Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 32
Filtrar
1.
Exp Dermatol ; 33(3): e15044, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38465766

RESUMO

Polycyclic aromatic hydrocarbons with the key substance benzo[a]pyrene (B[a]P) are widespread pollutants in the environment and at working places. Nonetheless, the exact underlying mechanisms of toxicological effects caused by B[a]P especially in absence and presence of UV irradiation remain uncertain. This study examines variations in exposure conditions: low B[a]P (4 nM), low B[a]P + UV and high B[a]P (4 µM), selected based on pertinent cytotoxicity assessments. Following cell viability evaluations post-treatment with varied B[a]P concentrations and UV irradiation, the identified concentrations underwent detailed metabolomic analysis via gas chromatography-mass spectrometry. Subsequently, resulting changes in metabolic profiles across these distinct exposure groups are comprehensively compared. Chemometric analyses showed modest regulation of metabolites after low B[a]P exposure compared to control conditions. High B[a]P and low B[a]P + UV exposure significantly increased regulation of metabolic pathways, indicating that additional UV irradiation plus low B[a]P is as demanding for the cells as higher B[a]P treatment alone. Further analysis revealed exposure-dependent regulation of glutathione-important for oxidative defence-and purine metabolism-important for DNA base synthesis. Only after low B[a]P, oxidative defence appeared to be able to compensate for B[a]P-induced perturbations of the oxidative homeostasis. In contrast, purine metabolism already responded towards adversity at low B[a]P. The metabolomic results give an insight into the mechanisms leading to the toxic response and confirm the strong effects of co-exposure on oxidative defence and DNA repair in the model studied.


Assuntos
Benzo(a)pireno , Hidrocarbonetos Policíclicos Aromáticos , Benzo(a)pireno/toxicidade , Benzo(a)pireno/metabolismo , Queratinócitos/metabolismo , Raios Ultravioleta , Glutationa/metabolismo , Purinas/farmacologia
2.
Exp Dermatol ; 31(5): 700-714, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-35030266

RESUMO

The skin's ability to function optimally is affected by many diverse factors. Metabolomics has a great potential to improve our understanding of the underlying metabolic changes and the affected pathways. Therefore, the objective of this study was to review the current state of the literature and to perform further metabolic pathway analysis on the obtained data. The aim was to gain an overview of the metabolic changes under altered conditions and to identify common and different patterns as a function of the investigated factors. A cross-study comparison of the extracted studies from different databases identified 364 metabolites, whose concentrations were considerably altered by the following factor groups: irradiation, xenobiotics, ageing and skin diseases (mainly psoriasis). Using metabolic databases and pathway analysis tools, the individual metabolites were assigned to the corresponding metabolic pathways and the most strongly affected signalling pathways were identified. All factors induced oxidative stress. Thus, antioxidant defence systems, especially coenzyme Q10  (ageing) and the glutathione system (irradiation, ageing, xenobiotics), were impacted. Lipid metabolism was also impacted by all factors studied. The carnitine shuttle as part of ß-oxidation was activated by all factor groups except ageing. Glycolysis, Krebs (TCA) cycle and purine metabolism were mainly affected by irradiation and xenobiotics. The pentose phosphate pathway was activated, and Krebs cycle was downregulated in response to oxidative stress. In summary, it can be ascertained that mainly energy metabolism, lipid metabolism, antioxidative defence and DNA repair systems were impacted by the factors studied.


Assuntos
Metabolômica , Xenobióticos , Ciclo do Ácido Cítrico , Metabolismo Energético , Via de Pentose Fosfato/fisiologia
3.
J Dtsch Dermatol Ges ; 20(9): 1179-1186, 2022 09.
Artigo em Inglês | MEDLINE | ID: mdl-36075872

RESUMO

Squamous cell carcinomas (SCC) of the skin can be induced by occupational exposures to polycyclic aromatic hydrocarbons (PAHs), as in tar and soot, or to UV radiation and can be recognized and compensated as occupational diseases. A possible syncarcinogenic effect of these exposures in the development of SCC in humans is under discussion. For the scientific validation of this question, a systematic literature search was conducted using the databases PubMed, Web of Science, and Scopus. Studies on individuals with SCC of the skin and their precursors as well as occupational, non-occupational, or therapeutic exposure to UV radiation and PAHs were selected. In addition, animal studies with exposure to UV radiation and PAHs were evaluated. After screening the abstracts of 510 identified studies, the full texts of 131 studies were reviewed. None of the epidemiological studies provided robust evidence for a syncarcinogenesis of PAHs and UV radiation in the development of SCC of the skin in humans. Nevertheless, as there are indications for a (super-)additive effect of UV radiation and PAH exposure from animal studies and mechanistic investigations, syncarcinogenesis seems possible. However, quantitative dose-response relationships are lacking which would allow comparison of the onset of an adverse effect between the different exposure levels.


