpose of private study or research. • You may not further distribute the material or use it for any profit-making activity or commercial gain • You may freely distribute the URL identifying the publication in the public portal. If the publication is distributed under the terms of Article 25fa of the Dutch Copyright Act, indicated by the "Taverne" license above, please follow below link for the End User Agreement:
N-nitroso compounds (NOCs) may be implicated in human colon carcinogenesis, but the toxicological... more N-nitroso compounds (NOCs) may be implicated in human colon carcinogenesis, but the toxicological mechanisms involved have not been elucidated. Because it was previously demonstrated that nitrosamines and nitrosamides, representing two classes of NOC, induce distinct gene expression effects in colon cells that are particularly related to oxidative stress, we hypothesized that different radical mechanisms are involved. Using electron spin resonance spectroscopy, we investigated the radical-generating properties of genotoxic NOC concentrations in human colon adenocarcinoma cells (Caco-2). Cells were exposed to nitrosamides (N-methyl-N#-nitro-Nnitrosoguanidine and N-methyl-N-nitrosourea) or nitrosamines (Nnitrosodiethylamine, N-nitrosodimethylamine, N-nitrosopiperidine, and N-nitrosopyrrolidine). Nitrosamines caused formation of reactive oxygen species (ROS) and carbon-centered radicals, which was further stimulated in the presence of Caco-2 cells. N-methyl-Nnitrosourea exposure resulted in a small ROS signal, and formation of nitrogen-centered radicals (NCRs), also stimulated by Caco-2 cells. N-methyl-N#-nitro-N-nitrosoguanidine did not cause radical formation at genotoxic concentrations, but at increased exposure levels, both ROS and NCR formation was observed. By associating gene expression patterns with ROS formation, several cellular processes responding to nitrosamine exposure were identified, including apoptosis, cell cycle blockage, DNA repair, and oxidative stress. These findings suggest that following NOC exposure in Caco-2 cells, ROS formation plays an important role in deregulation of gene expression patterns that may be relevant for the process of chemical carcinogenesis in the human colon, in addition to the role of DNA alkylation.
Honey is known for its wound healing potential. In this study we investigated free radical format... more Honey is known for its wound healing potential. In this study we investigated free radical formation and inhibition in different honeys and the influence on bacterial growth inhibition. α-Oxaldehydes and free amino acids were shown to be present in manuka and non-manuka honeys. With electron spin resonance we measured that methylglyoxal (MG) and glyoxal (GO) form different carbon centered radicals under the influence of either hydrogen peroxide or arginine and lysine. These radicals were also formed in a combination of α-oxaldehydes and amino acids in artificial honey, which was quantified by the cytochrome C assay. Also, a strong bactericidal activity for MG and GO (minimum concentration 0.6 and 1.25 mM) was shown against different bacterial strains. Interestingly, honeys with a high MG content were shown to be better free radical scavengers for hydroxyl and superoxide radicals, implying a complex mechanism for free radical donation and scavenging in honey.
The Journal of Maternal-Fetal & Neonatal Medicine, 2016
Aims: Oxidative stress is crucial in the pathophysiology of preeclampsia. Reactive oxygen species... more Aims: Oxidative stress is crucial in the pathophysiology of preeclampsia. Reactive oxygen species (ROS) like the superoxide radical O2-, are produced by the ischemic placenta, causing systemic inflammation and endothelial dysfunction. EPR uses microwave radiation and a high magnetic field and is the most direct and reliable method to detect free radicals. We explored the feasibility of determining O2-concentration in maternal serum and compared pre-eclamptic and uncomplicated pregnancies. Methods: In this study a CMH (1-hydroxyl-3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine) spin trap stabilizing free O2-radicals for EPR detection, has been used. Twenty-four patients were included of which 12 pre-eclamptic pregnancies and 12 uncomplicated pregnancies. Results: EPR successfully detects O2-concentration in maternal serum. Results are presented in Figure 1. Since there was an overt difference (p ¼ 0.007) between patients with HELLP syndrome versus pre-eclamptic patients with only hypertension and proteinuria, these patients were observed as a distinct group. A significant difference between uncomplicated and pre-eclamptic pregnancies (p ¼ 0.005) was observed. Discussion: We hypothesize that an increased protective scavenger system for ROS is activated in pre-eclampsia. In case of further decompensation as in HELLP syndrome, this system collapses resulting in increased vascular endothelial damage.
