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Item Identificação proteômica, expressão heteróloga, citolocalização, estudos de regulação transcricional e traducional da Aconitase Mitocondrial de Paracoccidioides brasiliensis(Universidade Federal de Goiás, 2009-11-18) BRITO, Wesley de Almeida; SOARES, Célia Maria de Almeida; http://lattes.cnpq.br/8539946335852637Paracoccidioides brasiliensis is a thermal-dimorphic fungus, the causative agent of Paracoccidioidomycosis (PCM), an important endemic mycosis in Latin America. A protein species preferentially expressed in yeast cells with a molecular mass of 80kDa and isoeletric point (pI) of 7.79 was isolated from the proteome of P. brasiliensis and characterized as an aconitase (E.C. 4.2.1.3). Aconitase is an enzyme that catalyzes the isomerization of citrate to isocitrate in both the Krebs cycle (KC) and the glyoxylate cycle (GC). We report the cloning and characterization of the cDNA encoding the aconitase of P. brasiliensis (PbACO). The cDNA showed a 2337 bp open reading frame (ORF) and encoded a predicted protein with 779 amino acids. A polyclonal antibody against the purified recombinant PbACO was obtained in order to analyze the subcellular localization of the molecule in P. brasiliensis. The protein is present in the extracellular fluid, cell wall, mitochondria, cytosol and peroxisomes of yeast cells as demonstrated by western blot and immunocytochemistry analysis. The expression analysis of the Pbaco gene was performed through quantitative real time RT-PCR and results demonstrated increasing expression during differentiation from mycelium to yeast cells. Real time RT-PCR assays was also used to evaluate the Pbaco expression when the fungus grows on media with acetate and ethanol as sole carbon sources and in different iron levels. The results demonstrated that Pbaco transcript is over expressed in acetate and ethanol as sole carbon sources and in highiron conditions.Item Caracterização do plasmídeo pVCM 04 extraídos de Salmonella enterica isolada de carcaçãs de frangos(Universidade Federal de Goiás, 2010-05-31) CARNEIRO, Lílian Carla; BATAUS, Luiz Artur Mendes; http://lattes.cnpq.br/5637230378599476A small cryptic plasmid isolated from Salmonella enterica Enteritidis called pVCM04 was sequenced and characterizated. pVCM04 is a 3583 pb circle molecule that showed no homology with other plasmids deposited in the GenBank. 12 ORF with more than 50 aminoacids were predicted using the ORF finder program. ORF1 and ORF2 showed homology with replication proteins of different plasmids. ORF 3-5 showed homology with mobilization proteins present in several plasmids; the others seven ORF showed no homology with genes deposited in GenBank. The pVCM04 possess a region with more than 500 pb that is not associated with none of the predicted proteins. This region is organizated in a G+C rich, A+T rich and two repeat direct sequences. The second repeat direct sequence contains a region of DnaA box connection (TTTACAC). This region is probably associated to the replication origin theta type. The phylogenetic relationship among replicase and mobilization deduced protein showed highest similarity of replicase proteins than mobilization proteins. Conjugation experiments showed that the pVCM04/pUC18 fusion not have a good ability to transfer, the plasmid stability test showed that the cells lost 60% of pVCM04/pUC18 on the first day of cultivation. The characterization suggests that the pVCM04 probably would be a cryptic plasmid from fusion of different ancestral plasmid.Item Morfologia, taxonomia, filogenia, anatomia foliar e titoquímica de espécies do gênero Hyptis Jacq. (Labiatae) ocorrentes em em Goiás e Tocantins(Universidade Federal de Goiás, 2010-04-29) FERREIRA, Heleno Dias; FERRI, Pedro Henrique; http://lattes.cnpq.br/2129799749473005; REZENDE, Maria Helena; http://lattes.cnpq.br/5093753722360659The genus Hyptis, with 280 species worldwide, belongs to the family Labiatae (Lamiaceae), subfamily Nepetoideae, tribe Ocimae, and subtribe Hyptidinae. The genus was divided in 27 sections, with 13 of them (Apodotes, Cephalohyptis, Cyanocephalanthus, Cyrta, Eriosphaeria, Gymneia, Induratae, Mesosphaeria, Polydesmia, Pusilae, Trichosphaeria, and Hylodontes) occur in the states of Goiás and Tocantins (Brazil). The collection of botanical material for morphological, taxonomical, anatomical, phytochemical studies, as