Teses em Genética e Biologia Molecular (Doutorado) - PPGBM/ICB

URI Permanente para esta coleçãohttps://repositorio.ufpa.br/handle/2011/8840

O Doutorado Acadêmico pertence ao Programa de Pós-Graduação em Genética e Biologia Molecular (PPGBM) do Instituto de Ciências Biológicas (ICB) da Universidade Federal do Pará (UFPA).

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Agora exibindo 1 - 9 de 9

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Alterações genômicas quantitativas com potencial clínico no adenocarcinoma gástrico
(Universidade Federal do Pará, 2016-05-31) ARAÚJO, Taíssa Maíra Thomaz; KHAYAT, André Salim; http://lattes.cnpq.br/6305099258051586
Gastric cancer is the fifth most frequent type of cancer and the third cause of cancer mortality worldwide. Despite progression in treatment of advanced gastric cancer, the prognosis of patients remains poor, in part due to the low rate of diagnosis during its early stages. This paradigm implies the necessity to search and identify molecular biomarkers for early gastric cancer diagnosis, as well as for disease monitoring, thus contributing to the development of new therapeutic approaches. Therefore, this study aimed at investigating copy number variations in gastric adenocarcnoma through array Comparative Genomic Hybridization (aCGH) technique and selecting genes for validation in a larger sample size by using real-time PCR, in order to find potential molecular biomarkers for this tumor type. The aCGH results demonstrated 22 gene alterations never described before as correlated with gastric carinogenesis, as well as several alterations significantly associated with serosal extravasation and patients aged less than or equal 50 years. Given that most of the genes had never been described in gastric cancer, we selected for validation four gene alterations that showed some consistency with studies published in the literature for other types of cancer. Thus, we investigated by real-time PCR the amplifications of RTEL1, B4GALT5, TRPV2 and ABCA13 genes. The results showed a high frequency of amplification of these genes, but the statistical associations with clinicopathological data of TRPV2 gene with younger patients and ABCA13, with serosal extravasation, observed by aCGH, were not confirmed. Moreover, new significant associations were demonstrated, including RTEL1 recurrent amplification associated with advanced age and intestinal type of gastric adenocarcinoma; B4GALT5 recurrent amplification associated with intestinal type of gastric adenocarcinoma; TRPV2 recurrent amplification associated with lymph node metastasis; ABCA13 recurrent amplification associated with lymph node metastasis and male patients and co-amplification of RTEL1 and ABCA13 associated with advanced staging. Therefore, the aCGH proved to be a useful tool for the investigating new genes associated with carcinogenesis. Additionally, recurrent amplification of RTEL1, B4GALT5, TRPV2 and ABCA13 seem to have an important role in the development and progression of gastric cancer and can be considered as potential biomarkers for this disease.
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Biologia molecular aplicada à hanseníase: estudo de parâmetros genéticos e epigenéticos em uma amostra do estado do Pará
(Universidade Federal do Pará, 2016-09-02) PINTO, Pablo Diego do Carmo; SANTOS, Ândrea Kely Campos Ribeiro dos; http://lattes.cnpq.br/3899534338451625
Leprosy is caused by Mycobacterium leprae and patients can be grouped in Paucibacillary and Multibacillary. Alternatively, according by Ridley-Jopling (1966), using immune-hystogical criteria, grouped in two distinct pole: (i) Tuberculóide (TT); and (ii) lepromatous (LL), and your intermediaries. Independently these classification, the disease can be affected by molecules that modulates immune response, like genes that encode these molecules, and by small RNA (micro-RNA), wich regulated these genes, thus these study can improve the knowledge about the mechanism of response to infectious process, as well as enable the identification of new possibles biomarkers to assist diagnosis in leprosy. The objective of this study was to investigate eight INDEL polymorphisms on genes CYP19A1, NFKβ1, IL1α, CASP8, UGT1A1, PAR1, CYP2E1, and IL4, to identify possible susceptibility markers of leprosy and evaluate the influence of genetic ancestry on disease risk. Besides was performed the first genome wide miRNA profiling of Leprosy by next generation sequencing (NGS), assessing and describing the expression standard in leprosy. Our study shows that the NFKβ1, CASP8, PAR1, IL4 and CYP19A1 genes are possible markers for the susceptibility to development of leprosy and the severe clinical form MB. Moreover, after correcting for population structure within an admixture population, the results show that different levels of ethnic group composition can generate different OR rates for leprosy susceptibility. The differential expression profile from tissue samples reveal 67 miRNAs differentially expression, with 43 down and 24 upregulated and from blood sample were found a total of 10 miRNAs differentially expression with 9 down and one upregulated. Moreover was performed in silico target analysis and detect the genes (IL1β, IL6, IL8, IL12, TLR2, TLR4, IL17RB, IFNGR1, TGFBR1, NFκβ, família SMAD, STAT3, CASP8, CYP19A1, BCL-2, in others) involved on pathological of leprosy. Lastly, was showed for the first time the genome wide microRNA of leprosy.
