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1.
J Cell Sci ; 135(5)2022 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-33975358

RESUMO

Lipid droplets (LDs) are lipid-rich organelles universally found in most cells. They serve as a key energy reservoir, actively participate in signal transduction and dynamically communicate with other organelles. LD dysfunction has been associated with a variety of diseases. The content level, composition and mobility of LDs are crucial for their physiological and pathological functions, and these different parameters of LDs are subject to regulation by genetic factors and environmental inputs. Coherent Raman scattering (CRS) microscopy utilizes optical nonlinear processes to probe the intrinsic chemical bond vibration, offering label-free, quantitative imaging of lipids in vivo with high chemical specificity and spatiotemporal resolution. In this Review, we provide an overview over the principle of CRS microscopy and its application in tracking different parameters of LDs in live cells and organisms. We also discuss the use of CRS microscopy in genetic screens to discover lipid regulatory mechanisms and in understanding disease-related lipid pathology.


Assuntos
Microscopia , Análise Espectral Raman , Biologia , Gotículas Lipídicas , Lipídeos
2.
EMBO Rep ; 21(11): e50214, 2020 11 05.
Artigo em Inglês | MEDLINE | ID: mdl-33034119

RESUMO

Lipid droplets (LDs) are dynamic cytoplasmic organelles present in most eukaryotic cells. The appearance of LDs in neurons is not usually observed under physiological conditions, but is associated with neural diseases. It remains unclear how LD dynamics is regulated in neurons and how the appearance of LDs affects neuronal functions. We discovered that mutations of two key lipolysis genes atgl-1 and lid-1 lead to LD appearance in neurons of Caenorhabditis elegans. This neuronal lipid accumulation protects neurons from hyperactivation-triggered neurodegeneration, with a mild decrease in touch sensation. We also discovered that reduced biosynthesis of polyunsaturated fatty acids (PUFAs) causes similar effects and synergizes with decreased lipolysis. Furthermore, we demonstrated that these changes in lipolysis and PUFA biosynthesis increase PUFA partitioning toward triacylglycerol, and reduced incorporation of PUFAs into phospholipids increases neuronal protection. Together, these results suggest the crucial role of neuronal lipolysis in cell-autonomous regulation of neural functions and neurodegeneration.


Assuntos
Gotículas Lipídicas , Lipólise , Animais , Ácidos Graxos Insaturados/metabolismo , Gotículas Lipídicas/metabolismo , Metabolismo dos Lipídeos/genética , Neurônios , Triglicerídeos
3.
PLoS One ; 15(2): e0229104, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32106243

RESUMO

Tyrosine kinase inhibitor (TKI) resistance is a major problem in chronic myeloid leukemia (CML). We generated a TKI-resistant K562 sub-population, K562-IR, under selective imatinib-mesylate pressure. K562-IR cells are CD34-/CD38-, BCR-Abl-independent, proliferate slowly, highly adherent and form intact tumor spheroids. Loss of CD45 and other hematopoietic markers reveal these cells have diverged from their hematopoietic origin. CD34 negativity, high expression of E-cadherin and CD44; decreased levels of CD45 and ß-catenin do not fully confer with the leukemic stem cell (LSC) phenotype. Expression analyses reveal that K562-IR cells differentially express tissue/organ development and differentiation genes. Our data suggest that the observed phenotypic shift is an adaptive process rendering cells under TKI stress to become oncogene independent. Cells develop transcriptional instability in search for a gene expression framework suitable for new environmental stresses, resulting in an adaptive phenotypic shift in which some cells partially display LSC-like properties. With leukemic/cancer stem cell targeted therapies underway, the difference between treating an entity and a spectrum of dynamic cellular states will have conclusive effects on the outcome.