well as geographical distribution and phylogenetic relationships, was realized with several field trips in the states of Goiás and Tocantins. For morphological studies, several loans of herbarium specimens of Hyptis were requested from Brazilian and international institutions. The genus Hyptis is mostly American, with Neotropical distribution, ranging from southern United States to Argentina. In South America, most species of Hyptis are associated with open areas, and occur from 300 m altitude in the valley of the Rio Araguaia, in Goiás, to 3300 m in the western slopes of the Peruvian Andes. In Goiás and Tocantins they can be found in several vegetation types: cerrado, rocky outcrops, margins of cow pastures, cultivated fields, wet fields, humid forests, and flooded areas with Mauritia palms. In these states they occur in a variety of soils: sandy, sandygravelly, with rocky outcrops, hydromorphic, and latossoils. Specie of the section Cyrta are exclusive of humid and flooded soils. The two main centers of diversity of Hyptis in Brazil are in the states of Minas Gerais and Goiás. Many herbarium specimens were analyzed and many species were studied and collected during field trips in Goiás and Tocantins. The geographic position of each collection was verified with the use of a GPS instrument. The specimens collected were deposited in the UFG Herbarium of the Federal University of Goiás. Thirty-two distribution maps of Hyptis in Goiás and Tocantins were elaborated from the specimens studied. A total of 89 species was catalogued, and 18 of them reported from Goiás and Tocantins were not found during the collecting trips. A key for the identification of the species was elaborated using morphological characters. A parsimony analysis using 35 morphological characters was realized, obtaining 1864 most parsimonious trees and a strict consensus tree. The parsimony analysis supports the monophyly of the sections Gymneya, Cyrta, Apodotes, and Cyanocephalus (except for the H. nitidula - H. peduncularis clade), and indicates that the sections Mesospheria, Xylodontes, Hyptis, Polydesmia, and Eriosphaeria are not monophyletic. Section Trichosphaeria is not well resolved. An anatomical study of 60 species, representing 11 sections of Hyptis, was undertaken. Some characters, as hypodermis, schlerenchimatic sheath extensions, trichomes, stomatal cripts, supply useful taxonomic for inferring phylogenetic relationships within the genus. For the analyses of chemical components, 29 species and two varieties of Hyptis were selected. With the chemical analises, 216 constituents of essential oils were identified, mostly monotherpenes and sesquiterpenes. Average percentage and pattern of each chemical component were calculated. The most common chemical components were arbitrarily selected, as present in nine or more species: sabinene (9 spp.), 1, alpha-tujene, alpha-cubebene, beta-selinene, and 14-hidroxy-(Z)-cariophyllene (10 spp.). alpha-muurolol and 8(13)-dien- 5beta-ol (11 spp.), mircene, cariophil-4 and germacrene B (12 spp.), gamma-cadinene, epialpha- caninol (13 spp.) and 14-hydroxy-9-epi-(E)-cariofilene (13 spp.), 1-epi-cubenol- and gamma-muurolene (14 spp.), humulene poxide II and bicyclogermacrene (16 spp.), betaelemene and alpha-cadinol (17 spp.), beta-pinene (18 spp.), limonene (19 spp.), espatulenol and germacrene D (24 spp.), alpha copaene and beta-bourbonene (25 spp.), delta-cadinene (26 spp.), alpha-humulene (27 spp.), cariofilene oxide and (E)-cariofilene (28 spp.). Two dendrograms were obtained with two multivariate analyses of chemical constituents, one with quantitative data and the other with qualitative data (presence/absence). Species of section Cyanocephalus were the only ones found in a consistent group in both dendrograms based on chemical constituents. The data obtained in the present study contribute important information for the taxonomy and phylogenetic relationships of the genus Hyptis and the subtribe Hyptidinae, and ultimately for the family Labiatae.Item Biotividade de extratos e frações das folhas da Eugenia uniflora L. e da Hyptidendron canum (Pohl ex Benth.) Harley em microrganismos (Bactérias e fungo) e em Oreochromis nilotius(Universidade Federal de Goiás, 2009-01-22) FIUZA, Tatiana de Sousa; PAULA, José Realino de; http://lattes.cnpq.br/3191837532986128; SABÓIA-MORAIS, Simone Maria Teixeira de; http://lattes.cnpq.br/6723881044959716The