RESUMO
Interest in life-cycle assessment (LCA) for foodstuffs has increased over recent years. In the same period, there could also be noticed an increased focus on food losses and waste (FLW); hence the need for guidance or a method that assesses the environmental impacts of FLW when analyzing agrifood systems with the use of LCA. This study introduces a method of assessing FLW in the context of supply chain LCA through: i) calculating the impacts of the food supply chain with the associated FLW, in order to account for the overall environmental impacts; and ii) assessing the benefits of FLW prevention strategies. After the method is presented, a practical application follows. The results of this study show that this method is able to aid LCA practitioners to include FLW on their LCA for food studies so as not to underestimate the impacts. Furthermore, the method is able to aid decision makers to assess the benefits of implementing a FLW prevention action in comparison to a baseline scenario. This method has some limitations: the attributional approach, lack of guidelines on how to estimate prevention potentials, as well as lack of guidelines to estimate additional impacts due to prevention actions.
Assuntos
Gerenciamento de Resíduos , Tomada de Decisões , Meio Ambiente , Alimentos , Abastecimento de AlimentosRESUMO
The morphology as well as the electric and electronic properties of aryl diazonium, in particular 4-nitrobenzene-diazonium (NBD), films on HOPG surfaces have been studied at the nanoscale level. By controlling the 2,2-diphenyl-1-picrylhydrazyl concentration during the NBD film growth, we have been able to control the thickness of the layer. The implications of NBD submonolayer adsorption on the electrical properties of this system have been analysed through Density Functional Theory (DFT) calculations, Atomic Force (AFM), Electric Force (EFM) and Kelvin Probe Force (KPFM) microscopies. DFT simulations showed that the NBD molecule adsorbs almost perpendicularly to the HOPG surface, which was confirmed experimentally through AFM imaging in the dynamic mode. In addition, DFT calculations showed that the adsorbed NBD has an appreciable dipole moment directed towards the HOPG surface and along the vertical direction of the HOPG surface. The existence of this dipole is the origin of the EFM contrast observed between the NBD-free and NBD-covered regions when a bias of -2 V was applied to the tip. Besides, the KPFM measurements show that the NBD adsorption leads to higher work function values, which is in agreement with the DFT calculations. Noticeably, our studies show that the KPFM signal is sensitive to the partial NBD coverage of the HOPG surface below the monolayer level.
RESUMO
Gold nanoparticle electrodeposition on a modified HOPG surface with a monolayer organic film based on aryl diazonium chemistry has been studied. This organic monolayer is electrochemically grown with the use of 2,2-diphenyl-1-picrylhydrazyl (DPPH), a radical scavenger. The electrodeposition of gold on this modified surface is highly favored resulting in an AuNP surface density comparable to that found on glassy carbon. AuNPs grow only in the areas covered by the organic monolayer leaving free clean HOPG zones. A progressive mechanism for the nucleation and growth is followed giving hemispherical AuNPs, homogeneously distributed on the surface and their sizes can be well controlled by the applied electrodeposition potential. By using AFM, C-AFM and electrochemical measurements with the aid of two redox probes, namely Fe(CN)6(4-)/Fe(CN)6(3-) and dopamine, relevant results about the electrochemical modified surface as well as the gold nanoparticles electrodeposited on them are obtained.
RESUMO
Reductive electrodesorption has been used to produce "naked" gold nanoparticles (AuNPs) 3 nm in size on HOPG from different thiolate-capped AuNPs. The clean AuNPs transform the electrocatalytic inert HOPG into an active surface for hydrogen peroxide electroreduction, causing a lowering of the cathodic overpotential of 0.25 V with respect to the Au(111) surface. Compared to the plain gold substrates, the nanostructures promote only a slight increase in the hydrogen evolution reaction. In a second modification step a â¼1 nm thick melanin-iron coating is electrochemically formed around the AuNPs. This ultrathin melanin-iron coating largely improves the catalytic activity of the bare AuNPs for both hydrogen peroxide electroreduction and hydrogen evolution reaction. This strategy, which integrates electrochemistry and nanotechnology, can be applied to the preparation of efficient "naked" AuNPs and organic-iron capped AuNPs catalysts.