Background During normal pregnancy, placental oxidative stress (OS) is present during all three t... more Background During normal pregnancy, placental oxidative stress (OS) is present during all three trimesters and is necessary to obtain normal cell function. However, if OS reaches a certain level, pregnancy complications might arise. In preeclampsia (PE), a dangerous pregnancy specific hypertensive disorder, OS induced in the ischemic placenta causes a systemic inflammatory response and activates maternal endothelial cells. In this study, we aimed to quantify superoxide concentrations (as a measure of systemic OS) using electron paramagnetic resonance (EPR) and correlate them to markers of systemic inflammation, iron status and vascular function. Methods Fifty-nine women with a healthy pregnancy (HP), 10 non-pregnant controls (NP) and 28 PE patients (32±3.3weeks) were included. During HP, blood samples for superoxide, neutrophil to lymphocyte ratio (NLR), mean platelet volume (MPV) and iron status were taken at 10, 25 and 39 weeks. Vascular measurements for arterial stiffness (carotid-femoral pulse wave velocity (CF-PWV), augmentation index (AIx), augmentation Pressure (AP)) and microvascular endothelial function (reactive hyperemia index (RHI)) were performed at 35 weeks. In PE, all measurements were performed at diagnosis. CMH (1-hydroxy-3-methoxycarbonyl
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
Hereditary spherocytosis (HS) is a congenital haemolytic anaemia which is characterized by a grea... more Hereditary spherocytosis (HS) is a congenital haemolytic anaemia which is characterized by a great variety of structural defects in the red cell's membrane skeleton and/or deficiencies in particular membrane (skeletal) proteins. Enhanced (Mg2+)‐dependent adenosine triphosphatase (Mg2+‐ATPase) activities, varying from 115% to 160%, were invariably found in erythrocyte ghosts derived from 13 HS patients. Similarly, an enhancement of Mg2+‐ATPase activity by 30% is observed in normal red cell ghosts that have been stripped of the greater part of their membrane skeletal proteins by treatment with a low ionic strength buffer. Reassociation of those stripped ghosts with spectrin reduces the enhanced Mg2+‐ATPase activity to its original level. Since in both cases, HS ghosts and stripped normal ghosts, the stabilizing effects that the membrane skeleton exerts on the maintenance of an endofacial localization of the aminophospholipids are impaired, the enhanced Mg2+‐ATPase activity is interpreted to reflect an increased activity of the aminophospholipid translocase. The present observations therefore support a role of the membrane skeleton in the stabilization of phospholipid asymmetry in the red cell membrane and consequently in reducing the energy consumption of the translocase.
Hereditary spherocytosis (HS) is a congenital haemolytic anaemia which is characterized by a grea... more Hereditary spherocytosis (HS) is a congenital haemolytic anaemia which is characterized by a great variety of structural defects in the red cell's membrane skeleton and/or deficiencies in particular membrane (skeletal) proteins. Enhanced (Mg2+)‐dependent adenosine triphosphatase (Mg2+‐ATPase) activities, varying from 115% to 160%, were invariably found in erythrocyte ghosts derived from 13 HS patients. Similarly, an enhancement of Mg2+‐ATPase activity by 30% is observed in normal red cell ghosts that have been stripped of the greater part of their membrane skeletal proteins by treatment with a low ionic strength buffer. Reassociation of those stripped ghosts with spectrin reduces the enhanced Mg2+‐ATPase activity to its original level. Since in both cases, HS ghosts and stripped normal ghosts, the stabilizing effects that the membrane skeleton exerts on the maintenance of an endofacial localization of the aminophospholipids are impaired, the enhanced Mg2+‐ATPase activity is inte...
