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EN
Environmental pollution causes a variety of health problems, including cancer. Many known pollutants have carcinogenic properties and polycyclic aromatic hydrocarbons (PAH) belong to this group. In this study, an in vitro culture of V79 cells of the Chinese hamster was subjected to three tested PAHs: 5-amino-2,3-dihydro-1H-cyclopenta-[c]phenanthrene (ACP[c]Ph), 5-amino-9-methoxy-2,3-dihydro-1H-cyclopenta[c]phenanthrene (AMCP[c]Ph) and cyclopenta[c]phenanthrene (CP[c]Ph). The in vitro micronucleus (MN) assay was applied in order to evaluate the genotoxic properties of the studied compounds. The highest genotoxic effect was observed for AMCP[c]Ph in a concentration of 0.02μg·ml-1. The genotoxic effect of the other two compounds was slightly lower.
EN
Proximate to the environment, adipose fin of fish may be considered as a lipid storing tissue, and thus can be a target for either waterborne or dietary polycyclic aromatic compounds (PACs). We determined the effects of benzo[a]pyrene (B[a]P), a model PAC member, on CYP1A gene expression in adipose fin and compared that with the effects in gill of juvenile rainbow trout (Oncorhynchus mykiss Walbaum) using the quantitative reverse transcription polymerase chain reaction (Q-RT-PCR). The results of the study demonstrated that constitutive CYP1A mRNA was present in adipose fin of rainbow trout, but the transcripts were far less abundant than those in gill tissue. We confirmed high CYP1A gene induction potential of the gills in rainbow trout injected with benzo[a]pyrene, but also showed moderately and transiently induced CYP1A mRNA in adipose fin. The modest and transitory gene expression may preclude rainbow trout adipose fin CYP1A mRNA levels from using it as an indicator of sustained exposure of fish to the polycyclic aromatic compounds.
EN
The expression of CYP1A (cytochrome P4501A) can be induced by a number of aromatic compounds in teleost fishes. We developed a real-time PCR assay for measuring relative quantities (RQ) of CYP1A mRNA in whitefish (Coregonus lavaretus). To test for the usefulness of the assay we performed a treatment study, using benzo[a]pyrene (B[a]P) a model CYP1A inducer. Primers for the CYP1A gene were adapted from the literature, whereas those for [beta]-actin (endogenous control) were designed from a region that was found to be conserved among salmonid [beta]-actin genes. A group of hatchery raised whitefish, with an average body mass of 15 g and total length of 12 cm were given an intraperitoneal injection (10 mg/kg) of B[a]P in corn oil (2 mg B[a]P/ml corn oil) or corn oil alone (Control). After 48 h, whitefish liver, head kidney and brains were collected for mRNA isolation and analysis. In all three tissues sampled, CYP1A mRNA was affected by treatment with B[a]P. Head kidney tissue showed the greatest induction potential (RQ=11.00) from base levels (RQ=1.00), followed by liver (RQ=9.45), and brain (RQ=3.76). These results demonstrated that CYP1A was highly inducible by B[a]P in whitefish head kidney and liver, and to some extent, in brain tissue. The approach presented here has the advantage of providing rapid and accurate measures of CYP1A induction in various tissues of fish responding to PAH contaminant exposure.
EN
Extracts from strawberry and raspberry leaves, carrot pomace, and spent grains, were tested as bioreducing agents for the synthesis of nanosilver particles (AgNP). Based on UV vis spectra, the leaf extracts produced the most AgNP, carrot pomace was less effective, and spent grains did not produce AgNP. The dynamic light scattering method revealed that AgNP ranged from 1 to 92nm in size, and that over 80% of the particles were about 10nm. Energy dispersive X ray spectroscopy showed that elements that typically stabilize nanoparticles were present. The well diffusion method (nutrient agar medium) indicated that AgNP synthesized with raspberry leaf extract exerted strong bacteriostatic and bactericidal activity against Gram negative bacteria and weaker activity against Gram positive bacteria. Although further analysis is needed to determine the mechanism of their synthesis, the results of this study show that plant extract based synthesis can produce nanoparticles with controlled size and morphology.