Assuntos
Ouro/química , Grafite/química , Melaninas/química , Nanoestruturas/química , Nanoestruturas/ultraestrutura , Catálise , Substâncias Macromoleculares/química , Teste de Materiais , Conformação Molecular , Tamanho da Partícula , Propriedades de SuperfícieRESUMO
Disinfection by-products (DBPs) are chemicals that are produced as a result of chlorine being added to water for disinfection. As well as the halogenated DBPs, N-nitrosamines have recently been identified as DBPs, especially when amines and ammonia ions are present in raw water. In this work, the genotoxicity of two nitrosamines, namely nitrosodimethylamine (NDMA) and nitrosodiethylamine (NDEA), has been studied in cultured human cells. To evaluate their genotoxic potential two assays were used, the comet assay and the micronucleus test. The comet assay measures the induction of single and double-strand breaks, and also reveals the induced oxidative DNA damage by using endoIII and FPG enzymes. Chromosomal damage was evaluated by means of the cytokinesis-blocked micronucleus test. The results of the comet assay show that both compounds are slightly genotoxic but only at high concentrations, NDEA being more effective than NDMA. Enzyme treatments revealed that only NDEA was able to produce increased levels of oxidized bases, mainly in purine sites. The results obtained in the micronucleus assay, which measures the capacity of the tested agents to induce clastogenic and/or aneugenic effects, are negative for both of the nitrosamines evaluated, either using TK6 cells or human peripheral blood lymphocytes. Taking into account the very high concentrations needed to produce DNA damage, our data suggest a low, if existent, genotoxic risk associated with the presence of these compounds in drinking water.
Assuntos
Dietilnitrosamina/toxicidade , Dimetilnitrosamina/toxicidade , Desinfetantes/toxicidade , Mutagênicos/toxicidade , Animais , Linhagem Celular , Ensaio Cometa , Dano ao DNA , Humanos , Masculino , Testes para Micronúcleos , Estresse Oxidativo , Ratos , Ratos Sprague-DawleyRESUMO
A comparative study on the thermodynamic stability of the lying down (LD) and standing up (SU) phases of alpha,omega-butanedithiol (BDT) on unreconstructed (U) and on reconstructed (R) Au(111) surfaces is presented. The R surface is made of dithiol-Au adatom units. Density functional calculations (DFT) allow the estimation of the adsorption energy of the LD and SU BDT phases on both substrates. Surface free energies based on the DFT calculations show the coverage of the clean Au(111) surface by the LD phase, and the LD to SU phase transition as the chemical potential of the BDT molecule is increased. The LD and SU phases are more stable on R than on U substrates, suggesting that the Au(111) surface should reconstruct upon BDT adsorption. The stability analysis is extended to longer alpha,omega-dithiols. Results reveal that the LD to SU phase transition is favored as the hydrocarbon chain length of the dithiol molecule is increased. Changes in the hydrogen pressure affect the formation of the LD phase, while they have only minor effects on the LD to SU phase transitions. Our calculations explain the influence of the number of carbon atoms in the hydrocarbon chains, hydrogen pressure and dithiol pressure (or concentration) on dithiol adsorption, and phase transitions. This information is relevant to control the coverage, reactivity, and surface chemistry of the alpha,omega-dithiol self-assembled monolayers on Au surfaces.
RESUMO
The redox potential of the T1 copper site of laccase from Fusarium proliferatum was determined by titration to be about 510 mV vs. SCE (750 mV vs. NHE), which makes it a high redox potential enzyme. Anaerobic electron transfer reactions between laccase and carbon and gold electrodes were detected, both in solution and when the enzyme was adsorbed on these surfaces. In solution, a single high-potential signal (660 mV vs. SCE) was recorded at the carbon surfaces, attributable to the T1 copper site of the enzyme. However, a well-defined oxidative process at about 660 mV and an anodic wave at 350 mV vs. SCE were recorded at the gold electrode, respectively associated with the T1 and T2 copper sites. Laccase-modified carbon electrodes behaved analogously when the enzyme was in solution, unlike laccase adsorbed on gold, which showed only a low-potential signal. Laccase molecules were successfully imaged by AFM; obtaining a thick compact stable film on Au(111), and large aggregates forming a complex network of small branches leaving voids on the HOPG surface. Laccase-modified carbon electrodes retained significant enzymatic activity, efficiently oxidising violuric acid and reducing molecular oxygen. Explanations are proposed for how protein-film organisation affects the electrode function.