Assuntos
Carcinoma de Células Escamosas , Exposição Ocupacional , Hidrocarbonetos Policíclicos Aromáticos , Animais , Carcinoma de Células Escamosas/etiologia , Humanos , Exposição Ocupacional/análise , Hidrocarbonetos Policíclicos Aromáticos/toxicidade , Pele , Fuligem , Raios Ultravioleta/efeitos adversos
4.
Arch Toxicol ; 95(6): 2007-2018, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33772346

RESUMO

Toxicological studies propose that exposure to carbon black nanoparticles induces organ injuries and inflammatory responses. Besides, current understanding of the molecular mechanisms implies that carbon black nanoparticles (CBNP) exposure induces the production of reactive oxygen species (ROS) causing inflammation, mitochondrial dysfunction or disturbance in calcium homeostasis. However, the precise mechanisms whereby CBNP exert these effects in the lung are still not fully understood. To gain insight into the possible mechanism of CBNP exerted toxicity, human alveolar epithelial cells (A549) were exposed to different concentrations of CBNP and for different timepoints. The reaction of the cells was monitored by the systematic use of cell-based measurements of calcium and ROS, in the presence and absence of calcium (Ca2+) pump inhibitors/chelators and antioxidants. Followed by an in-depth PCR analysis of 84 oxidative stress-related genes. The measurements revealed, as compared to the control, that exposure to CBNP nanoparticles leads to the generation of high ROS levels, as well as a disturbance in calcium homeostasis, which remained primarily unchanged even after 24 h of exposure. Nevertheless, in presence of antioxidants N-acetylcysteine (NAC) and Trolox, ROS formation was considerably reduced without affecting the intracellular calcium concentration. On the other hand, Ca2+ pump inhibitors/chelators, BAPTA (1,2-bis(o-amino phenoxy)ethane-N, N, N', N'-tetraacetic acid) and verapamil not only decreased the Ca2+ overload, but also further decreased the ROS formation, indicating its role in CBNP-induced oxidative stress. Further, a PCR array analysis of A549 cells in presence and absence of the calmodulin (CaM) antagonist W7, indicated toward nine altered oxidative stress-related genes which further confirmed our cytotoxicity results. Obtained data suggested that CBNP exposure elevates calcium ion concentration, which further contributes to oxidative stress, via the calcium-binding protein CaM. Its inhibition with W7 leads to downregulation in gene expression of nine oxidative stress-related genes, which otherwise, as compared to control, show increased gene expression. The results of the study thus confirm that exposure of lung epithelial cells to CBNP leads to oxidative stress; however, the oxidative stress itself is a result of a disturbance in both calcium and ROS homeostasis, and should be considered while searching for a new strategy for prevention of CBNP-induced lung toxicity.


Assuntos
Células Epiteliais Alveolares/efeitos dos fármacos , Nanopartículas/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Fuligem/toxicidade , Células A549 , Células Epiteliais Alveolares/patologia , Antioxidantes/farmacologia , Cálcio/metabolismo , Calmodulina/metabolismo , Relação Dose-Resposta a Droga , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Estresse Oxidativo/genética , Espécies Reativas de Oxigênio/metabolismo , Fuligem/administração & dosagem , Fatores de Tempo
5.
Contact Dermatitis ; 84(5): 317-325, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33320367

