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Visible-light responsive, stable, and abundant absorbers are required for the rapid integration of green, clean, and renewable technologies in a circular economy. Photoactive solid-solid heterojunctions enable multiple charge pathways, inhibiting recombination through efficient charge transfer across the interface. This study spotlights the physico-chemical synergy between titanium dioxide (TiO 2 ) anatase and carbon nitride (CN) to form a hybrid material. The CN(10%)-TiO 2 (90%) hybrid outperforms TiO 2 and CN references and literature homologs in four photo and photoelectrocatalytic reactions. CN-TiO 2 achieved a four-fold increase in benzylamine conversion, with photooxidation conversion rates of 51, 97, and 100 % at 625, 535, and 465 nm, respectively. The associated energy transfer mechanism was elucidated. In photoelectrochemistry, CN-TiO 2 exhibited 23 % photoactivity of the full-spectrum measurement when using a 410 nm filter. Our findings demonstrate that CN-TiO 2 displayed a band gap of 2.9 eV, evidencing TiO 2 photosensitization attributed to enhanced charge transfer at the heterointerface boundaries via staggered heterojunction type II., 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 © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Hemp ( Cannabis sativa L.) is an important source of fibre and seed oil and protein. By-products of industrial hemp fibre production, like hemp seeds and cakes, can be used as feed for all animal species as fat and protein source and the whole hemp plant (including stalk and leaves) might be a suitable fibre source for ruminants. However, a previous feeding experiment with leaf-flower-seed hemp silage, made from an industrial hemp variety, demonstrated detrimental effects on cow health parameters and a significant transfer of several cannabinoids, including the psychoactive tetrahydrocannabinol (∆9-THC), into cow's milk, posing a potential risk to the safety of consumers. Based on those observations, the present study tested the hypothesis that anaerobic fermentation, as during ensiling, increases the content of ∆9-THC in hemp. Therefore, silages of whole plants from the industrial hemp Cannabis sativa L. var. Ivory were prepared in a multifactorial design, with the four treatments 1) untreated control (CON), 2) addition of 10 mL per kg fresh weight homofermentative lactobacilli at 10 5 cfu/mL (LBAC), 3) addition of 10 mL per kg fresh weight homofermentative lactobacilli at 10 5 cfu/mL plus 30 g molasses (LBACmol) and 4) addition of propionic acid (10 mL/kg fresh weight) (PRO). Ultra high performance liquid chromatography coupled with tandem mass spectrometry with electrospray ionisation (UHPLC-MS/MS) was performed for analysis of cannabinoids in fresh hemp material and after 10 and 90 days of ensiling. The study revealed that ensiling decreased all acid forms of analysed cannabinoids in hemp at about 40-65% of the initial values after 90 days of storage, with the exception of cannabinolic acid (CBNA), and CBGA, the acidic form of cannabigerol (CBG). This decrease in most acidic forms was accompanied by an increase of the corresponding non-acidic forms of all cannabinoids, including the psychoactive ∆9-THC. Thus, although ensiling decreases the total cannabinoid content, psychoactive compounds like ∆9-THC can increase, enhancing the risk for animal health and a transfer of these substances into animal derived products.

The increasing spread of ragworts is observed with concern. Ragworts like tansy ragwort ( Jacobaea vulgaris Gaertn.) or marsh ragwort ( J. aquatica ) contain pyrrolizidine alkaloids (PA) which may induce hepatotoxic effects. Grazing animals usually avoid ragworts if their pasture management is appropriate. Preserved feed prepared from ragworts contaminated meadows may, however, lead to a significant exposure to PA. Previous studies on toxicity of PA for dairy cows revealed inconsistent results due to feeding ragwort plant material which was associated with heterogeneous PA exposure and thus failed to conclusively deduce critical PA doses. Therefore, the aim of the present study was to expose dairy cows ( n  = 4 per group) in a short-term scenario for 28 days with increasing PA doses (PA1: 0.47 mg PA/kg body weight (BW)/day (d); PA2: 0.95 mg PA/kg BW/d; PA3: 1.91 mg PA/kg BW/d) via oral administration by gavage of a defined PA-extract. While group PA3 was dosed with the PA-extract alone, groups PA2 and PA1 received PA-extracts blended in similar volumes with molasses to provide comparable amounts of sugar. Additionally, two control groups were treated either with water (CON Water ) or with molasses (CON Molasses ) to assess the effects of sugar without PA interference. While clinical traits including dry matter intake, milking performance, rectal body temperature, ruminal activity and body condition score (BCS) were not influenced by PA exposure, activities of enzymes indicative for liver damages, such as gamma-glutamyltransferase (GGT), aspartate aminotransferase (AST) and glutamate dehydrogenase (GLDH), increased significantly over time at an exposure of 1.91 mg total PA/kg BW/d.

This study focused on the investigation of cannabinoid profiles and contents of 23 different hemp teas and on the individual transfer of 16 cannabinoids from hemp teas into their tea infusions. The total cannabinoid content in the dry products averaged 14,960 mg kg -1 , with CBD&CBDA (sum of cannabidiol (CBD) and cannabidiolic acid (CBDA)) being the major component, accounting for 87% of the total cannabinoid content. The Δ9-tetrahydrocannabinol (Δ9-THC) content ranged from 16 mg kg -1 to 935 mg kg -1 and was on average 221 mg kg -1 . For each hemp tea, an infusion was prepared according to a standardized protocol issued by the German Standardisation body DIN and transfer rates per cannabinoid were estimated by comparing the contents in the dry material with the concentrations in the aqueous infusion. The limited water solubility of cannabinoids results in limited extraction efficiency for cannabinoids using boiling water to prepare a tea infusion and the average transfer rate of the psychoactive Δ9-THC was only 0.5%.

Retrorsine is a hepatotoxic pyrrolizidine alkaloid (PA) found in herbal supplements and medicines, food and livestock feed. Dose-response studies enabling the derivation of a point of departure including a benchmark dose for risk assessment of retrorsine in humans and animals are not available. Addressing this need, a physiologically based toxicokinetic (PBTK) model of retrorsine was developed for mouse and rat. Comprehensive characterization of retrorsine toxicokinetics revealed: both the fraction absorbed from the intestine (78%) and the fraction unbound in plasma (60%) are high, hepatic membrane permeation is dominated by active uptake and not by passive diffusion, liver metabolic clearance is 4-fold higher in rat compared to mouse and renal excretion contributes to 20% of the total clearance. The PBTK model was calibrated with kinetic data from available mouse and rat studies using maximum likelihood estimation. PBTK model evaluation showed convincing goodness-of-fit for hepatic retrorsine and retrorsine-derived DNA adducts. Furthermore, the developed model allowed to translate in vitro liver toxicity data of retrorsine to in vivo dose-response data. Resulting benchmark dose confidence intervals (mg/kg bodyweight) are 24.1-88.5 in mice and 79.9-104 in rats for acute liver toxicity after oral retrorsine intake. As the PBTK model was built to enable extrapolation to different species and other PA congeners, this integrative framework constitutes a flexible tool to address gaps in the risk assessment of PA., (© 2023. The Author(s).)

We present a rare reversible cause of bradycardia. A 49-year-old man who suffered from syncope was administered to our emergency department. In preclinical ECG recordings, a sinus node arrest was documented. All following examinations documented normal sinus node function. Finally, grayanotoxin poisoning, which can be present in honey from the Black Sea region, was proven. A pacemaker implantation could be avoided in this reversible cause of bradyarrhythmia., (© 2022. The Author(s), under exclusive licence to Springer Medizin Verlag GmbH, ein Teil von Springer Nature.)

Rumen metabolism of Senecio pyrrolizidine alkaloids (PAs) and their N -oxide forms was studied by mass spectrometry in in vitro batch culture incubates and confirmed in in vivo samples. Most N -oxides were found to undergo rapid conversion to their corresponding free bases, followed by biotransformation to metabolites hydrogenated at both the necine base and the necic acid moiety. Therefore, rumen metabolism can be considered a detoxification step, as saturated necine base structures are known as the platyphylline type, which is regarded as less or nontoxic. Individual Senecio PAs, such as jacoline, are metabolized slowly during rumen fermentation. PAs that showed limited biotransformation in the rumen in this study also showed limited transformation and CYP-mediated bioactivation in the liver in other studies. This could not only explain why PAs that are comparatively metabolically stable can pass into milk but also suggest that such PAs might be considered compounds of lesser concern.

