RESUMO
The pharmacological potential of drugs must be evaluated to establish their potential therapeutic benefits and side effects. This evaluation includes assessment of the effects of hepatic enzymes that catalyse their metabolic activation. Previously, our research group synthesized and characterized a set of synthetic 3-alkyl pyridine alkaloid (3-APA) analogues that cause in vitro cytotoxic, genotoxic, and mutagenic effects in various human cancer cell lines. The present study aimed to evaluate these activities with the two most promising synthetic 3-APAs (3-APA 1 and 3-APA 2) against cell lines derived from breast cancer (MDA-MB-231), ovarian cancer (TOV-21 G) and lung fibroblasts (WI-26-VA4) with and without metabolic activation (S9 fraction). The cytotoxicity of the compounds was evaluated employing MTT and clonogenic assays. In addition, comet assays, γH2AX immunocytochemistry labelling assays and cytokinesis-block micronucleus tests were carried out to evaluate the potential of these compounds to induce chromosomal damage. The results obtained in the MTT assay showed that compound 3-APA 2 exhibited high selectivity index (SI) values (ranging between 21.0 and 92.6). In addition, the cytotoxicity of the compounds was clearly enhanced by metabolic activation. Moreover, both compounds were genotoxic and induced double-strand breaks in DNA and chromosomal lesions with and without S9. The cancer cell lines tested showed higher genotoxic sensitivity to the compounds than did the non-tumour cell line used as a reference. The genotoxic and mutagenic effects of the compounds were potentiated in experiments with metabolic activation. The data obtained in this study indicate that compound 3-APA 2 is more active against the human cancer cell lines tested, both with and without metabolic activation, and can therefore be considered a candidate drug to treat human ovarian and breast cancer.
Assuntos
Ativação Metabólica , Alcaloides/farmacologia , Antineoplásicos/farmacologia , Citocinese/efeitos dos fármacos , Dano ao DNA , Mutagênicos/farmacologia , Neoplasias/patologia , Ensaio Cometa , Humanos , Testes para Micronúcleos , Neoplasias/tratamento farmacológico , Neoplasias/genética , Células Tumorais CultivadasRESUMO
Organophosphate (OP) pesticides are biotransformed into metabolites such as dialkylphosphates (DAPs). We have evaluated the genotoxicity of malathion and its metabolite dimethylthiophosphate (DMTP) in the human hepatic cell lines HepG2 and WRL-68 and in peripheral blood mononuclear cells (PBMC). In the Cytokinesis-Block Micronucleus assay (CBMN), malathion and DMTP increased the frequencies of micronuclei (MN) and nucleoplasmic bridges (NPB). Malathion was primarily clastogenic whereas DMTP was aneuploidogenic. When HepG2 or WRL-68 cells were treated with DMTP in the presence of sulconazole, a non-specific cytochrome P450 inhibitor, MN frequency was reduced, indicating that DMTP genotoxicity requires P450-cataliyzed metabolism.
Assuntos
Citocinese/efeitos dos fármacos , Malation/farmacologia , Testes de Mutagenicidade , Mutagênicos/farmacologia , Núcleo Celular/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Humanos , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/patologia , Malation/toxicidade , Mutagênicos/toxicidade , Praguicidas/farmacologia , Praguicidas/toxicidadeRESUMO
Metallic nanoparticles such as silver (Ag NPs) and iron oxide (Fe3O4 NPs) nanoparticles are high production volume materials due to their applications in various consumer products, and in nanomedicine. However, their inherent toxicities to human cells remain a challenge. The present study was aimed at combining lipidomics data with common phenotypically-based toxicological assays to gain better understanding into cellular response to Ag NPs and Fe3O4 NPs exposure. HepG2 cells were exposed to different concentrations (3.125, 6.25, 12.5, 25, 50 and 100 µg/ml) of the nanoparticles for 24 h, after which they were assayed for toxic effects using toxicological assays like cytotoxicity, mutagenicity, apoptosis and oxidative stress. The cell membrane phospholipid profile of the cells was also performed using shotgun tandem mass spectrometry. The results showed that nanoparticles exposure resulted in concentration-dependent cytotoxicity as well as reduced cytokinesis-block proliferation index (CBPI). Also, there was an increase in the production of ROS and superoxide anions in exposed cells compared to the negative control. The lipidomics data revealed that nanoparticles exposure caused a modulation of the phospholipidome of the cells. A total of 155 lipid species were identified, out of which the fold changes of 23 were significant. The high number of differentially changed phosphatidylcholine species could be an indication that inflammation is one of the major mechanisms of toxicity of the nanoparticles to the cells.