RESUMO

BACKGROUND: People are exposed to mixtures containing allergens and irritants often causing contact dermatitis. Therefore, regulatory authorities require systematic information on the effects of mixtures on the sensitization threshold. In this study a moderate (cinnamal) and a weak (ethylene glycol dimethacrylate) allergen were combined with irritants covering different mechanisms of action (sodium dodecyl sulfate, salicylic acid, and α-pinene). For a systematic approach, the single substances were initially tested using the KeratinoSens assay. Thereafter, each allergen was combined with noncytotoxic concentrations of the irritants. METHOD: The KeratinoSens assay was applied for the single substances according to OECD (Organisation for Economic Co-operation and Development) Test Guideline 442D. Based on these results, three noncytotoxic concentrations of the irritants were selected and applied simultaneously with 12 concentrations of the allergens to the KeratinoSens cells. Sensitization threshold and cytotoxicity were measured and compared with the individual testing. RESULTS: The combinations of allergens and irritants differed from the effects of the single substances and lowered the sensitization threshold. The quantitative approach allowed a clear description of the changes which varied by factors between 1.1 and 10.3. CONCLUSIONS: Overall, the allergen was the prominent compound in the mixture and its nature appeared to determine the degree of the response.


Assuntos
Alérgenos/farmacologia , Irritantes/farmacologia , Queratinócitos/efeitos dos fármacos , Pele/efeitos dos fármacos , Acroleína/análogos & derivados , Acroleína/farmacologia , Linhagem Celular , Dermatite Alérgica de Contato/diagnóstico , Dermatite Irritante/diagnóstico , Relação Dose-Resposta a Droga , Humanos , Metacrilatos/farmacologia
6.
Arch Toxicol ; 94(9): 3347, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32696078

RESUMO

The author would like to thank N. Bakhiya, S. Hessel-Pras, B. Sachse, and B. Dusemund for their support in the chapter about pyrrolizidine alkaloids.

7.
Arch Toxicol ; 94(6): 1787-1877, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32542409

RESUMO

The risk assessment of chemical carcinogens is one major task in toxicology. Even though exposure has been mitigated effectively during the last decades, low levels of carcinogenic substances in food and at the workplace are still present and often not completely avoidable. The distinction between genotoxic and non-genotoxic carcinogens has traditionally been regarded as particularly relevant for risk assessment, with the assumption of the existence of no-effect concentrations (threshold levels) in case of the latter group. In contrast, genotoxic carcinogens, their metabolic precursors and DNA reactive metabolites are considered to represent risk factors at all concentrations since even one or a few DNA lesions may in principle result in mutations and, thus, increase tumour risk. Within the current document, an updated risk evaluation for genotoxic carcinogens is proposed, based on mechanistic knowledge regarding the substance (group) under investigation, and taking into account recent improvements in analytical techniques used to quantify DNA lesions and mutations as well as "omics" approaches. Furthermore, wherever possible and appropriate, special attention is given to the integration of background levels of the same or comparable DNA lesions. Within part A, fundamental considerations highlight the terms hazard and risk with respect to DNA reactivity of genotoxic agents, as compared to non-genotoxic agents. Also, current methodologies used in genetic toxicology as well as in dosimetry of exposure are described. Special focus is given on the elucidation of modes of action (MOA) and on the relation between DNA damage and cancer risk. Part B addresses specific examples of genotoxic carcinogens, including those humans are exposed to exogenously and endogenously, such as formaldehyde, acetaldehyde and the corresponding alcohols as well as some alkylating agents, ethylene oxide, and acrylamide, but also examples resulting from exogenous sources like aflatoxin B1, allylalkoxybenzenes, 2-amino-3,8-dimethylimidazo[4,5-f] quinoxaline (MeIQx), benzo[a]pyrene and pyrrolizidine alkaloids. Additionally, special attention is given to some carcinogenic metal compounds, which are considered indirect genotoxins, by accelerating mutagenicity via interactions with the cellular response to DNA damage even at low exposure conditions. Part C finally encompasses conclusions and perspectives, suggesting a refined strategy for the assessment of the carcinogenic risk associated with an exposure to genotoxic compounds and addressing research needs.