CsPbBr 3 single crystals have potential for application in ionizing-radiation detection devices due to their optimal optoelectronic properties. Yet, their mixed ionic-electronic conductivity produces instability and hysteretic artifacts hindering the long-term device operation. Herein, we report an electrical characterization of CsPbBr 3 single crystals operating up to the time scale of hours. Our fast time-of-flight measurements reveal bulk mobilities of 13-26 cm 2 V -1 s -1 with a negative voltage bias dependency. By means of a guard ring (GR) configuration, we separate bulk and surface mobilities showing significant qualitative and quantitative transport differences. Our experiments of current transients and impedance spectroscopy indicate the formation of several regimes of space-charge-limited current (SCLC) associated with mechanisms similar to the Poole-Frenkel ionized-trap-assisted transport. We show that the ionic-SCLC seems to be an operational mode in this lead halide perovskite, despite the fact that experiments can be designed where the contribution of mobile ions to transport is negligible.

Pyrrolizidine alkaloid (PA) producing plants like Senecio jacobaea or Senecio vernalis are undesirable in fields for forage production, since PA are toxic to animals and humans. Previous studies have shown that ensiling can decrease the PA content in forages; however, no direct comparison of diverse PA from different Senecio spp. under various ensiling conditions has been made. Therefore, it was hypothesised that individual PA might react differently to ensiling, and silage inoculation with Lactobacillus will affect PA degradation because of a quick drop in pH, contrastingly to poor silage qualities resulting from contamination with soil. Laboratory scale grass silages were prepared in a multifactorial design with two levels of dry matter contents, four ensiling treatments and two storage durations (10 and 90 d). For each combination, four replicates were prepared individually. Ensiling treatments were (1) 10 ml water per kg fresh matter as control (CON), (2) 10 ml heterofermentative Lactobacillus buchneri strain LN4637 at 3 · 10 5 cfu/kg fresh matter plus 25 g molasses/kg fresh matter (LBHE), (3) 10 ml homofermentative lactobacilli at 3 · 10 5 cfu/kg fresh matter plus 25 g molasses/kg fresh matter (LBHO) and (4) 10 g soil/kg fresh matter (SOIL). Treatments affected formation of fermentation acids. Acetic acid was highest with treatment LBHE, and butyric acid was highest with treatment SOIL. All ensiling treatments effectively reduced total PA content by degrading the PA N -oxide (PANO) fraction. In parallel, though, the fraction of the tertiary base forms increased by around one-tenth of the original PANO content. Contents of jaconine and senkirkine were higher after ensiling than before, with regards to the sum of PA and PANO for jaconine, indicating higher stability or new formation through degradation of other PA. Overall, ensiling offers opportunities to decrease the PA-PANO content in feed and therefore lowers the risk of intoxication by Senecio in livestock.

Background: Extensively used grasslands are frequently utilised for hay production for equines. Especially, extensive meadows have a great variety of plant species, which may include plants that are poisonous for equines such as meadow saffron (Colchicum autumnale L.). To authors' knowledge investigations about horses` avoidance behaviour towards dried meadow saffron in hay are missing. Reports of farmers are contrary to clinical symptoms described in case reports and associated with meadow saffron in hay., Objectives: The aim of this study was to determine the rejection behaviour of horses for hay contaminated with meadow saffron (MS) when fed ad libitum., Study Design: An 18-day feeding trial with six adult geldings to observe the rejection behaviour for hay contaminated with MS., Methods: The horses were fed a basal diet containing hay ad libitum and a mineral supplement during the feeding trial. At six different daytimes, hay contaminated with 1% or 2% dried MS was provided to the horses over a duration of 1 h. The rejection behaviour was observed personally and by video recordings. If a horse ingested more than two plants of MS during one observation period, the observation was stopped and repeated at another day. When the observation period had to be stopped twice, the horse was excluded from the experiment., Results: Five of six horses ingested MS during the first feeding periods. One horse rejected leaves and capsules at the beginning of the study, but it showed repeated ingestion of MS after the seventh observation period., Main Limitations: Lack of knowledge about secondary plant metabolites affecting taste and their variability between fresh and dried plants., Conclusions: The intake of MS in hay by horses could not be ruled out with certainty. Therefore, feeding hay contaminated with MS should be avoided for equids., (© 2021 The Authors. Journal of Animal Physiology and Animal Nutrition published by Wiley-VCH GmbH.)

This paper reports on the major contributions and results of the 2nd International Workshop of Pyrrolizidine Alkaloids held in September 2020 in Kaiserslautern, Germany. Pyrrolizidine alkaloids are among the most relevant plant toxins contaminating food, feed, and medicinal products of plant origin. Hundreds of PA congeners with widespread occurrence are known, and thousands of plants are assumed to contain PAs. Due to certain PAs' pronounced liver toxicity and carcinogenicity, their occurrence in food, feed, and phytomedicines has raised serious human health concerns. This is particularly true for herbal teas, certain food supplements, honey, and certain phytomedicinal drugs. Due to the limited availability of animal data, broader use of in vitro data appears warranted to improve the risk assessment of a large number of relevant, 1,2-unsaturated PAs. This is true, for example, for the derivation of both toxicokinetic and toxicodynamic data. These efforts aim to understand better the modes of action, uptake, metabolism, elimination, toxicity, and genotoxicity of PAs to enable a detailed dose-response analysis and ultimately quantify differing toxic potencies between relevant PAs. Accordingly, risk-limiting measures comprising production, marketing, and regulation of food, feed, and medicinal products are discussed., Competing Interests: The authors declare that they have no conflict of interest., (Thieme. All rights reserved.)

Background: Senecio jacobaea contains pyrrolizidine alkaloids that can induce severe hepatic intoxication in horses, either acute when ingested in high amounts or chronic when consumed over a long period. The aim of this study was to determine horses' rejection behaviour towards the presence of Senecio jacobaea in hay when fed ad libitum. We hypothesized that adult horses can sort Senecio jacobaea out of the contaminated hay when hay is fed ad libitum. Six warmblood geldings with a mean (±SD) age of 15 ± 2 years were included. In a randomized study, Senecio jacobaea contaminated hay (5% or 10% contamination level) was provided at several timepoints over the day for 1 hour to six. Hay was provided ad libitum for the rest of the day. The horses' rejection behaviour towards Senecio jacobaea was observed. If a horse ingested two Senecio jacobaea plants twice at different timepoints, then the horse was excluded from the experiment., Results: Two out of six horses had to be excluded from the study after three out of 12 observation periods due to repeated Senecio jacobaea intake. Two other horses had to be excluded after nine and 11 out of 12 observation periods. Only two horses were able to sort out the various amounts (5 and 10% contamination level) of Senecio jacobaea during the whole experiment., Conclusions: Horses' intake of Senecio jacobaea cannot be avoided despite being fed with hay ad libitum. Due to the risk of chronic intoxication by pyrrolizidine alkaloids intake, feeding Senecio jacobaea contaminated hay must be avoided, and pastures with Senecio jacobaea growth are considered inappropriate for feed production., (© 2022. The Author(s).)

Scope: 1,2-unsaturated pyrrolizidine alkaloids (PAs) are secondary plant metabolites that are found in many plant species throughout the world. They are of concern for risk assessment as consumption of contaminated foodstuff can cause severe liver damage. Of late, transporter-mediated uptake and transport has advanced as a vital determinant of PA toxicity. In this study, the authors investigate a transporter-mediated uptake of PAs and its implications in PA toxicity., Methods and Results: We show that transporter expression levels are significantly affected by treatment with the PAs senecionine (Sc) and retrorsine (Re) in the human hepatoma cell line HepaRG. Furthermore, the specific contribution to PA uptake of the two transporters Na + /taurocholate co-transporting polypeptide (SLC10A1) and organic cation transporter I (SLC22A1), both belonging to the heterogeneous solute carrier super family, is investigated by means of a siRNA-mediated knockdown approach. Knockdown of both uptake transporters result in reduced uptake of Re and Sc in a time-dependent manner and attenuated PA-mediated cytotoxic effects in HepaRG cells., Conclusion: Our results confirm previous findings of active transport mechanisms of PAs into hepatocytes and highlight the importance of toxicokinetic studies for the risk assessment of PAs., (© 2021 Wiley-VCH GmbH.)