Oxidative stress is an important pathophysiological mechanism in the development of numerous card... more Oxidative stress is an important pathophysiological mechanism in the development of numerous cardiovascular disorders, but few studies have examined the levels of oxidative stress in adults with congenital heart disease (CHD). The objective of this study was to investigate oxidative stress levels in adults with CHD and the association with inflammation, exercise capacity and endothelial function. To this end, 36 adults with different types of CHD and 36 age- and gender-matched healthy controls were enrolled. Blood cell counts, hs-CRP, NT-proBNP, fasting glucose, cholesterol levels, iron saturation and folic acid concentrations were determined in venous blood samples. Levels of superoxide anion radical in whole blood were determined using electron paramagnetic resonance spectroscopy in combination with the spin probe CMH. Physical activity was assessed with the IPAQ-SF questionnaire. Vascular function assessment (EndoPAT) and cardiopulmonary exercise testing were performed in the pat...
Titanium dioxide as a food additive (E171) has been demonstrated to facilitate growth of chemical... more Titanium dioxide as a food additive (E171) has been demonstrated to facilitate growth of chemically induced colorectal tumours in vivo and induce transcriptomic changes suggestive of an immune system impairment and cancer development. The present study aimed to investigate the molecular mechanisms behind the tumour stimulatory effects of E171 in combination with azoxymethane (AOM)/ dextran sodium sulphate (DSS) and compare these results to a recent study performed under the same conditions with E171 only. BALB/c mice underwent exposure to 5 mg/kg bw /day of E171 by gavage for 2, 7, 14, and 21 days. Whole genome mRNA microarray analyses on the distal colon were performed. The results show that E171 induced a downregulation of genes involved in the innate and adaptive immune system, suggesting impairment of this system. In addition, over time, signalling genes involved in colorectal cancer and other types of cancers were modulated. In relation to cancer development, effects potentially associated with oxidative stress were observed through modulation of genes related to antioxidant production. E171 affected genes involved in biotransformation of xenobiotics which can form reactive intermediates resulting in toxicological effects. These transcriptomics data reflect the early biological responses induced by E171 which precede tumour formation in an AOM/DSS mouse model. Titanium dioxide (TiO 2) is the most produced mineral worldwide 1 and is used for its white colouring properties and stability over time 2. One of the main applications of TiO 2 is its use as a food additive in processed food such as cookies, sweets, toppings and coffee creamer 1-3. TiO 2 was accepted as a food colorant and registered under the code of E171 by the European Union (EU) in 1969 4. This approval followed a risk assessment made earlier that year by the joint Food Agriculture Organisation and World Health Organisation (FAO/WHO) that identified no risk concerning the ingestion of TiO 2 4. Therefore, E171 is permitted by the EU in food at quantum satis, which means that there are no maximum intake levels for ingestion. In the USA, the regulation by the food and drug administration limits titanium dioxide to a maximum of 1% of the weight of the food 5. Over the past decade, characteristics have been studied in order to assess possible adverse effects. The shape and size of E171 have been measured by several groups. E171 consist of approximately 40% of nanoparticles (NPs) (<100 nm) and 60% of microparticles (MPs) (>100 nm) 3,6,7. Studies on possible adverse effects have primarily been focused on inhalation of TiO 2. Based on the outcome of these studies, the International Agency for Research in Cancer (IARC) changed in 2010 the classification of TiO 2 from non-carcinogenic to probable carcinogenic to humans (Group 2B) 8. More recently, there are increasing number of studies to detect and quantify E171 in foods for human consumption 9 , pharmaceutical products 10 , to distinguish between NPs and MPs in foods, and to estimate the daily intake in adults and children 11. In addition, a number of both in vivo and in vitro studies report on the adverse effects of exposure to TiO 2 by the oral route and the biological mechanisms involved. In vitro studies have