Assuntos
Carcinógenos/toxicidade , Dano ao DNA , Mutagênicos/toxicidade , Animais , Testes de Carcinogenicidade , Humanos , Testes de Mutagenicidade , Medição de Risco , Toxicogenética
8.
Environ Res ; 173: 157-164, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30909101

RESUMO

It is still a major challenge to protect humans at workplaces and in the environment. To cope with this task, it is a prerequisite to obtain detailed information on the extent of chemical perturbations of biological pathways, in particular, adaptive vs. adverse effects and the dose-response relationships. This knowledge serves as the basis for the classification of non-carcinogens and carcinogens and for further distinguishing carcinogens in genotoxic (DNA damaging) or non-genotoxic compounds. Basing on quantitative dose-response relationships, points of departures can be derived for chemical risk assessment. In recent years, new methods have shown their capability to support the established rodent models of carcinogenicity testing. In vitro high throughput screening assays assess more comprehensively cell response. In addition, omics technologies were applied to study the mode of action of chemicals whereby the term "toxicogenomics" comprises various technologies such as transcriptomics, epigenomics, or metabolomics. This review aims to summarize the current state of toxicogenomic approaches in risk science and to compare them with established ones. For example, measurement of global transcriptional changes generates meaningful information for toxicological risk assessment such as accurate classification of genotoxic/non-genotoxic carcinogens. Alteration in mRNA expression offers previously unknown insights in the mode of action and enables the definition of key events. Based on these, benchmark doses can be calculated for the transition from an adaptive to an adverse state. In short, this review assesses the potential and challenges of transcriptomics and addresses the impact of other omics technologies on risk assessment in terms of hazard identification and dose-response assessment.


Assuntos
Carcinógenos , Toxicogenética , Testes de Carcinogenicidade , Relação Dose-Resposta a Droga , Humanos , Medição de Risco
9.
Arch Toxicol ; 93(9): 2593-2602, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31342136

RESUMO

Exposure to xenobiotic such as benzo[a]pyrene (B[a]P) induces metabolic changes, which have a considerable impact on the cellular response. Nevertheless, we are just in the beginning to reach an understanding of these processes. In this study, a gas chromatography-mass spectrometry (GC-MS)-based metabolomics approach was applied to distinguish the metabolic changes that bladder epithelia cells undergo upon B[a]P exposure. To closely reflect the epithelia cell conditions in vivo, freshly isolated primary porcine urinary bladder epithelial cells (PUBEC) were utilized for the current study. An untargeted metabolomics approach was used to characterize the time- (6 h, 24 h, 48 h) and dose-dependent (0.5 µM, 5 µM, 10 µM B[a]P) changes in the metabolome of PUBEC upon B[a]P exposure, which led to the profiling of more than 200 metabolites that differed significantly between control and exposed samples. Multivariate analysis of the data highlighted that in the experimental setup/model used other than the exposure concentration, it is the exposure time which seems to be most important for distinguishing between different groups and hence may have a bigger role in B[a]P-mediated toxicity but may be specific for cell model used and hence requires further investigations. Further, enrichment and pathway analysis using MetaboAnalyst highlighted that exposure to B[a]P mainly alters the cellular amino acid metabolism. Particularly, 1-pyrroline-5-carboxylic acid (P5C), an intermediate of the cycling of the amino acid proline, was identified as a differentially altered metabolite at all concentrations and exposure times used in the experiment. An increase in the activity of proline dehydrogenase/proline oxidase (PRODH/POX), which oxidizes proline to P5C, was also observed, further supporting our metabolomic data. Our findings contribute to an improved knowledge about the reprogramming of metabolism which is a fundamental element of the cellular response to B[a]P and draw attention to the role of proline in this context.


Assuntos
Benzo(a)pireno/toxicidade , Poluentes Ambientais/toxicidade , Células Epiteliais/efeitos dos fármacos , Metaboloma/efeitos dos fármacos , Prolina/metabolismo , Bexiga Urinária/efeitos dos fármacos , Animais , Relação Dose-Resposta a Droga , Células Epiteliais/metabolismo , Metabolômica , Cultura Primária de Células , Prolina Oxidase/metabolismo , Suínos , Fatores de Tempo , Bexiga Urinária/metabolismo
10.
Chem Res Toxicol ; 30(10): 1855-1864, 2017 10 16.
Artigo em Inglês | MEDLINE | ID: mdl-28922594