Assuntos
Hepatócitos/efeitos dos fármacos , Nanopartículas Magnéticas de Óxido de Ferro/toxicidade , Nanopartículas Metálicas/toxicidade , Compostos de Prata/toxicidade , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Citocinese/efeitos dos fármacos , Relação Dose-Resposta a Droga , Células Hep G2 , Hepatócitos/metabolismo , Hepatócitos/patologia , Humanos , Lipidômica , Necrose , Estresse Oxidativo/efeitos dos fármacos , Fosfolipídeos/metabolismo , Espectrometria de Massas por Ionização por Electrospray , Superóxidos/metabolismo , Espectrometria de Massas em TandemRESUMO
BACKGROUND/AIM: Car painting is considered an occupational exposure job with high risk for cancer development, due to the association with harmful chemicals and mutagens. This study aimed to profile car painters occupationally exposed and determine its association with DNA damage and genomic instability. MATERIALS AND METHODS: We collected a questionnaire and buccal cells of 74 individuals (37 car painters and 37 non-exposed workers) paired by age, alcohol and smoking habits. The number of pyknotic cells, karyolitic cells, karyorrhetic cells, condensed chromatin, binucleated cells, basal cells, differentiated cells (DIFF), micronucleated cells and nuclear buds were evaluated using the Buccal Micronucleus Cytome Assay protocol. RESULTS: A statistically significant increase was observed in all parameters (p<0.05) in the exposed group, but DIFF showed a statistically significant decrease (p<0.001), compared to the control group. CONCLUSION: In association with the poor work environment and lack of personal and collective protective equipment, occupational exposure of car painters leads to high DNA damage, genomic instability and alterations in cellular kinetics.
Assuntos
Citocinese/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Instabilidade Genômica/efeitos dos fármacos , Mutagênicos/efeitos adversos , Exposição Ocupacional/efeitos adversos , Pintura/efeitos adversos , Adulto , Automóveis , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/genética , Citocinese/genética , Dano ao DNA/genética , Humanos , Masculino , Micronúcleos com Defeito Cromossômico/efeitos dos fármacos , Testes para Micronúcleos/métodos , Mucosa Bucal/efeitos dos fármacos , Neoplasias/induzido quimicamente , Neoplasias/genéticaRESUMO
Giardia trophozoites have developed resistance mechanisms to currently available compounds, leading to treatment failures. In this context, the development of new additional agents is mandatory. Sirtuins, which are class III NAD+-dependent histone deacetylases, have been considered important targets for the development of new anti-parasitic drugs. Here, we evaluated the activity of KH-TFMDI, a novel 3-arylideneindolin-2-one-type sirtuin inhibitor, on G. intestinalis trophozoites. This compound decreased the trophozoite growth presenting an IC50 value lower than nicotinamide, a moderately active inhibitor of yeast and human sirtuins. Light and electron microscopy analysis showed the presence of multinucleated cell clusters suggesting that the cytokinesis could be compromised in treated trophozoites. Cell rounding, concomitantly with the folding of the ventro-lateral flange and flagella internalization, was also observed. These cells eventually died by a mechanism which lead to DNA/nuclear damage, formation of multi-lamellar bodies and annexin V binding on the parasite surface. Taken together, these data show that KH-TFMDI has significant effects against G. intestinalis trophozoites proliferation and structural organization and suggest that histone deacetylation pathway should be explored on this protozoon as target for chemotherapy.
Assuntos
Antiprotozoários/farmacologia , Giardia lamblia/efeitos dos fármacos , Inibidores de Histona Desacetilases/farmacologia , Trofozoítos/efeitos dos fármacos , Células CACO-2 , Citocinese/efeitos dos fármacos , Giardia lamblia/citologia , Giardia lamblia/crescimento & desenvolvimento , Humanos , Concentração Inibidora 50 , Microscopia , Microscopia Eletrônica , Testes de Sensibilidade Parasitária , Trofozoítos/citologia , Trofozoítos/crescimento & desenvolvimentoRESUMO
The in vitro effect of enrofloxacin (EFZ) was tested on two experimental somatic bovine cells in vitro: peripheral lymphocytes (PLs) and cumulus cells (CCs). The cytotoxicity and genotoxicity of this veterinary antibiotic were assessed using 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assays, single-cell gel electrophoresis (SCGE) assay, and cytokinesis-block micronucleus cytome (CBMN cyt) assay. Cells were treated during 24 h, and three concentrations were tested (50 µg/mL, 100 µg/mL, 150 µg/mL). When EFZ was tested in PLs, the results demonstrated that the antibiotic was able to induce cell death and DNA damage with all concentrations. In addition, 50 µg/mL and 100 µg/mL EFZ increased frequencies of micronuclei (MNi). On the other hand, the highest EFZ concentration occasioned cellular cytotoxicity in CCs as evidenced by mitochondrial activity alterations. Nevertheless, EFZ was not able to induce DNA damage and MNi in CCs. These results represent the first experimental evidence of genotoxic and cytotoxic effects exerted by EFZ in bovine PLs and CCs.