EN
Although p53, a protein of important tumor suppressive function, has been extensively studied in mammals, relatively little is known about the p53 pathways in lower vertebrates. Particularly, limited information exists on possible influences of environmental contaminants on the expression of the p53 gene in fish. In the current study, we assessed the effects of benzo[a]pyrene (B[a]P; potent tumor promoter) and cyclopenta[c]phenanthrene (CP[c]Ph; clastogenic agent) exposure on a 24h profile of p53 gene expression in head kidney of juvenile rainbow trout (Oncorhynchus mykiss). To analyze the p53 transcription rate, we developed protocol for the examination of both mRNA and heterogeneous nuclear (hn) RNA of the gene, using Real-Time RTPCR approach. The results show that both compounds are capable of suppressing p53 transcriptional activity within 12h of the treatment. Our finding supports the idea that structurally different PAHs may influence cell physiologic functions controlled by p53 in fish, in part, by down-regulating its RNA expression levels.
EN
MicroRNAs (miRNAs) are small, highly conserved, non-coding RNAs that regulate gene expression of target mRNAs through cleavage or translational inhibition. In the field of toxicology, the relationship between toxicity and microRNA expression is poorly understood. In the present study we analyzed the abundance of 9 selected miRNAs (omy-miR-21, omy-miR-21t, omy-miR-122, omy-miR-125a, omy-miR-125b, omy-miR-125t, omy-miR-199-5a, omy-miR-295, omy-let-7a) and mRNA of 3 genes (histone H2A, ribosome protein rpl19, and Dicer which is a miRNA processing enzyme) in liver samples of whitefish exposed to Microcystin-LR (MC-LR) at a dose of 100µg*kg-1body weight for 24 or 48h. In the examined liver tissue, omymiR-122 showed the highest relative constitutive level, what is consistent with data obtained from fish and mammals. Unexpectedly, the reference H2A mRNA level was consistently up-regulated (over 20-fold; P<0.05) in fish liver after both 24 and 48h of exposure to MC-LR. The result may suggest that MC-LR acts as an initiator of specific cell-physiologic signals triggering DNA replication in fish liver cells. MC-LR treatment had no effect on the examined miRNAs levels, except for omy-miR-125a and omy-let-7a. Whereas omy-miR-125a was up-regulated (ER=2.68; S.E. 1.61-6.78; P<0.05), omy-let-7a was down-regulated (ER=0.55; S.E. 0.32-0.79; P<0.05) in whitefish liver after 48h of the treatment with MC-LR, when compared to controls. More work with the fish is essential for understanding the crosstalk of the regulatory network controlled by the two miRNAs in the context of MC-LR toxicity.
EN
While estrogenic properties of mycotoxin zearalenone (ZEA) has been an extensively studied issue, little is known about molecular background of its biological responses that cannot be simply explained by the estrogenic potential. The present study describes effects of ZEA (10mg*kg-1 body weight) in binary mixtures either with benzo[a]pyrene (B[a]P), or with 17β-estradiol (E2) on ER- and AhR-dependent gene expression in juvenile rainbow trout liver, evaluated using Real-Time qPCR. The study revealed dual nature of ZEA, as the treatment with this compound alone increased mRNA levels of both ER- and AhR-mediated gene expression. However, our results did not show any synergistic or additive effect of ZEA in binary mixures with E2 or B[a]P on studied gene expression levels. Whether the intriguing potential of ZEA to elicit distinct signals was a result of binding affinity to AhR or/and ER and AhR mutual receptor interactions, should be investigated in further experiments.
EN
This attempt to synthesize the present state of knowledge on the correlation between the structure of chosen PAHs and their mutagenic or carcinogenic activity shows that the dependencies cannot be treated univocally. Extreme complexity of the subject, typical for investigations on the boundary between chemistry and biology, inclines one to search for new ways of synthesis of new compounds, which would be of interest for biologists studying the problem of chemical carcinogenesis. On the other hand, there is a strong need for elaborating a standardized test that would explicitly define the carcinogenic properties of a specific compound. The use of a variety of methods for the determination of carcinogenic potency of chemical compounds combined with simultaneous differentiation of structure of examined compounds, sometimes leads to contradictory conclusions. Correlation between mutagenic and carcinogenic activity of some compounds is also sometimes questionable. Modern molecular biology should allow for elaborating a method which would give a satisfactory and reliable answer to the fundamental question: is a specific compound carcinogenous for a human organism? And if so - to what extent?