RESUMO

A product of incomplete combustion of diesel fuel, 3-nitrobenzanthrone (3-NBA), has been classified as a cancer-causing substance. It first gained attention as a potential urinary bladder carcinogen due to the presence of its metabolite in urine and formation of DNA adducts. The aim of the present study was to characterize the dose-response relationship of 3-NBA in human urothelial cancer cell line (RT4) exposed to concentrations ranging from 0.0003 µM (environmentally relevant) to 80 µM by utilizing toxicological and metabolomic approaches. We observed that the RT4 cells were capable of bioactivation of 3-NBA within 30 min of exposure. Activity measurements of various enzymes involved in the conversion of 3-NBA in RT4 cells demonstrated NAD(P)H:quinone oxidoreductase (NQO1) as the main contributor for its bioactivation. Moreover, cytotoxicity assessment exhibited an initiation of adaptive mechanisms at low dosages, which diminished at higher doses, indicating that the capacity of these mechanisms no longer suffices, resulting in increased levels of intracellular reactive oxygen species, reduced proliferation, and hyperpolarisation of the mitochondrial membrane. To characterize the underlying mechanisms of this cellular response, the metabolism of 3-NBA and metabolomic changes in the cells were analyzed. The metabolomic analysis of the cells (0.0003, 0.01, 0.08, 10, and 80 µM 3-NBA) showed elevated levels of various antioxidants at low concentrations of 3-NBA. However, at higher exposure concentrations, it appeared that the cells reprogrammed their metabolism to maintain the cell homeostasis via activation of pentose phosphate pathway (PPP).


Assuntos
Benzo(a)Antracenos/administração & dosagem , Benzo(a)Antracenos/farmacologia , Neoplasias da Bexiga Urinária/induzido quimicamente , Benzo(a)Antracenos/química , Benzo(a)Antracenos/toxicidade , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Humanos , Estrutura Molecular , Via de Pentose Fosfato/efeitos dos fármacos , Relação Estrutura-Atividade , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/patologia
12.
J Proteome Res ; 14(1): 202-13, 2015 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-25348606

RESUMO

A proteomic analysis of the interaction among multiprotein complexes involved in 2,3,7,8-dibenzo-p-dioxin (TCDD)-mediated toxicity in urinary bladder epithelial RT4 cells was performed using two-dimensional blue native SDS-PAGE (2D BN/SDS-PAGE). To enrich the protein complexes, unexposed and TCDD-exposed cells were fractionated. BN/SDS-PAGE of the resulting fractions led to an effective separation of proteins and protein complexes of various origins, including cell membrane, mitochondria, and other intracellular compartments. Major differences between the proteome of control and exposed cells involved the alteration of many calcium-regulated proteins (calmodulin, protein S100-A2, annexin A5, annexin A10, gelsolin isoform b) and iron-regulated proteins (ferritin, heme-binding protein 2, transferrin). On the basis of these findings, the intracellular calcium concentration was determined, revealing a significant increase after 24 h of exposure to TCDD. Moreover, the concentration of the labile iron pool (LIP) was also significantly elevated in TCDD-exposed cells. This increase was strongly inhibited by the calmodulin (CaM) antagonist W-7, which pointed toward a possible interaction between iron and calcium signaling. Because nitric oxide (NO) production was significantly enhanced in TCDD-exposed cells and was also inhibited by W-7, we hypothesize that alterations in calcium and iron homeostasis upon exposure to TCDD may be linked through NO generated by CaM-activated nitric oxide synthase. In our model, we propose that NO produced upon TCDD exposure interacts with the iron centers of iron-regulatory proteins (IRPs) that modulate the alteration of ferritin and transferrin, resulting in an augmented cellular LIP and, hence, increased toxicity.


Assuntos
Cálcio/metabolismo , Poluentes Ambientais/toxicidade , Ferro/metabolismo , Óxido Nítrico/fisiologia , Dibenzodioxinas Policloradas/toxicidade , Proteoma/metabolismo , Calmodulina/metabolismo , Fracionamento Celular , Linhagem Celular , Núcleo Celular/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Homeostase , Humanos , Eletroforese em Gel de Poliacrilamida Nativa , Proteômica , Bexiga Urinária/citologia
13.
Amino Acids ; 47(6): 1155-66, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25715757