Assuntos
Carbono/química , Fusarium/enzimologia , Ouro/química , Lacase/química , Lacase/metabolismo , Adsorção , Anaerobiose , Barbitúricos/metabolismo , Biocatálise , Domínio Catalítico , Cobre , Eletroquímica , Eletrodos , Transporte de Elétrons , Estabilidade Enzimática , Grafite/química , Concentração de Íons de Hidrogênio , Microscopia de Força Atômica , Oxigênio/metabolismo , SoluçõesRESUMO
Oral delivery of IL-10 by genetically modified Lactococcus lactis (LL-pTmIL10) has been shown to efficiently reduce intestinal inflammation in mice with chronic colitis, but the mechanisms involved have not been elucidated. It has been suggested that IL-10 controls intestinal inflammation by inhibiting microbe-induced activation of dendritic cells. We therefore investigated whether LL-pTmIL10 can modulate the functions of bone marrow-derived dendritic cells (BM-DC) responding to LPS. Incubation of these cells with LL-pTmIL10 or with the control strain LL-pTREX reduced their ability to activate allogeneic T-cell proliferation. However, in contrast to LL-pTREX, LL-pTmIL10 inhibited the LPS-stimulated secretion of MCP-1 by BM-DC and reduced the synergistic up-regulation of IL-12/IL-23p40. In addition, LL-pTmIL10 treatment of LPS-stimulated BM-DC significantly inhibited their capacity to induce strong secretion of IL-17 by CD4+ T cells. Our data suggest that the beneficial effects of LL-pTmIL10 treatment during chronic colitis might involve inhibition of CD4+ Th17 cells and a reduced accumulation of these cells as well as other immune cells at the site of inflammation.
Assuntos
Células da Medula Óssea/efeitos dos fármacos , Células Dendríticas/efeitos dos fármacos , Interleucina-10/fisiologia , Lactococcus lactis/genética , Lipopolissacarídeos/farmacologia , Probióticos/farmacologia , Animais , Células da Medula Óssea/fisiologia , Quimiocina CCL2/metabolismo , Células Dendríticas/fisiologia , Feminino , Engenharia Genética , Interleucina-12/metabolismo , Interleucina-17/metabolismo , Interleucina-23/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BLRESUMO
Genetic polymorphisms have shown to be susceptibility factors playing an important role in the development of most cancers. Nevertheless, as far as we know, only few studies have been conducted linking thyroid cancer incidence and GST polymorphisms, and no data are available on the possible association between NAT2 polymorphisms and thyroid cancer risk. The possible relationship between polymorphism at the GSTM1, GSTT1, GSTP1, and NAT2 genes and increased susceptibility to thyroid cancer has been evaluated in 176 thyroid cancer patients and 167 healthy controls, all from the urban district of Barcelona (Spain). The results indicate a clear role of the C481T change, present in several NAT2*5 alleles [odds ratio (OR)=0.58; 95% confidence interval (95% CI)=0.35-0.98]. Thus, those individuals carrying this change are less prone to develop thyroid cancer, mainly of the papillary type. In addition, there is a tendency towards the over-representation of the GSTM1 null genotype among thyroid cancer patients, particularly in those patients with papillary type tumor. The same is observed for the GSTM1 and GSTT1 null genotypes combination, and for other combinations with different NAT2 polymorphisms. The combinations involving the NAT2*6 and NAT2*7 genotypes showed the most important effect, and individuals carrying both alleles present a higher risk of thyroid cancer (OR=7.36; 95% CI=0.85-63.47), mainly for the follicular type (OR=17.94; 95% CI=1.34-238.70). The combination of NAT2*5 with NAT2*7 was also found to increase 5.26 (95% CI=1.07-25.76) times the risk of thyroid cancer. In conclusion, our results show that NAT2 polymorphisms play a significant role in thyroid cancer risk modulation.