The application of transcriptome analyses in molecular epidemiology studies has become a promisin... more The application of transcriptome analyses in molecular epidemiology studies has become a promising tool in order to evaluate the impact of environmental exposures. These analyses have a great value in establishing the exposome, the totality of human exposures, both by identifying the chemical nature of the exposures and the induced molecular responses. Transcriptomic signatures can be regarded as biomarker of exposure as well as markers of effect which reflect the interaction between individual genetic background and exposure levels. However, the biological interpretation of modulated gene expression profiles is a challenging task and translating affected molecular pathways into risk assessment, for instance in terms of cancer promoting or disease preventing responses, is a far from standardised process. Here, we describe the in-depth analyses of the gene expression responses in a human dietary intervention in which the interaction between genotype and exposure to a blueberry-apple juice containing a complex mixture of phytochemicals is investigated. We also describe how data on differences in genetic background combined with different effect markers can provide a better understanding of gene-environment interactions. Pathway analyses of differentially expressed genes in combination with gene were used to identify complex but strong changes in several biological processes like immune response, cell adhesion, lipid metabolism and apoptosis. These observed changes may lead to upgraded growth control, induced immunity, reduced platelet aggregation and activation, diminished production of reactive oxidative species by platelets, blood glucose homeostasis, regulation of blood lipid levels and increased apoptosis. Our findings demonstrate that applying transcriptomics to well-controlled human dietary intervention studies can provide insight into mechanistic pathways involved in disease prevention by dietary factors. Analysing 'omics-' responses to dietary and environmental exposures in human population studies With the introduction of genomics-based technologies, providing large scale data on biological responses to dietary or environmental factors, our understanding of the molecular mechanisms behind increased cancer risk as well as the preventive action of specific dietary factors has started to grow. This understanding strengthens the biological plausibility of the observed links between exposures and cancer risk in epidemiological studies, whereas comparison of gene
The biological response to four well-characterized amorphous silica nanoparticles was investigate... more The biological response to four well-characterized amorphous silica nanoparticles was investigated in RAW 264.7 macrophages in view of their potential application as drug carriers to sites of inflammation. All silica nanoparticles-induced cell membrane damage, reduced metabolic activity, generated ROS and released various cytokines, but to different extents. Two silica nanoparticles of 34 nm (A and B) with different zetapotentials were more cytotoxic than (aggregated) 11 and 248 nm nanoparticles, while cytokines were mostly induced by the (aggregated) 11 nm and only one of the 34 nm nanoparticles (34A). The results indicate that specific silica nanoparticles may have counterproductive effects, for example when used as carriers of anti-inflammatory drugs. The physicochemical properties determining the response of nanoparticles vary for different responses, implying that a screening approach for the safe development of nanoparticles needs to consider the role of combinations of (dynamic) physicochemical properties and needs to include multiple toxicity endpoints. Keywords Nanoparticles Á Silica Á Cytotoxicity Á Reactive oxygen species Á Inflammation Á Cytokines Á Environmental and health effects 2010; Roy et al. 2008). An interesting aspect of nanoparticles biodistribution is that they tend to accumulate at sites of inflammation due to increased vascular permeability (Moulari et al. 2008). In
Food-derived heterocyclic aromatic amines (HCAs) have proved to be carcinogenic in both rodents a... more Food-derived heterocyclic aromatic amines (HCAs) have proved to be carcinogenic in both rodents and nonhuman primates. Two different metabolic pathways are suggested for the metabolic activation of HCA. The hepatic pathway proceeds via a two-step process involving N-hydroxylation by cytochrome P4501A2 and subsequent O-acetylation by N-acetyltransferase-2. An alternative pathway may be of particular interest in extrahepatic tissues and proceeds via one-electron oxidation catalyzed by prostaglandin H synthase (PHS), rendering free-radical metabolites. In this study, we investigated the metabolic activation of two HCAs, 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), by two different enzyme systems in vitro, generating different primary and secondary reactive metabolites. Rat liver S9 mix and PHS were used as the activating system and represent