Assuntos
Antibacterianos/toxicidade , Células do Cúmulo/efeitos dos fármacos , Enrofloxacina/toxicidade , Linfócitos/efeitos dos fármacos , Animais , Bovinos , Ensaio Cometa , Citocinese/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Feminino , Testes para Micronúcleos , Mitocôndrias/efeitos dos fármacos , Testes de Toxicidade/métodosRESUMO
Ribavirin is an important component of the treatment for hepatitis C virus (HCV) infection and, in combination with the new direct-acting antiviral (DAA) agents, comprises the major current therapeutic regimens. This study evaluated the cytotoxicity and chromosomal instability induced by ribavirin using the in vitro cytokinesis-block micronucleus cytome (CBMN-Cyt) assay in two cell lines with different expression levels of drug-metabolizing enzymes: human hepatocellular carcinoma cells (HepG2) and Chinese hamster ovary (CHO-K1) cells. HepG2 cells were treated with nine concentrations (from 15.3 µg/ml to 3.9 mg/ml) and CHO-K1 cells were exposed to eight concentrations (from 15.3 µg/ml to 1.9 mg/ml) of ribavirin for 24 h. Ribavirin inhibited cell proliferation in both cell lines, but at different concentrations: 3.9 mg/ml in HepG2 and 244.2 µg/ml in CHO-K1 cells. No significant differences were observed regarding aspects of cell death in HepG2 and CHO-K1 cells, reflecting the absence of cytotoxic effects associated to ribavirin. Ribavirin did not increase the frequency of nucleoplasmic bridges (NPBs) and nuclear bud (NBUD). However, when compared to the negative control, a significant increase in micronuclei (MNi) frequency was observed in both cell lines. However, chromosomal instability was induced by higher concentrations of ribavirin in HepG2 cells (from 61.1 to 976.8 µg/ml), compared with CHO-K1 cells (15.3 and 30.5 µg/ml). These results demonstrate the potential of ribavirin to promote chromosomal instability, and suggest that cells with different expressions of drug-metabolizing enzymes show different susceptibility to ribavirin effects.
Assuntos
Antivirais/toxicidade , Proliferação de Células/efeitos dos fármacos , Instabilidade Cromossômica/efeitos dos fármacos , Citocinese/efeitos dos fármacos , Micronúcleos com Defeito Cromossômico/induzido quimicamente , Ribavirina/toxicidade , Animais , Antivirais/metabolismo , Apoptose/efeitos dos fármacos , Células CHO , Cricetulus , Relação Dose-Resposta a Droga , Células Hep G2 , Humanos , Inativação Metabólica , Testes para Micronúcleos , Ribavirina/metabolismoRESUMO
Tuberculosis is a top infectious disease killer worldwide, caused by the bacteria Mycobacterium tuberculosis. Increasing incidences of multiple drug-resistance (MDR) strains are emerging as one of the major public health threats. However, the drugs in use are still incapable of controlling the appalling upsurge of MDR. In recent years a marked number of research groups have devoted their attention toward the development of specific and cost-effective antimicrobial agents against targeted MDR-Tuberculosis. In previous studies, ruthenium(II) complexes (SCAR) have shown a promising activity against MDR-Tuberculosis although few studies have indeed considered ruthenium toxicity. Therefore, within the preclinical requirements, we have sought to determine the cyto-genotoxicity of three SCAR complexes in this present study. The treatment with the SCARs induced a concentration-dependent decrease in cell viability in CHO-K1 and HepG2 cells. Based on the clonogenic survival, SCAR 5 was found to be more cytotoxic while SCAR 6 exhibited selectivity action on tumor cells. Although SCAR 4 and 5 did not indicate any mutagenic activity as evidenced by the Ames and Cytokinesis block micronucleus cytome assays, the complex SCAR 6 was found to engender a frameshift mutation detected by Salmonella typhimurium in the presence of S9. Similarly, we observed a chromosomal damage in HepG2 cells with significant increases of micronuclei and nucleoplasmic bridges. These data indicate that SCAR 4 and 5 complexes did not show genotoxicity in our models while SCAR 6 was considered mutagenic. This study presented a comprehensive genotoxic evaluation of SCAR complexes were shown to be genotoxic in vitro. All in all, further studies are required to fully elucidate how the properties can affect human health.