Contamination with 1,2-unsaturated pyrrolizidine alkaloids (PAs) is a serious problem for certain phytomedicines, foods, and animal feeds. Several of these PAs are genotoxic and carcinogenic, primarily in the liver, upon cytochrome P450 (CYP)-catalyzed activation into reactive (pyrrolic and pyrrole-like) metabolites. Here we investigated the metabolism of selected PAs (echimidine, europine, lasiocarpine, lycopsamine, retrorsine, and senecionine) in rat hepatocytes in primary culture and in human CYP3A4-transfected HepG2 cells. The open-chained diesters echimidine and lasiocarpine and the cyclic diester senecionine were extensively metabolized in rat hepatocytes into a broad spectrum of products released into the medium. A large portion of unidentified, possibly irreversibly bound, products remained in the cells while detectable amounts of reactive and other metabolites were found in the incubation media. In HepG2-CYP3A4 cells, lasiocarpine was more extensively metabolized than echimidine and senecionine which also gave rise to the release of pyrrolic metabolites. In human cells, no pyrrolic metabolites were detected in retrorsine or lycopsamine incubations, while no such metabolites were detected from europine in both cell types. Other types of metabolic changes comprised modifications such as side chain demethylation or oxygenation reactions like the formation of N -oxides. The latter, considered as a detoxification step, was a major pathway with cyclic diesters, was less distinctive for echimidine and lycopsamine and almost negligible for lasiocarpine and europine. Our data are in agreement with previously published cyto- and genotoxicity findings and suggests that the metabolic pattern may contribute substantially to the specific toxic potency of a certain congener. In addition, marked differences were found for certain congeners between rat hepatocytes and transfected human HepG2 cells, whereby a high level of bioactivation was found for lasiocarpine, whereas a very low level of bioactivation was observed for monoesters, in particular in human cells.

Pyrrolizidine alkaloids (PA) exert their toxic effects only after bioactivation. Although their toxicity has already been studied and metabolic pathways including important metabolites were described, the quantification of the latter revealed a large unknown portion of the metabolized PA. In this study, the qualitative and quantitative metabolite profiles of structurally different PAs in rat and human liver microsomes were investigated. Between five metabolites for europine and up to 48 metabolites for lasiocarpine were detected. Proposals for the chemical structure of each metabolite were derived based on fragmentation patterns using high-resolution mass spectrometry. The metabolite profiles of the diester PAs showed a relatively good agreement between both species. The metabolic reactions were summarized into three groups: dehydrogenation, oxygenation, and shortening of necic acid(s). While dehydrogenation of the necine base is considered as bioactivation, both other routes are considered as detoxification steps. The most abundant changes found for open chained diesters were dealkylations, while the major metabolic pathway for cyclic diesters was oxygenation especially at the nitrogen atom. In addition, all diester PAs formed several dehydrogenation products, via the insertion of a second double bond in the necine base, including the formation of glutathione conjugates. In rat liver microsomes, all investigated PAs formed dehydropyrrolizidine metabolites with the highest amount formed by lasiocarpine, whereas in human liver microsomes, these metabolites could only be detected for diesters. Our findings demonstrate that an extensive analysis of PA metabolism can provide the basis for a better understanding of PA toxicity and support future risk assessment.

Pyrrolizidine alkaloids (PAs) and their corresponding N -oxides (PANOs) have been determined in food and feed at levels relevant for consumer health. More than 660 different PAs have been detected, but few are available as reference substances for analytical demands. In the context of the European legislation on maximum levels of PAs in food products, a defined analytical scope of 21 PAs for determination has been suggested. An expansion of the scope from 21 to 35 PAs, including 14 structural isomers, is currently under discussion. In the present study, a target screening method was established for a comprehensive characterisation of PA profiles of the species Echium vulgare, Heliotropium europaeum, Cynoglossum officinale and Symphytum spp. to assess whether an expansion of the analytical scope is required to quantitatively cover the total PA contents of Boraginaceae species. The scope of the method comprised known and unknown PAs previously screened and confirmed in the respective plant extracts. A total of 176 PAs and PANOs were detected. The toxic 1,2-unsaturated PAs represent the predominant PA type with about 98% of the mean total content. This PA profiling demonstrates that an expansion of the scope from 21 to 35 PAs is required to adequately cover the mean total PA contents of Cynoglossum officinale and H. europaeum , whereas in the case of Symphytum spp. and Echium vulgare an expansion would not be necessary., Abbreviations: Pyrrolizidine alkaloid: PA, Pyrrolizidine alkaloid N -oxide: PANO, European Food Safety Authority: EFSA, German Federal Institute for Risk Assessment: BfR, ultra-high performance liquid chromatography: UHPLC, high-resolution: HR, tandem mass spectrometer: MS/MS, multiple reaction monitoring: MRM, data-independent MS2: ddms2, electrospray ionisation: ESI, limit of detection: LOD, limit of quantification: LOQ, gas chromatography coupled with mass spectrometry: GC-MS, fragmentation (data): MS2 (data), full scan: MS1, variable data-independent acquisition: vDIA, monoester esterified with a necic acid at position C9 of the necine base: O 9 -monoester, relative proportion: rel. prop., intraperitoneal: i.p., intravenous: i.v., higher-energy C-trap dissociation: HCD, all-ion fragmentation: AIF, parallel reaction monitoring: PRM.

Tansy ragwort, Jacobaea vulgaris Gaertn. (syn. Senecio jacobaea L.), is a common Asteraceae in Europe and Asia and known to be an invasive pest in several regions in the world. Recently it is also spreading immensely in native regions like Northern Germany. Pyrrolizidine alkaloids (PAs), which are found in high amounts in Jacobaea vulgaris, are toxic for humans and potentially lethal for grazing animals. In this study we investigated 27 populations of tansy ragwort in Northern Germany for their PA concentration and composition using liquid chromatography coupled to high resolution mass spectrometry. Furthermore, we investigated the genetic structure of selected populations using amplified length polymorphism markers. We detected 98 different PAs in the samples and considerable differences of PA composition between populations. In contrast, PA content of populations did not differ significantly. Genetic (4%) differentiation among populations was low while average genetic diversity was high (0.35). There was no correlation between genetic and geographic distance. Neither genetic markers nor chemical composition revealed any connection to the geographic pattern. As we could not detect any pattern in genetic or chemical diversity, we suggest that the existence of this diversity is a result of a broad interaction with the environment rather than that of evolutionary constraints in the current selection process driving PA composition in J. vulgaris in certain chemotypes., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Among naturally occurring plant constituents, the 1,2-unsaturated pyrrolizidine alkaloids (in the following termed 'PAs') play a distinct role because of the large number of congeners occurring in nature and the pronounced toxicity of some congeners. Several PAs are hepatotoxic in humans, experimental and farm animals and were shown to be potent hepatocarcinogens in laboratory rodents. Although the general mode of action leading to toxicity has been elucidated, i.e., being mediated by metabolic conversion of the parent molecule into a highly reactive electrophile capable of attacking cellular target molecules, major questions related to the risk assessment of PAs remain unresolved. It was the aim of a workshop held in September 2018 to shed more light on the occurrence, exposure, mode of action, toxicokinetics and -dynamics of PAs to improve the scientific basis for an advanced toxicological risk assessment. The contributions in nine chapters describe the scientific progress using advanced analytical methods, studies in subcellular fractions, cell culture, experimental animals and humans and the use of PBPK modeling and structure-activity relationship considerations aiming at a better understanding of PA toxicity and genotoxicity. Since PAs differ considerably in their toxic potencies and substantial species differences in sensitivity towards PA exposure exist, a special emphasis was placed on these issues., 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 © 2020 Elsevier Ltd. All rights reserved.)

