Made available in DSpace on 2014-12-17T15:32:21Z (GMT). No. of bitstreams: 1 EdilmarMS_DISSERT.pdf: 790528 bytes, checksum: 570c185c018d55b199d467de6ca18465 (MD5) Previous issue date: 2012-02-09 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / The most common malignant neoplasm of the oral cavity and oropharynx are squamous cell carcinoma. Injuries to the same stage and subjected to the same treatment protocol have sometimes different evolutionary courses. The scope of this study was to investigate, through a retrospective cohort, associations between the number of CD8 + T cells and natural killer, identified immunohistochemically in the inflammatory infiltrate in a series of cases of oral squamous cell carcinoma and orofaringeano, and the level of tumor response to radiotherapy and chemotherapy, overall survival and relapse-free survival of patients. We identified 54 patients with unresectable disease were treated exclusively with radiotherapy and chemotherapy. The median follow-up was 22 months. The sample was characterized by the predominance of male subjects, median age 60 years, all were smokers. The most frequent site was the tongue and 81.5% were in stage IV. Patients with disease in the oral cavity had a worse response to treatment (p = 0.006), worse relapse-free survival (p = 0.007), worse overall survival (p = 0.007). The advanced T stage was shown a negative prognostic factor (p= 0.006) for the clinical treatment response made. Immunohistochemistry was performed to select CD8 + cells (anti-CD8) and NK cells (anti-CD57). Lymphocytes positive and negative markings were counted using the program ImageJ ?. Two groups were created for each marking evaluated: Group I patients with more than 50% cells positive, Group II: less than 50% of labeled cells. For CD8 + cells detected in 38 (70.3%) of Group I were CD8 + and 16 (29.7%) Group II CD8 +. For NK cells, 26 (48.15%) Group I NK and 28 (51.85%) Group II NK. Regarding the clinical response to treatment, we observed that 39% of patients achieved a complete response and 25.9% remained without recurrence at the end of follow-up. These results were better in Group I CD8 + (p = 0.2). Identified that 72.2% of patients progressed to death, this finding had no association with the immunohistochemical data. There was no statistically significant differences between the number of CD8 + and NK cells and the ability of tumor response to radiotherapy and chemotherapy, or with overall survival and relapse-free survival of patients. However, especially in relation to a learned response, we found that this group of patients with advanced disease have a low count of CD8 + T cells active. Believing in the role that the immune response plays in the local fight against neoplastic cells, however, our results do not support the use of quantitative analysis of CD8 + T cells and NK cells as a prognostic factors for oral squamous cell carcinoma and oropharynx / A neoplasia maligna mais frequente da cavidade oral e da orofaringe ? o carcinoma epiderm?ide. Les?es com o mesmo estadiamento e submetidas ao mesmo protocolo terap?utico apresentam, por vezes, cursos evolutivos diferentes. O escopo do presente trabalho foi investigar, atrav?s de um coorte retrospectivo, associa??es entre a quantidade de c?lulas TCD8+ e natural killer, identificadas imuno-histoquimicamente no infiltrado inflamat?rio de uma s?rie de casos de carcinoma epiderm?ide oral e orofaringeano, e o n?vel de resposta tumoral ao tratamento radioter?pico e quimioter?pico, a sobrevida global e sobrevida livre de recidiva dos pacientes. Foram identificados 54 pacientes com doen?a irressec?vel, tratados exclusivamente com radioterapia e quimioterapia. A mediana de seguimento foi de 22 meses. A amostra se caracterizou pelo predom?nio de indiv?duos masculinos, com idade mediana de 60 anos; todos eram tabagistas. O s?tio mais frequente foi a l?ngua oral e 81,5% encontravam-se no est?dio IV. Os pacientes com doen?a na cavidade oral tiveram uma pior resposta ao tratamento (p=0,006), pior sobrevida livre de recidiva (p=0,007), pior sobrevida global (p=0,007). O est?dio T avan?ado se demonstrou um fator progn?stico negativo (p=0,006) para a resposta ao tratamento cl?nico efetuado. Foi realizada imuno-histoqu?mica para marcar c?lulas CD8+ (anti-CD8) e c?lulas NK (anti-CD57). Os linf?citos positivos e negativos para as marca??es foram contados atrav?s do programa ImageJ?. Dois grupos foram criados para cada marca??o avaliada: Grupo I: pacientes com mais de 50% das c?lulas positivas; Grupo II: menos de 50% das c?lulas marcadas. Para as c?lulas CD8+ detectamos que 38 (70,3%) eram do Grupo I CD8+ e 16 (29,7%) do Grupo II CD8+. Para as c?lulas NK, 26 (48,15%) Grupo I NK e 28 (51,85%) Grupo II NK. Em rela??o ? resposta cl?nica ao tratamento, observamos que 39% dos pacientes obtiveram resposta completa e 25,9% permaneceram sem recidiva ao final do seguimento. Esses resultados foram melhores no Grupo I CD8+ (p=0,2). Identificamos que 72,2% dos pacientes evolu?ram para o ?bito, esse achado n?o teve associa??o com os dados imuno-histoqu?micos. N?o se observou diferen?as estatisticamente significantes entre a quantidade de c?lulas CD8+ e NK e a capacidade de resposta tumoral ao tratamento radioter?pico e quimioter?pico, nem com a sobrevida global e sobrevida livre de recidiva dos pacientes. Contudo, principalmente em rela??o a resposta adquirida, detectamos que este grupo de pacientes com doen?a avan?ada tem uma baixa contagem de c?lulas TCD8+ ativas. Acreditando no papel fundamental que a resposta imune exerce no combate local ?s c?lulas neopl?sicas; no entanto, nossos resultados n?o suportam a utiliza??o da an?lise quantitativa das c?lulas TCD8+ e NK como um dos fatores progn?sticos para o carcinoma epiderm?ide oral e de orofaringe