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Evolução cromossômica e mapeamento genômico comparativo em morcegos da subfamília Phyllostominae (Mammalia, Chiroptera)
(Universidade Federal do Pará, 2016-07-11) SILVA, Natalia Karina Nascimento da; PIECZARKA, Julio Cesar; http://lattes.cnpq.br/6644368250823351
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Farmacogenômica das fluoropirimidinas no tratamento oncológico personalizado
(Universidade Federal do Pará, 2016-12-29) FERNANDES, Marianne Rodrigues; BURBANO, Rommel Mario Rodriguéz; http://lattes.cnpq.br/4362051219348099
Recently, cancer has become an obvious public health problem worldwide. The Fluoropyrimidine-based regimen has been the most widely used chemotherapy regimen worldwide in several types of solid tumors, including gastric and colorectal cancer. Of the total number of patients treated with 5-Fluorouracil (5-FU), 10-40% have severe toxicities, which usually result in prolonged and costly hospitalizations. The principle of personalized medicine is to study responses to medications based on individual genomic information. The high degree of miscegenation is a challenge for the worldwide implementation of personalized medicine in clinical practice. Many studies in the specialized literature have reported the influence of pharmacogenomic markers in mixed populations such as the Brazilian population. The aim of this study was to investigate the pharmacogenomic variability of different biomarkers in pharmacogenes involved in the metabolism pathway of Fluoropyrimidines in patients with gastric cancer or colorectal cancer, which are sub-strutured according to response and toxicity to treatment. To perform the research we used 216 patients with colorectal or gastric cancer who received 5-FU chemotherapy treatment. We investigated 33 genetic polymorphisms in 17 pharmacogens (ABCB1, ABCC2, ABCC4, ABCG2, CYP2A6, DPYD, FPSG, ITGB5, MTHFR, SLC22A7, SLC29A1, TP53, TYMS, UMPS, GGH, RRM1, TYMP) involved in the metabolism pathway of fluoropyrimidines. Our results showed that 77.3% of the patients presented some type of toxicity related to 5-FU treatment, of which 22% presented severe toxicities classified in grade 3 and 4. Death occurred in 23 patients, where three cases were related to toxicity and four cases with tumor progression and chemotherapeutic toxicity. Population substructuration was not influential in the association results for pharmacogenetic polymorphisms with the use of 5-FU. The FPGS gene (rs4451422) was shown to be significant in association with overall toxicity (p = 0.0052; OR 0.32) and toxicity events (p = 0.0004; OR 0.22). The ABCC4 gene (rs148551) had a significant association with the clinical response (p = 0.0056; OR 0.28). The SLC29A1 gene (rs760370) was shown to be significant for grade 3 and 4 toxicities (p = 0.0033; OR 4.73). In conclusion, due to the high degree of miscegenation in the Brazilian population, and particularly in the North of Brazil, the generated 5-FU pharmacogenomics data are particularly unique when compared to the homogenous populations investigated to date. The ABCC4, FPGS and SLC29A1 genes have been shown to be important biomarkers predictive of personalized medicine therapy using 5-FU.