RESUMO

Deoxyhypusine hydroxylase (DOHH) is a dinuclear iron enzyme required for hydroxylation of the aminobutyl side chain of deoxyhypusine in eukaryotic translation initiation factor 5A (eIF-5A), the second step in hypusine biosynthesis. DOHH has been recently identified in P. falciparum and P. vivax. Both enzymes have very peculiar features including E-Z type HEAT-like repeats and a diiron centre in their active site. Both proteins share only 26 % amino acid identity to the human paralogue. Hitherto, no X-ray structure exists from either enzyme. However, structural predictions based on the amino acid sequence of the active site in comparison to the human enzyme show that four conserved histidine and glutamate residues provide the coordination sites for chelating the ferrous iron ions. Recently, we showed that P. vivax DOHH is inhibited by zileuton (N-[1-(1-benzothien-2-yl)ethyl]-N-hydroxyurea), a drug that is known for inhibiting human 5-lipoygenase (5-LOX) by the complexation of ferrous iron. A novel discovery program was launched to identify inhibitors of the P. falciparum DOHH from the Malaria Box, consisting of 400 chemical compounds, which are highly active in the erythrocytic stages of Malaria infections. In a first visual selection for potential ligands of ferrous iron, three compounds from different scaffold classes namely the diazonapthyl benzimidazole MMV666023 (Malaria Box plate A, position A03), the bis-benzimidazole MMV007384 (plate A, position B08), and a 1,2,5,-oxadiazole MMV665805 (plate A, position C03) were selected and subsequently evaluated in silico for their potential to complex iron ions. As a proof of principle, a bioanalytical assay was performed and the inhibition of hypusine biosynthesis was determined by GC-MS. All tested compounds proved to be active in this assay and MMV665805 exhibited the strongest inhibitory effect. Notably, the results were in accordance with the preliminary quantum-mechanical calculations suggesting the strongest iron complexation capacity for MMV665805. This compound might be a useful tool as well as a novel lead structure for inhibitors of P. falciparum DOHH.


Assuntos
Antimaláricos , Inibidores Enzimáticos , Quelantes de Ferro , Oxigenases de Função Mista/antagonistas & inibidores , Plasmodium falciparum/enzimologia , Proteínas de Protozoários/antagonistas & inibidores , Antimaláricos/química , Antimaláricos/farmacologia , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Humanos , Quelantes de Ferro/química , Quelantes de Ferro/farmacologia
14.
Arch Toxicol ; 88(4): 913-34, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24464499

RESUMO

Epidemiological studies suggest that environmental exposure to airborne particulate matter may promote cardiovascular diseases; however, it is not clear whether this observation actually reflects exposure to nanosized particles in the environment. In the present study, the human endothelial cell line EA.hy926 was exposed to pure carbon black and, to mimic exposure to diesel exhaust, carbon black loaded with benzo[a]pyrene to ascertain effects of these particles on the cell proteome and metabolom. Particular emphasis was laid on an extended exposure period (14 days) and a low particle concentration (100 ng/mL). While ROS production essentially remained unaffected, exposure of the cells to the particles resulted in a significantly enhanced cell proliferation. Evaluation of the obtained proteomic and phosphoproteomic data revealed modulations of proteins involved in catalytic processes and cytoskeleton maintenance. The bioinformatic evaluation of the data revealed the possible involvement of the transcription factor peroxisome proliferator-activated receptor gamma. The further analysis of the cytoskeleton indicated changes of the cell motility, which is in agreement with an observed increase in the cellular migration and invasion, and macroscopic changes of the cytoskeleton of the exposed cells.


Assuntos
Benzo(a)pireno/toxicidade , Movimento Celular/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Metabolômica , Proteômica , Fuligem/toxicidade , Cálcio/metabolismo , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Biologia Computacional , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/metabolismo , Citoesqueleto/patologia , Relação Dose-Resposta a Droga , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Humanos , Metabolômica/métodos , Estresse Oxidativo/efeitos dos fármacos , Proteômica/métodos , Espécies Reativas de Oxigênio/metabolismo , Integração de Sistemas , Fatores de Tempo
15.
Mutat Res ; 828: 111855, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38569440