purpose of this work is to perform the pharmacognostic studies of the species Hyptidendron canum (Pohl ex Benth.) Harley (Lamiaceae) and Eugenia uniflora L. (Myrtaceae), investigate the population variability of the constituents of the essential oil from H. canum leaves and inflorescences, and assess the bioactivity of the crude ethanol extracts and fractions from leaves of this plants against microorganisms (bacteria and the fungus Candida albicans) and on Oreochromis niloticus Linnaeus. For the variability study of the H. canum essential oil, samples obtained from leaves and inflorescences from Hidrolândia, Silvânia, Bela Vista and Goiânia cities were analyzed by gas chromatography and gas chromatography with mass spectrometry. The anatomic analysis and the phytochemical screening of the leaves from the two species were performed using conventional techniques. The antimicrobial activity was assessed with Gram-positive and Gram-negative bacteria and with the Candida albicans fungus using the well diffusion test and the agar dilution method to determine the minimal inhibitory concentration (MIC). The biological activities of the crude ethanol extract and ethyl acetate, hexanic and chloroformic fractions of the leaves from the two species were tested in O. niloticus fish hepatopancreas and gill. The H. canum essential oils analysis indicated high chemovariability in the oils from different locations. The phytochemical screening and the thin layer chromatography (TLC) analysis of the H. canum leaves evidenced the presence of flavonoids, saponins, terpenes and lignans, while the E. uniflora leaves evidenced the presence of tannins, steroids, triterpenes, anthraquinonic heterosides, saponinic and flavonoids. As for the antimicrobial activity testing, the crude ethanol extracts from both species presented antimicrobial activity against all the microorganisms tested and the E. uniflora ethyl acetate fraction presented satisfactory activity against resistant Pseudomonas aeruginosa. The H. canum crude extract and fractions presented vasoactive activity in the hepatopancreas and gill of the tilapias, causing varying degrees of vasodilation and vascular congestion. An apparent proliferation of blood capillaries was detected in the hepatopancreas of the fish that ingested the hexanic fraction. The E. uniflora crude extract and the ethyl acetate, chloroform and hexanic fractions caused vasodilation, vascular congestion and tissue alterations in the O. niloticus hepatopancreas and gillsItem Caracterização do estado indiferenciado de células tronco embrionárias murinas expandidas na presença de nanopartículas magnéticas e isolamento de células tronco embrionárias a partir de blastócitos bovinos(Universidade Federal de Goiás, 2009-08-14) FREITAS, Erika Regina Leal de; GUILLO, Lídia Andreu; http://lattes.cnpq.br/3401436781775091Magnetic nanoparticles (MNPs) have been used in a great variety of biomedical applications, especially in cancer treatment, drug delivery, and diagnosis by magnetic resonance imaging. Embryonic stem cells (ESCs), due to its capacity of auto-renewal and differentiation in some types of cells, offer a great potential for its use in tissue regeneration and in alternative treatments for many degenerative diseases. The study had as aims: i) to evaluate in vitro citotoxicity of maghemite nanoparticles functionalized with lauric acid, DMSA, and citrate in human melanoma cells (SK-MEL-37) by the MTT assay, electronic microscopy, and DNA electroforesis by agarose gel; ii) to develop a culture system using the previously selected MNPs and a magnet to expand in vitro murine embryonic stem cells (mESCs) in the absence of a co-culture of murine embryonic fibroblasts (MEF) (the indifferentiated state of the mES was analyzed by alkaline phosphatase cytochemistry, electronic microscopy, and analysis of Oct-4 and Nanog gene expression by RT-PCR); iii) to isolate and expand ESCs from bovine blastocysts, and to characterize its pluripotency by analysis of Oct-4 and STAT-3 gene expression by RT-PCR. The MNPs coated with lauric acid, citrate, and DMSA showed no citotoxicity, judging by the high values of IC50 found (254, 433 and 2260 μg-iron/ml, respectively), and that the nanoparticles coated with citrate was chosen to expand the mESCs. The doubling time for the cells cultivated in the presence of MNPs was slightly higher than in the presence of MEF (20.67 ± 0.19 