Assuntos
Resistencia a Medicamentos Antineoplásicos/genética , Proteínas de Fusão bcr-abl/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Inibidores de Proteínas Quinases/farmacologia , Células 3T3 , Animais , Antígenos CD/genética , Antígenos CD/metabolismo , Caderinas/genética , Caderinas/metabolismo , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Proteínas de Fusão bcr-abl/antagonistas & inibidores , Proteínas de Fusão bcr-abl/metabolismo , Perfilação da Expressão Gênica , Humanos , Mesilato de Imatinib/farmacologia , Mesilato de Imatinib/uso terapêutico , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Camundongos , Mutação/efeitos dos fármacos , Análise de Sequência com Séries de Oligonucleotídeos , Domínios Proteicos/genética , Inibidores de Proteínas Quinases/uso terapêutico
4.
Int Urol Nephrol ; 48(10): 1699-704, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27473155

RESUMO

PURPOSE: The purpose of the current prospective study was to evaluate the effects of low sodium dialysate on oxidative stress parameters, blood pressure (BP) and endothelial dysfunction in maintenance hemodialysis (HD) patients. METHODS: After baseline measurements were taken, the dialysate sodium concentration was reduced from 140 to 137 mEq/L. Oxidative stress parameters and flow-mediated dilatation (FMD %) were measured before and after 6 months of HD with low sodium dialysate. Interdialytic weight gain (IDWG) and pre- and post-dialysis BP were monitored during the study. RESULTS: A total of 52 patients were enrolled and 41 patients completed the study. There was a significant reduction in systolic blood pressure at the end of the study [130.00 (90.00-190.00) vs. 120.00 (90.00-150.00), p < 0.001]. Similarly, there were significant improvements in IDWG [2670.00 (1670.00-4300.00) vs. 1986.00 (1099.00-3998.00), p < 0.001] and FMD % [7.26 (4.55-8.56) vs. 9.56 (6.55-12.05), p < 0.001]. Serum MDA levels (p < 0.001) were significantly decreased; serum SOD (p < 0.001) and GPx (p < 0.001) activities were significantly increased after low sodium HD compared to standard sodium HD. CONCLUSION: Our data seem to suggest a potential role of 137 mEq/L sodium dialysate for improving hemodynamic status, endothelial function and reducing oxidative stress than 140 mEq/L sodium dialysate in maintenance HD patients.


Assuntos
Pressão Sanguínea/efeitos dos fármacos , Endotélio Vascular/efeitos dos fármacos , Soluções para Hemodiálise/farmacologia , Falência Renal Crônica , Estresse Oxidativo/efeitos dos fármacos , Diálise Renal , Sódio , Adulto , Endotélio Vascular/fisiopatologia , Feminino , Humanos , Falência Renal Crônica/diagnóstico , Falência Renal Crônica/metabolismo , Falência Renal Crônica/terapia , Masculino , Pessoa de Meia-Idade , Monitorização Fisiológica/métodos , Diálise Renal/efeitos adversos , Diálise Renal/métodos , Sódio/sangue , Sódio/farmacologia , Estatística como Assunto
5.
BMC Bioinformatics ; 16 Suppl 18: S9, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26679222

RESUMO

BACKGROUND: Neddylation is a reversible post-translational modification that plays a vital role in maintaining cellular machinery. It is shown to affect localization, binding partners and structure of target proteins. Disruption of protein neddylation was observed in various diseases such as Alzheimer's and cancer. Therefore, understanding the neddylation mechanism and determining neddylation targets possibly bears a huge importance in further understanding the cellular processes. This study is the first attempt to predict neddylated sites from protein sequences by using several sequence and sequence-based structural features. RESULTS: We have developed a neddylation site prediction method using a support vector machine based on various sequence properties, position-specific scoring matrices, and disorder. Using 21 amino acid long lysine-centred windows, our model was able to predict neddylation sites successfully, with an average 5-fold stratified cross validation performance of 0.91, 0.91, 0.75, 0.44, 0.95 for accuracy, specificity, sensitivity, Matthew's correlation coefficient and area under curve, respectively. Independent test set results validated the robustness of reported new method. Additionally, we observed that neddylation sites are commonly flexible and there is a significant positively charged amino acid presence in neddylation sites. CONCLUSIONS: In this study, a neddylation site prediction method was developed for the first time in literature. Common characteristics of neddylation sites and their discriminative properties were explored for further in silico studies on neddylation. Lastly, up-to-date neddylation dataset was provided for researchers working on post-translational modifications in the accompanying supplementary material of this article.