EN
The mutagenicity of two, newly synthesized amino-derivatives of cyclopenta[c]phenanthrene (CP[c]Ph) was investigated in the Ames test using TA98 and TA100 histidine dependent Salmonella typhimurium strains. Neither of the examined compounds showed mutagenic activity without metabolic activation. The incorporation of an activation system caused high mutagenicity of CP[c]Ph derivatives in both S. typhimurium strains. The results are compared with the previous data on the mutagenic activity of unsubstituted CP[c]Ph and possible mechanisms of activation are discussed.
EN
We assessed the effects of cyclopenta[c]phenanthrene (CP[c]Ph) and benzo[a]pyrene (B[a]P; positive control) on CYP1A gene expression in brain of juvenile rainbow trout (Oncorhynchus mykiss) using the quantitative reverse transcription polymerase chain reaction (Q-RT-PCR). A group of hatchery raised rainbow trout, with an average body mass of 49.4 g and total length of 15.5 cm were given an intraperitoneal injection (10 mg*kg-1) of either CP[c]Ph or B[a]P in corn oil (2 mg*mi-1 corn oil) or corn oil alone (control). After 24 and 48 h, trout brains were collected for mRNA isolation and analysis. After 24 hours of the exposure, only B[a]P-treated rainbow trout had 10-fold higher number of CYP1A transcripts (mean = 3.63*106 transcripts*µg-1 total RNA) than control fish (3.24*105 transcripts*µg-1 total RNA; Tukey test, P<0.05). After 48 hrs, significantly higher levels of CYP1A expression (Tukey test, P<0.001) were found in either CP[c]Ph- or B[a]P- induced group (1.45*106 and 6.92*106 transcriptsźµg-1 total RNA, respectively) over a control group (mean=1.41*105 transcripts*µg-1 total RNA). The finding that CYP1A in brain tissue was inducible by CP[c]Ph, a polycyclic aromatic hydrocarbon (PAH) of different than B[a]P planar characteristics, may further validate the use of rainbow trout brain CYP1A mRNA levels as a biomarker of PAH exposure.
EN
4-Aryl-substituted 1-aminonaphthalene-2-carbonitriles and 3-phenyl-1-oxo-1H-indene-2-carbonitriles were prepared in a two--step synthesis from appropriate carbonyl compounds and were evaluated in vitro for growth-inhibiting activity against some phytopathogenic fungi: Fusarium culmorum, Penicillium expansum, Botrytis cinerea and Alternaria species. The results were compared with the activity of a commercial fungicide - captan. The highest fungistatic activity was revealed by 1-amino-4-phenylnaphthalene-2-carbonitrile. An efficient synthesis of 1-amino-4-arylnaphthalene-2-carbonitriles involves condensation of diarylacetaldehydes or 1,1-diarylacetones with malonodinitrile and cyclization of the obtained aryl-ylidenemalonodinitriles in concentrated sulfuric acid. The benzannulation reaction is accompanied by a quasi-aromatic rearrangement. The indenone derivatives were synthesized in the same manner from substituted benzophenones. As the formation of five-membered ring during the cyclization of diarylmethylidenemalonodinitriles was less effective, the rearrangement was not observed.
EN
Mutagenic properties of four different diphenylthiophenes: 3,4-diphenylthiophene, 3,4-di(4'-methylphenyl)thiophene, 3,4-di(4'-methoxyphenyl)thiophene and 3,4-di(4'-pentoxyphenyl)thiophene were investigated applying the Salmonella test. The research was done on two strains of Salmonella Typhimurium: TA98 and TA100, tested in two variants: with (+S9) and without (-S9) enzymatic activation. Only one compound 3,4-di(4'-methylphenyl)thiophene showed mutagenic activity when studied with metabolic activation (+S9) and its mutagenic rate (MR) score was 3.41 for the dose of 10.00µg*plate-1. Other studied compounds did not show any mutagenic activity (+/-S9) and their MR score did not exceed the threshold value of 2.0.