the hepatic and extrahepatic pathways, respectively. Electron-spin resonance spectroscopy showed that both IQ and PhIP exerted inhibiting effects on PHS-mediated formation of hydroxyl radicals during the conversion of arachidonic acid to prostaglandins. Evidence for the formation of HCA free radicals was presented in an indirect way by the formation of glutathione-derived thiyl radicals, with purified PHS as the activating system. Activation by S9 mix did not result in the formation of detectable radical metabolites, showing that the two metabolic routes primarily led to the formation of different metabolites. In all electron-spin resonance experiments, IQ appeared to be more effective than PhIP. In contrasts, DNA adduct analysis by means of (32)P-postlabeling showed similar adduct patterns for S9 and PHS in single-stranded and double-stranded salmon testes DNA after incubation with PhIP, indicating the ultimate formation of a common reactive intermediate. For IQ, activation by PHS led to an additional adduct spot that was not present after S9 activation. Furthermore, activation of IQ resulted in higher adduct levels compared with PhIP for both activation pathways. Overall, adduct levels were higher in single-stranded DNA than double-stranded DNA. Our results showed that the hepatic and extrahepatic pathways resulted in different primary metabolites, while the ultimate formation of a similar reactive intermediate for PhIP, possibly an arylnitrenium ion, suggested that both pathways could play an important role in the onset of carcinogenesis.
Reactive oxygen species-induced oxidative stress in the colon is involved in inflammatory bowel d... more Reactive oxygen species-induced oxidative stress in the colon is involved in inflammatory bowel diseases and suggested to be associated with colorectal cancer risk. However, our insight in molecular responses to different oxygen radicals is still fragmentary. Therefore, we studied global gene expression by an extensive time series (0.08, 0.25, 0.5, 1, 2, 4, 8, 16, or 24 h) analyses in human colon cancer (caco-2) cells after exposure to H 2 O 2 or the superoxide anion donor menadione. Differences in gene expression were investigated by hybridization on two-color microarrays against nonexposed time-matched control cells. Next to gene expression, correlations with related phenotypic markers (8-oxodG levels and cell cycle arrest) were investigated. Gene expression analysis resulted in 1404 differentially expressed genes upon H 2 O 2 challenge and 979 genes after menadione treatment. Further analysis of gene expression data revealed how these oxidant responses can be discriminated. Time-dependent coregulated genes immediately showed a pulse-like response to H 2 O 2 , while the menadione-induced expression is not restored over 24 h. Pathway analyses demonstrated that H 2 O 2 immediately influences pathways involved in the immune function, while menadione constantly regulated cell cycle-related pathways Altogether, this study offers a novel and detailed insight in the similarities and differences of the time-dependent oxidative stress responses induced by the oxidants H 2 O 2 and menadione and show that these can be discriminated regarding their modulation of particular colon carcinogenesis-related mechanisms.
Journal of toxicology and environmental health, Feb 1, 2006
The mechanisms by which particulate matter (PM) produces adverse effects on the respiratory syste... more The mechanisms by which particulate matter (PM) produces adverse effects on the respiratory system, such as pulmonary dysfunction in children, are largely unknown. However, oxidative stress is thought to play an important role. Various chemical compounds in ambient particulate matter, including transition metals and aromatic organic compounds, may contribute to adverse effects through intrinsic generation of reactive oxygen species (ROS). It was hypothesized that ROS generation by PM, as determined through electron spin resonance (ESR) spectroscopy, may be negatively associated with pulmonary function in school children. PM(2.5), PM(10), and total suspended particulates (TSP) were sampled at the playgrounds of six elementary schools in the city of Maastricht, the Netherlands. All children (8-13 yr) from the six schools were asked to undergo spirometry. Multivariate linear regression models were constructed to evaluate associations between oxygen radical formation by PM and lung function. The radical-generating capacity per microgram PM correlated negatively to forced expiratory volume in 1 s (FEV(1)) and forced expiratory flow at 50% (FEF(50%)) of forced vital capacity (FVC). The data indicate that chemical features that contribute to intrinsic generation of ROS may be relevant for PM risk assessment.