Assuntos
Biologia Computacional , Processamento de Proteína Pós-Traducional , Sequência de Aminoácidos , Área Sob a Curva , Lisina/química , Lisina/metabolismo , Matrizes de Pontuação de Posição Específica , Curva ROC , Máquina de Vetores de Suporte , Ubiquitinas/química , Ubiquitinas/metabolismo
6.
PLoS One ; 10(8): e0136212, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26287606

RESUMO

The outcome of H. pylori infection is closely related with bacteria's virulence factors and host immune response. The association between T cells and H. pylori infection has been identified, but the effects of the nine major H. pylori specific virulence factors; cagA, vacA, oipA, babA, hpaA, napA, dupA, ureA, ureB on T cell response in H. pylori infected patients have not been fully elucidated. We developed a multiplex- PCR assay to detect nine H. pylori virulence genes with in a three PCR reactions. Also, the expression levels of Th1, Th17 and Treg cell specific cytokines and transcription factors were detected by using qRT-PCR assays. Furthermore, a novel expert derived model is developed to identify set of factors and rules that can distinguish the ulcer patients from gastritis patients. Within all virulence factors that we tested, we identified a correlation between the presence of napA virulence gene and ulcer disease as a first data. Additionally, a positive correlation between the H. pylori dupA virulence factor and IFN-γ, and H. pylori babA virulence factor and IL-17 was detected in gastritis and ulcer patients respectively. By using computer-based models, clinical outcomes of a patients infected with H. pylori can be predicted by screening the patient's H. pylori vacA m1/m2, ureA and cagA status and IFN-γ (Th1), IL-17 (Th17), and FOXP3 (Treg) expression levels. Herein, we report, for the first time, the relationship between H. pylori virulence factors and host immune responses for diagnostic prediction of gastric diseases using computer-based models.


Assuntos
Infecções por Helicobacter/diagnóstico , Infecções por Helicobacter/imunologia , Helicobacter pylori , Proteínas de Bactérias/genética , Simulação por Computador , Diagnóstico por Computador , Sistemas Especialistas , Gastrite/diagnóstico , Gastrite/imunologia , Gastrite/microbiologia , Genes Bacterianos , Infecções por Helicobacter/microbiologia , Helicobacter pylori/genética , Helicobacter pylori/imunologia , Helicobacter pylori/patogenicidade , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Imunidade Celular , Reação em Cadeia da Polimerase Multiplex , Subpopulações de Linfócitos T/imunologia , Fatores de Virulência/genética
7.
Curr Biol ; 25(5): 621-627, 2015 Mar 02.
Artigo em Inglês | MEDLINE | ID: mdl-25702577

RESUMO

Identification of nutritious compounds is dependent on expression of specific taste receptors in appropriate taste-cell types [1]. In contrast to mammals, which rely on a single, broadly tuned heterodimeric sugar receptor [2], the Drosophila genome harbors a small subfamily of eight, closely related gustatory receptor (Gr) genes, Gr5a, Gr61a, and Gr64a-Gr64f, of which three have been proposed to mediate sweet taste [3-6]. However, expression and function of several of these putative sugar Gr genes are not known. Here, we present a comprehensive expression and functional analysis using Gr(LEXA/GAL4) alleles that were generated through homologous recombination. We show that sugar Gr genes are expressed in a combinatorial manner to yield at least eight sets of sweet-sensing neurons. Behavioral investigations show that most sugar Gr mutations affect taste responses to only a small number of sugars and that effective detection of most sugars is dependent on more than one Gr gene. Surprisingly, Gr64a, one of three Gr genes previously proposed to play a major role in sweet taste [3, 4], is not expressed in labellar taste neurons, and Gr64a mutant flies exhibit normal sugar responses elicited from the labellum. Our analysis provides a molecular rationale for distinct tuning profiles of sweet taste neurons, and it favors a model whereby all sugar Grs contribute to sweet taste. Furthermore, expression in olfactory organs and the brain implies novel roles for sugar Gr genes in olfaction and internal nutrient sensing, respectively. Thus, sugar receptors may contribute to feeding behavior via multiple sensory systems.


Assuntos
Fenômenos Fisiológicos da Nutrição Animal/fisiologia , Proteínas de Drosophila/metabolismo , Drosophila/metabolismo , Comportamento Alimentar/fisiologia , Receptores de Superfície Celular/metabolismo , Olfato/fisiologia , Percepção Gustatória/fisiologia , Animais , Encéfalo/citologia , Encéfalo/metabolismo , Drosophila/fisiologia , Componentes do Gene , Perfilação da Expressão Gênica , Técnicas de Introdução de Genes , Recombinação Homóloga/genética , Neurônios/metabolismo , Neurônios Receptores Olfatórios/metabolismo , Sensilas/metabolismo
8.
Adv Mater ; 27(14): 2367-76, 2015 Apr 08.
Artigo em Inglês | MEDLINE | ID: mdl-25722118

RESUMO

A multilayered triboelectric nanogenerator (MULTENG) that can be actuated by acoustic waves, vibration of a moving car, and tapping motion is built using a 3D-printing technique. The MULTENG can generate an open-circuit voltage of up to 396 V and a short-circuit current of up to 1.62 mA, and can power 38 LEDs. The layers of the triboelectric generator are made of polyetherimide nanopillars and chalcogenide core-shell nanofibers.

9.
BMC Genomics ; 15 Suppl 9: S18, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25521314

RESUMO

BACKGROUND: Sumoylation, which is a reversible and dynamic post-translational modification, is one of the vital processes in a cell. Before a protein matures to perform its function, sumoylation may alter its localization, interactions, and possibly structural conformation. Abberations in protein sumoylation has been linked with a variety of disorders and developmental anomalies. Experimental approaches to identification of sumoylation sites may not be effective due to the dynamic nature of sumoylation, laborsome experiments and their cost. Therefore, computational approaches may guide experimental identification of sumoylation sites and provide insights for further understanding sumoylation mechanism. RESULTS: In this paper, the effectiveness of using various sequence properties in predicting sumoylation sites was investigated with statistical analyses and machine learning approach employing support vector machines. These sequence properties were derived from windows of size 7 including position-specific amino acid composition, hydrophobicity, estimated sub-window volumes, predicted disorder, and conformational flexibility. 5-fold cross-validation results on experimentally identified sumoylation sites revealed that our method successfully predicts sumoylation sites with a Matthew's correlation coefficient, sensitivity, specificity, and accuracy equal to 0.66, 73%, 98%, and 97%, respectively. Additionally, we have showed that our method compares favorably to the existing prediction methods and basic regular expressions scanner. CONCLUSIONS: By using support vector machines, a new, robust method for sumoylation site prediction was introduced. Besides, the possible effects of predicted conformational flexibility and disorder on sumoylation site recognition were explored computationally for the first time to our knowledge as an additional parameter that could aid in sumoylation site prediction.


Assuntos
Biologia Computacional/métodos , Proteínas/química , Proteínas/metabolismo , Sumoilação , Máquina de Vetores de Suporte , Sequência de Aminoácidos , Sítios de Ligação
10.
Fly (Austin) ; 8(4): 189-96, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25984594

RESUMO

Arthropods employ a large family of up to 100 putative taste or gustatory receptors (Grs) for the recognition of a wide range of non-volatile chemicals. In Drosophila melanogaster, a small subfamily of 8 Gr genes is thought to mediate the detection of sugars, the fly's major nutritional source. However, the specific roles for most sugar Gr genes are not known. Here, we report the generation of a series of mutant sugar Gr knock-in alleles and several composite sugar Gr mutant strains, including a sugar blind strain, which will facilitate the characterization of this gene family. Using Ca(2+) imaging experiments, we show that most gustatory receptor neurons (GRNs) of sugar blind flies (lacking all 8 sugar Gr genes) fail to respond to any sugar tested. Moreover, expression of single sugar Gr genes in most sweet GRNs of sugar-blind flies does not restore sugar responses. However, when pair-wise combinations of sugar Gr genes are introduced to sweet GRNs, responses to select sugars are restored. We also examined the cellular phenotype of flies homozygous mutant for Gr64a, a Gr gene previously reported to be a major contributor for the detection of many sugars. In contrast to these claims, we find that sweet GRNs of Gr64a homozygous mutant flies show normal responses to most sugars, and only modestly reduced responses to maltose and maltotriose. Thus, the precisely engineered genetic mutations of single Gr genes and construction of a sugar-blind strain provide powerful analytical tools for examining the roles of Drosophila and other insect sugar Gr genes in sweet taste.


Assuntos
Carboidratos/fisiologia , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Comportamento Alimentar/fisiologia , Regulação da Expressão Gênica/fisiologia , Receptores de Superfície Celular/metabolismo , Animais , Carboidratos/química , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Mutação , Receptores de Superfície Celular/genética , Percepção Gustatória/fisiologia
11.
Curr Biol ; 23(15): 1466-71, 2013 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-23850280

RESUMO

Evaluation of food chemicals is essential to make appropriate feeding decisions. The molecular genetic analysis of Gustatory receptor (Gr) genes and the characterization of the neural circuits that they engage has led to a broad understanding of taste perception in adult Drosophila [1, 2]. For example, eight relatively highly conserved members of the Gr gene family (Gr5a, Gr61a, and Gr64a-f), referred to as sugar Gr genes, are thought to be involved in sugar taste in adult flies [3-8], while the majority of the remaining Gr genes are likely to encode bitter taste receptors [9-11], albeit some function as pheromone [12-14] and carbon dioxide [15, 16] receptors. In contrast to the adult fly, relatively little is known about the cellular and molecular basis of taste perception in larvae. Here, we identify Gr43a, which was recently shown to function as a hemolymph fructose sensor in adult flies [17], as the major larval sugar receptor. We show that it is expressed in taste neurons, proventricular neurons, as well as sensory neurons of the brain. Larvae lacking Gr43a fail to sense sugars, while larvae mutant for all eight sugar Gr genes exhibit no obvious defect. Finally, we show that brain neurons are necessary and sufficient for sensing all main dietary sugars, which probably involves a postingestive mechanism of converting carbohydrates into fructose.


Assuntos
Encéfalo/fisiologia , Carboidratos/fisiologia , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/fisiologia , Receptores de Superfície Celular/metabolismo , Animais , Animais Geneticamente Modificados , Encéfalo/citologia , Proteínas de Drosophila/genética , Larva/metabolismo , Larva/fisiologia , Neurônios/metabolismo , Receptores de Superfície Celular/genética , Paladar/fisiologia
12.
Turkiye Parazitol Derg ; 35(3): 140-4, 2011.
Artigo em Turco | MEDLINE | ID: mdl-22203502

RESUMO

OBJECTIVE: This study was carried out to determine the molecular characterization of msa-2c gene of one Babesia bovis isolate from cattle in the Aegean Region and to compare identities with similar isolates from the World and Turkey. METHODS: Between 2008-2010 blood samples were collected from a total of 235 cattle localized in 9 provinces of the Marmara and Aegean Regions. Smears were prepared, genomic DNA's were extracted from the blood samples and investigated for Babesia species by RLB. PCR was performed on one sample determined as B. bovis, the obtained amplicon was purified, sequenced and deposited to GenBank. Identities with similar isolates from Turkey and the World were investigated. RESULTS: Bovine babesiosis was not determined in the microscopic examination. According to the RLB results there was no B. bovis positivity in cattle from the Marmara Region, while only one B. bovis positivity was detected in cattle from the Aegean Region. The molecular prevalence of B. bovis was determined as 0.42% in the total of the examined 235 cattle. The sequenced B. bovis isolate shared 91-92% and 89-96% identities with the isolates from Turkey and the World, respectively. CONCLUSION: Molecular characterization of msa-2c gene region of B. bovis detected from cattle in the Aegean Region was carried out in this study.