vs. 15.95 ± 0.21) (p= 0.001). The mESCs cultivated in the presence of MNPs showed morphology similar to ESCs, and its pluripotency was confirmed by expression of indifferentiation markers Oct-4 and Nanog by RT-PCR and high alkaline phosphatase activity. One bovine embryonic stem cell (bESCs) line was obtained and maintained by six subcultures for 60 days period. The pluripotency of the bESCs was confirmed morphologically as well as by Oct-4 e STAT-3 gene expressionItem Isolamento e identificação de Klebsiella de solo de cerrado e o uso de suas enzimas para modificação química de fungicidas(Universidade Federal de Goiás, 2008-10-23) LOPES, Flavio Marques; SILVEIRA, Sydnei Mitidieri; http://lattes.cnpq.br/9042287508998087; FERNANDES, Kátia Flávia; http://lattes.cnpq.br/9737543228759171In this research, open ground samples (Brazilian Savannah Cerrado) proceeding from treated cultures of soy with Opera® (epoxiconazol and piraclostrobin) had been used as source of resistant microorganisms to fungicide. The sample were collected in July 2006, in three points of planted area and two depths, 0 to 20 cm, and from 80 cm to 1m, next to the state highway GO-139, in the towns of São Miguel do Passa Quatro, in a producing farm model of soy seeds, where Opera® was used since its launching in 2002. The collection point was characterized as a risk place for ecological accidents, presenting area with extensive agriculture activity, intense use of pest management, soil with good level of infiltration and storage of water, and areas of declivity. The microorganism s selection was carried through using selective solid J.E. supplemented with 0.1% of Opera® with microorganisms collected in depth of 80cm. In a previous selection, 9 microorganisms were isolated, and from these, the one that showed a better growth within the selective J.E. liquid supplemented with 0.03% of Opera® (1805) was chosen for the development of this research. The analysis for coloration of Gram-negative showed that the isolated microorganism is one bacillococcus Gram-negative with poliphormic characteristics, being sensible to all the tested antibiotics. Biochemical identification made through the method of Bactray®, presented a correlaction of 99% with the genus Klebsiella. Through molecular identification by the gene 16S rDNA, it was possible to find a correlaction of 99% of similarity with the species K.pneumoniae and K. oxytoca. The proteins released by the Klebsiella in a supplemented environment with 0.03% of Opera had been precipitated with acetone, obtaining two proteinic fractions, a soluble (FS) and the other insoluble (FI) in a 0,1 mol L-1 phosphate buffer, pH 6.0. In the soluble solution (FS), it was found an oxirredutase from the peroxidases family, which uses hydrogen peroxide as an agent and several substrates as reducing agents, presenting greater activity for pyrogallol, TMB, o-dianisidine and catechol. In a chromatogram obtained through separation by molecular exclusion in Sephadex G- 100, two peaks with peroxidasic activities had been observed. Peak 1 presented two bands in SDS-PAGE, one with39.3 kDa and another one with 24.6 kDa. In the FI, it was possible to determine the presence of one hydrolysis with capacity to break the bonds between the ester molecule, from the family of carboxylesterases esterase B. The esterase B presented activity against casein substrates (9.3 UE), BApNA (1.31 UE), pNPP (0.01), presenting 5,4% activity of inhibition after incubation with E64, 6.5% activity of inhibition after incubation with protease inhibitors cocktail containing AEBSF and 7.5% of inhibition after incubation with TLCK. The electrophoreses of FI presented two protein bands with molecular mass of approximately 61.7 kDa for superior band and 60.7 kDa for the inferior band. The enzymatic characterization using pNPP showed high activity in pH 5.0 and pH 7.0. Esterase B kept 100% of activity after 120 minutes of incubation at 60oC. The presence of ZnCl2, EDTA, MgSO4 and CuSO4 had presented an inhibition effect of 16%, 16%, 23.2% and 40.8% respectively, within the enzyme activity. Esterase B slightly presented higher activity in the presence of DMSO and acetone, and a reduction of up to 87% comparing to isopropyl alcohol (lower polarity). The vale of Km using pNPP as substratum was of 3.4 mmol L-1, with a Vmax of 0.019 UE. When the selective J.E. supplemented with 0.03% Opera was treated with