RESUMO
There is a growing body of evidence indicating that the mechanisms that control genome stability are of key importance in the development and function of the nervous system. The major threat for neurons is oxidative DNA damage, which is repaired by the base excision repair (BER) pathway. Functional mutations of enzymes that are involved in the processing of single-strand breaks (SSB) that are generated during BER have been causally associated with syndromes that present important neurological alterations and cognitive decline. In this review, the plasticity of BER during neurogenesis and the importance of an efficient BER for correct brain function will be specifically addressed paying particular attention to the brain region and neuron-selectivity in SSB repair-associated neurological syndromes and age-related neurodegenerative diseases.
Assuntos
Encéfalo/metabolismo , Dano ao DNA , Reparo do DNA , Doenças do Sistema Nervoso/genética , Neurônios/metabolismo , Estresse Oxidativo , Animais , Quebras de DNA de Cadeia Simples , Humanos , Neurogênese/genéticaRESUMO
Endocrine-disrupting compounds are chemicals that alter the normal functioning of the endocrine system of living organisms. They can be natural (N-EDCs) or synthetic compounds (S-EDCs). N-EDCs can belong to different groups, such as phytoestrogens (PEs), including flavonoids, or mycotoxins originating from plants or fungi, and cyanotoxins, derived from bacteria. Humans encounter these substances in their daily lives. The aim of this rapid review (RR) is to provide a fine mapping of N-EDCs and their toxicological effects on human health in terms of various medical conditions or adverse consequences. This work is based on an extensive literature search and follows a rigorous step-by-step approach (search strategy, analysis strategy and data extraction), to select eligible papers published between 2019 and 2023 in the PubMed database, and to define a set of aspects characterizing N-EDCs and the different human target systems. Of the N-EDCs identified in this RR, flavonoids are the most representative class. Male and female reproductive systems were the targets most affected by N-EDCs, followed by the endocrine, nervous, bone and cardiovascular systems. In addition, the perinatal, pubertal and pregnancy periods were found to be particularly susceptible to natural endocrine disruptors. Considering their current daily use, more toxicological research on N-EDCs is required.
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The survival of the honey bee (Apis mellifera), which has a crucial role in pollination and ecosystem maintenance, is threatened by many pathogens, including parasites, bacteria, fungi and viruses. The ectoparasite Varroa destructor is considered the major cause of the worldwide decline in honey bee colony health. Although several synthetic acaricides are available to control Varroa infestations, resistant mites and side effects on bees have been documented. The development of natural alternatives for mite control is therefore encouraged. The study aims at exploring the effects of cinnamon and oregano essential oils (EOs) and of a mixed fruit cocktail juice on mite infestation levels and bee colony health. A multi-method study including hive inspection, mite count, molecular detection of fungal, bacterial and viral pathogens, analysis of defensin-1, hymenoptaecin and vitellogenin immune gene expression, colony density and honey production data, was conducted in a 20-hive experimental apiary. The colonies were divided into five groups: four treatment groups and one control group. The treatment groups were fed on a sugar syrup supplemented with cinnamon EO, oregano EO, a 1:1 mixture of both EOs, or a juice cocktail. An unsupplemented syrup was, instead, used to feed the control group. While V. destructor affected all the colonies throughout the study, no differences in mite infestation levels, population density and honey yield were observed between treatment and control groups. An overexpression of vitellogenin was instead found in all EO-treated groups, even though a significant difference was only found in the group treated with the 1:1 EO mixture. Viral (DWV, CBPV and BQCV), fungal (Nosema ceranae) and bacterial (Melissococcus plutonius) pathogens from both symptomatic and asymptomatic colonies were detected.
Assuntos
Infestações por Ácaros , Varroidae , Animais , Varroidae/efeitos dos fármacos , Varroidae/fisiologia , Abelhas/parasitologia , Abelhas/virologia , Abelhas/efeitos dos fármacos , Óleos Voláteis/farmacologiaRESUMO
Fungal osteomyelitis, especially associated with septic arthritis, is uncommon in Brazil; therefore, sometimes it is difficult to diagnose and treat it. We report the case of a young patient, with no immunosuppressive risk factor, with osteomyelitis leading to septic arthritis of the hip. The diagnosis was performed after surgical drainage and visualization of Cryptococcus neoformans at pathological anatomy. Antifungal treatment resulted in complete remission of the symptoms. Since there is no consensus on the treatment of fungal osteomyelitis, this case report aims to inform orthopedists about the importance of hip arthritis differential diagnosis and the good evolution of clinical treatment after drainage and pathogen isolation.
