Toxicological Characterization of the Inorganic and Organic Arsenic Metabolite Thio-DMAV in Cultured Human Lung Cells

We synthesised and toxicologically characterised the arsenic metabolite thiodimethylarsinic acid (thio-DMAV). Successful synthesis of highly pure thio-DMAV was confirmed by state-of-the-art analytical techniques including 1H-NMR, HPLC-FTMS, and HPLC-ICPMS. Toxicological characterization was carried out in comparison to arsenite and its well-known trivalent and pentavalent methylated metabolites. It comprised cellular bioavailability as well as different cytotoxicity and genotoxicity end points in cultured human A549 lung cells. Of all arsenicals investigated, thio-DMAV exerted the strongest cytotoxicity. Moreover, thio-DMAV did not induce DNA strand breaks and an increased induction of both micronuclei and multinucleated cells occurred only at beginning cytotoxic concentrations, indicating that thio-DMAV does not act via a genotoxic mode of action. Finally, to assess potential implications of thio-DMAV for human health, further mechanistic studies are urgently necessary to identify the toxic mode of action of this highly toxic, unusual pentavalent organic arsenical

Molecular and functional changes in neutrophilic granulocytes induced by nicotine: a systematic review and critical evaluation

Background: Over 1.1 billion people smoke worldwide. The alkaloid nicotine is a prominent and addictive component of tobacco. In addition to tumors and cardiovascular disorders, tobacco consumption is associated with a variety of chronic-inflammatory diseases. Although neutrophilic granulocytes (neutrophils) play a role in the pathogenesis of many of these diseases, the impact of nicotine on neutrophils has not been systematically reviewed so far. Objectives: The aim of this systematic review was to evaluate the direct influence of nicotine on human neutrophil functions, specifically on cell death/damage, apoptosis, chemotaxis, general motility, adhesion molecule expression, eicosanoid synthesis, cytokine/chemokine expression, formation of neutrophil extracellular traps (NETs), phagocytosis, generation of reactive oxygen species (ROS), net antimicrobial activity, and enzyme release. Material and methods: This review was conducted according to the PRISMA guidelines. A literature search was performed in the databases NCBI Pubmed (R) and Web of Science (TM) in February 2023. Inclusion criteria comprised English written research articles, showing in vitro studies on the direct impact of nicotine on specified human neutrophil functions. Results: Of the 532 originally identified articles, data from 34 articles were finally compiled after several evaluation steps. The considered studies highly varied in methodological aspects. While at high concentrations (>3 mmol/l) nicotine started to be cytotoxic to neutrophils, concentrations typically achieved in blood of smokers (in the nmol/l range) applied for long exposure times (24-72h) supported the survival of neutrophils. Smoking-relevant nicotine concentrations also increased the chemotaxis of neutrophils towards several chemoattractants, elevated their production of elastase, lipocalin-2, CXCL8, leukotriene B4 and prostaglandin E2, and reduced their integrin expression. Moreover, while nicotine impaired the neutrophil phagocytotic and anti-microbial activity, a range of studies demonstrated increased NET formation. However, conflicting effects were found on ROS generation, selectin expression and release of beta-glucuronidase and myeloperoxidase. Conclusion: Nicotine seems to support the presence in the tissue and the inflammatory and selected tissue-damaging activity of neutrophils and reduces their antimicrobial functions, suggesting a direct contribution of nicotine to the pathogenesis of chronic-inflammatory diseases via influencing the neutrophil biology

Molecular and functional changes in neutrophilic granulocytes induced by nicotine: a systematic review and critical evaluation

BackgroundOver 1.1 billion people smoke worldwide. The alkaloid nicotine is a prominent and addictive component of tobacco. In addition to tumors and cardiovascular disorders, tobacco consumption is associated with a variety of chronic-inflammatory diseases. Although neutrophilic granulocytes (neutrophils) play a role in the pathogenesis of many of these diseases, the impact of nicotine on neutrophils has not been systematically reviewed so far.ObjectivesThe aim of this systematic review was to evaluate the direct influence of nicotine on human neutrophil functions, specifically on cell death/damage, apoptosis, chemotaxis, general motility, adhesion molecule expression, eicosanoid synthesis, cytokine/chemokine expression, formation of neutrophil extracellular traps (NETs), phagocytosis, generation of reactive oxygen species (ROS), net antimicrobial activity, and enzyme release.Material and methodsThis review was conducted according to the PRISMA guidelines. A literature search was performed in the databases NCBI Pubmed® and Web of Science™ in February 2023. Inclusion criteria comprised English written research articles, showing in vitro studies on the direct impact of nicotine on specified human neutrophil functions.ResultsOf the 532 originally identified articles, data from 34 articles were finally compiled after several evaluation steps. The considered studies highly varied in methodological aspects. While at high concentrations (>3 mmol/l) nicotine started to be cytotoxic to neutrophils, concentrations typically achieved in blood of smokers (in the nmol/l range) applied for long exposure times (24-72h) supported the survival of neutrophils. Smoking-relevant nicotine concentrations also increased the chemotaxis of neutrophils towards several chemoattractants, elevated their production of elastase, lipocalin-2, CXCL8, leukotriene B4 and prostaglandin E2, and reduced their integrin expression. Moreover, while nicotine impaired the neutrophil phagocytotic and anti-microbial activity, a range of studies demonstrated increased NET formation. However, conflicting effects were found on ROS generation, selectin expression and release of β-glucuronidase and myeloperoxidase.ConclusionNicotine seems to support the presence in the tissue and the inflammatory and selected tissue-damaging activity of neutrophils and reduces their antimicrobial functions, suggesting a direct contribution of nicotine to the pathogenesis of chronic-inflammatory diseases via influencing the neutrophil biology