Assuntos
Arilamina N-Acetiltransferase/genética , Glutationa Transferase/genética , Polimorfismo Genético , Neoplasias da Glândula Tireoide/genética , Carcinoma Papilar/genética , Feminino , Predisposição Genética para Doença , Glutationa S-Transferase pi/genética , Humanos , Masculino , EspanhaRESUMO
The molecular basis of the interindividual differences of normal individuals to ionizing radiation is poorly understood. Several studies in telomerase KO mice with short telomeres have uncovered an inverse relationship between telomere length and radiation sensitivity. The present work aims to determine if chromosome radiosensitivity is correlated with telomere length in healthy individuals. With this purpose, individual radiosensitivity was determined by the micronucleus assay in peripheral blood lymphocytes from two groups of individuals of the same age but with highly heterogeneous telomere length, selected from a population of 181 individuals where we previously measured telomere length. Our study demonstrates that telomere length modulates chromosome in vitro radiosensitivity in healthy individuals as the group with short telomeres presented higher frequencies of ionizing radiation-induced micronuclei when compared to the long telomeres group. This result supports the conclusion that individual telomere length acts as biomarker of individual chromosome instability upon exposure to ionizing radiation.
Assuntos
Instabilidade Cromossômica/efeitos da radiação , Cromossomos Humanos/efeitos da radiação , Linfócitos/efeitos dos fármacos , Tolerância a Radiação/genética , Telômero , Adulto , Células Cultivadas , Humanos , Testes para Micronúcleos , Valores de ReferênciaRESUMO
Peptides of the trefoil factor family (TFF) are expressed along the gastro-intestinal tract. They protect mucous epithelia from damage and contribute to mucosal repair, which is essential for preventing inflammation. Moreover, it has been suggested that TFF2 and TFF3, in particular, play a role in regulating immune responses. Depending on their activation status, dendritic cells (DC) can initiate either tolerance or immunity. This study, by comparing LPS-induced maturation of mTFF3-treated DC and non-treated DC, investigated whether murine TFF3 directly regulated DC function. mTFF3-treated DC and non-treated DC did not differ phenotypically or functionally. Both populations expressed, both before and after LPS-stimulation, similar levels of co-stimulatory molecules and cytokines, and were both efficient stimulators of T-cells. Our results suggest that mTFF3 does not govern immune responses on the level of DC function.
Assuntos
Células Dendríticas/imunologia , Lipopolissacarídeos/metabolismo , Mucinas/metabolismo , Animais , Diferenciação Celular/imunologia , Ciclo-Oxigenase 2/biossíntese , Citocinas/biossíntese , Células Dendríticas/citologia , Células Dendríticas/metabolismo , Feminino , Imunidade nas Mucosas , Ativação Linfocitária/imunologia , Camundongos , Mucinas/imunologia , RNA Mensageiro/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Linfócitos T/imunologia , Linfócitos T/metabolismo , Fator Trefoil-3RESUMO
The alkaline single-cell gel electrophoresis (or Comet) assay was applied to evaluate the eventual DNA damage induced by the triphenolic metabolite of benzene, 1,2,4-benzenetriol (BT), in isolated human lymphocytes. Prior to BT treatment, ranging from 5 to 50 microM, a supplementation with glutathione (GSH, 350 microg/ml) was carried out to assess whether GSH may have a modulating effect on the Comet response. The effect of a fixed dose of BT was also evaluated in the presence of the exogenous antioxidant vitamin C (40 and 200 microM). Additionally, we investigated whether the polymorphism of glutathione S-transferase T1 (GSTT1) gene may affect the individual level of BT-induced DNA damage in vitro. For all donors included in the present study, BT produced a significant dose-response relationship. No clear effect of GSH preincubation was seen on the BT-induced response. On the contrary, a significant reduction of DNA damage was observed in the presence of vitamin C (at least at 200 microM). Although our data suggest some individual differences according to the GSTT1 genotype in the outcome of the Comet assay, a large number of individuals should be studied in further investigations to obtain reliable conclusions.
Assuntos
Ácido Ascórbico/farmacologia , Benzeno/toxicidade , Dano ao DNA/efeitos dos fármacos , Glutationa/farmacologia , Linfócitos/efeitos dos fármacos , Adulto , Benzeno/metabolismo , Separação Celular , Ensaio Cometa/métodos , Dano ao DNA/fisiologia , Relação Dose-Resposta a Droga , Humanos , Linfócitos/fisiologia , MasculinoRESUMO
To determine the genotoxic risk associated to environmental arsenic exposure, the frequency of micronuclei in buccal cells (BCMN) of people drinking arsenic-contaminated water has been evaluated. A group of 105 individuals from the Antofagasta region (north Chile), and 102 individuals from the area of Concepcion, used as reference group, were included in the study. Arsenic concentration in drinking water was high (0.75 mg/L) in the Antofagasta area, 75-fold the maximum recommended level by WHO (0.01 mg/L), while the values obtained in Concepcion were significantly lower (0.002 mg/L). Individual measures of arsenic exposure were also determined in fingernails, which clearly confirm the existence of chronic exposure in the sampled populations from the Antofagasta region (10.15 microg/g versus 3.57 microg/g). The cytogenetic results indicate that, although the BCMN frequency is higher in exposed than in controls, this increase does not attain statistical significance. When the exposure biomarkers were related with the cytogenetic values, no correlations were observed between BCMN and arsenic content in water or in fingernails. In addition, the genotoxicity values do not seem to be related to the ethnic origin from people belonging to the exposed group. As a conclusion it appears that, in the studied population, the chronic ingestion of arsenic-contaminated water does not induce cytogenetic damage, measured as micronuclei, in the cells of the oral mucous in a significant extent.
Assuntos
Arsenicais/efeitos adversos , Exposição Ambiental/efeitos adversos , Monitoramento Ambiental , Micronúcleos com Defeito Cromossômico/induzido quimicamente , Mucosa Bucal/efeitos dos fármacos , Poluentes Químicos da Água/efeitos adversos , Adulto , Chile , Feminino , Humanos , Masculino , Testes para Micronúcleos , Mucosa Bucal/citologia , Unhas/química , Abastecimento de Água/normasRESUMO
In the present study we have evaluated whether or not environmental exposure to arsenic in ground drinking-water results in a significant increase in the frequency of micronuclei (MN) in peripheral blood lymphocytes. Thus, 106 individuals from the Antofagasta region (North Chile), together with 111 individuals from the area of Concepción, were used in this investigation. In the Antofagasta area, arsenic levels in drinking-water as high as 0.750 mg/L were measured. In Concepción, located about 2500 km towards the south and used as reference area, arsenic levels in tap water were as low as 0.002 mg/L. The total content of arsenic in fingernails was determined as a biomarker of individual exposure. The cytogenetic results obtained in this study indicate that in the exposed group the overall frequency of binucleated micronucleated cells (BNMN) is higher than in the reference group, the difference being statistically significant. In addition, no differences were found between the exposed and the reference groups, regarding the cytokinesis-block proliferation index (CBPI). No association was observed between BNMN and arsenic content in water or arsenic in fingernails. On the other hand, when the exposed group was divided according to their Atacameno or Caucasian ethnicity, no significant differences were observed between them. In addition, as usually found in other human biomonitoring studies, sex and age are factors that modulate the frequency of MN in both exposed and reference populations.