Klebsiella for 120 h, it had a reduction of 43.7% in the absorbance of the compounds with a peak of 200 nm, 55.7% - 220 nm and 28.7% - 260 nm. Using released enzymes by Klebsiella for treatment of the epoxiconazol standards in the reduction of the absorbance 200, 220 and 260 nm the observed results were 99.7%, 95.7% and 99.5% respectively. Whereas for the pyraclostrobin, the observed reduction were 95.5% and 80.2% for the wave lengths 200 and 260 nm respectivelyItem Detecção e caracterização molecular de talassemia alfa(Universidade Federal de Goiás, 2009-03-27) PENNA, Karlla Greick Batista Dias; BATAUS, Luiz Artur Mendes; http://lattes.cnpq.br/5637230378599476Alpha thalassemia is a syndrome resulting from disturbances in the synthesis of alpha globin chain that forms the tetramer of the hemoglobin molecule. Alpha thalassemia is classified into four types according to the number of alpha genes affected: silent carrier (-α/αα), alpha thalassemia trait (--/αα or -α/-α) Hemoglobin H disease (-- /-α), and fetalis hydrops (--/--). The decrease in synthesis of alpha globin causes inadequate production of hemoglobin resulting in hypochromic and microcytic anaemia. Also it causes accumulation of beta chains, inside the erythrocytes, resulting in formation of beta chain tetramer of hemoglobin called Hb H. Clinically the individual with thalassemia can be asymptomatic or present severe anemia. Asymptomatic forms of thalassemia, silent carrier and alpha thalassemia trait, are more difficult to diagnose because of the inclusions bodies of Hb H are not always present. In these situations it is necessary to research the molecular characterization of the genotype and confirming the presence of alpha thalassemia. This is mainly because the diagnosis by conventional methods, although important, is limited and imprecise. This study evaluated some of the traditional laboratory methods in the detection of alpha thalassemia and associated molecular characterization of the more prevalent deletion forms α3,7 and α4,2. For confirmation and characterization of alpha thalassemia, new oligonucleotides were designed. By conventional PCR technique, using 3.7F/KGB01 primers it was possible to detect the deletion α3,7, differentiating the normal genotype (αα/αα), the heterozygote (-α3,7/αα), and homozygous (-- α3,7/- α3,7). Although it was designed to detect the deletion α3,7, this primers also identified the deletion α4,2 when in homozigose (-α4,2/- α4,2). The primers KGB04/KGB05 detected the deletion α4,2, but without differentiating between the heterozygous and homozygous genotype. The most prevalent deletion founded was the α4,2 (20.0%) which represents 9.2% in the homozygous form (- α4,2/-α4,2). The deletion α3,7 in the heterozygous form was detected in 12.3% of patients. The data demonstrate that the importance of molecular detection for alpha thalassemia is not limited only to the definition of the genotype, but also confirmation of the presence in patients with abnormal erythrogram values, with regular erythrogram values, with values closed to the boundary values and in neonates.Item Estudo da associação do polimorfismo genético em carcinomas da tiróide(Universidade Federal de Goiás, 2010-02-24) REIS, Angela Adamski da Silva; CRUZ, Aparecido Divino da; http://lattes.cnpq.br/7868817504129985Thyroid nodules are common in clinical practice and the incidence of thyroid cancer is increasing throughout the world. Certainly, an important factor for the increase of the incidence is the use of ultrasound and PAFF. The identification of genetic polymosphism is important for understanding the potential mechanisms involved in thyroid carcinogenesis. We hypothesized that polymorphisms of xenobiotic enzyme system (CYP1A1, GSTM1 and GSTT1) and the common germline polymorphism of TP53 gene at codon 72 may be associated with the risk of thyroid cancer. To evaluate the role of such polymorphisms, we investigated 122 cases of thyroid nodules, classified according to the following: 35 malignant neoplasic nodules (MNN), 20 benign neoplasic nodules (BNN) and 67 non-neoplasic nodules (NNN) compared with 134 controls of the healthy individuals randomly selected. The PCR-RFLP was used in the analysis of the CYP1A1m1 and CYP1Am2 genotypes; the multiplex PCR was used in the deletion analysis of the GSTM1 and GSTT1; and for the determination of the polymorphism in the gene TP5372, the samples were