EN
Heterocyclic aromatic compounds containing sulfur (S-HET), have been detected in air, soil, marine environment and freshwater sediment. Toxicity and mutagenicity data of this class of substances are scarce. The present study focuses on implications of two aryl thiophenes and their mutagenic properties in Salmonella/microsome test. In our experiment only 2,4-diphenylthiophene showed little mutagenic effect in both variants of activaction (+/-S9) in strain TA100. Thiophene ring joined to K-region of phenanthrene did not change the biological activity of 3,6-dimetoxyphenanthro [9,10-c]thiophene and this compound did not show mutagenic potency.
EN
Toxic effects of polycyclic aromatic hydrocarbons (PAHs) have been extensively studied in fish, although knowledge concerning biological activities of phenanthrene and its derivatives remains still incomplete. The aim of this work was to evaluate lethal and sublethal effects of benzo(a)pyrene, phenanthrene and phenanthrene derivatives (1-methylphenanthrene, 4-methylphenanthrene, 1-phenylphenanthrene and 4-phenylphenanthrene) on zebrafish (Danio rerio) larvae. We conducted acute toxicity test, using 96h static renewal exposure to a series of the PAH concentrations (0.05, 0.50, 5.00, 50.00µmol*l-1), to determine the No Effect Concentration (NEC) value for each compound examined. The mean NEC estimates obtained in the study were 5.16۪.45µmol*l-1 (B[a]P), 4.88۪.13µmol*l-1 (Ph), 40.24䔰.93µmol*l-1 (1P-Ph), 47.92ۭ.61µmol*l-1 (1M-Ph), 24.31۱.33µmol*l-1 (4P-Ph) and 3.11۫.01µmol*l-1 (4M-Ph) and suggested the following order of PAH toxicities on Danio rerio larvae: 4M-Ph>Ph˜B[a]P>4PPhᲙP-Ph>1M-Ph. To gain insight into possible molecular mechanisms of apparent toxicity of phenanthrene derivatives on zebrafish larvae, we examined mRNA expression of cyp1a, cyp1b1, and vtg genes in the larvae exposed for 48h to a PAH concentration of 0.50µmol*l-1. Whereas the larvae exposed to each tested PAH displayed many developmental abnormalities (i.e. pericardial and yolk sac edema, dorsal curvature, or tail malformations), no significant upregulation of cyp1a and cyp1b1 mRNA was observed, except for zebrafish exposed to B[a]P. However, significant reduction of vtg mRNA was observed in the larvae exposed to phenanthrene and its 4P- derivative. The results may contribute to the development of a new knowledge about effects of structurally diverse phenanthrene derivatives on vertebrate organisms.
EN
Polycyclic aromatic hydrocarbons (PAHs) are widely spread environmental pollutants mainly originating from anthropogenic sources such as fossil fuel combustion, industries, and others. Although a large body of literature exists on the toxicity and carcinogenicity of PAHs, primarily benzo[a]pyrene, toxicity data for phenanthrene deriveratives are very limited. The main aim of the experiment was to investigate if there exists correlation between molecular structure and mutagenic activity of four phenanthrene derivatives: 1 methylphenanthrene, 4 methylphenanthrene, 1 phenylphenanthrene, and 4 phenylphenanthrene. An Ames assay using two strains of histidine dependent Salmonella Typhimurium (TA98 and TA100) was conducted to assess the mutagenic activity of studied compounds both in the presence (+S9) and in the absence (-S9) of an exogenous source of metabolic activation. The compounds were also tested in an in vitro chromosome aberration assay in which V-79 cells were exposed to the phenanthrene derivatives investigated both in the presence and in the absence of metabolic activation. The phenylphenanthrenes showed no mutagenic effect. These compounds occasionally induced significant decrease in the number of revertants in the Ames test. The greatest mutagenic effects were observed for 1 methylphenanthrene after metabolic activation (+S9). In the micronucleus test the greatest mutagenic effect was observed for 4 methylphenanthrene also in the presence of metabolic activation system. The results obtained are comparable to those reported earlier for the methylphenanthrenes.
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