Made available in DSpace on 2014-12-17T15:32:22Z (GMT). No. of bitstreams: 1 KaruzaMAP.pdf: 544780 bytes, checksum: e0fadeed7d2d1ec7568970935306d05c (MD5) Previous issue date: 2006-02-22 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / The Human Papillomavirus (HPV) has been strongly implicated on development of some cases of oral squamous cell carcinoma (OSCC). However, the immunological system somehow reacts against the presence of this virus. Among the cells involved on such mechanism of defense detaches the Langerhans cells (LC), which are responsible for processing and presenting antigens. The purpose of this study was to evaluate the immunohistochemical reactivity for Langerhans cells between HPV positive and HPV negative OSCC, as well as, the relation of the immunoreactivity for this cells and the histological grading of malignancy proposed by Bryne (1998) and modified by Miranda (2002). Additionally, HPV infection was evaluated in relation to sex, age, lesion localization and histological grading of malignancy. In the total, 27 cases of OSSC were evaluated, 09 of them HPV positive and 18 HPV negative. Anti S-100 antibody was utilized for the immunohistochemical labelling, followed by the counting of LCs in 5 highpower fields (400x). No statistically significant difference was verified between the variables sex, age, lesion localization, histological grading of malignancy and HPV presence in OSSC. There was neither association between the immunohistochemical labeling for LCs (S-100+) and HPV infection nor correlation between the quantity of LCs labeled and the histological grading of malignancy of OSSC. The results suggest that despite the absence of statistically significant difference, the presence of HPV in such cases of OSCC can alter the immunological system, particularly the Langerhans cells / O Papilomav?rus Humano (HPV) tem sido implicado fortemente no desenvolvimento de alguns carcinomas epiderm?ides orais (CEOs). Contudo, o sistema imunol?gico reage de alguma forma ? presen?a desse v?rus. Dentre as c?lulas envolvidas nesse mecanismo de defesa, destaca-se a c?lula de Langerhans (CL), por serem c?lulas processadoras e apresentadoras de ant?genos. O objetivo desse estudo foi avaliar a marca??o imuno-histoqu?mica das c?lulas de Langerhans entre os casos de CEOs HPV positivos e negativos, bem como a rela??o da imunomarca??o dessas c?lulas e a grada??o histol?gica de malignidade proposta por Bryne (1998) e modificada por Miranda (2002). Adicionalmente, a infec??o pelo HPV foi estudada com rela??o ao sexo, idade, localiza??o da les?o e a grada??o histol?gica de malignidade. Foram analisados 27 casos de CEOs, sendo 09 destes HPV positivos e 18 casos negativos. Para a marca??o imuno-histoqu?mica utilizou-se o anticorpo anti S-100, sendo as CLs quantificadas em 5 campos de maior aumento (400x). A an?lise estat?stica revelou n?o existir rela??o das vari?veis, sexo, idade, localiza??o da les?o e grada??o histol?gica, com a presen?a do HPV nos CEOs estudados. N?o existiu associa??o entre a marca??o imuno-histoqu?mica das CLs(S-100+) e a infec??o pelo HPV, e tamb?m n?o houve correla??o entre as CLs imunomarcadas e a grada??o histol?gica nos casos de CEOs analisados. Diante desses resultados, pode-se sugerir que mesmo n?o havendo diferen?a significativa, a presen?a do HPV nos casos de carcinoma epiderm?ide oral pode alterar o sistema imune, particularmente as c?lulas de Langerhans

Made available in DSpace on 2014-12-17T15:32:24Z (GMT). No. of bitstreams: 1 JoaoAVGF.pdf: 1413564 bytes, checksum: f5d8ea2da8907cd551169a4091430007 (MD5) Previous issue date: 2006-02-17 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / Squamous cell carcinoma is the most common malignant neoplasm in the oral cavity, accounting for more than 90% of all malignancies in this location. Cyclooxygenases (COX s) are key enzymes on arachidonic acid metabolism and prostaglandin synthesis, being expressed basically in two forms: the constitutive (COX-1) and the inducible (COX-2). Increased levels on the expression of COX-2 have been implicated in the pathogenesis tumor progression of various forms of human cancer, including oral squamous cell carcinoma, some of what suggesting a possible interaction between COX-2 and the protein expressed by the tumor suppressor gene p53, mutated in more than 50% of all human cancers. The mean of the present research consisted in analyze the correlation between the expression of COX-2 and p53, at the protein level, as well as evaluate the difference on the expression of these two proteins with the histological grading of malignancy. 34 cases of oral squamous cell carcinoma were selected and graded according to the histological grading system proposed by Bryne (1998) and the labeling indexes (LI s) for COX-2 and p53 evaluated using immunohistochemistry method. The results revealed that COX-2 was expressed in increased levels in most of the specimens, although there was no statistic significant correlation between LI s from COX-2 and p53 (p>0.05), and there were no statistical differences on the expression of these proteins between tumors of high and low grade of malignancy (p>0.05). Interestingly, the expression of COX-2 and p53 was detected in fragments of dysplastic oral epithelium adjacent to tumor areas, on basal and suprabasal layers. The absence of statistical correlation between the expression of COX-2 and p53 proteins do not rule ot the existence of a relation between them, were it may reflect the diversity of regulatory pathways between both, different direct and indirect inhibitory effects of COX-2 over p53, as well as the wide range of activation macheenisms for COX-2 and mutational status of the p53 gene Another conclusion point that the increased expression of COX-2 observed in oral squamous cell carcinomas suggest a role for this protein in the processes of pathogenesis and tumoral evolution of