Pyrrolizidine alkaloid (PA) forming plants are found worldwide and may contaminate food products at levels being of concern for human health. Due to the high biodiversity of PA producing plants many different types of PA structures are formed. PAs themselves are not toxic but require metabolic activation to exert toxicity. To investigate if the structure of the PAs affects their in vitro metabolism, we incubated a set of 22 PAs and compared the degradation rates and the amount of formed glutathione (GSH) conjugates. With human liver microsomes, no metabolic degradation of monoesters was found. Degradation rates of diester PAs tended to correlate with their hydrophilicity, whereby the more polar and branched-chained PAs exhibited lower degradation. There was a trend towards higher degradation rates in the presence of rat liver microsomes, but the GSH conjugate levels were similar. Although an effective degradation seems to be related with high GSH conjugate levels, no clear correlation between both parameters could be deduced. For both species no GSH conjugates, or only trace amounts, were formed from monoesters. However, for both open-chained as well as cyclic diesters GSH conjugates were detected and determined levels were comparable for both ester types without major structure-dependent differences., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Pyrrolizidine alkaloids (PA) are widely distributed phytotoxins contaminating food and feed. Hepatic enzymes are considered to bioactivate PA. Previous studies showed differences in the metabolism rate in liver homogenates of different species. Thus, uncertainty remains with respect to the relevance of human metabolism. Our study aimed to analyze whether the PA representative lasiocarpine is toxified by human cytochrome P450 (CYP) enzymes. We compared the metabolic elimination of lasiocarpine in the presence of rat and human S9 fractions and liver microsomes. Experiments with the potent CYP3A/Cyp3a inhibitor ketoconazole and supersomes containing individual human and rat CYPs revealed that enzymes of the CYP3A/Cyp3a family of both species are of major relevance for lasiocarpine metabolism. To assess if metabolism by human CYP3A4 results in a toxification of lasiocarpine we performed experiments with V79 cells. γH2AX and micronucleus formation were analyzed as endpoints for genotoxicity. No effects were observed in the wildtype cells, which lack CYP activity. By contrast, a V79 clone engineered for expression of human CYP3A4 showed concentration-dependent γH2AX and micronucleus formation. Concluding, our results showed the CYP3A4-dependent formation of genotoxic metabolites of lasiocarpine. The results confirm previous data indicating the need to include metabolism of PA for human risk assessment., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Background: Data from the "Corinthia" study, an observational, cross-sectional survey of 2,043 participants were analyzed. Subjects were classified into 4 groups: group A: systolic pressure (SBP) 120-129 and diastolic pressure (DBP) < 80 mmHg, group B: SBP 130-139 or DBP 80-89 mmHg, group B1: SBP 130-139 and DBP < 80 mmHg and group B2: SBP 130-139 and DBP 80-89 mmHg. Aortic stiffness was assessed by carotid-to-femoral pulse wave velocity (PWV). A value of PWV > 10m/s was consider indicative of asymptomatic organ damage while values of PWV exceeded the 90 % percentile for each age group were consider as abnormal. Patients and methods. Groups B, B1 and B2 have significantly increased PWV compared to group A, independently from age and other risk factors (PWV: 9.2 ± 2.8 vs 9.4 ± 2.7 vs 8.6 ± 2.5 vs 8.1 ± 2.3 m/s, p < 0.01, respectively). The prevalence of PWV > 10 m/s and abnormal PWV values in group A was significantly lower than the corresponding prevalence in randomly selected, age-matched subjects from group B (13.5 % vs 24.4 %, p = 0.027 and 5.6 % vs 14.2 %, p = 0.022, respectively). Results: The reclassified subjects as stage 1 hypertensive by the new guidelines have a significantly increased aortic stiffness and greater prevalence in asymptomatic aortic damage compared to subjects with elevated BP. This finding may indirectly explain the increased cardiovascular risk of this group.Conclusions: The reclassified subjects as stage 1 hypertensive by the new guidelines have a significantly increased aortic stiffness and greater prevalence in asymptomatic aortic damage compared to subjects with elevated BP. This finding may indirectly explain the increased cardiovascular risk of this group.

Objective: The purpose of the present study was the analysis of the association between consumption of candy and fruit gums, diagnosis of attention deficit hyperactivity disorder (ADHD), and behavioural problems. Methods: In total, 1,187 children and adolescents of the German Health Interview and Examination Survey for Children and Adolescents (KiGGS) were analyzed. Results: Because with the present design no conclusions on causality or directionality of the found associations could be drawn, results are discussed quite broadly in the light of several previously published interpretations, also to serve as a generator for further research. One more innovative speculation is that children and adolescents with ADHD may consume more frequently candy and fruit gums in order i) to compensate for their higher needs of energy resulting from hyperactive behaviour and/or ii) to compensate for the ADHD-typical deficits in the "reward cascade".Conclusions: Because with the present design no conclusions on causality or directionality of the found associations could be drawn, results are discussed quite broadly in the light of several previously published interpretations, also to serve as a generator for further research. One more innovative speculation is that children and adolescents with ADHD may consume more frequently candy and fruit gums in order i) to compensate for their higher needs of energy resulting from hyperactive behaviour and/or ii) to compensate for the ADHD-typical deficits in the "reward cascade".

Background: Isolated superior mesenteric artery dissection (ISMAD) is rare, especially when associated with intestinal ischaemia. We report our clinical experience managing this condition., Patients and Methods: Medical records from 22 patients with ISMAD and intestinal ischaemia were retrospectively analysed. Conservative treatment was given to all patients as first line therapy. Subsequently, 15 patients received endovascular stent placement and three patients received endovascular stent placement plus intestinal resection and anastomosis., Results: After conservative treatment, the symptoms of three patients were remarkably relieved; however, a repeat contrast CT showed that stenosis was aggravated. Hence, endovascular stent placement was performed in all 15 patients. Enteral nutrition was successfully restored in 12 patients. Three patients showed signs of chronic intestinal ischaemia, including peritonitis and ileus. These patients underwent intestinal resection and anastomosis. Enteral nutrition was restored at postoperative week two. No signs of intestinal ischaemia recurred during two-years of follow-up., Conclusions: We recommend endovascular stent placement as a feasible, effective, and minimally invasive procedure in patients with ISMAD and symptoms of intestinal ischaemia.

Background: Vascular access site-related complications are frequent in the context of transfemoral transcatheter aortic valve replacement (TAVR). The implantation of a covered stent graft is an effective treatment option for bleeding control. However, the external iliac and common femoral arteries are exposed to flexion of the hip joint. Therefore, stent compression and stent/strut fractures may occur, facilitating stent occlusion., Patients and Methods: In all 389 patients who received transfemoral TAVR from 2013-2015 at the Düsseldorf Heart Centre, we monitored the management of vascular access site-related complications. Our analyses focused on immediate technical success and bleeding control, primary patency, and the occurrence of stent/strut fractures after six to 12 months of follow-up., Results: Vascular access site-related complications occurred in 13 % (n = 51), whereof in 10 patients, the bleeding was successfully managed by prolonged compression. In 40 out of 51 patients, a covered stent graft was implanted in the common femoral artery, leading to 100 % immediate bleeding control. After a mean follow-up of 334 ± 188 days, 28 stents out of 29 patients with completed follow-up (excluding e. g. death) were without flow-limiting stenosis (primary patency 97 %) or relevant stent compression (diameter pre/post 8.6/8.1 mm, p = 0.048, late lumen loss 1.1 ± 0.2 mm, mean flow velocity 92 ± 34 cm/s). In four asymptomatic patients, stent/strut fractures were detected (14 %) without flow-limiting stenosis., Conclusions: The implantation of a covered stent graft is highly effective and safe to control vascular access site-related complications after TAVR. Stent/strut fractures in the flexible segment of the common femoral artery may occur, as consequently verified by X-ray visualization, but show no impairment on flow or clinical parameters after six to 12 months.

Background: Microarray analysis has been carried out in this pilot study to compare delineated gene expression profiles in the biopsies of skeletal muscle taken from patients with chronic critical limb ischaemia (CLI) and non-ischaemic control subjects., Patients and Methods: Biopsy of gastrocnemius muscle was obtained from six patients with unreconstructed CLI referred for surgical major amputation. As control, biopsies of six patients undergoing elective knee arthroplasty without evidence of peripheral arterial occlusive disease were taken. The differences in gene expression associated with angiogenic processes in specimens obtained from ischaemic and non-ischaemic skeletal muscle were confirmed by quantitative real-time polymerase chain reaction (PCR) analysis., Results: Compared with non-ischaemic skeletal muscle biopsy of chronic-ischaemic skeletal muscle contained 55 significantly up-regulated and 45 down-regulated genes, out of which 64 genes had a known genetic product. Tissue samples of ischaemic muscle were characterized by increased expression of cell survival factors (e. g. tissue factor pathway inhibitor 2) in combination with reduced expression of cell proliferation effectors (e. g. microfibrillar-associated protein 5 and transferrin receptor). The expression of growth factors (e. g. early growth response 3 and chemokine receptor chemokine C-X-C motif ligand 4) which play a central role in arterial and angiogenic processes and anti-angiogenetic factors (e. g. pentraxin 3) were increased in chronic ischaemic skeletal muscle. An increased expression of extracellular matrix proteins (e. g. cysteine-rich angiogenic inducer 61) was also observed., Conclusions: Gene expression profiles in biopsies of gastrocnemius muscle in patients with chronic critical limb ischaemia showed an increase in pro-survival factors, extracellular matrix protein deposition, and impaired proliferation, compared with non-ischaemic controls. Further studies are required to analyse the endogenous repair mechanism.