Acesso aberto (Open Access)
Integração dos estudos cromossômicos e DNA barcoding em Rhamphichthys (Pisces: Gymnotiformes)
(Universidade Federal do Pará, 2016-05-16) SILVA, Patrícia Corrêa da; PIECZARKA, Julio Cesar; http://lattes.cnpq.br/6644368250823351
The Order Gymnotiformes is composed by 219 valid species, which are distributed in five families. The most investigated genera are Eigenmannia and Gymnotus. Our work focused on family Rhamphichthyidae, genus Rhamphichthys that, like other Gymnotiformes, present greater abundance and diversity in the Amazon region. Sampling was carried out in the municipalities of Abaetetuba, Barcarena and Belém (Pará) and Tefé, Ecological Reserve Mamirauá (Amazonas), in order to better define the species, through the integration of classical cytogenetic data, cytogenomic analysis (probes for repetitive DNA sequences) and DNA Barcoding and thus understand the evolution of this fish in the Amazon. A new karyotype was identified for R. rostratus with the presence of B chromosomes and karyotype formula FC = 48m / sm + 2st / a + (5-10) B, as well as a new cytotype from the Amazon region, in Rhamphichthys sp. FC = 44m / sm + 6st / a, and also in R. marmoratus, FC = 46 + 4st / a in the state of Pará. The analysis of repetitive sequences in the new cytotypes demonstrated that probes 18S coincided with the regions of constriction secondary that are marked with silver nitrate in the classical NOR staining technique. The DNA probes 5S mark multiple sites, letting clear that the evolution of the ribosomal gene family is independent, at least in the genus Rhamphichthys. Retroelements REX1 and REX3 marked in a dispersed fashion throughout the genome, as already described in literature for other fishes. The REX1 element also marks the secondary constriction in R. rostratus, which has also been described in other species of fishes that inhabit polluted environments, exposed to environmental stresses and also in hybrid individuals. The barcoding DNA analysis allowed the construction of a Bayesian tree, which is in agreement with the cytogenetic data. Thus, populations of R. rostratus with and without B chromosomes are separate taxa. In turn, the sample from Mamirauá, herein called Rhamphichthys sp., since it was not been formally described, it is more similar in both karyotypic data as the barcoding analysis with R. hanni from southeastern Brazil. Our data let clear that the number of species in Rhamphichthys is underestimated, which reinforces the need for a taxonomic revision of the genus.
Acesso aberto (Open Access)
Parâmetros de saúde metabólica e visual em pacientes diagnosticadas com câncer de mama em tratamento anticâncer
(Universidade Federal do Pará, 2021-09) SIQUEIRA, Maria Lúcia Souza; SOUZA, Givago da Silva; http://lattes.cnpq.br/5705421011644718; https://orcid.org/0000-0002-4525-3971; MONTEIRO, Marta Chagas; http://lattes.cnpq.br/6710783324317390; https://orcid.org/0000-0002-3328-5650
Female breast cancer, according to current estimates, is still reaching a very high level of incidence and prevalence in Brazil and Pará. The need for studies that expand knowledge about therapeutic options and mitigate the damage suffered by cancer becomes innovative in our region. Thus, we aim to determine and assess metabolic and visual health parameters in patients diagnosed with breast cancer exposed to anti-cancer therapies. The specific objectives were divided into three sections: Section I: determine lipid, anthropometric, oxidative stress and pro-inflammatory cytokine markers to assess metabolic health; Section II: determine tamoxifen metabolites (4-hydroxy tamoxifen and endoxifen) and correlate with blood lipids to assess drug interaction and metabolism with lipids; Section III: Perform visual tests to assess the effect of anticancer therapies on retinal thickness. Research approved by the ethics committee with opinion number 1.915.051 (CEP-HOL) and 1.897.057 (CEP-ICS-UFPA) from March 2017 to September 2019. It consisted of 40 women diagnosed with breast cancer (Ductal Carcinoma Grade II and III) in a public cancer reference hospital in Belém of Pará and who consented to participate in the research. A group of 20 patients formed the chemotherapy group (GQt), 20 patients the tamoxifen hormone therapy group (GTam) and another group of women without cancer was constituted as a comparative control (GC) based on the inclusion criteria. For the determination of biochemical markers, the automated method was used; anthropometry followed the Ministry of Health manual; for oxidative stress the colorimetric method; for the measurement of cytokines the ELISA method; for the determination of tamoxifen and metabolites in plasma, High Performance Liquid Chromatography (HPLC) was used; for retinal analyses, optical coherence tomography (OCT) was used. The results obtained showed that the patients were predominantly women aged between 40 and 50 years, brown, with an income of 1 to 3 minimum wages, were in pre- and post-menopause; were overweight/obese with high BMI, between 25 to 29.9 kg/m2, waist circumference above 80 cm and did not perform regular physical activity; the mean levels of lipids were altered in both groups studied, but with emphasis on the chemotherapy group with elevation of total cholesterol, LDL and triglycerides and low levels of HDL cholesterol, (p<0.05); the antioxidant parameters (TEAC, GSH, CAT and SOD) showed low levels of GSH and high CAT activity for the chemotherapy group and less activity for the tamoxifen group compared to the control (p<0.01); the pro-oxidant parameters (MDA and NO) showed that chemotherapy patients had a higher level of lipid peroxidation than tamoxifen patients compared to control (p<0.05); the GSH/MDA ratio showed greater sensitivity to oxidative damage for chemotherapy patients (p<0.01); as for cytokines, TNF-α and IL-6, they were elevated both in the chemotherapy group and in the tamoxifen group (p<0.05). Mean plasma concentrations of tamoxifen, 4-hydroxy tamoxifen, and endoxifen were 62 ng/mL, 1.04 ng/mL and 8.79 ng/mL; triglyceride levels ranged from 59 to 352 mg/dL, total cholesterol from 157 to 321 mg/dL, LDL-c from 72 mg/dL to 176 mg/dL and HDL-c from 25.1 mg/dL to 62.8 mg/dL; there was no significant association between tamoxifen and metabolites with cholesterol and triglyceride levels; there was a weak association between tamoxifen and its active metabolites with HDLc, LDLc and VLDLc. Mean retinal macular thickness revealed no significant difference between patients who received chemoradiotherapy and control tamoxifen (p>0.05); regional macular thickness revealed that only one macular field showed a significant difference between two groups; in the external nasal macular field, the chemoradiotherapy patients showed thinner retinal thickness compared to the control. We conclude that breast cancer patients exposed to chemotherapy and hormone therapy presented: unfavorable lipid profile with high levels of total cholesterol, LDL, triglycerides and low HDL, overweight and obesity, lower antioxidant capacity for patients who received chemotherapy and systemic levels of inflammatory cytokines, TNF-α and IL-6, elevated; there was a weak association between plasma concentrations of tamoxifen and its active metabolites with levels of HDL-c, LDL-c and VLDL-c, with a low impact of lipoprotein levels on exposure to tamoxifen, 4-hydroxytamoxifen and endoxifen; a macular field was altered in the chemotherapy group, which had a thinner retina compared to the control group, however the retinal structure was sensitive to the presence of tamoxifen metabolites.
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Perfil de expressão de microRNAs no câncer oral
(Universidade Federal do Pará, 2016-03-11) LOPES, Camile de Barros; SANTOS, Ândrea Kely Campos Ribeiro dos; http://lattes.cnpq.br/3899534338451625
Oral squamous cell carcinoma (OSCC) is a multifactorial disease involving genetic, epigenetic and environmental factors. It is characterized by a pattern of aggressive growth, high metastatic potential and high mortality rates. Despite advances in technology in the surgical treatment, radiotherapy and chemotherapy, the five-year survival rate has no significant improvement in recent years. By regulating the expression of target genes, microRNAs play an important role in the initiation and progression of human cancers. Therefore, they are a promising tool for the identification of biomarkers of risk, as well as prognostic and therapeutic targets. In order to investigate the miRNAs expression profile in ten tissues of OSCC were characterized based on data generated from high-performance sequencing. Furthermore, by qPCR we assessed the adjacent tissue to the OSCC for four miRNAs. The results showed 17 differentially expressed miRNAs were able to discriminate the tissue without cancer. Among these, we found seven new miRNAs (hsalet- 7c, hsa-miR-10a, hsa-miR-199a, hsa-miR-381, hsa-miR-501, hsa-miR-654 and hsamiR- 941) which are not described in the literature their participation in OSCC. Additionally, we found that the four miRNAs hsa-miR-221, hsa-miR-21, hsa-miR-135b and hsa-miR-29c presented overexpression in the adjacent tissue, confirming the field cancerization effect. The results revealed that these miRNAs are potential biomarkers occurring in OSCC with the ability to identify individuals at high risk of developing this type of cancer, and indicate their utility as potential therapeutic targets.