this malignant neoplasm / O carcinoma epiderm?ide ? a neoplasia maligna mais comum na cavidade oral, representando mais de 90% das malignidades nesta localiza??o. As cicloxigenases (COX s) s?o enzimas chave no metabolismo do ?cido aracd?nico e s?ntese de prostaglandinas, sendo expressas basicamente sob duas formas: uma constitutiva (COX-1) e uma induzida (COX-2). N?veis elevados na express?o da COX-2 t?m sido implicados na patog?nese e progress?o tumoral em diversos tipos de c?ncer em humanos, incluindo o carcinoma epiderm?ide oral, alguns dos quais sugerindo uma poss?vel intera??o entre a COX-2 e a prote?na expressa pelo gene supressor tumoral p53, mutado em mais de 50% de todos c?nceres humanos. O prop?sito da presente pesquisa consistiu em analisar a correla??o entre a express?o de COX-2 e p53, em n?vel de prote?na, bem como avaliar a diferen?a na express?o destas duas prote?nas em rela??o ao grau histol?gico de malignidade. Para tal, foram selecionados 34 casos de carcinoma epiderm?ide oral, os quais foram classificados de acordo com o sistema de grada??o histol?gica de malignidade proposto por Bryne (1998) e cujos ?ndices de positividade para COX-2 e p53 foram avaliados atrav?s da t?cnica imuno-histoqu?mica. O resultados revelaram que a COX-2 esteve expressa em n?veis elevados na maior parte dos esp?cimes analisados, embora n?o se tenha verificado correla??o estatisticamente significativa entre os IP s da COX-2 e da p53 (p>0,05), tampouco diferen?a estatisticamente significativa entre a express?o destas prote?nas entre tumores de alto e baixo grau de malignidade (p>0,05). Interessantemente, foi detectada a express?o da COX-2 e da p53 em fragmentos de epit?lio oral displ?sico, nas camadas basal e parabasal, adjacentes ao tumor. A aus?ncia de correla??o estat?stica entre a express?o das prote?nas COX-2 e p53 n?o descarta a exist?ncia de uma rela??o entre as mesmas, podendo refletir a diversidade de vias regulat?rias entre ambas, os diferentes efeitos inibit?rios diretos e indiretos da COX-2 sobre a p53, bem como os in?meros mecanismos de ativa??o da COX-2 e o estado mutacional do gene p53. Conclui-se ainda que a elevada express?o da COX-2 observada em carcinomas epiderm?ides orais sugere um papel desta prote?na dentro dos processos de patog?nese e evolu??o tumoral desta neoplasia maligna

Made available in DSpace on 2014-12-17T15:32:25Z (GMT). No. of bitstreams: 1 AndreaFS_tese.pdf: 610478 bytes, checksum: 22934e3fba7a401686d3b0057e0e7f35 (MD5) Previous issue date: 2007-07-11 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / The expression of bone morphogenetic proteins (BMPs) is altered in a variety of human canceres. The BMP-2/4 and BMPR-IA were recently shown to be overexpressed in high-risk premalignant and malignant lesions of oral epithelium. The present study analysed the expression of BMP-2/4 and BMPR-IA in Oral Squamous Cell Carcinoma (OSCC) such as their implications in disease prognostic using munohistochemistry. Ten cases of Oral Fibroepithelial Hiperplasia were selected as a control group. The experimental group included 16 cases of OSCC without metastases and 7 cases of OSCC metastatic. The presence or absence of nodal metastases was used as parameter to evaluated the disease prognostic. The results demonstrated weak immunoreactivity for BMP-2/4 and BMPR-IA in every case of the control group. In the cases of OSCC with metastases an overexpression of BMP-2/4 (71,4%) was observed while the BMPR-IA showed weak expression (85,7%). In the cases of OSCC without metastases BMP-2/4 (62,5%) and BMPR-IA showed strong immunostaining standing out an overexpression of the receptor in all the specimens. Observed statistical significance for correlation between the oral cancer prognostic and the staining intensity of the BMP-2/4 (p=0,002). There wasn t statistical significance for association between the staining intensity of the BMPR-IA and the disease prognostic (p<0,001). In conclusion, this findings suggest that the overexpression of BMP-2/4 associated with the loss of expression of the BMPR-IA in OSCC metastatic has prognostic relevance, as the loss of sensitivity to BMPs can be an indicative of metastases development in OSCC / A express?o das prote?nas morfogen?ticas ?sseas (BMPs) est? alterada em v?rios c?nceres humanos. A BMP-2/4 e o BMPR-IA foram recentemente encontrados superexpressos em les?es malignas e pr?-malignas de alto risco em epit?lio oral. Este estudo analisou a express?o da BMP-2/4 e seu receptor BMPR-IA em 23 esp?cimes de Carcinoma Epiderm?ide Oral (CEO), utilizando a imuno-histoqu?mica. O grupo controle constou de 10 casos de Hiperplasia Fibro-epitelial da mucosa oral. O grupo experimental foi constitu?do por 16 casos de CEO n?o metast?tico e 7 casos de CEO metast?tico. Utilizou-se o par?metro presen?a ou aus?ncia de met?stase nodal para avaliar o progn?stico da doen?a. Os resultados demonstraram imunorreatividade fraca para a BMP-2/4 e o BMPR-IA em todos os esp?cimes do grupo controle. No grupo experimental com met?stase, a BMP-2/4 exibiu forte expressividade (71,4%), enquanto que o BMPR-IA mostrou fraca express?o (85,7%). No grupo experimental sem met?stase, evidenciou-se forte express?o para a BMP-2/4 (62,5%) e para o BMPR-IA (100%). Encontrou-se signific?ncia estat?stica para a associa??o entre o progn?stico do CEO e a intensidade de marca??o da BMP-2/4 (p=0,002). Para o BMPR-IA n?o houve signific?ncia estat?stica ? sua associa??o com o progn?stico da doen?a (p<0,001), em fun??o do tamanho da amostra. Portanto, os resultados sugerem que a fraca expressividade do BMPR-IA associada ? forte express?o da BMP-2/4, no grupo experimental com met?stase, tem relev?ncia progn?stica, j? que a perda de sensibilidade ?s BMPs, atrav?s da perda de express?o de seus receptores pode ser indicativo de desenvolvimento de met?stase em CEO