Background: Genome-wide association studies identified several gene variants associated with peripheral arterial disease (PAD). Among them, rs6584389 A>C was significantly associated with PAD defined by decreased ankle-brachial index (ABI). The aim of this study was to investigate whether the rs6584389 variant is also associated with the earlier stages of atherosclerosis assessed by intima-media thickness (IMT) or pulse-wave velocity (PWV) in clinically asymptomatic subjects with type 2 diabetes (T2DM), a group of patients with a high cardiovascular risk., Patients and Methods: In total, 111 patients with T2DM (56 females, 55 males) with a mean age 63.0 ± 9.1 years were consecutively included in the study. IMT was measured by ultrasound using 7 MHz linear transducer. PWV was measured using a piezoelectric method. Genotyping for rs6584389 was performed by PCR-HRMA method., Results: The carriers of the risk C-allele of rs6584389 variant had significantly higher mean left-side IMT (AA: 0.67 ± 0.12, AC 0.77 ± 0.21, CC 0.78 ± 0.22 mm; p = 0.04). In multiple linear regression analysis, rs6586389 genotype was significantly associated with all measured IMT parameters. The presence of each risk C-allele predicted an increase in left-side IMT by 0.056 mm (p = 0.017), right-side IMT by 0.053 mm (p = 0.039), average IMT by 0.054 mm (p = 0.023), and maximal IMT by 0.058 mm (p = 0.021). Age and HbA1c levels were also significantly associated with increased IMT in all multivariate models., Conclusions: Gene variant rs6584389 A>C near to PAX2 gene was associated with increased carotid IMT in patients with type 2 diabetes independently of the other main risk factors for atherosclerosis.

Background: The aim of this study was to investigate whether patients undergoing elective endovascular aneurysm repair (EVAR) with loco-regional anaesthetic techniques have better outcomes than those treated with general anaesthesia (GA)., Patients and Methods: We retrospectively evaluated outcomes of EVAR performed with regional anaesthesia (RA) or GA over a five-year period. Furthermore, we searched electronic bibliographic sources (MEDLINE, EMBASE, CINAHL, and CENTRAL) to identify studies comparing different anaesthetic methods in EVAR. We defined perioperative mortality and morbidity as well as length of hospital stay (LOS) as the primary outcome measures. Pooled effect estimates were calculated using fixed-effect or random-effects models. Results are reported as the odds ratio (OR) or mean difference (MD) and 95 % confidence interval (CI)., Results: Three hundred and fifty-five patients underwent standard EVAR over the study period (RA, 215 patients; GA 140 patients). Patients in both groups had comparable baseline demographics and clinical characteristics. Perioperative mortality was significantly lower in the RA group (0.5 % vs. 4.3 %, P = 0.017). No difference was found in perioperative morbidity (P = 0.370), LOS (P = 0.146), postoperative destination (P = 0.799), reoperation (P = 0.355) or readmission within 30 days (P = 0.846). Meta-analysis of data on 15,472 patients from 15 observational studies found a significantly lower perioperative mortality (OR 0.70, 95 % CI 0.52-0.95, P = 0.02) and morbidity (OR 0.73, 95 % CI 0.55-0.96, P = 0.02) in patients treated with loco-regional anaesthetic techniques compared to those treated with GA. Our sub-group analysis demonstrated that both local anaesthesia (LA) (P = 0.003) and RA (P < 0.0001) were associated with a significantly shorter LOS compared to GA., Conclusions: Local and/or regional anaesthetic techniques may be advantageous over GA in elective EVAR, as indicated by reduced perioperative mortality and morbidity and a shorter hospital stay. Considering the current level of evidence, LA or RA should be considered in selected patients. Further clinical research is required to provide high level evidence on the optimal anaesthetic technique in EVAR.

Pyrrolizidine alkaloids (PA) are secondary metabolites of certain flowering plants. The ingestion of PAs may result in acute and chronic effects in man and livestock with hepatotoxicity, mutagenicity, and carcinogenicity being identified as predominant effects. Several hundred PAs sharing the diol pyrrolizidine as a core structure are formed by plants. Although many congeners may cause adverse effects, differences in the toxic potency have been detected in animal tests. It is generally accepted that PAs themselves are biologically and toxicologically inactive and require metabolic activation. Consequently, a strong relationship between activating metabolism and toxicity can be expected. Concerning PA susceptibility, marked differences between species were reported with a comparatively high susceptibility in horses, while goat and sheep seem to be almost resistant. Therefore, we investigated the in vitro degradation rate of four frequently occurring PAs by liver enzymes present in S9 fractions from human, pig, cow, horse, rat, rabbit, goat, and sheep liver. Unexpectedly, almost no metabolic degradation of any PA was observed for susceptible species such as human, pig, horse, or cow. If the formation of toxic metabolites represents a crucial bioactivation step, the found inverse conversion rates of PAs compared to the known susceptibility require further investigation.

Background: Osteoprotegerin plays a role in the development of several bone diseases. In addition, osteoprotegerin may contribute to the development of vascular disease. Little is known about the association between serum osteoprotegerin levels and the presence or severity of peripheral arterial disease (PAD). The aim of this study was to examine the association between serum osteoprotegerin levels and both the presence as well as the severity of lower extremity arterial disease in patients with type 2 diabetes (T2DM)., Patients and Methods: The study included 165 consecutive patients with T2DM (57 % males, mean age 65.0 ± 0.7 years). PAD was diagnosed by measurement of the toe-brachial index (TBI). Serum osteoprotegerin was measured using ELISA., Results: The mean osteoprotegerin level was significantly higher in patients with PAD in comparison to patients without PAD (18.2 ± 1.0 vs. 13.1 ± 2.0 pmol/L, p = 0.014). Significant univariate correlations between TBI and osteoprotegerin level (r = -0.308; p < 0.001), age, body mass index, and HDL cholesterol were observed. In the multivariate linear regression analysis, serum osteoprotegerin (β = -0.005; p = 0.020), higher age, and male gender were significant predictors of TBI. When 25(OH) vitamin D was introduced into the mentioned model, OPG was no longer a significant predictor of TBI and was replaced in the model with vitamin D (β = 0.009, p = 0.001). This finding suggests a role of OPG as a mediator of the effects of 25(OH) vitamin D., Conclusions: Serum osteoprotegerin level is significantly associated with both the presence and severity of PAD in patients with T2D. Osteoprotegerin might be a biomarker for the presence of atherosclerotic disease in patients with T2DM.

Background: Inflammation is associated with an increased risk of thrombotic events and complete blood count (CBC) is an easily measured test. The purpose of this study was to evaluate the value of CBC relative parameters including mean platelet volume (MPV), platelet-to-lymphocyte ratio (PLR), mean platelet volume-to-lymphocyte ratio (MPVLR), and neutrophil-to-lymphocyte ratio (NLR) for patients with acute deep vein thrombosis (DVT)., Patients and Methods: A total of 115 patients with unprovoked DVT of the lower extremities and 105 controls were recruited in this study. Blood samples were drawn from all participants to obtain the concentrations of CBCs and D-dimers., Results: MPVs (P = 0.044), PLRs (P = 0.005), MPVLRs (P = 0.001), and NLRs (P < 0.0001) were significantly higher in acute DVT patients compared to controls. The MPV was inversely correlated with platelet count (P < 0.0001) and the NLR was positively associated with D-dimers (P = 0.002) and the PLR (P < 0.0001). Notably, on multivariate logistic regression analysis, NLRs and D-dimers were independent risk factors of acute DVT (OR: 1.889, P = 0.024; OR: 1.009, P < 0.0001, respectively)., Conclusions: MPV, PLR, MPVLR, and NLR have potential diagnostic values for patients with unprovoked DVT. NLR is an independent risk factor related to DVT.

