Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2017-10-04T21:05:45Z No. of bitstreams: 1 ClelioDiogoSoares_DISSERT.pdf: 3089611 bytes, checksum: d4a3b5a01c7dc8a828197d92e650d07e (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2017-10-11T22:38:17Z (GMT) No. of bitstreams: 1 ClelioDiogoSoares_DISSERT.pdf: 3089611 bytes, checksum: d4a3b5a01c7dc8a828197d92e650d07e (MD5) / Made available in DSpace on 2017-10-11T22:38:17Z (GMT). No. of bitstreams: 1 ClelioDiogoSoares_DISSERT.pdf: 3089611 bytes, checksum: d4a3b5a01c7dc8a828197d92e650d07e (MD5) Previous issue date: 2012-08-30 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico (CNPq) / Introdu??o: H? evidencias consider?veis sugerindo que genes relacionados ao metabolismo do folato possuem um papel importante na etiologia das fendas orofaciais. A express?o de genes que possam estar ligados ?s fendas orais requer a presen?a de apropriadas causas ambientais em combina??o com fatores gen?ticos que culminam com a falha na fus?o dos processos faciais. Objetivo: Realizar estudo de associa??o da express?o dos genes da via do metabolismo do acido f?lico - MTHFR, MTR, RFC1, MTRR, com a ocorr?ncia das fendas orais. Metodologia: Foram estudados 50 filhos casos e suas respectivas m?es, e 50 indiv?duos controles e suas respectivas m?es. Inicialmente foram realizadas as an?lises bioqu?micas (Glicose, ALT, AST, Creatinina, Folato, Vitamina B12, Homociste?na), hematol?gicas (hemoglobina, hemat?crito, contagem de hem?cias, os ?ndices hematol?gicos, VCM, HCM e CHCM) e estudo de suas caracter?sticas cl?nicas. Para a realiza??o do estudo de express?o g?nica foi extra?do o RNA total a partir das c?lulas do sangue perif?rico, o qual foi quantificado e analisado quanto a sua pureza e integridade e encaminhado para a obten??o do cDNA a ser utilizado para o estudo de express?o utilizando ensaios pr?-desenhados. Finalmente foi avaliada a influ?ncia de determinados gen?tipos (MTRR A66G; MTHFR C677T; MTHFR A1298C; MTR A2756G; RFC1 A80G) sobre a express?o de RNAm dos respectivos genes estudados. A an?lise estat?stica dos dados foi realizada considerando o n?vel de signific?ncia de 95% (P<0,050) Resultados: Foi evidenciada a presen?a do consumo de ?lcool como fator de risco significativo presente para as m?es caso (P=0,001). Em rela??o as dosagens bioqu?micas n?o foram observadas diferen?as significativas entre os casos e respectivos controles os quais apresentaram valores de AST, ALT e creatinina dentro dos valores de refer?ncia. A dosagem de ?cido f?lico apresentou-se reduzida significativamente para o grupo dos filhos caso (P=0,010) e para suas m?es (P=0,001). Na an?lise hemat?logica n?o foram observadas altera??es em nenhum dos par?metros avaliados como hemoglobina, hemat?crito, contagem de hem?cias, os ?ndices hematol?gicos, VCM, HCM e CHCM dentre os grupos avaliados. A avalia??o da express?o g?nica para o grupo das m?es caso mostrou uma redu??o significativa na express?o do RNAm em todos os genes avaliados: para o gene da metionina sintase (MTR, p=0,008), da metionina sintase redutase (MTRR, p=0,015), do RFC1 (P=0,004) e da MTHFR (P=0,017) comparados com o grupo de m?es controle. No grupo de filhos fissurados, houve uma redu??o significativa na express?o do RNAm para os genes da metionina sintase (MTR, p=0,010) e da metionina sintase redutase (MTRR, p=0,034). Para a an?lise da influencia dos gen?tipos na express?o observou-se que o gen?tipo recessivo (CC) para o polimorfismo A1298C do gene MTHFR poderia estar associada a uma redu??o da express?o de seu RNAm. Conclus?o: Genes do metabolismo de folato relacionados