Made available in DSpace on 2014-12-17T15:32:28Z (GMT). No. of bitstreams: 1 MartaRP.pdf: 1904281 bytes, checksum: 2912c6b8ea9a773c553d0776245f93a5 (MD5) Previous issue date: 2009-02-27 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / The Oral Epithelial Dysplasia (OED) is the lesion that precedes or co-exists with the Oral Squamous Cell Carcinoma (OSCC), presenting molecular and/or histological similar alterations. The divergences about the malignization potential of OEDs and the role of inflammation in this process make hard the early diagnosis and evaluation of OSCCs aggressiveness. Thus, it became the goal of this study to evaluate the role of inflammation in oral carcinogenesis and tumoral aggressiveness. For this purpose a morphological study was performed in 20 OED cases and 40 OSCC cases to detect the malignization potential of OEDs and the histologic malignancy grading (HMG) of OSCCs, analyzing superficial masses for dismorphism evaluation and the invasive front for evaluation of tumoral growing; and immunohistochemical, using anti-CD8, anti-FOXP3, anti-TGF?, anti-TNF? and anti-NF-?B antibodies, comparing their with the types lesion, histological degree and intensity of the inflammatory infiltrate. The results were statistically significant for the parameters: cell maturity (p=0,0001), masses presence (p=0,038) and dismorphism (p=0,037), when associated to HMG. To compare the expression of the markers with the types lesion, a significantly higher expression of CD8 (p=0,001) and NF-?B (p=0,002) in the OED, and also a smaller expression of the epithelial TGF? in the severe OEDs (p=0,011), without significant expression between OSCC degrees. By relating the expression of the studied markers with the inflammatory infiltrate intensity, a positive relation was observed with: inflammatory TNF?(p=0,003), epithelial TNF? and NF-?B (p=0,051 and p=0,004), in OEDs; and with CD8 (p=0,021) and TNF? (p=0,015) in conjunctive OSCCs; and a negative relation with epithelial TNF? (p=0,034) in OSCCs. No significant relation was found between FOXP3 with any of the studied variables. These findings lead to the conclusion that, the study of the invasive front is as important as the study of superficial masses for the evaluation of tumoral aggressiveness; the intensity of the inflammatory infiltrate has no use as a parameter for prognostic evaluation of OSCC in routine exams, but, the molecular events detected in this study may be necessary to give basis for determining the malignant potential in OEDs and aggressiveness in OSCCs / A Displasia Epitelial Oral (DEO) ? a les?o que precede ou co-existe com o Carcinoma Epiderm?ide Oral (CEO), apresentando altera??es moleculares e/ou histol?gicas semelhantes. As diverg?ncias sobre o potencial de maligniza??o das DEO e o papel da inflama??o nestes processos t?m dificultado o diagn?stico precoce e a avalia??o da agressividade dos CEO. Sendo assim, tornou-se objetivo deste estudo avaliar o papel da inflama??o na carcinog?nese oral e agressividade tumoral. Para isso foi realizado estudo morfol?gico em 20 casos de DEO e 40 casos de CEO para detectar o potencial de maligniza??o das DEO e o Grau Histol?gico de Malignidade (GHM) dos CEO, analisando as massas superficiais para avalia??o do dismorfismo e o front invasivo para avalia??o do crescimento tumoral; e imuno-histoqu?mico, utilizando os anticorpos anti-CD8, anti-FOXP3, anti-TGF?, anti-TNF-? e anti-NF-?B, para comparar a express?o dos mesmos com o tipo de les?o, grau histol?gico e intensidade do infiltrado inflamat?rio. Os resultados foram estatisticamente significantes para os par?metros, maturidade celular (p=0,0001), presen?a de massas (p=0,038) e dismorfismo (p=0,037), quando associados aos GHM. Ao comparar a express?o dos marcadores com o tipo de les?o, encontrou-se uma express?o significativamente maior do CD8 (p=0,001) e do NF-?B (p=0,002) nas DEO, assim como uma menor express?o do TGF? epitelial nas DEO severas (p=0,011), n?o tendo express?o significativa entre os graus dos CEO. Ao relacionar a express?o dos marcadores estudados com a intensidade do infiltrado inflamat?rio, observou-se uma rela??o positiva com o TNF? inflamat?rio (p=0,003), o TNF? e o NF-?B epiteliais (p=0,051 e p=0,004), nas DEO; com o CD8 (p=0,021) e o TNF? (p=0,015) no conjuntivo dos CEO; e uma rela??o negativa com o TNF? (p=0,034) epitelial dos CEO. N?o foi encontrada rela??o significativa da FOXP3 com nenhuma das vari?veis estudadas. Esses achados levaram a concluir que, o estudo do front invasivo ? t?o importante quanto o estudo das massas superficiais para avalia??o da agressividade tumoral; a intensidade do infiltrado inflamat?rio n?o pode ser utilizado como par?metro para avalia??o progn?stica do CEO no exame de rotina; mas os eventos moleculares detectados neste estudo podem ser necess?rios para embasar a determina??o do potencial de malignidade nas DEO e da agressividade nos CEO

Made available in DSpace on 2014-12-17T15:32:28Z (GMT). No. of bitstreams: 1 Tese_GeorgeJFN.pdf: 2720479 bytes, checksum: 6d72db8b12a0e5f743d69a67430d66b7 (MD5) Previous issue date: 2010-02-18 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / Matrix metalloproteinase-7 (MMP-7) and -9 (MMP-9) modulate important functions strictly related to the development, invasion and metastasis of several human cancers among them the squamous cell carcinoma of the tongue (SCCT). However, individual genetic factors such as the functional single nucleotide polymorphisms (SNPs) influence the pattern of protein expression of these MMPs and thus may be related to the variability observed in the clinical behavior of patients with SCCT. In this context, the present cross-sectional study aimed to evaluate the association between the frequency of the functional SNPs MMP-7 -181 A/G and MMP-9 -1562 C/T and the clinical (age, gender and metastasis) and