Posterior nutcracker syndrome (PNCS) is the entrapment of the left renal vein between the aorta and the vertebral column. Although uncommon, it is still an important diagnosis due to the high morbidity associated with the risk of secondary anaemia from haematuria, from long-term left renal vein hypertension, vascular thrombosis, and even blood clots in the urinary system. A literature search of PubMed and EMBASE databases was performed and 27 publications containing 27 cases were included for the final analysis. The following frequency of clinical signs and symptoms was noted: twenty-five patients had haematuria, 13 patients had flank pain, and two had hypertension. Overall, male-female distribution was balanced and there were more adult than paediatric (age < 18 years) patients. All symptoms of patients with conservative treatment were either well-controlled or under spontaneous resolution. Conservative management instead of surgical treatment should be preferred in most cases. Taken together, despite the low incidence of PNCS, its recognition and management are highly important. This systematic study explores the evidence base for conservative and medical options.

Objective: A considerable number of adolescents exhibit severe self-regulation deficits in affect and behavior, which are referred to as affective dysregulation (AD). AD may be conceptualized as a dimensional trait that, in its extreme form, resembles the diagnostic categories of severe mood dysregulation (SMD) or disruptive mood dysregulation disorder (DMDD). Assuming a shared pathway of psychopathology in AD and SMD, similar underlying dysfunctional mechanisms in emotion processing, particularly emotion recognition (RECOG) and regulation (REGUL), may be postulated., Method: Adolescent inpatients with AD (CAD, N = 35), without AD (CCG, N = 28), and nonclinical controls (NCG; N = 28) were administered a morphed facial recognition task (RECOG). REGUL abilities, levels of irritability as well as depressive symptoms were also assessed., Results: We found no significant group differences in accuracy and thresholds for RECOG abilities. Patients with AD reported more dysfunctional REGUL strategies than did CCG and NCG. Both depression and AD, but not irritability, influenced the overall degree of maladaptive REGUL., Conclusion: The broad phenotype of AD does not involve the deficits in RECOG reported for adolescents with a narrow phenotype (SMD); regarding REGUL strategies, AD seems to be associated with specific impairments.

Background: Endovascular aortic repair (EVAR) has emerged as standard of care for abdominal aortic aneurysm (AAA). Real-world evidence is limited to compare this technology to open repair (OAR). Major gaps exist related to short-term and long-term outcomes, particularly in respect of gender differences., Materials and Methods: Health insurance claims data from Germany's third largest insurance provider, DAK-Gesundheit, was used to investigate invasive in-hospital treatment of intact (iAAA) and ruptured AAA (rAAA). Patients operated between October 2008 and April 2015 were included in the study., Results: A total of 5,509 patients (4,966 iAAA and 543 rAAA) underwent EVAR or OAR with a median follow-up of 2.44 years. Baseline demographics, comorbidities, and clinical characteristics of DAK-G patients were assessed. In total, 84.6 % of the iAAA and 79.9 % of the rAAA were male. Concerning iAAA repair, the median age (74 vs. 73 years, p < .001) compared to men was higher in females, but their EVAR-rate (66.8 % vs. 71.1 %, p = .018) was lower. Besides higher age of female patients (80 vs. 75 years, p < .001), no further statistically significant differences were seen following rAAA repair. In-hospital mortality was slightly lower in males compared to females following iAAA (2.3 % vs. 3.1 %, p = .159) and rAAA (37.3 % vs. 43.1 %, p = .273) repair. Concerning iAAA repair, a higher rate of female patients was transferred to another hospital (3.7 % vs. 2.0 %, p = 0.008) or discharged to rehabilitation (6.0 % vs. 2.7 %, p < .001) compared to male patients., Conclusions: In this large German claims data cohort, women are generally older and more often transferred to another hospital or discharged to rehab following iAAA repair. Nonetheless, no significantly increased risk of in-hospital or late death appeared for women in multivariate analyses. Further studies are necessary to evaluate the impact of recent gender-specific treatment strategies on overall outcome under real-world settings.

Pyrrolizidine alkaloids (PAs) are secondary metabolites of plant families such as Asteraceae or Boraginaceae and are suspected to be genotoxic carcinogens. Recent investigations revealed their frequent occurrence in honey and particularly in tea. To obtain a comprehensive overview of the PA content in animal- and plant-derived food from the European market, and to provide a basis for future risk analysis, a total of 1105 samples were collected in 2014 and 2015. These comprised milk and milk products, eggs, meat and meat products, (herbal) teas, and (herbal) food supplements collected in supermarkets, retail shops, and via the internet. PAs were detected in a large proportion of plant-derived foods: 91% of the (herbal) teas and 60% of the food supplements contained at least one individual PA. All types of (herbal) teas investigated were found to contain PAs, with a mean concentration of 460 µg kg -1 dry tea (corresponding to 6.13 µg L -1 in [herbal] tea infusion). The highest mean concentrations were found in rooibos tea (599 µg kg -1 dry tea, 7.99 µg L -1 tea infusion) and the lowest in camomile tea (274 µg kg -1 dry tea, 3.65 µg L -1 tea infusion). Occurrence of PAs in food supplements was found to be highly variable, but in comparable ranges as for (herbal) tea. The highest concentrations were present in supplements containing plant material from known PA-producing plants. In contrast, only 2% of the animal-derived products, in particular 6% of milk samples and 1% of egg samples, contained PAs. Determined levels in milk were relatively low, ranged between 0.05 and 0.17 µg L -1 and only trace amounts of 0.10-0.12 µg kg -1 were found in eggs. No PAs were detected in the other animal-derived products.

Background: We performed a meta-analysis to assess the presence of an annual rhythmic variability of ruptured abdominal aortic aneurysm (RAAA) onset., Patients and Methods: Eligible studies were observational studies enrolling patients with RAAA and reporting monthly incidence of RAAA. Study-specific estimates, i. e. monthly incidence of RAAA, were combined using the random-effects model. Chronobiological analysis was performed by applying a partial Fourier series to pooled monthly incidence by using the weighted least-squares method., Results: We identified 14 eligible studies enrolling a total of 3,798 patients with RAAA. Pooled monthly incidence of RAAA was 8.7 % in January, 7.7 % in February, 8.7 % in March, 7.3 % in April, 7.8 % in May, 7.2 % in June, 7.0 % in July, 7.0 % in August, 8.1 % in September, 8.8 % in October, 8.4 % in November, and 8.3 % in December. Chronobiological analysis identified a significant (p = 0.0020) annual pattern in the occurrence of RAAA with a peak in December to January and a nadir in June to July. Pooled analysis demonstrated significantly more incidence in December than in June (p = 0.03) as well as in January than in July (p = 0.05)., Conclusions: A significant annual pattern in the occurrence of RAAA with a peak in December to January and a nadir in June to July was identified with significantly more incidence in December than in June and in January than in July.

Background: This study reports the mid-term prevalence and therapeutic consequences of anastomotic leaks after surgery for Stanford type A aortic dissections., Patients and Methods: From July 2007 to July 2013, 93 patients survived surgery for acute type A dissections at our center and underwent a standardized follow-up. The pre-, peri-, and postoperative as well as the midterm results were collected prospectively. Follow-up computed tomography (CT) imaging was performed 7 days, 3, and 12 months after surgery, and yearly thereafter, to assess the presence or progression of anastomotic leaks at the aorto-prosthesis anastomotic sites., Results: The mean follow-up was 4 years (1534 ± 724 days). Follow-up CT revealed anastomotic leaks in 4 patients (4.3 %). All leaks developed during midterm follow-up and half of them did not increase with time. Two patients required redo surgery for an increase in periaortic extravasation and compression of neighboring structures. Further analysis was not able to reveal independent risk factors for development or deterioration of leaks., Conclusions: Anastomotic leaks after surgery for Stanford Type A aortic dissection can develop in midterm follow-up, even after initially excellent results. Meticulous follow-up is mandatory to detect possible deterioration and a need for redo surgery.