Structural annotation of electro- and photochemically generated transformation products of moxidectin using high-resolution mass spectrometry

Moxidectin (MOX) is a widely used anthelmintic drug for the treatment of internal and external parasites in food-producing and companion animals. Transformation products (TPs) of MOX, formed through metabolic degradation or acid hydrolysis, may pose a potential environmental risk, but only few were identified so far. In this study, we therefore systematically characterized electro- and photochemically generated MOX TPs using high-resolution mass spectrometry (HRMS). Oxidative electrochemical (EC) TPs were generated in an electrochemical reactor and photochemical (PC) TPs by irradiation with UV-C light. Subsequent HRMS measurements were performed to identify accurate masses and deduce occurring modification reactions of derived TPs in a suspected target analysis. In total, 26 EC TPs and 59 PC TPs were found. The main modification reactions were hydroxylation, (de-)hydration, and derivative formation with methanol for EC experiments and isomeric changes, (de-)hydration, and changes at the methoxime moiety for PC experiments. In addition, several combinations of different modification reactions were identified. For 17 TPs, we could predict chemical structures through interpretation of acquired MS/MS data. Most modifications could be linked to two specific regions of MOX. Some previously described metabolic reactions like hydroxylation or O-demethylation were confirmed in our EC and PC experiments as reaction type, but the corresponding TPs were not identical to known metabolites or degradation products. The obtained knowledge regarding novel TPs and reactions will aid to elucidate the degradation pathway of MOX which is currently unknown

Kombinationswirkungen umweltrelevanter Metallverbindungen in Lungenzellen

This project aims to investigate the genotoxicity of environmentally relevant particulate nickel and cadmium compounds in human lung cells as primary targets of metal-induced carcinogenicity. Special focus will be given to the question whether direct DNA damage or inhibition of DNA repair processes causing enhancing effects with other DNA damaging agents are predominant. In a first step, a sensitive procedure to quantify DNA adducts of benzo[a]pyrene (B[a]P) was established. This method is based on HPLC with fluorescence detection and allows the specific and reproducible quantification of 1 DNA-adduct of benzo[a]pyrene diol epoxide (BPDE) per 108 base pairs in cultured cells. By this approach, we demonstrated the dose-dependent generation of DNA lesions and their repair in HeLa and A549 lung cells. The removal of DNA adducts of BPDE was inhibited by both NiCl2 and NiO in the non-cytotoxic concentration range. The results obtained so far indicate that the repair inhibition by Ni(II) observed previously also applies for lung cells and for both soluble and insoluble nickel compounds. Zusammenfassung Gesamtziel des Vorhabens ist die Abklärung der Genotoxizität von partikulären, umweltrelevanten Nickel- und Cadmiumverbindungen in menschlichen Lungenzellen als Zielzellen der metallinduzierten Kanzerogenese. Insbesondere soll abgeklärt werden, ob primäre gentoxische Effekte oder DNA-Reparaturinhibitionen im Vordergrund stehen, die in der Folge zu Wirkungsverstärkungen in Kombination mit anderen DNA-schädigenden Agentien führen. Hierfür wurde zunächst ein auf HPLC mit Fluoreszenzdetektion basierendes Testsystem zum empfindlichen Nachweis von DNA-Schäden des Benzo[a]pyrens (B[a]P) etabliert, welches nunmehr den spezifischen und reproduzierbaren Nachweis von 1 DNA-Addukt des Benzo[a]pyren 7,8-diol 9,10-epoxids (BPDE) pro 108 Basenpaare in Kulturzellen ermöglicht. Mit Hilfe dieses Testsystems wurde die dosisabhängige Induktion der DNA-Schäden und ihre Reparatur in HeLa- und A549-Zellen gezeigt. Die Entfernung der DNA-Läsionen wurde sowohl durch NiCl2 als auch durch NiO in nicht-cytotoxischen Konzentrationen gehemmt. Die bislang erhaltenen Ergebnisse weisen darauf hin, daß die in Vorarbeiten gezeigten Reparaturinhibitionen durch NiCl2 auch in Lungenzellen zu beobachten sind und daß sowohl wasserlösliche als auch weitgehend wasserunlösliche Nickelverbindungen inhibitorisch auf die Nucleotid-Exzisionsreparatur wirken