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Metabolic syndrome (MetS) represents one of the greatest challenges to public health given its serious consequences on cardiovascular diseases and type 2 diabetes. A carbohydrate-restricted, low-fat diet is the current therapy for MetS. Natural mineral waters (NMWs) are known to exert beneficial effects on human health. Our primary objective was to shed light on the potential therapeutic properties of NMWs in MetS. A total of 125 C57BL/6 male and female mice were included in the study. Of these, 10 were left untreated. They were fed a standard diet with tap water throughout the study period, and stayed healthy. The remaining 115 mice were initially fed a high-calorie diet (HCD) consisting of a high-fat feed (60% of energy from fat) with 10% fructose in tap water, served ad libitum over a period of 4 months to induce MetS (the MetS induction phase). Mice were then randomly divided into six treatment groups and a control group, all of which received a low-calorie diet (LCD), but with a different kind of drinking water, for 2 months (the treatment phase). Five groups were each treated with a different kind of NMW, one group by alternating the five NMWs, and one group - the control group - was given tap water. Body weight and blood biochemistry were monitored over the 6-month trial. After 4 months, male and female mice on HCD developed obesity, hypercholesterolaemia and hyperglycaemia, although gains in body weight, total cholesterol, and blood glucose in males were greater than those observed in females (P < 0.0001). When combined with an LCD, the NMWs rich in sulphate, magnesium and bicarbonate, and the minimally mineralised one were the most effective in reducing the blood levels of total cholesterol, high-density lipoprotein (HDL) cholesterol, and glucose. Sex differences emerged during both the MetS induction phase and the treatment phase. These results suggest that NMWs rich in specific macronutrients, such as bicarbonate, sulphate and magnesium, and minimally mineralised water, in combination with an LCD, may contribute to controlling blood lipid and glucose levels in subjects with MetS. Further studies are needed to confirm these results and to extend them to humans.
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The genotoxicity of nano-structured synthetic amorphous silica (SAS), a common food additive, was investigated in vivo in rats. A 90-day oral toxicity study was performed according to OECD test guideline 408 and the genotoxicity of pyrogenic SAS nanomaterial NM-203 was assessed in several organs, using complementary tests. Adult Sprague-Dawley rats of both sexes were treated orally for 90 days with 0, 2, 5, 10, 20, or 50 mg SAS/kg bw per day. Dose levels were selected to approximate expected human dietary exposures to SAS. DNA strand breaks were evaluated by the comet assay in blood, bone marrow, liver, and spleen according to OECD test guideline 489; mutations induced in bone marrow precursors of erythrocytes were assessed by the Pig-a assay and chromosome/ genome damage by the micronucleus assay in blood (OECD test guideline 474) and colon. No treatment-related increases of gene (Pig-a) or chromosome/genome (micronucleus) mutations were detected in the blood. The percentage of micronucleated cells was not increased in the colon of treated rats. Among the organs analyzed by the comet assay, the spleen was the only target showing a weak but biologically relevant genotoxic effect.
Assuntos
Dano ao DNA , Dióxido de Silício , Animais , Ensaio Cometa , Feminino , Masculino , Testes para Micronúcleos , Ratos , Ratos Sprague-Dawley , Dióxido de Silício/toxicidadeRESUMO
Several lines of evidence suggest an association between oxidative DNA-damage repair capacity and cancer risk. In particular, a DNA-glycosylase assay for removal of 8-oxoguanine (8-oxoG) in peripheral blood mononuclear cells (PBMC) has been successfully applied to identify populations with increased risk for lung cancer and squamous cell carcinomas of head and neck. In order to verify whether EBV-transformed lymphoblastoid cell lines (LCL) are a suitable surrogate for PBMC in specific DNA-repair phenotypic assays, a validation trial was conducted. PBMC from 20 healthy subjects were collected and an aliquot was transformed with EBV to obtain LCL. The ability of cell-free extracts from both cell types to incise a 3'-fluorescently labelled duplex oligonucleotide containing a single 8-oxoG (OGG assay) was evaluated. Since this activity is mediated predominantly by OGG1, the OGG1 gene expression was also measured. 8-oxoG DNA-glycosylase activity and OGG1 expression were significantly higher (p<0.0001) in LCL than in PBMC. However, while this assay was shown to be robust and reproducible when used on PBMC (intra-assay CV=8%), a high intra-culture variability was observed with LCL (intra-culture CV=16.8%). Neither differences on OGG1 gene expression nor the cell-cycle distribution seemed to account for this variability. Inter-individual variability of OGG activity in PBMC and LCL was not associated with OGG1 gene expression. We have therefore established a non-radioactive cleavage assay that can be easily applied to measure OGG activity in human PBMC. The use of LCL for DNA-repair genotype-phenotype correlation studies seems to be inappropriate, at least with cell-free based functional assays.