submitted to conventional PCR reaction. We included case-control studies that compare the incidence of germline polymorphism of TP5372 in patients with thyroid cancer by DerSimonian-Laird method. Our results demonstrated that CYP1A1m1 and CYP1A1m2 genotypes were frequent not only as neoplasic thyroid nodules and non-neoplasic thyroid nodules but also in the control group, which suggests that those are not associated with thyroid nodules. The null genotype for the GSTT1 gene was predominant in benign nodules compared with the control group (p<0,005) indicated that individuals that possess such null genotypes presented a predisposition to benign thyroid diseases than malignant. The risk analysis, done by Odds Ratio, suggests that the risk genotypes GSTM1 (OR=12,82; p=0,004) and GSTT1 (OR=4,53; p<0,0001) contribute to the development of the BNN and NNN, respectively. The frequency of the p53 Arg allele was significantly higher in both patient and control groups. The genotype p53Arg Arg presents a lower risk to thyroid cancer, indicating that the allele arginine in homozygosis can present a protective effect against thyroid carcinogenesis (OR: 0.15; p<0.0001). The data of the meta-analysis demonstrates that the relation between the genotype and phenotype from the TP5372 polymorphism is not associated with the genetic susceptibility at thyroid cancer. The high incidence of thyroid pathologies among women is a characteristic not completely understood. Therefore, some factors, such as the imbalance of sexual hormones; nutritional deficiencies, especially in iodine; therapeutic exposure to radiation; failures in the control systems of the cellular cycle; and the genetic polymorphism could explain the high incidence of thyroid disease among women. Although the interindividual variation in the susceptibility to thyroid diseases could indicate new perspectives to an early diagnostic and prognostic, the polymorphic profile requires additional studies.Item Avaliação das atividades genotóxica, antigenotóxica, angiogênica e potencial de cicatrização do látex da Synadenium umbellatum Pax(Universidade Federal de Goiás, 2009-11-27) REIS, Paulo Roberto de Melo; CHEN, Lee Chen; http://lattes.cnpq.br/4621907105842007Synadenium umbellatum Pax (1894) is a vegetable species from Euphorbiaceae‟s family. The species of this family produce latex. The latex extracted from S. umbellatum has been used by the Brazilian people as anti-tumoral, anti-inflammatory and wound healing agents. However, this latex presents toxic substances and proteolic enzymes. The aim of this work was to evaluate the possible genotoxic, antigenotoxic, cytotoxic, angiogenic and wound healing activities of S. umbellatum latex (SuL) in rats. The genotoxic and antigenotoxic activities were evaluated by mouse bone marrow micronucleus test. An alkylant agent mitomycin C (4 mg.kg-1 body weight) was used as a positive control. For all doses, micronucleated polychromatic erythrocytes (MNPCE) frequency was evaluated after 24 hours of treatment. To evaluate the antimutagenic activity, animals were treated with 10, 30, 50 and 100 mg.kg-1 and 4 mg. kg-1 of mitomycin C (MMC) simultaneously. The frequency of MNPCE was evaluated 24 hours after exposure. Cytotoxicity was evaluated by the polychromatic and normochromatic erythrocytes ratio (PCE/NCE). Angiogenic activity of the SuL was evaluated in chorioallantoic membrane (CAM) of fertilized chicken eggs. The concentrations of the SuL were of 10 and 20 mg/mL. Wound healing activity was evaluated in skin of rats. The skin fragment was removed with area 3 x 2,5 cm and submitted to topic application with 5 mL of SuL (concentration 10 mg/mL) and positive and negative controls. The results showed that the SuL was strongly mutagenic, cytotoxic, and antimutagenic, and a moderate activity of anticytotoxicity. In the evaluation of the angiogenesis, the SuL has promoted the increase of the proliferation of blood vessels in the chorioallantoic membrane of fertilized chicken eggs and it also induced the increase of velocity of wound healing in rat skin lesion.Item O polimorfismo do gene p5372(RP) no câncer de cabeça e pescoço: estudo de associação e meta-análise(Universidade Federal de Goiás, 2009-10-20) SILVA, Antonio Márcio Teodoro Cordeiro; CRUZ, Aparecido Divino da; http://lattes.cnpq.br/7868817504129985Head and neck cancer arises in the oral cavity and nearby regions, larynx, pharynx, including