Toxicological Characterization of the Inorganic and Organic Arsenic Metabolite Thio-DMAV in Cultured Human Lung Cells

We synthesised and toxicologically characterised the arsenic metabolite thiodimethylarsinic acid (thio-DMAV). Successful synthesis of highly pure thio-DMAV was confirmed by state-of-the-art analytical techniques including 1H-NMR, HPLC-FTMS, and HPLC-ICPMS. Toxicological characterization was carried out in comparison to arsenite and its well-known trivalent and pentavalent methylated metabolites. It comprised cellular bioavailability as well as different cytotoxicity and genotoxicity end points in cultured human A549 lung cells. Of all arsenicals investigated, thio-DMAV exerted the strongest cytotoxicity. Moreover, thio-DMAV did not induce DNA strand breaks and an increased induction of both micronuclei and multinucleated cells occurred only at beginning cytotoxic concentrations, indicating that thio-DMAV does not act via a genotoxic mode of action. Finally, to assess potential implications of thio-DMAV for human health, further mechanistic studies are urgently necessary to identify the toxic mode of action of this highly toxic, unusual pentavalent organic arsenical.

Spezies der KFZ-emittierten Platingruppenelemente (PGE) und ihre toxische Wirkung

Vergleichsweise hohe Platingruppenelement-Emissionen aus Autokatalysatoren sind in den letzten Jahren entlang von Autobahnen und in Städten nachgewiesen worden, über deren Toxizität bislang nur wenig bekannt ist. Ziel der Untersuchungen im Rahmen dieses Projektes ist die Abklärung der Bioverfügbarkeit sowie eines möglichen gentoxischen Potentials von Platin-, Palladium- und Rhodiumverbindungen in Säugerzellen. Im Rahmen dieser Vorstudie sollten anhand von metallischen Platin- und Palladiumverbindungen zunächst die geplanten Methoden auf ihre Anwendbarkeit für diese Fragestellung geprüft und erste Abschätzungen über die Bioverfügbarkeit der PGE getroffen werden. Dabei wurden zunächst die Größe und physikalischen Eigenschaften der eingesetzten Partikel durch Rasterelektronenmikroskopie und Röntgendiffraktometrie bestimmt. Ihre Phagozytose durch A549 Lungenzellen konnte qualitativ durch Lichtmikroskopie nachgewiesen werden. Schließlich gelang es, eine Methode zum quantitativen Nachweis von Platin- und Palladium-induzierten DNA-Addukten mit Hilfe der HR-ICP-MS zu etablieren und es konnte in ersten Versuchen gezeigt werden, daß die Exposition von A549 Lungenzellen gegenüber metallischen Palladium- und Platinpartikeln zu einer deutlich meßbaren DNA-Adduktbildung führt. Somit muß von einer Bioverfügbarkeit der Partikel ausgegangen werden. Summary During the last years, comparatively high concentrations of platinum group elements derived from automobile catalysts have been detected along high-traffic streets and in cities. Nevertheless, only little is known about their toxic potentials. Aim of this project is to investigate the bioavailability and potential genotoxic effects of platinum, palladium and rhodium compounds in mammalian cells in culture. By applying metallic platinum and palladium particles, we examined the suitability of different methods to estimate their bioavailabilty. The physical characteristics of the particles were evaluated by electron microscopy and X-ray diffractometry. Phagocytosis of the particles by A549 lung cells was demonstrated by light microscopy. Finally, a procedure was established to quantitate platinum- and palladium-induced DNA adducts by HR-ICP-MS. In preliminary experiments, there was a dose-dependent induction of DNA adducts after exposure of A549 cells towards both types of particles, indicating their bioavailability

N-Acetylcysteine as modulator of the essential trace elements copper and zinc

N-acetylcysteine (NAC) is a frequently prescribed drug and known for its metal chelating capability. However, to date it is not well characterized whether NAC intake affects the homeostasis of essential trace elements. As a precursor of glutathione (GSH), NAC also has the potential to modulate the cellular redox homeostasis. Thus, we aimed to analyze effects of acute and chronic NAC treatment on the homeostasis of copper (Cu) and zinc (Zn) and on the activity of the redox-sensitive transcription factor Nrf2. Cells were exposed to 1 mM NAC and were co-treated with 50 µM Cu or Zn. We showed that NAC treatment reduced the cellular concentration of Zn and Cu. In addition, NAC inhibited the Zn-induced Nrf2 activation and limited the concomitant upregulation of cellular GSH concentrations. In contrast, mice chronically received NAC via drinking water (1 g NAC/100 mL). Cu and Zn concentrations were decreased in liver and spleen. In the duodenum, NQO1, TXNRD, and SOD activities were upregulated by NAC. All of them can be induced by Nrf2, thus indicating a putative Nrf2 activation. Overall, NAC modulates the homeostasis of Cu and Zn both in vitro and in vivo and accordingly affects the cellular redox balance