Assuntos
Intoxicação por Arsênico/epidemiologia , Arsênio/análise , Arsênio/toxicidade , Carcinógenos/análise , Exposição Ambiental , Micronúcleos com Defeito Cromossômico , Adulto , Biomarcadores , Células Cultivadas , Chile , Feminino , Testes Hematológicos , Humanos , Linfócitos/citologia , Linfócitos/fisiologia , Masculino , Testes para Micronúcleos , Unhas/química , Poluentes Químicos da Água/análise , Poluentes Químicos da Água/toxicidade , Poluição Química da Água , Abastecimento de ÁguaRESUMO
Fanconi anaemia is an autosomal recessive disease characterized by chromosome fragility, multiple congenital abnormalities, progressive bone marrow failure and a high predisposition to develop malignancies. Most of the Fanconi anaemia patients belong to complementation group FA-A due to mutations in the FANCA gene. This gene contains 43 exons along a 4.3-kb coding sequence with a very heterogeneous mutational spectrum that makes the mutation screening of FANCA a difficult task. In addition, as the FANCA gene is rich in Alu sequences, it was reported that Alu-mediated recombination led to large intragenic deletions that cannot be detected in heterozygous state by conventional PCR, SSCP analysis, or DNA sequencing. To overcome this problem, a method based on quantitative fluorescent multiplex PCR was proposed to detect intragenic deletions in FANCA involving the most frequently deleted exons (exons 5, 11, 17, 21 and 31). Here we apply the proposed method to detect intragenic deletions in 25 Spanish FA-A patients previously assigned to complementation group FA-A by FANCA cDNA retroviral transduction. A total of eight heterozygous deletions involving from one to more than 26 exons were detected. Thus, one third of the patients carried a large intragenic deletion that would have not been detected by conventional methods. These results are in agreement with previously published data and indicate that large intragenic deletions are one of the most frequent mutations leading to Fanconi anaemia. Consequently, this technology should be applied in future studies on FANCA to improve the mutation detection rate.
Assuntos
Proteínas de Ligação a DNA/genética , Anemia de Fanconi/genética , Deleção de Genes , Reação em Cadeia da Polimerase/métodos , Elementos Alu , Análise Mutacional de DNA , Éxons/genética , Anemia de Fanconi/epidemiologia , Proteína do Grupo de Complementação A da Anemia de Fanconi , Heterogeneidade Genética , Heterozigoto , Humanos , Espanha/epidemiologiaRESUMO
UNLABELLED: Lemierre syndrome is a severe postanginal sepsis complicated by internal jugular thrombophlebitis. We report on a 14 y-old girl affected by high fever, shivering chills, headache, severe lateral neck pain, left ocular proptosis and general malaise. Magnetic resonance imaging of the head and neck showed right internal jugular vein and sigmoid sinus thrombosis. Fusobacterium sp. was identified in the blood culture. CONCLUSION: Our report is a reminder that Lemierre syndrome still exists and remains potentially life threatening. A high index of suspicion is necessary to prompt diagnosis and treatment.
Assuntos
Infecções por Fusobacterium/diagnóstico , Infecções por Fusobacterium/terapia , Fusobacterium/isolamento & purificação , Veias Jugulares/diagnóstico por imagem , Sepse/diagnóstico , Sepse/terapia , Tromboflebite/diagnóstico , Tromboflebite/terapia , Adolescente , Feminino , Infecções por Fusobacterium/microbiologia , Humanos , Veias Jugulares/microbiologia , Radiografia , Sepse/microbiologia , Síndrome , Tromboflebite/microbiologiaRESUMO
Possible mechanisms by which liver antigen presenting cells (APC) may facilitate tolerance induction are discussed. Tolerance may be facilitated by a distinctive combination of factors, linked to the unique anatomical and microenvironmental features of the liver.