s?o reduzidamente expressos em ambos os grupos caso deste estudo, uma vez que todos os quatro genes (RFC1, MTHFR, MTR, MTR), estavam reduzidos nas m?es e dois genes (MTR, MTRR) em seus filhos. A redu??o da express?o desses genes representa um aumento do risco associado com a presen?a de fendas orais nestes indiv?duos. / Introduction: There is considerable evidence suggesting that folate-related genes play a role in the etiology of oral facial clefts. Clefts are known to have a strong genetic component. The expression profile of genes involved on the pathway and folic acid metabolism which is linked to oral clefts requires appropriate environmental causes in combination with genetic factors that culminate in the failure of fusion of facial processes. Objective: The objective is to perform the association of differential gene expression of folate metabolism genes (MTHFR, MTR, RFC1, MTRR) with the occurrence of oral clefts. Methods: We studied 50 subjects with oral clefts and their mothers, and 50 individuals absent of oral clefts and their mothers, totaling 100 individuals referred cases and 100 controls respectively. For gene expression study, total RNA was extracted from peripheral blood cells, then it was quantified and analyzed for purity by absorbance relation and integrity in MOPS gel. The mRNA samples with good purity and integrity were transcribed to cDNA using reverse transcription kit. The cDNA was used in pre-designed gene expression assays. In a previous study performed by our group were evaluated by PCR-RFLP the MTRR A66G, MTHFR C677T, MTHFR A1298G, MTR A2756G, RFC1 A80G polymorphisms, in this study we evaluated the influence of these polymorphisms on gene expression. It was also performed biochemical, hematological analyzes and a clinical characteristics study of these individuals. We performed statistical analysis considering the significance level of 95% (P <0.050) Results: The consumption of Alcohol was reported as a significant risk factor for the present case mothers (p = 0.001). Regarding the biochemical there were no significant differences between children cases group and their controls which had values of AST, ALT and creatinine within the reference values. The folic acid dosage presented significantly reduced in case mothers group (P = 0.011). In haematological analysis was not observed significant changes in any of the evaluated parameters such as hemoglobin, hematocrit, erythrocyte count, and hematological indices, MCV, MCH and MCHC among the groups. The assessment of gene expression for case mother group showed a significant reduction in mRNA expression in all evaluated genes; for the methionine synthase gene (MTR, p = 0.008), the methionine synthase reductase (MTRR, p = 0.015), the reduced folate carrier 1 (RFC1, P= 0.004) and methylenetetrahydrofolate reductase (MTHFR, P= 0.017) compared with the control group mothers. In the case children group, similar results were obtained, with a significant reduction in mRNA expression of methionine synthase gene (MTR, p = 0.010) and methionine synthase reductase (MTRR, p = 0.034) analysis of genotypes in relation to expression was found that the recessive genotype (CC) for the MTHFR gene A1298C polymorphism is associated with reduced expression of its mRNA. Conclusion: Folate metabolism related genes are low expressed on both case groups of this study, since all four genes (RFC1,MTHFR, MTR, MTR,) were reduced on mothers and two genes (MTR, MTRR) in their children. These down regulated genes represent an increased risk associated with the presence of oral clefts in these individuals.