Assuntos
DNA Glicosilases/genética , DNA Glicosilases/metabolismo , Reparo do DNA , Guanina/análogos & derivados , Linfócitos/metabolismo , Linhagem Celular , Transformação Celular Viral , Criopreservação , Expressão Gênica , Estudos de Associação Genética , Guanina/metabolismo , Herpesvirus Humano 4 , Humanos , Técnicas In VitroRESUMO
Mancozeb (MZ) and zoxamide (ZOX) are fungicides commonly used in pest control programs to protect vineyards. Their toxic and genotoxic potential were investigated in vitro on HepG2 and A549 cell lines at environmentally relevant concentrations. Cytotoxicity, apoptosis, necrosis and intracellular reactive oxygen species (ROS), comet assay and a micronucleus test with CREST immunofluorescence were used. The expression of a panel of genes involved in apoptosis/necrosis (BAX/BCL2), oxidative stress (NRF2), drug metabolism (CYP1A1) and DNA repair (ERCC1/OGG1) was evaluated by real-time PCR. Both fungicides were cytotoxic at the highest tested concentrations (295.7 and 463.4 µM, respectively); MZ induced necrosis, ZOX did not increase apoptosis but modulated BAX and BCL2 expression, suggesting a different mechanism. Both compounds did not increase ROS, but the induction of CYP1A1 and NRF2 expression supported a pro-oxidant mechanism. The comet assay evidenced MZ genotoxicity, whereas no DNA damage due to ZOX treatment was observed. Positive micronuclei were increased in both cell lines treated with MZ and ZOX, supporting their aneugenic potential. ERCC1 and OGG1 were differently modulated, indicating the efficient activation of the nucleotide excision repair system by both fungicides and the inhibition of the base excision repair system by MZ. Overall, MZ confirmed its toxicity and new ZOX-relevant effects were highlighted.
Assuntos
Fungicidas Industriais , Maneb , Zineb , Amidas , Ensaio Cometa , Dano ao DNA , Fungicidas Industriais/toxicidade , Maneb/toxicidade , Estresse Oxidativo , Espécies Reativas de Oxigênio , Zineb/toxicidadeRESUMO
Bis(2-ethylhexyl) phthalate (DEHP) is a plasticizer used in several items, non-covalently bound to plastics and easily released, since metabolites were found in human matrices. DEHP is an endocrine disrupter and children are particularly vulnerable and susceptible to DEHP effects due to higher exposure levels and developmental stage. A juvenile toxicity study was performed to identify DEHP hazard and mode of action in Sprague-Dawley rats of both sexes during peri-pubertal period - corresponding to childhood phase - from weaning, post-natal day (PND) 23, to full sexual maturity (PND60); the dose levels of 0, 9, 21 and 48 mg/kg bw/day were derived from LIFE PERSUADED biomonitoring study in children. DEHP was administered by gavage for 28 days (5 days/week); timing of preputial separation and vaginal opening was observed during treatment. Histopathological analysis was performed on: adrenals, spleen, liver, thyroid and reproductive organs. The following serum biomarkers were assessed: estradiol, testosterone, anti-Mullerian hormone, tetraiodothyronine, thyroid stimulating hormone, adiponectin and leptin. Gene expression on hypothalamic-pituitary area was focused on follicle stimulating, luteinizing, and thyroid stimulating hormones. The results showed that main targets of DEHP during juvenile period were liver and metabolic system in both sexes, while sex-specific effects were recorded in reproductive system (male rats) and in thyroid (female rats). DEHP exposure during peri-pubertal period at dose levels derived from biomonitoring study in children can induce sex-specific imbalances identifying the juvenile animal model as a sound tool to identify hazards for a reliable risk assessment targeted to children.