Assuntos
Células Apresentadoras de Antígenos/imunologia , Transplante de Fígado/imunologia , Humanos , Tolerância Imunológica , Imunologia de TransplantesRESUMO
Fanconi anaemia (FA) is a rare autosomal recessive disease characterized by increased spontaneous and DNA crosslinker-induced chromosome instability, progressive pancytopenia and cancer susceptibility. An increasing number of genes are involved in FA, including the breast cancer susceptibility gene BRCA2. Five of the FA proteins (FANCA, FANCC, FANCE, FANCF and FANCG) assemble in a complex that is required for FANCD2 activation in response to DNA crosslinks. Active FANCD2 then interacts with BRCA1 and forms discrete nuclear foci. FANCD2 is independently phosphorylated by ATM (the protein whose gene is mutated in ataxia telangiectasia) in response to ionizing radiation. In addition, the FA proteins are interconnected with other nuclear and cytoplasmic factors all related to cellular responses to carcinogenic stress and to caretaker and gatekeeper functions. In this review, the most recently published data on the molecular biology of the FA pathway and its molecular crosstalk with ATM, BRCA1 and BRCA2, proteins involved in xenobiotic and reactive oxygen species metabolism, apoptosis, cell cycle control and telomere stability, are summarized. The currently available data indicate that FA is a central node in a complex nuclear and cytoplasmic network of tumour suppressor and genome stability pathways fully committed to prevent cancer.
Assuntos
Anemia de Fanconi/genética , Genes Supressores de Tumor , Apoptose , Proteínas Mutadas de Ataxia Telangiectasia , Ciclo Celular , Proteínas de Ciclo Celular , Núcleo Celular/metabolismo , Cromatina/genética , Reparo do DNA , Proteínas de Ligação a DNA , Anemia de Fanconi/metabolismo , Anemia de Fanconi/patologia , Proteína do Grupo de Complementação D2 da Anemia de Fanconi , Feminino , Genes BRCA1 , Genes BRCA2 , Humanos , Masculino , Modelos Biológicos , Mutação , Proteínas Nucleares/genética , Estresse Oxidativo , Proteínas Serina-Treonina Quinases/genética , Telômero/genética , Transcrição Gênica , Proteínas Supressoras de TumorRESUMO
Fanconi anaemia (FA) is a rare genetic syndrome of cancer susceptibility characterized by spontaneous and induced chromosome fragility, especially after treatment with cross-linking agents. Recent investigations showed interactions between FA proteins and chromatin remodelling factors. To investigate a potential uneven distribution of the FA pathway through the human genome depending on chromatin conformation, we have analysed chromosome breakage in the largest constitutively heterochromatic region in the human genome, the 1q12 band, in lymphocytes from FA patients, carriers and healthy controls after treatment with the cross-linking agents mitomycin-C (MMC) and diepoxybutane (DEB). As expected, a higher level of MMC-induced cytotoxicity and chromosome breakage was observed in cells from FA patients when compared with normal controls and carriers. However, the increase in 1q12 breakage after increasing concentrations of MMC was of a similar magnitude in FA patients, carriers and controls. Similarly, DEB induced a high level of overall genome chromosome fragility in cells from FA patients when compared with controls with no parallel increase in chromosome breaks specifically involving the heterochromatic band 1q12. We therefore conclude that, unlike the overall genome, the sensitivity of chromosome 1 constitutive heterochromatin to the chromosome breaking activity of cross-linking agents is independent of a functional FA pathway, indicating that the action of the FA pathway is unevenly distributed through the human genome.
Assuntos
Cromossomos Humanos Par 1 , Reagentes de Ligações Cruzadas/toxicidade , Compostos de Epóxi/toxicidade , Anemia de Fanconi/genética , Mitomicina/toxicidade , Mutagênicos/toxicidade , Estudos de Casos e Controles , Triagem de Portadores Genéticos , Humanos , Hibridização in Situ Fluorescente/métodosRESUMO
High inter- and intra-individual variability was reported in the level of DNA damage, both spontaneous and induced, when peripheral blood mononuclear leukocytes were used to perform the Comet assay. In order to find out the underlying causes for such variability, different subsets of T lymphocytes were isolated by immunomagnetic cell sorting. The level of DNA damage was evaluated with the alkaline version of the Comet assay by using three different parameters: tail moment, tail length and amount of DNA in the tail (%). Helper T cells (CD4+), cytotoxic T cells (CD8+), their negative fraction and the mixed cell population were evaluated both in untreated cells and after 10 and 20 microM H(2)O(2) treatments. Differences between cell subsets were only observed after H(2)O(2) treatment. The results indicate that, although CD4+ is the fraction with the highest induced level of genetic damage, this value is not high enough to explain the large inter- and intra-individual variability found.