Assuntos
Antígenos de Protozoários/genética , Babesia bovis/genética , Babesiose/veterinária , Doenças dos Bovinos/parasitologia , Proteínas de Membrana/genética , Proteínas de Protozoários/genética , Animais , Babesia bovis/classificação , Babesiose/epidemiologia , Babesiose/parasitologia , Sequência de Bases , Bovinos , Doenças dos Bovinos/epidemiologia , DNA de Protozoário/sangue , DNA de Protozoário/química , Epidemiologia Molecular , Dados de Sequência Molecular , Filogenia , Prevalência , Alinhamento de Sequência/veterinária , Turquia/epidemiologia
13.
Biomaterials ; 32(31): 7847-55, 2011 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-21820734

RESUMO

Microscale hydrogels find widespread applications in medicine and biology, e.g., as building blocks for tissue engineering and regenerative medicine. In these applications, these microgels are assembled to fabricate large complex 3D constructs. The success of this approach requires non-destructive and high throughput assembly of the microgels. Although various assembly methods have been developed based on modifying interfaces, and using microfluidics, so far, none of the available assembly technologies have shown the ability to assemble microgels using non-invasive fields rapidly within seconds in an efficient way. Acoustics has been widely used in biomedical arena to manipulate droplets, cells and biomolecules. In this study, we developed a simple, non-invasive acoustic assembler for cell-encapsulating microgels with maintained cell viability (>93%). We assessed the assembler for both microbeads (with diameter of 50 µm and 100 µm) and microgels of different sizes and shapes (e.g., cubes, lock-and-key shapes, tetris, saw) in microdroplets (with volume of 10 µL, 20 µL, 40 µL, 80 µL). The microgels were assembled in seconds in a non-invasive manner. These results indicate that the developed acoustic approach could become an enabling biotechnology tool for tissue engineering, regenerative medicine, pharmacology studies and high throughput screening applications.


Assuntos
Fibroblastos/citologia , Hidrogéis/farmacologia , Microesferas , Som , Animais , Sobrevivência Celular/efeitos dos fármacos , Células Imobilizadas/citologia , Células Imobilizadas/efeitos dos fármacos , Células Imobilizadas/metabolismo , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Camundongos , Células NIH 3T3
14.
PLoS One ; 6(4): e19344, 2011 Apr 28.
Artigo em Inglês | MEDLINE | ID: mdl-21552485

RESUMO

Decellularization and cellularization of organs have emerged as disruptive methods in tissue engineering and regenerative medicine. Porous hydrogel scaffolds have widespread applications in tissue engineering, regenerative medicine and drug discovery as viable tissue mimics. However, the existing hydrogel fabrication techniques suffer from limited control over pore interconnectivity, density and size, which leads to inefficient nutrient and oxygen transport to cells embedded in the scaffolds. Here, we demonstrated an innovative approach to develop a new platform for tissue engineered constructs using live bacteria as sacrificial porogens. E.coli were patterned and cultured in an interconnected three-dimensional (3D) hydrogel network. The growing bacteria created interconnected micropores and microchannels. Then, the scafold was decellularized, and bacteria were eliminated from the scaffold through lysing and washing steps. This 3D porous network method combined with bioprinting has the potential to be broadly applicable and compatible with tissue specific applications allowing seeding of stem cells and other cell types.


Assuntos
Escherichia coli/citologia , Viabilidade Microbiana , Engenharia Tecidual/métodos , Tecidos Suporte/química , Tecidos Suporte/microbiologia , Animais , Proliferação de Células/efeitos dos fármacos , Hidrogéis/química , Hidrogéis/metabolismo , Hidrogéis/toxicidade , Camundongos , Células NIH 3T3 , Porosidade
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