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Perfil de MicroRNAs hepáticos pode regular apoptose, lesões vasculares e inflamação na dengue hemorrágica
(Universidade Federal do Pará, 2016-06-30) OLIVEIRA, Layanna Freitas de; BURBANO, Rommel Mario Rodriguéz; http://lattes.cnpq.br/4362051219348099
Dengue is the most prevalent arbovirosis in the world caused by Dengue virus (DENV) and is present in all continents, for more than three decades has been a constant public health concern and often fatal by dengue hemorrhagic fever (DHF). The pathogenesis of dengue is closely related to the host immune response, reaching exacerbated inflammation and transient autoimmunity. All tissues are affected, which liver is one of the most important in severe conditions, due its intense viral replication and its significant role in metabolism. The study of microRNAs (miRNA) as regulatory elements of metabolism and immune response during infection is crucial to understanding the regulatory mechanisms of gene expression on DENV infection, and can help in diagnostic development of anti-viral therapies. We sequenced the miRNoma in MySeq platform (Illumina) to identify the miRNAs profile expressed in FFPE liver tissue, ten DHF fatal cases were compared to five control cases. Eight miRNAs exhibited differential expression in DHF liver, miR-126-5p, a regulatory molecule of endothelial cells, and miR-133a-3p are upregulated in dengue and miR-122-5p, a liver-specific miRNA, miR- 146a-5p, interferon regulator, miR-10b-5p, miR-204-5p, miR-148a-5p and miR-423-5p were downregulated. Functional analysis of KEGG pathways and GO terms with predicted target genes of overexpressed miRNAs found regulatory pathways of apoptosis and immune response, involving MAPK gene, RAS, CDK and FAS; immune response pathways showed NF- kB, CC and CX families, IL and TLR. The same analysis with target genes of downregulated miRNAs also identified in most pathways of apoptosis and biosynthetic pathways of metabolism. In our knowledge, this is the first description of the liver miRNA profile in DHF, the results together show a feasible relationship of miR-126-5p, miR-122-5p and miR-146a-5p with liver pathogenesis of DHF, through endothelial repair and vascular permeability regulation, control of homeostasis and liver expression regulation of inflammatory cytokines.
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Reconstrução e modelagem in silico da via de biossíntese de ácidos graxos da bactéria psicotrófica Exiguobacterium antarticum linhagem B7
(Universidade Federal do Pará, 2016-04-04) FRANCÊS, Regiane Silva Kawasaki; SCHNEIDER, Maria Paula Cruz; http://lattes.cnpq.br/3901112943859155
Mathematical modeling in silico based restrictions is an approach adopted by systems biology to analyze metabolic networks. The Gram-positive bacterium Exiguobacterium antarticum B7 is an extremophile organism able to survive in cold environments as glacial ice and permafrost. The ability of these microorganisms of adaptation to cold attracts great biotechnological interest. An important factor for the understanding of cold adaptation process is related to the chemical modification of fatty acids constituting the cell membrane of psicotrophic bacteria in order to maintain membrane fluidity to avoid freezing ofthe bacteria. In this work, the metabolic pathway of fatty acid biosynthesis of the bacterium E. antarticum B7 was rebuilt from its annotated genome. The software tools KEGG (Kyoto Encyclopedia of Genes and Genomes) and RAST (The Rapid Annotation Server) were used to generate a preliminary network model. The next step was to cure manually the genomic, biochemical and physiological informations available in different databases and specific literature. During this process, the FabZ and DesK enzymes responsible for adding carbon-carbon unsaturations in the fatty acid chain during synthesis have been identified in the genome, though in a truncated form. The fluxome metabolic pathway was defined, describing the routes of the main reactions since the first monomer, Acetyl-CoA, to the final product, the Hexadecenoic acid. A computational modeling was done using the software MATLAB® with toolboxes and specific tools for systems biology. The quantification of metabolites produced via was performed by the method constraint-based Flux Balance Analysis (FBA). To evaluate the influence of the gene expression in the fluxome analysis, the FBA method was also calculated using the log2FC values obtained in the transcriptome analysis at 0ºC and 37ºC. The fatty acid biosynthesis pathway showed a total of 13 elementary flux modes, four of which showed routes for the production of hexadecenoic acid. The reconstructed pathway demonstrated the capacity of E. antarcticum B7 to produce fatty acid molecules. Under the influence of the transcriptome, the fluxome was altered, promoting the production of short-chain fatty acids. The calculated models contributes to better understand the bacterial adaptation at cold environments.

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