RESUMO

Environmental and occupational exposure to polycyclic aromatic hydrocarbons (PAHs) is associated with adverse health effects in humans. Uncertainty exists regarding the causation of urinary bladder cancer by benzo[a]pyrene (B[a]P) due to a lack of sufficient data. In this work, we focused on in-vitro DNA damage and the formation of micronuclei and chromosomal aberrations as predictors of cancer risk, applying a wide range of dosages and time periods to quantify the onset, intensity, and duration of the response. We chose two urothelial cell types to compare susceptibility and the ability to increase the malignity of a pre-existing bladder cancer: a cancer cell line (T24) and a pooled sample of primary urinary bladder epithelia cells (PUBEC) from pigs. The highest level of DNA damage assessed by comet assay was observed following 24-h treatment in both cell types, whereas PUBEC cells were clearly more susceptible. Even 4-h treatment induced DNA damage in PUBEC cells with benchmark doses of 0.0027 µM B[a]P and 0.00023 µM after 4-h and 24-h exposure, respectively. Nearly no effect was observed for periods of 48 h. The frequency of micronucleus formation increased more markedly in T24 cells, particularly with 24-h treatment. In PUBEC cells, 48-h exposure notably induced the formation of nucleoplasmic bridges and nuclear buds. Even though only one biological replicate was studied due to the sophisticated study design, our results give a strong indication of the potential of B[a]P to induce and increase malignity in human-relevant cell types.


Assuntos
Benzo(a)pireno , Instabilidade Cromossômica , Dano ao DNA , Urotélio , Benzo(a)pireno/toxicidade , Dano ao DNA/efeitos dos fármacos , Projetos Piloto , Animais , Urotélio/efeitos dos fármacos , Urotélio/patologia , Instabilidade Cromossômica/efeitos dos fármacos , Humanos , Suínos , Testes para Micronúcleos , Relação Dose-Resposta a Droga , Aberrações Cromossômicas , Neoplasias da Bexiga Urinária/induzido quimicamente , Neoplasias da Bexiga Urinária/patologia , Neoplasias da Bexiga Urinária/genética , Fatores de Tempo , Ensaio Cometa , Linhagem Celular Tumoral , Bexiga Urinária/efeitos dos fármacos , Bexiga Urinária/patologia
16.
Electrophoresis ; 34(4): 501-4, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23192385

RESUMO

The recent introduction of the La(3+) precipitation method for the enrichment of phosphoproteins allows a gel-based analysis of these posttranslationally modified proteins. However, if this method is applied to cell lysates stored in urea-containing lysis buffer for an extended period of time, incomplete phosphoprotein recovery is observed. We ascribe this effect to the presence of urea in the lysis buffer. To overcome this problem various strategies were tested, where cell lysates stored at least for one year were utilized. By applying an optimized protocol approximately 250 proteins could be observed following separation by 2DE.


Assuntos
Eletroforese em Gel Bidimensional/métodos , Lantânio/química , Fosfoproteínas/isolamento & purificação , Tioureia/química , Ureia/química , Precipitação Química , Células Endoteliais/química , Humanos , Concentração de Íons de Hidrogênio , Fosfoproteínas/análise , Redobramento de Proteína
17.
Proteomics ; 12(11): 1731-55, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22623321

RESUMO

Benzo[a]pyrene (BaP), a five-ring polycyclic aromatic hydrocarbon, is a well-recognized environmental pollutant. Coal-processing waste products, petroleum sludge, asphalt, creosote, and tobacco smoke, all contain high levels of BaP. Exposure to BaP elicits many adverse biological effects, including tumor formation, immunosuppression, teratogenicity, and hormonal effects. In addition to the genetic damage caused by BaP exposure, several studies have indicated the disruption of protein-protein signaling pathways. However, contrary to the large number of studies on BaP-induced DNA damage, only few data have been gathered on its effects at the protein level. This review highlights all proteomic studies to date used for assessing the toxicity of BaP and its metabolites in various organ systems. It will also give an overview on the role proteomics may play to elucidate the mechanisms underlying BaP toxicity.


Assuntos
Benzo(a)pireno/metabolismo , Benzo(a)pireno/toxicidade , Poluentes Ambientais/toxicidade , Animais , Benzo(a)pireno/química , Dano ao DNA , Exposição Ambiental , Peixes , Humanos , Camundongos , Órgãos em Risco , Proteômica , Ratos
18.
Arch Toxicol ; 86(12): 1861-71, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22790669