Made available in DSpace on 2014-12-17T15:32:32Z (GMT). No. of bitstreams: 1 JoabeSP_TESE.pdf: 3616540 bytes, checksum: 2904dbfc0f83145c9569635cf5feba21 (MD5) Previous issue date: 2013-09-16 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / Oral squamous cell carcinoma (OSCC) is an important cause of morbidity and mortality worldwide despite recent advances in treatment. There are several studies aiming to find markers that may improve the assessment of this disease prognosis. Studies about genetic polymorphisms have gained prominence due to their influence on individual susceptibility to cancer development. The aim of this study was to evaluate the association between the frequency of polymorphisms XPD Lys751Gln and XRCC3 Thr241Met and clinicopathological features of OSCC cases, including age, sex, presence or absence of metastases, and histological grading of malignancy according to Bryne (1998). Sample consisted of 54 cases of OSCC and 40 cases of inflammatory fibrous hyperplasia (IFH). OSCC cases were classified as low or high grade. DNA samples were previously extracted from paraffin blocks. Genotypes for each case were determined through PCR-RFLP (polymerase chain reaction - restriction fragment length polymorphism). Results were analyzed by Fisher s exact test and Chi-square test and the odds ratio was calculated considering p < 0.05 to indicate statistical significance. For XPD, Lys/Gln genotype was more common in IFHs (n=28; 70%) than in OSCCs (n=24; 44.4%) (OR: 0.3; p<0.05). Frequency of Gln allele was higher in high-grade lesions when compared to low grade lesions (0.48 and 0.21, respectively) (OR: 3.4; p<0.05). For XRCC3, Met allele was more common in OSCC than in IFH (0.49 and 0.35, respectively) (OR: 2.6; p<0.05). Met/Met genotype was associated with presence of metastases (OR: 8.1; p<0.05). There was no statistically significant association between the genotypes and the age or sex of patients. In the present sample, the higher frequency of XPD Gln allele in IFH reveals a possible protective role of this variant against the development of OSCC. However, its association with high-grade lesions indicates that this allele could influence the tumor progression after the neoplasia development. The presence of XRCC3 Met allele, in turn, seems to contribute to the development of OSCC and metastases / O carcinoma oral de c?lulas escamosas (COCE) ? importante causa de morbidade e mortalidade em todo o mundo a despeito dos recentes avan?os nas formas de tratamento. Diante disto, v?rias s?o as pesquisas no intuito de se encontrar marcadores que possam melhorar o progn?stico desta doen?a. Neste sentido t?m se destacado os estudos dos polimorfismos gen?ticos, os quais podem influenciar a suscetibilidade individual para o desenvolvimento do c?ncer. O objetivo deste estudo foi avaliar a associa??o entre a frequ?ncia dos polimorfismos XPD Lys751Gln e XRCC3 Thr241Met e o perfil clinicopatol?gico em casos de COCE, incluindo idade, sexo, presen?a ou n?o de met?stase e grada??o histol?gica de malignidade de Bryne (1998). A amostra foi composta por 54 casos de COCE e 40 casos de hiperplasia fibrosa inflamat?ria (HFI). Os casos de COCE foram classificados como les?es de baixo ou de alto grau de malignidade. Foram utilizadas amostras de DNA previamente extra?do de blocos de parafina. Os gen?tipos para cada caso foram determinados atrav?s da t?cnica de PCR-RFLP (rea??o em cadeia da polimerase - polimorfismos de comprimento de fragmentos de restri??o). Os resultados foram submetidos aos testes estat?sticos Exato de Fisher e Qui-quadrado de Pearson e foi calculada a raz?o de chance (odds ratio) considerando o n?vel de signific?ncia quando p<0,05. Para o XPD, o gen?tipo Lys/Gln foi mais comum nas HFIs (n=28; 70%) que nos COCEs (n=24; 44,4%) (OR: 0,3; p<0,05). A frequ?ncia do alelo Gln foi maior nas les?es de alto grau, em compara??o ?s de baixo grau (0,48 e 0,21, respectivamente) (OR: 3,4; p<0,05). Para o XRCC3, o alelo Met foi mais frequente no COCE que na HFI (0,49 e 0,35, respectivamente) (OR: 2,6; p<0,05). O gen?tipo Met/Met foi associado ? presen?a de met?stases (OR: 8,1; p<0,05). N?o houve associa??o estat?stica significativa entre os gen?tipos e a idade ou sexo dos pacientes. Na amostra analisada, a maior frequ?ncia do alelo XPD Gln na HIF revela um poss?vel papel protetor dessa variante contra o desenvolvimento do COCE. Todavia, sua associa??o com les?es de alto grau, indica que esse alelo poderia influenciar no processo de progress?o ap?s o tumor instalado. A presen?a do alelo XRCC3 Met, por sua vez, parece contribuir com o desenvolvimento do COCE e de met?stases nessas les?es