Assuntos
Monitoramento Biológico/métodos , Dietilexilftalato/toxicidade , Reprodução/efeitos dos fármacos , Maturidade Sexual/efeitos dos fármacos , Glândula Tireoide/efeitos dos fármacos , Glândula Tireoide/metabolismo , Administração Oral , Fatores Etários , Animais , Animais Recém-Nascidos , Criança , Dietilexilftalato/administração & dosagem , Relação Dose-Resposta a Droga , Feminino , Humanos , Masculino , Plastificantes/administração & dosagem , Plastificantes/metabolismo , Plastificantes/toxicidade , Ratos , Ratos Sprague-Dawley , Reprodução/fisiologia , Maturidade Sexual/fisiologiaRESUMO
Synthetic amorphous silica (SAS) consists of agglomerates and aggregates of primary particles in the nanorange (<100 nm) and it is the E551 authorized food additive. The potential risks for human health associated to dietary exposure to SAS are not completely assessed; in particular, data on male and female reproductive systems are lacking. A 90-day oral toxicity study with pyrogenic SAS nanomaterial NM-203 was carried out on the basis of the OECD test guideline 408 in the frame of the NANoREG project. Adult Sprague-Dawley rats of both sexes were orally treated for 90 days with 0, 2, 5, 10, 20 and 50 mg SAS/kg bw per day. Dose levels were selected to be as close as possible to the expected human exposure to food additive E551. The present paper provides specific information on potential effects on male and female reproductive systems, through the evaluation of serum biomarkers, sperm count, histopathological analysis of testis, epididymis, ovary and uterus and real-time PCR on uterus; potential genotoxic alterations were evaluated by comet assay on testis, sperm and ovary. NM-203 did not induce histophatological and genotoxic effects in male reproductive system. In female rats, ovary is not target of NM-203 and only tissue-specific effects on uterus were recorded up to 10 mg/kg bw per day. To our best knowledge, this is the first study providing data on male and female reproductive systems after long-term, repeated oral exposure at dose levels close to dietary human exposure, which identifies a limited concern only for female reproductive health.
Assuntos
Dióxido de Silício/toxicidade , Administração Oral , Animais , Ensaio Cometa , Estradiol/sangue , Feminino , Expressão Gênica/efeitos dos fármacos , Genitália/efeitos dos fármacos , Genitália/metabolismo , Antígeno Ki-67/metabolismo , Masculino , Ratos Sprague-Dawley , Contagem de Espermatozoides , Testosterona/sangue , Testes de Toxicidade SubcrônicaRESUMO
Bisphenol A (BPA) is a plasticizer with endocrine disrupting properties particularly relevant for children health. Recently BPA has been associated with metabolic dysfunctions but no data are yet available in specific, long-term studies. This study aimed to evaluate BPA modes of action and hazards during animal juvenile life-stage, corresponding to childhood. Immature Sprague-Dawley rats of both sexes were orally treated with 0 (vehicle only-olive oil), 2, 6, and 18 mg/kg bw per day of BPA for 28 days, from weaning to sexual maturity. Dose levels were obtained from the PERSUADED biomonitoring study in Italian children. Both no-observed-adverse-effect-level (NOAEL)/low-observed-adverse-effect-level (LOAEL) and estimated benchmark dose (BMD) approaches were applied. General toxicity, parameters of sexual development, endocrine/reproductive/functional liver and kidney biomarkers, histopathology of target tissues, and gene expression in hypothalamic-pituitary area and liver were studied. No mortality or general toxicity occurred. Sex-specific alterations were observed in liver, thyroid, spleen, leptin/adiponectin serum levels, and hypothalamic-pituitary gene expression. Thyroid homeostasis and liver were the most sensitive targets of BPA exposure in the peripubertal phase. The proposed LOAEL was 2 mg/kg bw, considering as critical effect the liver endpoints, kidney weight in male and adrenal histomorphometrical alterations and osteopontin upregulation in female rats. The BMD lower bounds were 0.05 and 1.33 mg/kg bw in males and females, considering liver and thyroid biomarkers, respectively. Overall, BPA evaluation at dose levels derived from children biomonitoring study allowed to identify sex-specific, targeted toxicological effects that may have significant impact on risk assessment for children.
Assuntos
Compostos Benzidrílicos/toxicidade , Hormônios/análise , Hormônios/sangue , Fenóis/toxicidade , Adiponectina , Administração Oral , Glândulas Suprarrenais , Fatores Etários , Animais , Compostos Benzidrílicos/análise , Compostos Benzidrílicos/sangue , Monitoramento Biológico , Biomarcadores/sangue , Peso Corporal/efeitos dos fármacos , Disruptores Endócrinos/toxicidade , Feminino , Expressão Gênica , Genitália , Sistema Hipotálamo-Hipofisário , Fígado , Masculino , Pâncreas , Fenóis/análise , Fenóis/sangue , Ratos Sprague-Dawley , Baço , Glândula TireoideRESUMO
Food additive E551 consists of synthetic amorphous silica (SAS), comprising agglomerates and aggregates of primary particles in the nanorange (<100 nm), which potential nanospecific risks for humans associated to dietary exposure are not yet completely assessed. In NANoREG project, aim of the study was to identify potential hazards of pyrogenic SAS nanomaterial NM-203 by a 90-day oral toxicity study (OECD test guideline 408). Adult Sprague-Dawley rats of both sexes were orally treated with 0, 2, 5, 10, 20 and 50 mg SAS/kg bw per day; dose levels were selected to be as close as possible to E551 dietary exposure. Several endpoints were investigated, the whole integrative study is presented here along with the results of dispersion characterization, tissue distribution, general toxicity, blood/serum biomarkers, histopathological and immunotoxicity endpoints. No mortality, general toxicity and limited deposition in target tissues were observed. NM-203 affected liver and spleen in both sexes. Proposed NOAEL 5 mg/kg bw per day in male rats for enlarged sinusoids in liver. In female rats, TSH and creatinine levels were affected, proposed LOAEL 2 mg/kg bw per day. Overall, these data provide new insight for a comprehensive risk assessment of SAS exposure by the oral route.
Assuntos
Aditivos Alimentares/toxicidade , Nanoestruturas/toxicidade , Dióxido de Silício/toxicidade , Administração Oral , Animais , Biomarcadores/sangue , Feminino , Aditivos Alimentares/administração & dosagem , Fígado/patologia , Masculino , Nanoestruturas/administração & dosagem , Nível de Efeito Adverso não Observado , Ratos Sprague-Dawley , Medição de Risco , Silício/análise , Dióxido de Silício/administração & dosagemRESUMO
3-Hydroxy 3-methylglutaryl coenzyme A reductase (HMG-CoAR) is the key and rate-limiting enzyme of cholesterol biosynthetic pathway. Although HMG-CoAR activity has already been related to the differentiation of some cellular lines there are no studies that analyze the role of HMG-CoAR, and the pathway it is involved with in a fully characterized muscle differentiation model. Thus, the aim of this work is to evaluate such role and delineate the pathway involved in foetal rat myoblasts (L6) induced to differentiate by insulin -- a standard and feasible model of the myogenic process. The results obtained by biochemical and morphological approaches demonstrate that (i) HMG-CoAR increase is crucial for differentiation induction, (ii) p21waf, whose increase is a necessary requisite for differentiation to occur, rises downstream HMG-CoAR activation, (iii) the main role of p38/MAPK as key regulator also for HMG-CoAR. Pathologies characterized by muscle degeneration might benefit from therapeutic programmes committed to muscle function restoration, such as modulation and planning myoblast differentiation. Thus, the important role of HMG-CoAR in muscular differentiation providing new molecular basis for the control of muscle development can help in the design of therapeutic treatment for diseases characterized by the weakening of muscular fibers and aging-related disorders (sarcopenia).