RESUMO

More than 90 % of all bladder cancers are transitional cell carcinomas arising from the cells lining the inside of the hollow organ (uroepithelium). Cell cultures from primary urinary bladder epithelial cells (PUBEC) of pigs were established to assess the uptake, intracellular concentration, and subcellular distribution of the environmental pollutant benzo(a)pyrene (BaP). During treatment of the cells with 0.5 µM BaP for up to 24 h, intracellular concentration of BaP increased without saturation but with marked differences between various PUBEC pools. Analysis of BaP uptake by laser scanning microscopy indicated that BaP is rapidly partitioned into the cell membrane, while only a slight but significant increase in BaP fluorescence intensity was observed in the cytosol and nucleus. Spectrofluorometric quantification of BaP in PUBEC using ex situ calibration revealed a strong accumulation of BaP, leading to intracellular concentrations ranging from 7.28 to 35.70 µM in cells exposed to 0.5 µM BaP and from 29.9 to 406.64 µM in cells exposed to 10 µM BaP. These results were confirmed by gas chromatographic mass spectrometric analysis. Apoptotic cell nuclei were assessed by TUNEL analysis to see whether BaP exposure at the given concentrations results in a toxic effect. While apoptotic cells were barely detectable in control epithelial cells, there was a marked elevation in apoptosis in the BaP-exposed cells. In conclusion, a comprehensive study on uptake and quantification of BaP in epithelial cells from pig bladder is reported for the first time. The study may be helpful in understanding the pattern of BaP uptake and distribution in bladder and its possible implication in bladder cancer development.


Assuntos
Benzo(a)pireno/toxicidade , Carcinógenos/toxicidade , Células Epiteliais/efeitos dos fármacos , Urotélio/efeitos dos fármacos , Animais , Benzo(a)pireno/metabolismo , Carcinógenos/metabolismo , Linhagem Celular , Fragmentação do DNA , Cromatografia Gasosa-Espectrometria de Massas , Marcação In Situ das Extremidades Cortadas , Indicadores e Reagentes , Cinética , Microscopia Confocal , Frações Subcelulares/química , Frações Subcelulares/metabolismo , Suínos , Urotélio/citologia
19.
Proteomics ; 11(4): 644-56, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-21246732

RESUMO

The simultaneous analysis of a wide array of proteins may provide valuable information on the activation and suppression of cellular systems at different stages of the exposure-disease continuum. In this review, results of proteomic studies in the field of toxicology are covered, focusing on the effects of chemical carcinogens. So far, alterations of highly abundant proteins have been identified which, irrespective of the wide differences in study design and technologies used, can be grossly assigned to three functional classes: proteins related to cellular stress response, inflammation, and stimulation of the immune system. It is obvious that the observed protein alterations are not causal factors in the development of chemically induced cancer but rather reflect common reactions to cellular perturbations. In order to gain deeper insights into the process of chemical carcinogenesis, the previously applied "shotgun" analyses have to be abandoned in favour of targeted proteomic approaches focusing on the accurate identification and quantification of selected proteins. Advanced analytical techniques such as selective reaction monitoring (SRM) and multiple reaction monitoring (MRM) offer this opportunity. If toxicoproteomic research moves into that direction and takes advantage of such techniques it will have the potential to contribute to the elucidation of chemical carcinogenesis.


Assuntos
Testes de Carcinogenicidade/métodos , Carcinógenos/toxicidade , Proteínas de Neoplasias/análise , Neoplasias/induzido quimicamente , Neoplasias/metabolismo , Proteoma/análise , Proteômica/métodos , Animais , Humanos , Biossíntese de Proteínas/efeitos dos fármacos
20.
Proteomics ; 11(4): 776-93, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-21229584

RESUMO

Of the numerous animal models available for proteomic studies only a small number have been successfully used in understanding human biology. To date, rodents have been widely employed in proteomic and genomic studies but often these models do not truly mimic the relevant human conditions. On the other hand, the pig shows similarity in size, shape and physiology to human and has been used as a major mammalian model for many studies concerning xenotransplantation, cardiovascular diseases, blood dynamics, nutrition, general metabolic functions, digestive-related disorders, respiratory diseases, diabetes, kidney and bladder diseases, organ-specific toxicity, dermatology and neurological sequelae. With the substantially improved knowledge of the structure and function of the pig genome in the last two decades it has been found that this animal shares a high sequence and chromosomal structure homology with humans. Nevertheless, in comparison to other available model organisms, very little work has been devoted to pig proteomics until recently. Keeping this in mind, the present review will highlight some of the advantages and disadvantages of pig as a model system for proteomic studies.


Assuntos
Modelos Animais de Doenças , Proteômica , Suínos/fisiologia , Animais , Humanos , Suínos/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA