The role of benzo[a]pyrene (BaP), a prominent genotoxic carcinogen and aryl hydrocarbon receptor (AhR) ligand, in tumor progression remains poorly characterized. We investigated the impact of BaP on the process of epithelial-mesenchymal transition (EMT) in normal human bronchial epithelial HBEC-12KT cells. Early morphological changes after 2-week exposure were accompanied with induction of SERPINB2, IL1, CDKN1A/p21 (linked with cell cycle delay) and chemokine CXCL5. After 8-week exposure, induction of cell migration and EMT-related pattern of markers/regulators led to induction of further pro-inflammatory cytokines or non-canonical Wnt pathway ligand WNT5A. This trend of up-regulation of pro-inflammatory genes and non-canonical Wnt pathway constituents was observed also in the BaP-transformed HBEC-12KT-B1 cells. In general, transcriptional effects of BaP differed from those of TGFβ1, a prototypical EMT inducer, or a model non-genotoxic AhR ligand, TCDD. Carcinogenic polycyclic aromatic hydrocarbons could thus induce a unique set of molecular changes linked with EMT and cancer progression.
- MeSH
- benzopyren * toxicita MeSH
- epitelové buňky * metabolismus MeSH
- lidé MeSH
- ligandy MeSH
- poškození DNA MeSH
- receptory aromatických uhlovodíků genetika metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
OBJECTIVE: Our study aimed to evaluate the extent of polycyclic aromatic hydrocarbon (PAH) exposure in coke oven workers from Eastern Slovakia by cytogenetic analysis of human peripheral lymphocytes. METHODS: A total of 81 peripheral blood samples were collected from PAH-exposed workers (mean age 45.84 ± 9.73 years) and 30 samples constituted the control group (41.93 ± 15.39 years). The samples were processed using routine cytological analysis. Conventional cytogenetic analysis of human peripheral lymphocytes has been used to evaluate the effects of PAHs. RESULTS: Comparison of the aberrant cells in the total exposed with the controls showed a significant difference (p < 0.05). A high level of significance (p < 0.001) was observed when comparing the gaps between the exposed group and the control group. There was a significant difference (p < 0.01) in aberrant cells and chromatid breaks (p < 0.05) in the GR1 working subgroup compared with the control group. The results of the correlation analysis did not show a significant relationship between the length of occupational exposure and the frequency of aberrant cells (r = 0.071, p = 0.529). Similarly, no association was observed between smoking among coke plant workers and the frequency of aberrant cells (r = 0.117, p = 0.538). CONCLUSION: Cytogenetic analysis showed an increased frequency of chromosomal aberrations in coke oven workers in Eastern Slovakia.
- MeSH
- cytogenetické vyšetření MeSH
- dospělí MeSH
- koks * analýza MeSH
- lidé středního věku MeSH
- lidé MeSH
- lymfocyty chemie MeSH
- polycyklické aromatické uhlovodíky * toxicita analýza MeSH
- pracovní expozice * škodlivé účinky analýza MeSH
- pyreny analýza MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Slovenská republika MeSH
The testis is a priority organ for developing alternative models to assess male reproductive health hazards of chemicals. This study characterized a 3D in vitro model of murine prepubertal Leydig TM3 cells with improved expression of steroidogenesis markers suitable for image-based screening of testicular toxicity. This 3D scaffold-free spheroid model was applied to explore the impact of prototypical endocrine-disrupting chemicals (EDCs) and environmental reprotoxicants (benzo[a]pyrene, 2- and 9-methylanthracenes, fluoranthene, triclosan, triclocarban, methoxychlor) on male reproductive health. The results were compared to the male reprotoxicity potential of EDCs assessed in a traditional monolayer (2D) culture. The testicular toxicity was dependent not only on the type of culture (2D vs. 3D models) but also on the duration of exposure. Benzo[a]pyrene and triclocarban were the most active compounds, eliciting cytotoxic effects in prepubertal Leydig cells at low micromolar concentrations, which might be a mechanism contributing to their male reprotoxicity.
- MeSH
- benzopyren toxicita MeSH
- endokrinní disruptory * chemie MeSH
- Leydigovy buňky * MeSH
- myši MeSH
- rozmnožování MeSH
- testis MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Deciphering the role of the aryl hydrocarbon receptor (AhR) in lung cancer cells may help us to better understand the role of toxic AhR ligands in lung carcinogenesis, including cancer progression. We employed human lung carcinoma A549 cells to investigate their fate after continuous two-week exposure to model AhR agonists, genotoxic benzo[a]pyrene (BaP; 1 μM) and non-genotoxic 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; 10 nM). While TCDD increased proliferative rate of A549 cells, exposure to BaP decreased cell proliferation and induced epithelial-to-mesenchymal transition (EMT)-like phenotype, which was associated with enhanced cell migration, invasion, and altered cell morphology. Although TCDD also suppressed expression of E-cadherin and activated some genes linked to EMT, it did not induce the EMT-like phenotype. The results of transcriptomic analysis, and the opposite effects of BaP and TCDD on cell proliferation, indicated that a delay in cell cycle progression, together with a slight increase of senescence (when coupled with AhR activation), favors the induction of EMT-like phenotype. The shift towards EMT-like phenotype observed after simultaneous treatment with TCDD and mitomycin C (an inhibitor of cell proliferation) confirmed the hypothesis. Since BaP decreased cell proliferative rate via induction of p21 expression, we generated the A549 cell model with reduced p21 expression and exposed it to BaP for two weeks. The p21 knockdown suppressed the BaP-mediated EMT-like phenotype in A549 cells, thus confirming that a delayed cell cycle progression, together with p21-dependent induction of senescence-related chemokine CCL2, may contribute to induction of EMT-like cell phenotype in lung cells exposed to genotoxic AhR ligands.
- MeSH
- benzopyren toxicita MeSH
- epitelové buňky MeSH
- fenotyp MeSH
- karcinom * MeSH
- lidé MeSH
- nádory plic * genetika MeSH
- plíce MeSH
- receptory aromatických uhlovodíků genetika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Several sequences forming G-quadruplex are highly conserved in regulatory regions of genomes of different organisms and affect various biological processes like gene expression. Diverse G-quadruplex properties can be modulated via their interaction with small polyaromatic molecules such as pyrene. To investigate how pyrene interacts with G-rich DNAs, we incorporated deoxyuridine nucleotide(s) with a covalently attached pyrene moiety (Upy) into a model system that forms parallel G-quadruplex structures. We individually substituted terminal positions and positions in the pentaloop of the c-kit2 sequence originating from the KIT proto-oncogene with Upy and performed a detailed NMR structural study accompanied with molecular dynamic simulations. Our results showed that incorporation into the pentaloop leads to structural polymorphism and in some cases also thermal destabilization. In contrast, terminal positions were found to cause a substantial thermodynamic stabilization while preserving topology of the parent c-kit2 G-quadruplex. Thermodynamic stabilization results from π-π stacking between the polyaromatic core of the pyrene moiety and guanine nucleotides of outer G-quartets. Thanks to the prevalent overall conformation, our structures mimic the G-quadruplex found in human KIT proto-oncogene and could potentially have antiproliferative effects on cancer cells.
- MeSH
- deoxyuridin chemie MeSH
- G-kvadruplexy * MeSH
- lidé MeSH
- molekulární modely MeSH
- nukleární magnetická rezonance biomolekulární MeSH
- promotorové oblasti (genetika) MeSH
- protoonkogenní proteiny c-kit genetika MeSH
- pyreny chemie MeSH
- termodynamika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Sphingolipids (SLs), glycosphingolipids (GSLs), and eicosanoids are bioactive lipids, which play important roles in the etiology of various diseases, including cancer. However, their content and roles in cancer cells, and in particular in the exosomes derived from tumor cells, remain insufficiently characterized. In this study, we evaluated alterations of SL and GSL levels in transformed cells and their exosomes, using comparative HPLC-MS/MS analysis of parental human bronchial epithelial cells HBEC-12KT and their derivative, benzo[a]pyrene-transformed HBEC-12KT-B1 cells with the acquired mesenchymal phenotype. We examined in parallel SL/GSL contents in the exosomes released from both cell lines. We found significant alterations of the SL/GSL profile in the transformed cell line, which corresponded well with alterations of the SL/GSL profile in exosomes derived from these cells. This suggested that a majority of SLs and GSLs were transported by exosomes in the same relative pattern as in the cells of origin. The only exceptions included decreased contents of sphingosin, sphingosin-1-phosphate, and lactosylceramide in exosomes derived from the transformed cells, as compared with the exosomes derived from the parental cell line. Importantly, we found increased levels of ceramide phosphate, globoside Gb3, and ganglioside GD3 in the exosomes derived from the transformed cells. These positive modulators of epithelial-mesenchymal transition and other pro-carcinogenic processes might thus also contribute to cancer progression in recipient cells. In addition, the transformed HBEC-12KT-B1 cells also produced increased amounts of eicosanoids, in particular prostaglandin E2. Taken together, the exosomes derived from the transformed cells with specifically upregulated SL and GSL species, and increased levels of eicosanoids, might contribute to changes within the cancer microenvironment and in recipient cells, which could in turn participate in cancer development. Future studies should address specific roles of individual SL and GSL species identified in the present study.
- MeSH
- benzopyren toxicita MeSH
- bronchy cytologie MeSH
- buněčné linie MeSH
- exozómy metabolismus MeSH
- karcinogeny toxicita MeSH
- lidé MeSH
- nádorová transformace buněk * MeSH
- respirační sliznice účinky léků metabolismus MeSH
- sfingolipidy metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
BACKGROUND: Asthma represents a syndrome for which our understanding of the molecular processes underlying discrete sub-diseases (i.e., endotypes), beyond atopic asthma, is limited. The public health needs to characterize etiology-associated endotype risks is becoming urgent. In particular, the roles of polyaromatic hydrocarbon (PAH), globally distributed combustion by-products, toward the two known endotypes - T helper 2 cell high (Th2) or T helper 2 cell low (non-Th2) - warrants clarification. OBJECTIVES: To explain ambient B[a]P association with non-atopic asthma (i.e., a proxy of non-Th2 endotype) is markedly different from that with atopic asthma (i.e., a proxy for Th2-high endotype). METHODS: In a case-control study, we compare the non-atopic as well as atopic asthmatic boys and girls against their respective controls in terms of the ambient Benzo[a]pyrene concentration nearest to their home, plasma 15-Ft2-isoprostane (15-Ft2-isoP), urinary 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), and lung function deficit. We repeated the analysis for i) dichotomous asthma outcome and ii) multinomial asthma-overweight/obese (OV/OB) combined outcomes. RESULTS: The non-atopic asthma cases are associated with a significantly higher median B[a]P (11.16 ng/m3) compared to that in the non-atopic controls (3.83 ng/m3; P-value < 0.001). In asthma-OV/OB stratified analysis, the non-atopic girls with lean and OV/OB asthma are associated with a step-wisely elevated B[a]P (median,11.16 and 18.00 ng/m3, respectively), compared to the non-atopic lean control girls (median, 4.28 ng/m3, P-value < 0.001). In contrast, atopic asthmatic children (2.73 ng/m3) are not associated with a significantly elevated median B[a]P, compared to the atopic control children (2.60 ng/m3; P-value > 0.05). Based on the logistic regression model, on ln-unit increate in B[a]P is associated with 4.7-times greater odds (95% CI, 1.9-11.5, P = 0.001) of asthma among the non-atopic boys. The same unit increase in B[a]P is associated with 44.8-times greater odds (95% CI, 4.7-428.2, P = 0.001) among the non-atopic girls after adjusting for urinary Cotinine, lung function deficit, 15-Ft2-isoP, and 8-oxodG. CONCLUSIONS: Ambient B[a]P is robustly associated with non-atopic asthma, while it has no clear associations with atopic asthma among lean children. Furthermore, lung function deficit, 15-Ft2-isoP, and 8-oxodG are associated with profound alteration of B[a]P-asthma associations among the non-atopic children.
- MeSH
- 8-hydroxy-2'-deoxyguanosin moč MeSH
- benzopyren analýza MeSH
- bronchiální astma krev epidemiologie patofyziologie moč MeSH
- dinoprost analogy a deriváty krev MeSH
- dítě MeSH
- fenotyp MeSH
- kojenec MeSH
- kotinin moč MeSH
- látky znečišťující vzduch analýza MeSH
- lidé MeSH
- mladiství MeSH
- plíce patofyziologie MeSH
- předškolní dítě MeSH
- studie případů a kontrol MeSH
- vystavení vlivu životního prostředí analýza MeSH
- Check Tag
- dítě MeSH
- kojenec MeSH
- lidé MeSH
- mladiství MeSH
- mužské pohlaví MeSH
- předškolní dítě MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Česká republika MeSH
DNA damage caused by exogenous or endogenous factors is a common challenge for developing fish embryos. DNA damage repair (DDR) pathways help organisms minimize adverse effects of DNA alterations. In terms of DNA repair mechanisms, sturgeons represent a particularly interesting model due to their exceptional genome plasticity. Sterlet (Acipenser ruthenus) is a relatively small species of sturgeon. The goal of this study was to assess the sensitivity of sterlet embryos to model genotoxicants (camptothecin, etoposide, and benzo[a]pyrene), and to assess DDR responses. We assessed the effects of genotoxicants on embryo survival, hatching rate, DNA fragmentation, gene expression, and phosphorylation of H2AX and ATM kinase. Exposure of sterlet embryos to 1 µM benzo[a]pyrene induced low levels of DNA damage accompanied by ATM phosphorylation and xpc gene expression. Conversely, 20 µM etoposide exposure induced DNA damage without activation of known DDR pathways. Effects of 10 nM camptothecin on embryo development were stage-specific, with early stages, before gastrulation, being most sensitive. Overall, this study provides foundational information for future investigation of sterlet DDR pathways.
- MeSH
- benzopyren toxicita MeSH
- embryo nesavčí účinky léků embryologie metabolismus MeSH
- embryonální vývoj účinky léků genetika MeSH
- etoposid toxicita MeSH
- fragmentace DNA účinky léků MeSH
- kamptothecin toxicita MeSH
- kometový test MeSH
- mutageny toxicita MeSH
- oprava DNA * MeSH
- poškození DNA * MeSH
- ryby embryologie genetika MeSH
- testy genotoxicity metody MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
The aryl hydrocarbon receptor (AhR) transcription factor is activated by polycyclic aromatic hydrocarbons (PAH) and other ligands. Activated AhR binds to dioxin responsive elements (DRE) and initiates transcription of target genes, including the gene encoding prostaglandin endoperoxide synthase 2 (PTGS-2), which is also activated by the transcription factor NF-ĸB. PTGS-2 catalyzes the conversion of arachidonic acid (AA) into prostaglandins, thromboxanes or isoprostanes. 15-F2t-Isoprostane (IsoP), regarded as a universal marker of lipid peroxidation, is also induced by PAH exposure. We investigated the processes associated with lipid peroxidation in human alveolar basal epithelial cells (A549) exposed for 4 h or 24 h to model PAH (benzo[a]pyrene, BaP; 3-nitrobenzanthrone, 3-NBA) and organic extracts from ambient air particulate matter (EOM), collected in two seasons in a polluted locality. Both EOM induced the expression of CYP1A1 and CYP1B1; 24 h treatment significantly reduced PTGS-2 expression. IsoP levels decreased after both exposure periods, while the concentration of AA was not affected. The effects induced by BaP were similar to EOM except for increased IsoP levels after 4 h exposure and elevated AA concentration after 24 h treatment. In contrast, 3-NBA treatment did not induce CYP expression, had a weak effect on PTGS-2 expression, and, similar to BaP, induced IsoP levels after 4 h exposure and AA levels after 24 h treatment. All tested compounds induced the activity of NF-ĸB after the longer exposure period. In summary, our data suggest that EOM, and partly BaP, reduce lipid peroxidation by a mechanism that involves AhR-dependent inhibition of PTGS-2 expression. The effect of 3-NBA on IsoP levels is probably mediated by a different mechanism independent of AhR activation.
- MeSH
- benz(a)anthraceny toxicita MeSH
- benzopyren toxicita MeSH
- buňky A549 MeSH
- cyklooxygenasa 1 metabolismus MeSH
- cytochrom P-450 CYP1A1 metabolismus MeSH
- lidé MeSH
- mutageny toxicita MeSH
- nádorové buněčné linie MeSH
- NF-kappa B metabolismus MeSH
- peroxidace lipidů účinky léků MeSH
- pevné částice toxicita MeSH
- pneumocyty účinky léků MeSH
- polycyklické aromatické uhlovodíky toxicita MeSH
- receptory aromatických uhlovodíků metabolismus MeSH
- transkripční faktory bHLH metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
In this work, we compare labeling by two negatively charged fluorescent labels, 8-aminopyrene-1,3,6-trisulfonic acid (APTS) and 8-(2-hydrazino-2-oxoethoxy)pyrene-1,3,6-trisulfonic acid (Cascade Blue hydrazide [CBH]). Effectiveness of the labeling chemistries were investigated by 4-hydroxybenzaldehyde and maltoheptaose followed by LC/UV-MS and CE/LIF analysis, respectively. The reaction yield of APTS labeling was determined to be only ∼10%. This is due to reduction of almost 90% of the analyte by sodium cyanoborohydride to alcohol, which cannot be further labeled via reductive amination. However, the CBH labeling provides ∼90% reaction yield based on the LC/UV-MS measurements. The significantly higher labeling yield was also confirmed by CE/LIF measurements. Finally, the more effective hydrazone formation technique of CBH was characterized and applied for N-linked glycan analysis by CE/LIF.
- MeSH
- aminace MeSH
- chromatografie kapalinová metody MeSH
- elektroforéza kapilární metody MeSH
- fluorescenční barviva chemie MeSH
- hmotnostní spektrometrie metody MeSH
- hydrazony chemie MeSH
- oligosacharidy analýza chemie MeSH
- pyreny chemie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