Exposure to increased levels of ambient particulate matter (PM) are associated with several healt... more Exposure to increased levels of ambient particulate matter (PM) are associated with several health effects, including cardiopulmonary diseases. The formation of reactive oxygen species (ROS) is thought to play an important role in the induction of these health effects. To quantify the ROS generating capacity of PM, we developed an improved electron spin resonance (ESR) spectrometry-based method. ROS formation was measured directly on PMcontaining filters, thereby avoiding the selective extraction of components and loss of material or reactivity, which is likely to occur during filter extraction. Also, ascorbic acid was added to stimulate ROS formation. This method was applied to PM 10 samples originating from different sources. The radical generating capacity of PM 10 from both gasoline and diesel engine exhaust was significantly higher as compared to that of PM 10 from ambient or indoor air. Furthermore, in urban PM 10 and PM 2.5 , ROS-generating capacity significantly correlated with concentrations of polycyclic aromatic hydrocarbon content and particular transition metals. This indicates that this improved ESR method may be a valuable tool for evaluating the relationship between ROS formation by PM and the adverse health effects associated with this type of air pollution.
People interested in the research are advised to contact the author for the final version of the ... more People interested in the research are advised to contact the author for the final version of the publication, or visit the DOI to the publisher's website. • The final author version and the galley proof are versions of the publication after peer review. • The final published version features the final layout of the paper including the volume, issue and page numbers. Link to publication General rights Copyright and moral rights for the publications made accessible in the public portal are retained by the authors and/or other copyright owners and it is a condition of accessing publications that users recognise and abide by the legal requirements associated with these rights. • Users may download and print one copy of any publication from the public portal for the purpose of private study or research. • You may not further distribute the material or use it for any profit-making activity or commercial gain • You may freely distribute the URL identifying the publication in the public portal. If the publication is distributed under the terms of Article 25fa of the Dutch Copyright Act, indicated by the "Taverne" license above, please follow below link for the End User Agreement:
N-nitroso compounds (NOCs) may be implicated in human colon carcinogenesis, but the toxicological... more N-nitroso compounds (NOCs) may be implicated in human colon carcinogenesis, but the toxicological mechanisms involved have not been elucidated. Because it was previously demonstrated that nitrosamines and nitrosamides, representing two classes of NOC, induce distinct gene expression effects in colon cells that are particularly related to oxidative stress, we hypothesized that different radical mechanisms are involved. Using electron spin resonance spectroscopy, we investigated the radical-generating properties of genotoxic NOC concentrations in human colon adenocarcinoma cells (Caco-2). Cells were exposed to nitrosamides (N-methyl-N#-nitro-Nnitrosoguanidine and N-methyl-N-nitrosourea) or nitrosamines (Nnitrosodiethylamine, N-nitrosodimethylamine, N-nitrosopiperidine, and N-nitrosopyrrolidine). Nitrosamines caused formation of reactive oxygen species (ROS) and carbon-centered radicals, which was further stimulated in the presence of Caco-2 cells. N-methyl-Nnitrosourea exposure resulted in a small ROS signal, and formation of nitrogen-centered radicals (NCRs), also stimulated by Caco-2 cells. N-methyl-N#-nitro-N-nitrosoguanidine did not cause radical formation at genotoxic concentrations, but at increased exposure levels, both ROS and NCR formation was observed. By associating gene expression patterns with ROS formation, several cellular processes responding to nitrosamine exposure were identified, including apoptosis, cell cycle blockage, DNA repair, and oxidative stress. These findings suggest that following NOC exposure in Caco-2 cells, ROS formation plays an important role in deregulation of gene expression patterns that may be relevant for the process of chemical carcinogenesis in the human colon, in addition to the role of DNA alkylation.
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