Assuntos
Antituberculosos/toxicidade , Complexos de Coordenação/toxicidade , Mutagênicos/toxicidade , Rutênio/toxicidade , Animais , Células CHO , Cricetulus , Citocinese/efeitos dos fármacos , Células Hep G2 , Humanos , Testes para Micronúcleos , Rutênio/química , Salmonella typhimurium/efeitos dos fármacosRESUMO
Leishmaniasis, a complex of diseases caused by protozoa of the genus Leishmania, is endemic in 98 countries, affecting approximately 12 million people worldwide. Current treatments for leishmaniasis have many disadvantages, such as toxicity, high costs, and prolonged treatment, making the development of new treatment alternatives highly relevant. Several studies have verified the antileishmanial activity of ß-carboline compounds. In the present study, we investigated the in vitro antileishmanial activity of N-butyl-[1-(4-methoxy)phenyl-9H-ß-carboline]-3-carboxamide (ß-CB) against Leishmania amazonensis. The compound was active against promastigote, axenic amastigote, and intracellular amastigote forms of L. amazonensis, exhibiting high selectivity for the parasite. Moreover, ß-CB did not exhibit hemolytic or mutagenic potential. Promastigotes treated with the alkaloid presented rounding of the body cell, cell membrane projections, an increase in the number of promastigotes presenting two flagella, and parasites of abnormal phenotype, with three or more flagella and/or nuclei. Furthermore, we observed an increase in the subpopulation of cells in the G2/M stage of the cell cycle. Altogether, these results suggest that ß-CB likely prevents cytokinesis, although it does not interfere with the duplication of cell structures. We also verified an increase in O2(·-) production and the accumulation of lipid storage bodies. Cell membrane integrity was maintained, in addition to the absence of phosphatidylserine externalization, DNA fragmentation, and autophagosomes. Although the possibility of an apoptotic process cannot be discarded, ß-CB likely exerts its antileishmanial activity through a cytostatic effect, thus preventing cellular proliferation.
Assuntos
Antiprotozoários/farmacologia , Carbolinas/farmacologia , Citocinese/efeitos dos fármacos , Citostáticos/farmacologia , Leishmania mexicana/efeitos dos fármacos , Leishmaniose Cutânea/tratamento farmacológico , Estágios do Ciclo de Vida/efeitos dos fármacos , Animais , Antiprotozoários/síntese química , Cultura Axênica , Carbolinas/síntese química , Linhagem Celular , Citostáticos/síntese química , Eritrócitos/efeitos dos fármacos , Feminino , Flagelos/efeitos dos fármacos , Hemólise/efeitos dos fármacos , Humanos , Concentração Inibidora 50 , Leishmania mexicana/crescimento & desenvolvimento , Leishmaniose Cutânea/microbiologia , Macrófagos/efeitos dos fármacos , Macrófagos/parasitologia , Masculino , Camundongos , Camundongos Endogâmicos BALB CRESUMO
The compounds 6-dimethylaminopurine and cycloheximide promote the successful production of cloned mammals and have been used in the development of embryos produced by somatic cell nuclear transfer. This study investigated the effects of 6-dimethylaminopurine and cycloheximide in vitro, using the thiazolyl blue tetrazolium bromide colorimetric assay to assess cytotoxicity, the trypan blue exclusion assay to assess cell viability, the comet assay to assess genotoxicity, and the micronucleus test with cytokinesis block to test mutagenicity. In addition, the comet assay and the micronucleus test were also performed on peripheral blood cells of 54 male Swiss mice, 35 g each, to assess the effects of the compounds in vivo. The results indicated that both 6-dimethylaminopurine and cycloheximide, at the concentrations and doses tested, were cytotoxic in vitro and genotoxic and mutagenic in vitro and in vivo, altered the nuclear division index in vitro, but did not diminish cell viability in vitro. Considering that alterations in DNA play important roles in mutagenesis, carcinogenesis, and morphofunctional teratogenesis and reduce embryonic viability, this study indicated that 6-dimethylaminopurine and cycloheximide utilized in the process of mammalian cloning may be responsible for the low embryo viability commonly seen in nuclear transfer after implantation in utero.