Background: The OPG/RANKL/RANK (osteoprotegerin/receptor-activator of nuclear factor κB ligand/receptor-activator of nuclear factor κB) axis has been recently linked to the development of atherosclerosis and plaque destabilization. We have investigated whether polymorphism rs2073618 of the OPG gene is associated with subclinical markers of carotid atherosclerosis in subjects with type 2 diabetes mellitus (T2DM)., Patients and Methods: 595 subjects with T2DM were enrolled in the cross-sectional study. Subclinical markers of carotid atherosclerosis (carotid intima media thickness, plaque thickness, and plaques presence) were assessed with ultrasound at the time of recruitment. Genotyping for rs2073618 (a missense variant located in exon I of the OPG gene) was performed, and OPG serum levels were determined by ELISA., Results: Compared to the GG genotype, the CC genotype of the rs2073618 polymorphism had a significantly increased risk for the presence of carotid plaque (OR = 2.54, 95 % CI = 1.22-5.28, p = 0.01). No statistically significant difference could be detected (p = 0.68) upon comparing median values of serum OPG levels among studied genotype groups in subjects with T2DM. Multivariable linear regression analyses in T2DM subjects demonstrated that GC and CC genotypes (p = 0.03 and p = 0.003), together with statin therapy (p = 0.009), were independent predictors of the number of carotid segments with plaques., Conclusions: Despite the fact that OPG rs2073618 genotypes failed to predict the serum OPG levels as there was no statistical difference among compared genotypes, our results demonstrate that the rs2073618 polymorphism could be a possible genetic marker for the prediction of increased risk for carotid plaque burden as a measure of advanced subclinical atherosclerosis in T2DM subjects.

Background: Carotid intima-media thickness (CIMT), an ultrasonographic marker of cardiovascular risk, is increasingly used in adults and children. The choice of specific images used to quantify CIMT from a cine sequence is often based on image quality rather than on a consistent point in the cardiac cycle. This methodological study quantified the imprecision that may be introduced by variation of CIMT during the cardiac cycle., Probands and Methods: Data from four-year-olds, 11 to 12-year-olds, and adults (n=30 each age group) were selected retrospectively from two population-derived studies. Far wall CIMT of the right common carotid artery was measured at end-diastole and peak systole using standardized protocols. All images were analysed using semi-automated edge-detection software., Results: In all age groups CIMT varied significantly during the cardiac cycle and was largest at end-diastole. The mean difference in CIMT between end-diastole and peak systole was greater in four-year-olds (38 μm; 95 % confidence interval (CI) 33 to 43 μm) and 11 to 12-year-olds (31 μm; CI 26 to 36 μm) than in adults (18 μm; CI 16 to 22 μm). Carotid IMT increased by 8.8 % (CI 7.7 to 9.8 %), 6.9 % (CI 5.8 to 8.1 %), and 3.8 % (CI 3.1 to 4.5 %) between minimum and maximum arterial diameter in four-year-olds, 11 to 12-year-olds, and adults, respectively. The greatest variation in CIMT during the cardiac cycle was observed in children (up to 14 %)., Conclusions: Inconsistent timing of CIMT measurement during the cardiac cycle is an avoidable source of imprecision, especially in children, in whom inter-individual differences are smallest. As CIMT is largest at end-diastole, this is the most appropriate time point for consistent and comparable measurements to be made.

Background: Liver involvement is a common manifestation of hereditary haemorrhagic telangiectasia (HHT). Although a number of studies have been carried out in adult patients, no study has ever been focused on investigating HHT-related hepatic involvement in paediatric patients. The present study aimed for the first time to systematically estimate the prevalence of HHT-associated liver involvement and to characterize HHT-associated hepatic angiodynamic features in paediatric age., Patients and Methods: The study was designed as a cross-sectional survey in an HHT paediatric cohort, subclassified as HHT1 and HHT2 according to the mutated gene. The evaluation of the angiodynamic profile was performed by duplex ultrasound examination. Investigation by multi-slice computed tomography (MSCT) or magnetic resonance angiography (MRA) was performed in patients >12 years., Results: MSCT/MRA examination disclosed silent hepatic involvement in 7/20 (35.0 %) children, and nodular regenerative hyperplasia in two cases. Diameter of common hepatic artery was significantly larger in HHT2 (0.45 ± 0.15 cm) compared to HHT1 (0.33 ± 0.09, p < 0.01) and control children (0.32 ± 0.08, p < 0.05). None of the patients had clinical manifestations of liver involvement. Angiodynamic profiles were different between paediatric and adult HHT patients., Conclusions: Liver involvement can be detected in paediatric HHT patients, albeit with a lower frequency compared to adults. Paediatric HHT2 children show a higher frequency of liver involvement and a trend to hepatic artery dilation when compared to HHT1 children.

Background: Cardiovascular diseases are the leading cause of death in Germany. The knowledge of causal risk factors and their distribution is of utmost importance to design screening programs., Probands and Methods: In this cross-sectional study design we used STROBE criteria to achieve the highest comparability possible. Anthropometric measures (height and weight), total cholesterol, glucose level, and blood pressure were measured. Probands' history was collected by using a standardized questionnaire. The data was age- and gender-adjusted for the working population 16 to 70 years of age, derived from the micro census, the 1 %-sample census of the German statistical office. For each study year weight factors were calculated. Logistic regression analysis was conducted regarding the cardiovascular risk factors: smoking, arterial hypertension, diabetes, hypercholesterolemia, and obesity., Results: Between 2006 and 2015 a total of 28,293 employees took part in the ongoing company screenings. The mean age was 42.3 years for both sexes (median: 43 years). The mean body mass index (BMI) was 25.6 kg/m 2 (men: 26.5 kg/m 2 , women: 24.7 kg/m 2 ). A history of hypertension was present in 16 % of the employees (men: 17.8 %, women: 13.8 %). Of the respondents 2 % suffered from diabetes (men: 2.4 %, women: 1.6 %). Lipid-lowering drugs were taken by 2.8 % of all employees (3.6 % men and 1.9 % women). 23.3 % of the men and women indicated to be active smokers. In the regression analysis obesity was associated with a four times higher risk of hypertension and a three times higher risk of elevated glucose levels, thus manifesting as main contributor for vascular diseases. Meanwhile the risk for obesity was 140 % higher in probands who are former smokers., Conclusions: We regard obesity as the number one cardiovascular risk which should be assessed by various medical, legislative, and socio-economic actions to limit future mortality and health-care costs in Germany.

Background: The beneficial effect of statin therapy on the progress of atherosclerotic disease has been demonstrated by numerous studies. Vascular strain imaging is an arising method to evaluate arterial stiffness. Our study examined whether an influence of statin therapy on the vessel wall could be detected by vascular strain imaging., Patients and Methods: 88 patients with recently detected atherosclerosis underwent an angiological examination including ankle-brachial index (ABI), pulse wave index (PWI), central puls ewave velocity and duplex ultrasound. Captures for vascular strain analysis were taken in B-mode during ultrasound examination of the common carotid artery and evaluated using a workstation equipped with a speckle tracking based software. A statin therapy was recommended and after six months a follow-up examination took place. Meanwhile, the non-adherence of a group of patients (N = 18) lead to a possibility to observe statin effects on the vascular strain., Results: In the statin non-adherent group the ABI decreased significantly to a still non-pathological level (1.2 ± 0.2 vs. 1.0 ± 0.2; p = 0.016) whereas it stagnated in the adherent group (1.0 ± 0.2 vs. 1.0 ± 0.2; p = 0.383). The PWI did not differ in the non-adherent group (180.5 ± 71.9 vs. 164.4 ± 75.8; p = 0.436) but under statin therapy it decreased significantly (261.8 ± 238.6 vs. 196.4 ± 137.4; p = 0.016). In comparison to the adherent group (4.2 ± 2.0 vs. 4.0 ± 1.8; p = 0.548) under statin therapy the radial strain decreased significantly in the non-adherent group (4.7 ± 2.0 vs. 3.3 ± 1.1; p = 0.014)., Conclusions: Our findings reveal a beneficial influence of statin therapy on the arterial wall detected by vascular strain analysis.

Background: Pseudoxanthoma elasticum (PXE) is an autosomal recessive inherited multisystem disorder of the connective tissue caused by a loss-of-function mutation of the ABCC6 gene. It can affect the cardiovascular system, presumably leading to a high prevalence of atherosclerosis., Patients and Methods: 46 PXE patients and 18 controls underwent an angiological examination consisting of measurement of ankle-brachial index (ABI), strain-gauge arterial reserve (SGAR), arterial resting perfusion, pulse wave index (PWI), central pulse wave velocity, and ultrasound examination., Results: With an average age of 51.4 ± 12.4 years, 35/46 (76.1 %) of the PXE patients had atherosclerotic lesions, and 10 of them (28.6 %) had a chronic vascular occlusion of one or more peripheral vessels. 34/46 (73.9 %) had a pathologic ABI < 0.9, 15/42 (35.7 %) had a pathological SGAR < 10 mL/100 mL tissue/min, and 23/38 (60.5 %) had a pathological PWI > 180. The differences between the groups were statistically significant for ABI, arterial reserve, and PWI., Conclusions: In PXE patients atherosclerosis was found with a much higher prevalence than expected. Moreover, they were at very high risk for total vessel occlusions.
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Background: Individual shear rate therapy (ISRT) has been designed as a novel non-invasive treatment option for peripheral artery disease (PAD) patients and has been shown to improve endothelial function and walking distance. The aim of this study was to elucidate the impact of ISRT on the level of nitric oxide in patient blood plasma and the expression of related molecular markers in peripheral blood mononuclear cells (PBMCs). Molecular diagnostic tests were performed for two ISRT trials., Patients and Methods: In ISRT-1 26 healthy subjects underwent one session of treadmill training and one session of ISRT respectively in a cross-over design. In ISRT-2 14 PAD patients with a stable intermittent claudication underwent a 30 hours long-term treatment. Plasma nitrite release as well as the mRNA expression of NOS2 and key regulators of the kallikrein-kinin system were measured in PBMCs at different time points., Results: Short-term ISRT revealed significantly decreased NOS2 expression in PBMCs of healthy volunteers and PAD patients. Long-term ISRT, in turn, demonstrated a significant plasma nitrite increase in PAD patients., Conclusions: We verified that long-term ISRT stimulates the vascular system and exerts a comparable effect to physical exercise in regards to NO release, which coincide with recent findings regarding an improvement of endothelial function. However, further studies are necessary to investigate the role for circulating leukocytes.
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The ingestion of seafood contaminated with the marine biotoxin okadaic acid (OA) can lead to diarrhetic shellfish poisoning with symptoms like nausea, vomiting and abdominal cramps. Both rat and the human hepatic cytochrome P450 monooxygenases (CYP) metabolize OA. However, liver cell toxicity of metabolized OA is mainly unclear. The aim of our study was to detect the cellular effects in HepG2 cells exposed to OA in the presence of recombinant CYP enzymes of both rat and human for the investigation of species differences. The results should be set in correlation with a CYP-specific metabolite pattern. Comparative metabolite profiles of OA after incubation in rat and human recombinant CYP enzymes were established by using LC-MS/MS technique. Results demonstrated that metabolism of OA to oxygenated metabolites correlates with detoxification which was mainly catalyzed by human CYP3A4 and CYP3A5. Detoxification by rat Cyp3a1 was lower compared to human CYP3A enzymes and activation of OA by Cyp3a2 was observed, coincident with minor overall conversion capacity of OA. By contrast human and rat CYP1A2 seem to activate OA into cytotoxic intermediates. In conclusion, different mechanisms of OA metabolism may occur in the liver. At low OA doses, the human liver is likely well protected against cytotoxic OA, but for high shellfish consumers a potential risk cannot be excluded.

High-resolution mass spectrometry (HRMS) was applied for the detection of grayanotoxins (GrTx) in a contaminated honey sample. This sample was provided by a hospital due to a suspicion of intoxication after a patient had shown the typical symptoms of GrTx poisoning. Subsequent analysis proved the contamination with high amounts of GrTx and other toxins belonging to grayanane-type diterpenoids. This group of natural toxins is synthesised by the plant family Ericaceae and comprises more than 60 individual toxins, but only one compound is available as a reference standard. We applied a screening approach that easily confirms the presence or absence of GrTx without access to standards. By searching for predictable mass spectrometric fragment ions, including typical in-source fragments arising from collision-induced dissociation during electrospray ionisation, the complete toxin profile was screened and allowed the mass spectrometric identification of 15 individual GrTx. The potential of this approach is especially demonstrated by the fact that at least two of these toxins have not been previously described in the literature. A semi-quantitative estimation indicated a total toxin concentration of 358 mg kg(-1). An investigation of 49 honeys from the German retail market did not reveal the presence of GrTx.

Scope: 1,2-Unsaturated pyrrolizidine alkaloids (PA) are found in plants such as Asteraceae and Boraginaceae families. Acute PA poisoning via contaminated food or feed causes severe damage to liver depending on species-specific oral bioavailability. For assessing PA bioavailability, their passage across the intestinal barrier was investigated using Caco-2 cells., Methods: Differentiated Caco-2 cells were exposed in transport chambers to the PA heliotrine (Hn), echimidine (Em), senecionine (Sc), and senkirkine (Sk). Cell supernatants were analyzed by LC-MS/MS., Results: PA pass Caco-2 monolayer from the apical into basolateral compartment depending on their chemical structure. Compared to the cyclic diesters Sc and Sk with a passage rate of 47% ± 4 and 40% ± 3, respectively, the transferred amount of the monoester Hn (32% ± 3) and open-chained diester Em (13% ± 2) was substantially lower. This suggested an active transport of Hn and Em. Using Madin-Darby canine kidney II/P-glycoprotein (ABCB1)-overexpressing cells, the active excretion of Hn and Em by ABCB1 from the gastrointestinal epithelium into the gut lumen was shown., Conclusion: PA cross the intestinal barrier structure-dependently. The passage of the noncyclic PA Hn and Em is reduced by an ABCB1-driven efflux into the gastrointestinal lumen resulting in a decreased oral bioavailability., (© 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The consumption of okadaic acid (OA) contaminated shellfish can induce acute toxic symptoms in humans such as diarrhea, nausea, vomiting and abdominal pain; carcinogenic and embryotoxic effects have also been described. Toxicokinetic studies with mice have shown that high cytotoxic doses of OA can pass the gastrointestinal barrier presumably by paracellular passage. However, in vitro studies using human intestinal Caco-2 cell monolayers to represent the intestinal barrier have shown that at low-dose exposure OA is transported against a concentration gradient suggesting an active efflux mechanism. Since P-glycoprotein (P-gp) transports a wide variety of substrates, we investigated its possible influence on the observed elimination of OA. We used two different cellular transwell models: (i) Caco-2 cell monolayer endogenously expressing human P-gp and simulating the intestinal barrier and (ii) MDCK-II cell monolayer stably over-expressing P-gp. Our study demonstrates clearly that OA at non-cytotoxic concentrations passes the monolayer barrier only to a low degree, and that it is actively eliminated by P-gp over the apical membrane. Therefore, our in vitro data indicate that humans appear to have efficient defense mechanisms to protect themselves against low-dose contaminated shellfish by exhibiting a low bioavailability as a result of active elimination of OA by P-gp., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

Honey was previously considered to be one of the main food sources of human pyrrolizidine alkaloid (PA) exposure in Europe. However, comprehensive analyses of honey and tea sampled in the Berlin retail market revealed unexpected high PA amounts in teas. This study comprised the analysis of 87 honey as well as 274 tea samples including black, green, rooibos, melissa, peppermint, chamomile, fennel, nettle, and mixed herbal tea or fruit tea. Total PA concentrations in tea ranged from < LOD to 5647 µg kg(-1), while a mean value of about 10 µg kg(-1) was found in honey samples. Additionally, herbal drugs were investigated to identify the source of PA in teas. Results suggest that PA in tea samples are most likely a contamination caused by co-harvesting of PA-producing plants. In some cases such as fennel, anise or caraway, it cannot be excluded that these plants are able to produce PA themselves.

In tandem mass spectrometry the multiple reaction monitoring (MRM) mode is normally used for targeted analysis but this mode also has the potential to screen for structural similarities of analytes. On the basis of the fact that in general similar molecular structures result in similar fragments or losses of neutrals, this approach was used for pyrrolizidine alkaloid (PA) screening but could also be easily adapted to screen for other compound classes. PA are plant toxins of which several hundred individual compounds have been identified. Our MRM screening approach uses the structural relation and similar core structure of all PA which results in a common and thus predictable mass spectrometric fragmentation behaviour. On this basis a method was developed which screens for PA structures by MRM transitions and allows the detection of each individual PA down to a low microgram per kilogram concentration range. The approach was applied to investigate plants from the families of Asteraceae (several species of Senecio and Eupatorium), Boraginaceae (Echium, Cynoglossum, Borago and Anchusa officinalis as well as Heliotropium europaeum) and Fabaceae (Crotalaria incana) for a complete qualitative and quantitative PA characterisation. All analytes that were detected as possible PA by MRM screening were further investigated by recording product ion spectra. Analytes which exhibited a typical PA fragmentation pattern were either confirmed as PA or otherwise deleted as false positive signals (false positive rate was below 10 %). Sum formulas of confirmed PA were determined by additional measurements applying high resolution mass spectrometry. In that way 121 unknown PA were identified and for the first time complete PA profiles of different PA plants were delivered.