pathological (malignancy histological grading and immunohistochemistry expression) features of SCCT cases. Genotyping of these SNPs were performed by PCR-RFLP on DNA samples from 71 cases of SCCT and 60 individuals without cancer who constitute the control group. Among the results of this research, it was observed that the frequency of the polymorphic alleles MMP-7 -181 G and MMP-9 -1562 T in SCCT patients was 28% and 12%, respectively, and the frequency of the heterozygotes A/G (PR = 2.00; p < 0.001) and C/T (PR = 1.54; p = 0.014) were significantly higher in the patient group than in the controls. The prevalence of patients carrying the combination of SNPs studied was significantly associated with SCCT cases (PR = 2.00; p = 0.011) and metastasis (PR = 2.00; p < 0.001). Furthermore, with the frequency of SNPs analyzed, the age, gender, histological grading and immunoreactivity of MMP-7 and MMP-9 formed clinical and pathological parameters relevant to the identification of population subgroups more related to the development of SCCT and metastasis. Based on these results, it is suggested that the protein expression levels of MMP-7 and -9 substantially influence the balance between their pro- and anticancer biological functions and hence the clinicopathological profile of the squamous cell carcinoma of the tongue / As metaloproteinases da matriz extracelular-7 (MMP-7) e -9 (MMP-9) modulam importantes fun??es relacionadas ao desenvolvimento, invas?o e met?stase de diversos c?nceres humanos, dentre os quais o carcinoma epiderm?ide de l?ngua (CEL). Entretanto, fatores gen?ticos individuais, tais como polimorfismos de nucleot?deo ?nico (SNPs) funcionais, influenciam no padr?o de express?o proteica dessas MMPs, podendo estar relacionados ? variabilidade no comportamento cl?nico tumoral observado em pacientes com CEL. Neste contexto, o presente trabalho objetivou, atrav?s de an?lise em sec??o transversal, estudar a associa??o entre a frequ?ncia dos SNPs funcionais MMP-7 -181 A/G e MMP-9 -1562 C/T e as caracter?sticas cl?nicas (idade, sexo e met?stase) e patol?gicas (grada??o histol?gica e express?o imuno-histoqu?mica) em uma s?rie de casos de CEL. A genotipagem dos referidos SNPs foi executada por PCR-RFLP em amostras de DNA de 71 casos de CEL e de 60 indiv?duos sem c?ncer, que constitu?ram o grupo controle. Dentre os resultados da presente pesquisa, evidenciou-se que a frequ?ncia dos alelos polim?rficos MMP-7 -181 G e MMP-9 -1562 T nos pacientes com CEL foi de 28% e 12%, respectivamente, sendo as frequ?ncias dos heterozigotos A/G (RP = 2.00; p < 0.001) e C/T (RP = 1.54; p = 0.014) significativamente maiores neste grupo de pacientes que no grupo controle. A preval?ncia dos pacientes portadores da combina??o dos SNPs estudados associou-se significativamente aos casos de CEL (RP = 2.00; p = 0.011) e ? met?stase (RP = 2.00; p < 0.001). Ademais, junto ? frequ?ncia dos SNPs analisados, a idade, sexo, grada??o histol?gica e imunoexpress?o da MMP-7 e -9 constitu?ram par?metros clinicopatol?gicos relevantes para a identifica??o de subgrupos populacionais mais predispostos ao desenvolvimento do CEL e met?stase. Frente a estes resultados, sugere-se que os n?veis de express?o da MMP-7 e -9 influenciam consideravelmente no balan?o entre suas fun??es pr? e antineopl?sicas e, consequentemente, no perfil clinicopatol?gico do carcinoma epiderm?ide de l?ngua.

Made available in DSpace on 2014-12-17T15:32:31Z (GMT). No. of bitstreams: 1 AnaRLAM_TESE.pdf: 3774507 bytes, checksum: 2b9028dcadafe1a4c66e01ba9daab791 (MD5) Previous issue date: 2012-07-19 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / Squamous cell carcinoma of the lower lip is among the most common malignant tumors of the oral and maxillofacial region, with good prognosis in more than 90% of patients with 5-year survival. In these carcinomas, the development of lymph node metastasis decreases the prognosis and it has been associated with the formation of new lymphatic vessels. It has been suggested the important role of vascular endothelial growth factor-C (VEGF-C), the receptor type 3 VEGF (VEGFR-3) and hypoxia-induced factor 1 (HIF-1) in this process. The aim of this study was to evaluate the immunoexpression of VEGF-C, VEGFR-3 and HIF-1? and correlate with intra and peritumoral lymphatic density in squamous cell carcinomas of the lower lip metastatic and non-metastatic. The sample consisted of 50 cases of squamous cell carcinoma of lower lip, of which 25 had regional lymph node metastasis and 25, absence of metastasis. The percentages of cells immunostained for VEGF-C, VEGFR-3 and HIF-1? in front of tumor invasion and in the center of tumor were evaluated. Microvessel density lymphatic (MDL) was determined by the counting of lymph microvessels immunostained by the anti-D2-40 in five fields (200?), in an area of evaluation with 0.7386 mm2. The invasion of the lymph vessels by malignant cells was also evaluated. Immunostaining was correlated with the presence and absence of metastasis, TNM clinical stage, local recurrence, disease outcome (remission of injury or patient death) and histological grading. The analysis of intra and peritumoral lymphatic density showed no significant association with clinicopathological parameters and immunoexpressions of VEGF-C, VEGFR-3 and HIF-1? (p > 0,05). There was a weak positive correlation, significant, between intra and peritumoral lymphatic density (r = 0,405; p = 0,004). VEGF-C showed no significant association with clinicopathological and prognosis parameters (p > 0,05). For VEGFR-3, there was scarce membrane staining and intense and homogenous cytoplasmic staining in neoplastic cells. Percentage of positive cytoplasmic VEGFR-3 in center of tumor, exhibited a statistically significant association with metastasis (p = 0,009), patient death (p = 0,008) and histological grades of malignancy