Assuntos
Diferenciação Celular/fisiologia , Hidroximetilglutaril-CoA Redutases/metabolismo , Músculo Esquelético/enzimologia , Músculo Esquelético/fisiologia , Animais , Células Cultivadas , Colesterol/química , Colesterol/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Ativação Enzimática , Hidroximetilglutaril-CoA Redutases/genética , Inibidores de Hidroximetilglutaril-CoA Redutases/metabolismo , Insulina/metabolismo , Lovastatina/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Músculo Esquelético/citologia , Ratos , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Proteína rhoA de Ligação ao GTP/metabolismoRESUMO
Placenta is a target organ of Bisphenol A (BPA). To investigate possible effects on syncytiotrophoblast, the exchanging surface between mother and fetus, we exposed a trophoblast model (BeWo) to BPA concentrations occurring in humans (1 and 50â¯nM). We assessed the gene and protein expression of three human endogenous retroviral envelopes, specifically expressed in placenta (ERVW-1, ERVFRD-1 and ERV3-1), the secretion of ß-hCG, the extent of trophoblast fusion and the activity of apoptosis markers (caspases 8, 3, 9 and PARP); additionally, the gene expression of transcription factors regulating HERV expression (i.e. GCM1, PPARγ, ERα and ERß) was evaluated. At 50â¯nM, BPA induced ERVW-1, ERVFRD-1 and the corresponding syncytin proteins, ERV3-1, PPARγ, ERα and ERß expression, increased ß-hCG secretion and BeWo cells fusion, thus promoting the syncytiotrophoblast phenotype. The results support placenta as a target organ of BPA. Possible implications on fetal and pregnancy health should be carefully considered.
Assuntos
Compostos Benzidrílicos/toxicidade , Gonadotropina Coriônica Humana Subunidade beta/metabolismo , Disruptores Endócrinos/toxicidade , Produtos do Gene env/genética , Fenóis/toxicidade , Proteínas da Gravidez/genética , Trofoblastos/efeitos dos fármacos , Fusão Celular , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Feminino , Expressão Gênica/efeitos dos fármacos , Humanos , Gravidez , Trofoblastos/metabolismo , Trofoblastos/patologiaRESUMO
Base excision repair (BER) is the main pathway for repair of DNA damage in mammalian cells. This pathway leads to the formation of DNA repair intermediates which, if still unsolved, cause cell lethality and mutagenesis. To characterize mutations induced by BER intermediates in mammalian cells, an SV-40 derived shuttle vector was constructed carrying a site-specific lesion within the recognition sequence of a restriction endonuclease. The mutation spectra of abasic (AP) sites, 5'-deoxyribose-5-phosphate (5'dRp) and 3'-[2,3-didehydro-2,3-dideoxy-ribose] (3'ddR5p) single-strand breaks (ssb) in mammalian cells was analysed by RFLP/PCR and mutation frequency was estimated by quantitative PCR. Point mutations were the predominant events occurring at all BER intermediates. The AP site-induced mutation spectrum supports evidence for the 'A-rule' and is also consistent with the use of the 5' neighbouring base to instruct nucleotide incorporation (5'-rule). Preferential adenine insertion was also observed after in vivo replication of 5'dRp or 3'ddR5p ssb. We provide original evidence that not only the abasic site but also its derivatives 'faceless' BER intermediates are mutagenic, with a similar mutation frequency, in mammalian cells. Our findings support the hypothesis that unattended BER intermediates could be a constant threat for genome integrity as well as a spontaneous source of mutations.
Assuntos
Reparo do DNA , Desoxirribose/análogos & derivados , Desoxirribose/metabolismo , Mutagênese , Ribosemonofosfatos/metabolismo , Animais , Células COS , Chlorocebus aethiops , Enzimas Reparadoras do DNA/metabolismo , Enzimas de Restrição do DNA/metabolismo , Vetores Genéticos , Mutação Puntual , Reação em Cadeia da Polimerase , Vírus 40 dos Símios/genéticaRESUMO
Children show unique features concerning chemical hazards and risks, due to different exposure scenarios, age-related metabolic capacity and biological susceptibility linked to post-natal development. Chemical Regulatory frameworks state the need of children risk characterization. Current testing guidelines covering post-natal development are not routinely required by regulatory applications other than pesticides and biocides. Juvenile toxicity studies are foreseen for paediatric drugs: the toxicological repeated-dose tests don't allow accurate evaluations of effects upon direct exposure of immature organism. The paper discusses a testing approach aimed to address regulatory requirements of chemical hazard identification/characterization in a children-specific perspective. Juvenile toxicity test could be performed primarily on chemicals that may have relevant modes of action and/or age-related toxicokinetic differences and/or lead to important children exposure. This could be pursued by updating existing guidelines/test protocols with triggers for endpoints relevant to juvenile toxicity.