oropharynx, nasopharynx, and hipopharynx. Extensive epidemiologic studies have revealed that chronic tobacco smoking and alcohol consumption as the two main risk factors associated with the multifactorial etiology of head and neck cancers. Additionally, nutritional status, HPV infection, and genetic polymorphism were also related with the disease. A frequently studied polymorphism in Squamous Cell Carcinoma (SCC) of head and neck is a G-to-C SNP (Single Nucleotide Polymorphism) at codon 72 in the p53 gene, which codes for an Arg or Pro (Arginine or Proline) in P53 protein. In order to investigate the distribution and potential association of that SNP in SCC, biological samples were obtained from 331 cases of head and neck SCC from Araújo Jorge Hospital and 271 healthy control individuals from Goiânia (Brazil) population. DNA was isolated and subsequently used for PCR amplification to genotype cases and controls with respect to their p5372 SNP. Additionaly, a meta-analysis was carried out using 29 relevant case-control studies that used p5372 SNP genotyping in SCC of the head and neck. Allelic frequencies for cases were 73.3% and 27.7% for Arg and Pro, respectively. On the other hand, control allelic frequencies were 74.2% and 25.8% for Arg and Pro, respectively (p = 0.119). Genotypic frequencies were 56.8% Arg/Arg, 32.9% Arg/Pro, and 10.3% Pro/Pro for all cases. The control genotypic frequencies were 61.3% Arg/Arg, 25.8% Arg/Pro and 12.9% Pro/Pro (p = 0,137). According to the current data and meta-analysis, no association between p5372 SNP and the development of SCC of the head and neck was found. Although, the homozygous genotype Arg/Arg was found as an important oncogenic risk factor associated with the carcinoma of the oropharynx.Item Avaliação Genético-Molecular do Carcinoma das Células Escamosas da Laringe(Universidade Federal de Goiás, 2009-10-21) SILVA, Cláudio Carlos da; CRUZ, Aparecido Divino da; http://lattes.cnpq.br/7868817504129985The larynx is a structure of the upper aerodigestive tract responsible for the production of sounds as well as protecting the lower airways and helping during the normal act of swallowing. Any pathology which affects the larynx can impose several challenges that disrupt its normal physiological function, and consequently and directly resulting in reduction of the patient s quality of life. Among the different diseases that affect the larynx, cancer is one of the most serious. The squamous cell carcinoma (SCC) of the larynx is a multifactorial disease, influenced by environmental factors and individual behavioral, habits, and susceptibility. The current study describes the molecular and genetic assessment of 20 patients with the squamous cell carcinoma of the larynx. In summary, the study strategy included the analyses of the genetic polymorphism of codon 72 of the TP53, the detection and genotyping of HPV genome, assessing genomic instability (MIS and LOH) and random chromosomal imbalances using PCR and CGH approaches. Regarding to the polymorphism of the TP53 gene, arginine homozygous genotypes (p53AA) and arginine-proline heterozygous genotypes (p53PA) were found in 65% (13/20) and 35% (7 / 20) of cases, respectively. HPV genome was found in association with tumor cells in 20% of SCC cases. HPV 16, 11, and 45 were the genotypes identified. Moreover, co-infection of HPV 11 and 45 was also observed. Both samples found positive for HPV genome were associated with p53AA at the TP53 gene. Our results corroborated the data published in the literature that described an increased susceptibility to the virus-induced carcinogenesis of the larynx in arginine homozygous genotypes. Here in we report genomic instability for a panel of 8 microsatellite markers distributed in 5 chromosomes. One locus (D8S135) was found in homozygous for all cases. On the other hand, RH-92600 was not included in the analysis mainly because it was also found homozygous for most cases. We found 76.2% (244/320) of informative loci. Thus the mean frequency of MIS was 9.0% (22/244) and LOH was 6.1% (15/244). The frequency of MIS was found higher than the LOH, suggesting that the tumorigenesis of SCC of the larynx follows a mechanism of mutator phenotype. Using CGH, chromosomal gains were observed in 1q21qter. On the other hand, chromosomal losses occurred in chromosome 3p21p28, 11q23, 16p16, 16q12, 17p13pter and 22q13. The techniques