proposed by Bryne et al. (1992) (p = 0,002) and World Health Organization (p = 0,003). A low positive correlation was statistically significant between the immunoreactivity of VEGFC and VEGFR-3 cytoplasmic (r = 0,358; p = 0,011) and between the percentage of positive cytoplasmic VEGFR-3 in front of tumor invasion and in the center of the tumor (r = 0,387; p = 0,005) was also demonstrated. There was no association between HIF-1?, clinicopathological and prognosis parameters, and VEGF-C and VEGFR-3. The percentage of nuclear positivity for HIF-1? was significantly higher in cases without invasion of peritumoral lymphatic (p = 0,040). Based on the results we can conclude that most cytoplasmic expression of VEGFR-3 in center of tumor in metastatic cases, high degree of malignancy and poorly differentiated, contributes to poor outcome of squamous cell carcinoma of the lower lip, including patient death. Intra and peritumoral lymphatic density seems to be not associated with lymph node metastasis in these carcinomas / O carcinoma epiderm?ide de l?bio inferior est? entre as les?es malignas mais comuns da regi?o oral e maxilofacial, com progn?stico bom, em mais de 90% dos pacientes com sobrevida de 5 anos. Nestas les?es, o desenvolvimento de met?stase linfonodal diminui sobremaneira o progn?stico e tem sido associado ? forma??o de novos vasos linf?ticos. Tem sido sugerido o importante papel do fator de crescimento endotelial vascular-C (VEGF-C), do receptor tipo 3 do VEGF (VEGFR-3) e do fator 1 induzido por hip?xia (HIF-1) neste processo. O objetivo desta pesquisa foi avaliar as imunoexpress?es de VEGF-C, VEGFR-3, HIF-1? e a densidade linf?tica intra e peritumoral em carcinomas epiderm?ides de l?bio inferior metast?ticos e n?o-metast?ticos, correlacionando-as com par?metros clinicopatol?gicos e progn?sticos. A amostra foi constitu?da por 50 casos de carcinoma epiderm?ide de l?bio inferior, 25 com met?stase linfonodal regional e 25 sem met?stase. Foram avaliados os percentuais de c?lulas imunomarcadas para os anticorpos anti-VEGF-C, anti-VEGFR-3 e anti-HIF-1?, no front de invas?o e no centro tumoral. A densidade microvascular linf?tica (LMVD) foi estabelecida por meio da soma da contagem de microvasos linf?ticos imunomarcados pelo anticorpo anti-D2-40, em cinco campos (200?), em uma ?rea de avalia??o com 0,7386 mm2. A invas?o dos vasos linf?ticos por c?lulas neopl?sicas tamb?m foi avaliada. A imunomarca??o foi relacionada com a presen?a e aus?ncia de met?stase, estadiamento cl?nico TNM, recidiva local, desfecho da doen?a (remiss?o da les?o ou ?bito dos pacientes) e grada??o histol?gica. A an?lise das densidades linf?ticas intra e peritumorais n?o demonstrou associa??o significativa com os par?metros clinicopatol?gicos, progn?sticos e imunoexpress?es de VEGF-C, VEGFR-3 e HIF-1? (p > 0,05). Houve fraca correla??o positiva, significativa, entre as densidades linf?ticas intra e peritumorais (r = 0,405; p = 0,004). O VEGF-C n?o exibiu associa??o significativa entre os par?metros clinicopatol?gicos e progn?sticos avaliados (p > 0,05). Para o VEGFR-3, houve escassa marca??o membranar e intensa e homog?nea marca??o citoplasm?tica nas c?lulas neopl?sicas. O percentual de positividade citoplasm?tica do VEGFR-3, no centro tumoral, exibiu associa??o estatisticamente significativa com a presen?a de met?stase (p = 0,009), ?bito dos pacientes (p = 0,008) e grada??es histol?gicas de malignidade proposta por Bryne et al. (1992) (p = 0,002) e pela Organiza??o Mundial de Sa?de (p = 0,003). Uma fraca correla??o, estatisticamente significativa, entre a imunoexpress?o de VEGF-C e VEGFR-3 citoplasm?tica (r = 0,358; p = 0,011) e entre os percentuais de positividade citoplasm?tica de VEGFR-3 no front de invas?o e no centro tumoral (r = 0,387; p = 0,005) tamb?m foi demonstrada. N?o foi observada associa??o entre o HIF-1? os par?metros clinicopatol?gicos, progn?sticos e o VEGF-C e VEGFR-3. O percentual de positividade nuclear para HIF-1? foi significativamente maior nos casos sem invas?o dos linf?ticos peritumorais (p = 0,040). Com base nos resultados pode-se concluir que a maior express?o citoplasm?tica de VEGFR-3, no centro tumoral, nos casos metast?ticos, de alto grau de malignidade e pobremente diferenciados, contribui para pior evolu??o dos carcinomas epiderm?ides de l?bio inferior, incluindo o ?bito dos pacientes. As densidades linf?ticas intra e peritumorais parecem n?o estar associadas ao densenvolvimento de met?stase linfonodal nestes carcinomas

Made available in DSpace on 2014-12-17T15:32:33Z (GMT). No. of bitstreams: 1 CyntiaHPC_TESE.pdf: 1535929 bytes, checksum: 7d7c8298def3233365b9ad5eb617d015 (MD5) Previous issue date: 2014-02-27 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / Oral squamous cell carcinoma (OSCC) is the most prevalent malignancy in the oral cavity and reach a large number of individuals, has become an important public health problem. Studies have demonstrated changes in pathway components BMP in various types of cancers as prostate, colon, breast, gastric and OSCCs. Is the current knowledge that these proteins may exert pro-tumor effect in more advanced stages of neoplastic development coming to favor progression and invasion tumor. The inhibition of the signaling pathway BMP-2 through its antagonists, have shown positive results of antitumor activity and use of Noggin may be a novel therapeutic target for cancer. Given this evidence and the few studies with BMP-2, Noggin and OSCC, the objective of this research was to evaluate the effect of BMP-2 and its antagonist Noggin on proliferation and migration cell in line of cell cultures of human tongue squamous cell carcinoma (SCC25). The study was divided in three groups, a control group, where SCC25 cells suffered no treatment, a BMP-2 group, in which cells were treated with 100ng/ml of BMP-2 and a group of cells that were treated with 100ng/ml of Noggin. For the proliferation assay and cell cycle were established three time intervals (24, 48 and 72 hours). Proliferative activity was investigated by trypan blue and cell cycle analysis by staining with propidium iodide flow cytometry. The potential for migration / invasion of SCC25 cells was performing by a cell invasion assay using Matrigel in a 48-hour interval. The proliferation curve showed a higher proliferation in cells treated with BMP-2 in 72 hours (p < 0.05), and lower overgrowth and cell viability in Noggin group. Recombinant proteins favored a greater percentage of cells in cell cycle phase Go/G1 with a statistically significant difference in the interval of 24 hours (p < 0.05). BMP- 2 produced a greater invasion of cells studied as well as its antagonist Noggin inhibits invasion of cells (p < 0.05). Thus, these results indicate that BMP-2 promotes malignant phenotype, dues stimulates proliferation and invasion of SCC25 cells and, its antagonist Noggin may be an alternative treatment, due to inhibit the tumor progression / O carcinoma epiderm?ide oral (CEO) representa a neoplasia maligna mais prevalente na cavidade oral e por atingir um grande n?mero de indiv?duos, acaba se tornado um relevante problema de sa?de p?blica. Muitos estudos demonstram altera??es nos componentes da via BMP em v?rios tipos de tumores, como os de pr?stata, c?lon, mama, g?stricos e CEOs. ? do conhecimento atual que essas prote?nas podem exercer efeito pr?-tumoral em est?gios mais avan?ados do desenvolvimento neopl?sico vindo a favorecer a progress?o e invas?o tumoral. A inibi??o da via de sinaliza??o da BMP-2, atrav?s dos seus antagonistas, tem mostrado resultados positivos de a??o antitumoral e que assim, o uso do Noggin pode ser um novo alvo terap?utico contra o c?ncer. Diante destas evid?ncias e dos escassos trabalhos com BMP-2, Noggin e CEO, o objetivo desta pesquisa foi avaliar o efeito da BMP-2 e seu antagonista Noggin sobre a prolifera??o e migra??o celulares em culturas de c?lulas de carcinoma epiderm?ide de l?ngua humana (SCC25). Foi feita a divis?o em tr?s grupos de estudo, um grupo controle, onde as c?lulas SCC25 n?o sofriam tratamento com subst?ncia alguma, um grupo BMP-2, no qual as c?lulas eram tratadas com 100ng/ml de BMP-2 e um grupo de c?lulas que eram tratadas com 100ng/ml de Noggin. Para o ensaio de prolifera??o e ciclo celular foram estabelecidos tr?s intervalos de tempo (24, 48 e 72 horas). A atividade proliferativa foi investigada por azul de tripan e a an?lise do ciclo celular atrav?s da marca??o por iodeto de prop?dio em Citometria de fluxo. O potencial de migra??o/invas?o das c?lulas SCC25 foi avaliado atrav?s da realiza??o de um ensaio de invas?o celular utilizando o matrigel em um intervalo de 48 horas. A curva de prolifera??o revelou maior crescimento celular nas c?lulas tratadas com BMP-2 no intervalo de 72 horas (p<0.05) e menor crecimento e viabilidade celular no grupo Noggin. As prote?nas recombinantes favoreceram a maior porcentagem das c?lulas na fase do ciclo celular Go/G1 com diferen?a estatisticamente significativa no intervalo de 24 horas (p<0,05). A BMP-2 promoveu uma maior invas?o das c?lulas estudadas, assim como o seu antagonista Noggin inibiu a invas?o das c?lulas estudadas (p<0,05). Dessa forma, os resultados indicam que a BMP-2 favorece o fen?tipo maligno, pois estimula a prolifera??o e invas?o das c?lulas SCC25 e seu antagonista Noggin pode ser uma alternativa terap?utica pois inibiu essas caracter?sticas pr?-tumorais

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / A proteína óxido nítrico sintase induzível (iNOS) é a mais importante proteína da família das óxido nítrico sintases, a qual é capaz de produzir grandes quantidades de óxido nítrico. A indução permanente de iNOS foi levantada como potencialmente mutagênica, apresentando um papel central na biologia tumoral. O objetivo deste estudo é investigar o papel da iNOS no desenvolvimento de carcinomas de células escamosas bucais (CCEB), usando um modelo experimental de carcinogênese lingual em camundongos knockouts para NOS2 e amostras humanas derivadas da mucosa normal, leucoplasia bucal e CCEB. Camundongos selvagens (iNOS+/+) e knockout (iNOS-/-) para iNOS foram desafiados com 4-nitroquinolina-1-óxido (4NQO) diluída em água potável por 16 semanas e sacrificados após 0, 8 e 16 semanas de acompanhamento. As línguas foram removidas e o número de displasias e carcinomas foram contados. Sangue periférico destes camundongos foram analisados, por citometria de fluxo, em busca de células tumorais circulantes. Amostras humanas derivadas da mucosa normal, leucoplasia e CCEB, foram utilizadas para quantificar o RNA mensageiro (mRNA) de NOS2. As amostras humanas também foram imuno-coradas com anticorpos anti-iNOS e anti-nitrotirosina. Em ambos os grupos, foram observadas uma redução no número de displasias e um aumento de carcinomas da 16 semana até a 32 semanas. Camundongos iNOS-/- demonstraram menor número de lesões displásicas e neoplásicas em todos os períodos de avaliação, sendo menores e menos invasivas, quando comparados aos animais selvagens. Aparentemente, há uma tendência em ascensão nos níveis do mRNA e da proteína iNOS durante a carcinogênese bucal humana. Dados similares foram obtidos com a imuno-detecção de nitrotirosina. Além disso, as imuno-marcações de iNOS e nitrotirosina foram associadas a várias características clínicopatológicas dos CCEB (tamanho, presença de metástase, estadiamento e recidiva). Nossos resultados demonstraram que iNOS afeta o processo de carcinogênese bucal experimental e humana, estando associada com a gênese e progressão dos CCEB. Mais estudos devem ser feitos, para nos fornecer, uma compreensão profunda das funções de iNOS no desenvolvimento de câncer bucal. / Inducible nitric oxide synthase (iNOS) is