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Saúde da Criança , Medição de Risco , Testes de Toxicidade/métodos , Animais , HumanosRESUMO
Prostate function is critical for male fertility and its well-known oncological biomarker, namely Prostate-Specific Antigen (PSA), can be also used to monitor prostate epithelial human cells upon treatment with pharmaceutical drugs or natural bioactive compounds. The LNCaP human prostate cell line was previously set up as a model system to investigate chemicals affecting prostate epithelium functionality by means of a tiered approach integrating two different toxicological endpoints, cell viability (MTS) and PSA secretion assays. Here, the same approach has been used to characterize the biological effects of phytochemicals on prostate epithelium. The antiandrogenic ability of phytochemicals to inhibit DHT-induced PSA secretion has been investigated also characterizing their intracellular distribution, in the presence or absence of sex steroids. Intracellular distribution allows to verify whether and to which extent each phytochemical is able to enter the cell and to reach the nucleus, the latter being the target of the supposed transcriptional modulatory activity upon phytochemicals' binding to sex steroid receptors. Some phytochemicals, supposed to have a role in the functionality of the prostate epithelium, have been tested in a dose-dependent manner in both MTS and PSA secretion assays. In parallel, to establish the "effective concentration", in comparison to the "nominal one", the intracellular amount of each phytochemical has been assessed upon cell fractionation of LNCaP-treated cells and subsequent chromatographic measurements.
Assuntos
Adenocarcinoma/metabolismo , Antineoplásicos Fitogênicos/farmacologia , Flavonoides/farmacologia , Hormônios Esteroides Gonadais/metabolismo , Compostos Fitoquímicos/farmacologia , Neoplasias da Próstata/metabolismo , Adenocarcinoma/patologia , Antineoplásicos Fitogênicos/metabolismo , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Núcleo Celular/patologia , Proliferação de Células , Sobrevivência Celular , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Flavonoides/metabolismo , Humanos , Masculino , Compostos Fitoquímicos/metabolismo , Antígeno Prostático Específico/metabolismo , Neoplasias da Próstata/patologiaRESUMO
Cockayne syndrome (CS) is a rare hereditary multisystem disease characterized by neurological and development impairment, and premature aging. Cockayne syndrome cells are hypersensitive to oxidative stress, but the molecular mechanisms involved remain unresolved. Here we provide the first evidence that primary fibroblasts derived from patients with CS-A and CS-B present an altered redox balance with increased steady-state levels of intracellular reactive oxygen species (ROS) and basal and induced DNA oxidative damage, loss of the mitochondrial membrane potential, and a significant decrease in the rate of basal oxidative phosphorylation. The Na/K-ATPase, a relevant target of oxidative stress, is also affected with reduced transcription in CS fibroblasts and normal protein levels restored upon complementation with wild-type genes. High-resolution magnetic resonance spectroscopy revealed a significantly perturbed metabolic profile in CS-A and CS-B primary fibroblasts compared with normal cells in agreement with increased oxidative stress and alterations in cell bioenergetics. The affected processes include oxidative metabolism, glycolysis, choline phospholipid metabolism, and osmoregulation. The alterations in intracellular ROS content, oxidative DNA damage, and metabolic profile were partially rescued by the addition of an antioxidant in the culture medium suggesting that the continuous oxidative stress that characterizes CS cells plays a causative role in the underlying pathophysiology. The changes of oxidative and energy metabolism offer a clue for the clinical features of patients with CS and provide novel tools valuable for both diagnosis and therapy.
Assuntos
Síndrome de Cockayne/metabolismo , Fibroblastos/metabolismo , Estresse Oxidativo/fisiologia , Senilidade Prematura/genética , Senilidade Prematura/metabolismo , Senilidade Prematura/patologia , Síndrome de Cockayne/genética , Síndrome de Cockayne/patologia , Dano ao DNA , Reparo do DNA , Fibroblastos/patologia , Humanos , Mitocôndrias/metabolismo , Oxirredução , Fosforilação OxidativaRESUMO
Reproductive toxicity, with its many targets and mechanisms, is a complex area of toxicology; thus, the screening and identification of reproductive toxicants is a main scientific challenge for the safety assessment of chemicals, including the European Regulation on Chemicals (REACH). Regulatory agencies recommend the implementation of the 3Rs principle (refinement, reduction, replacement) as well as of intelligent testing strategies, through the development of in vitro methods and the use of mechanistic information in the hazard identification and characterization steps of the risk assessment process. The EU Integrated Project ReProTect (6th Framework Programme) implemented an array of in vitro tests to study different building blocks of the mammalian reproductive cycle: methodological developments and results on male and female germ cells, prostate and placenta are presented.