of comparative genomic hybridization have improved the understanding of genetic alterations in squamous cell carcinoma of the larynx, especially for characterizing the relationship between early precursors of laryngeal carcinomas, as well as to identify genomic regions that may contain oncogenes and tumor suppressor genes involved in the tumorigenesis of this anatomical site.Item Estudo morfoanatômico, avaliação da atividade biológica e composição química e variabilidade do óleo essencial da Memora nodosa (SILVA MANSO) Miers - Bignoniaceae(Universidade Federal de Goiás, 2008-12-18) TRESVENZOL, Leonice Manrique Faustino; PAULA, José Realino de; http://lattes.cnpq.br/3191837532986128Memora nodosa (Silva Manso) Miers is a native savannah plant popularly used for wound cicatrization (leaves and stems) and for treating abdominal pains and scabies (roots). The purpose of this work is to study the morphoanatomy and determine the composition and variability of the constituents of the essential oil obtained from the M. nodosa leaves, as well as its antimicrobial, leishmanicidal, cicatrizing activity and toxicity. Morphologically, M. nodosa is a bush with clumps of straight or curved stems. It has a deep, pivoting root and presents a chandelier shaped structure in the upper part from where the stems sprout. It presents compound, bipinnate, imparipinnate, oppositely crossed leaves. The inflorescence is racemose with golden-yellow flowers. It is characterized by septifrague capsule type fruit with replum and winged seeds. Anatomically the leaves are hypostomatic with predominantly paracytic stomates. The epidermis is uniseriate with a thick cuticle and glandular trichomes throughout the structure of the young leaves. The essential oil from the M. nodosa leaves collected from seven different locations in the Brazilian savannah was analyzed by GC(FID) and GC/MS. Five substances were identified, benzaldehyde and 1- octene-3-ol were the main constituents. Two different classes regarding sampling location and constituents were identified by means of the principal component and grouping analyses. The antimicrobial activity of the ethanolic extracts from leaves, stems, roots and of essential oil from leaves, and the fractions obtained from the roots, were assessed against Grampositive and Gram-negative bacteria, as well as the Candida albicans fungus, using well diffusion test and the agar dilution method. The ethanolic extract from the roots presented small antimicrobial activity while the hexane and dichloromethane fractions obtained from this extract presented MIC 0.39 mg/mL and 0.78 mg/mL against a number of Gram-positive bacteria and the Candida albicans fungus. The essential oil presented MIC 0.78 mg/mL to 1.56 mg/mL. The antimicrobial activity of the essential oil was the result of synergic action between the benzaldehyde, 1-octene-3-ol, 1-octanol, linalool and the mandelonitrile, constituents of the essential oil. Promastigote forms of Leishmania chagasi were used for the leishmanicidal assessment and it was verified that the ethanol extract from the leaves presented an IC50 of 93.2 μg/mL and from the roots presented an IC50 of 95.87 μg/mL. The hexane fraction from the roots presented an IC50 of 13.51μg/mL. A mixture of β-sitosterol and stigmasterol was isolated and identified in the hexane fraction obtained from the ethanol extract from the roots. This mixture was tested against L. chagasi resulting in an IC50 of 69.79 μg/mL. The cicatrizing activity of the ethanol extract from the leaves and roots was assessed on skin wounds in rats, and it be noted reepithelization occurred faster only in the group treated with a 2% aqueous solution of the ethanol extract from the roots. This action may be related to the alantoine isolated in the ethanol extract from the roots. The toxicity assessment of the ethanol extracts from roots, leaves and hexane fraction obtained from M . nodosa roots, tested on rats, mice and Artemia salina proved that this plant is not potentially toxic. In conclusion, according to the tests performed, the best results were obtained with ethanolic extract and it presented antimicrobial, wound cicatrization and leishmanicidal activities. Hexane and dichloromethane fractions obtained from roots extract presented antimicrobial activity against Gram-positive bacteria and the Candida albicans fungus and hexane fraction strong leishmanicidal activity.