most important protein of nitric oxide synthases family, which is capable to produce huge amounts of nitric oxide. Permanent induction of iNOS has been raised as potentially mutagenic; presenting pivotal roles in tumor biology. The aim of this study is to investigate the role of iNOS in the development of oral squamous cells carcinomas (OSCC), using a mouse model of oral carcinogenesis in NOS2 knockout animals and human samples derived from normal mucosa, leukoplakia and OSCC. Wild-type (iNOS+/+) and NOS2-knockout (iNOS-/-) mices, were challenged with 4- nitroquinoline- 1-oxide (4NQO) in drinking water for 16 weeks and killed after 0, 8 and 16 weeks of after treatment. Tongues were removed and the number of dysplasias and carcinomas was counted. Mice’s peripheral blood were analyzed by flux cytometry in search of circulating tumor cells. Human samples derived from normal mucosa, leukoplakia and OSCC, were utilized to relative quantify the amount of NOS2-mRNA. The human samples were also immune-stained with anti-iNOS and anti- Nitrotyrosine antibodies. In both groups, a reduction of dysplasias and an increase of carcinomas from week 16 to week 32 were observed. iNOS-/- mices had shown a small number of dysplastic and neoplastic lesion in all evaluated periods, moreover these lesions were usually smaller and less invasive when compared to wild type animals. Apparently, there is a rising tendency in the iNOS mRNA and protein levels during human oral carcinogenesis. Similar findings were obtained in the nitrotyrosine staining. Furthermore, iNOS and nitrotyrosine imuno-stained is associated with several clinic-pathological features of OSCC (site, presence of metastasis, staging and recidive). Our results shows that, iNOS affects the process of experimental and human oral carcinogenesis, being associated with OSCC genesis and progression. More studies should be done to provide us a deep understanding of the iNOS functions in oral cancer development. / Tese (Doutorado)

Conselho Nacional de Desenvolvimento Científico e Tecnológico / O carcinoma epidermoide de esôfago (CEE) representa 90% dos casos de câncer de esôfago no Brasil. O CEE tem detecção tardia, um comportamento extremamente agressivo e baixa sobrevida, sendo, portanto, um alvo interessante para o estudo dos mecanismos envolvidos em sua carcinogênese, a fim de se identificar possíveis alvos terapêuticos ou marcadores moleculares que ajudem na prática clínica. Mudanças no metabolismo energético da célula tumoral parecem ter papel de destaque na transformação maligna. Sabe-se que células tumorais consomem glicose avidamente produzindo ácido lático, mesmo em condições de normóxia. Dentre os fatores que podem contribuir para o estímulo da glicólise em células tumorais destacam-se as alterações em enzimas da via glicolítica tais como: as piruvato-cinases M1 e M2 (PKM1 e PKM2), a hexocinase II (HKII), isofoma 1 do transportador de glicose, GLUT-1, e o fator de transcrição induzido por hipóxia (HIF1&#945;), responsável pela transcrição das proteínas citadas. O objetivo do estudo é avaliar a relação entre a expressão de HIF1&#945;, HK2, PKM2, PKM1 e GLUT-1 e dados clínico-patológicos no CEE. Para tal, foram avaliados tumores conservados em parafina de 44 pacientes com CEE matriculados no INCA e no Hospital das Clínicas de Porto Alegre. Além disso, foram coletadas amostras de biópsia de esôfago em 67 pacientes sem doença esofágica, que foram submetidos à endoscopia no Hospital Universitário Pedro Ernesto (HUPE). A expressão das proteínas foi avaliada nos tecidos por imuno-histoquímica, enquanto que a expressão do mRNA de GLUT-1 também foi avaliada nas amostras controle. Foi observado que as amostras controle expressam HK2, PKM1, PKM2, HIF1&#945; nas camadas do epitélio esofágico. Já GLUT-1 e Ki-67 são vistos apenas na camada basal. Além disso, a expressão do mRNA de GLUT-1 não teve correlação com fatores etiológicos da doença. Em CEE a expressão de HK2, PKM2 e GLUT-1 foi vista em todos os tumores, já a expressão de HIF1&#945; e PKM1 foi variável. Além disso, observou-se que maior expressão de HIF-1&#945; apresenta correlação com invasão linfonodal e diferenciação, enquanto que a expressão de HK2 tem relação com sobrevida e PKM1 com diferenciação. As correlações clínicas encontradas sugerem que alterações no metabolismo energético é um alvo de estudo interessante para desenvolvimento de marcadores moleculares que auxiliem a prática clínica. / The esophageal squamous cell carcinoma (ESCC) represents 90% of cases of esophageal cancer in Brazil. The ESCC has late diagnosis, highly aggressive behavior and poor survival. ESCC is an interesting target to the study of mechanism involved in its carcinogenesis, in order to identify potential drug targets or biomarkers to help clinical practice. Changes in tumor cell energy metabolism appear to have a prominent role in malignant transformation. Tumor cells consume glucose avidly and produce lactic acid, even under normoxia. Among the factors that may contribute to the stimulation of glycolysis in tumor cells, there are changes in the glycolytic pathway enzymes such as: pyruvate kinase M1 and M2 (PKM2 and PKM1), hexokinase II (HKII), glucose transporter isoform 1, GLUT-1, and transcription factor induced by hypoxia (HIF1&#945;), responsible for the transcription of proteins cited. The goal of the study is to evaluate the relationship between the expression of HIF1&#945;, HK2, PKM2, PKM1 and GLUT-1 and clinicopathological data in ESCC. Biopsy of the esophagus in patients without esophageal disease were collected, who underwent endoscopy at University Hospital Pedro Ernesto (HUPE). Tissue samples were collected from 44 patients with a histologically confirmed diagnosis of ESCC recruted from Hospital Universitário Pedro Ernesto (HUPE-UERJ), and Instituto Nacional de Câncer (INCA). Tissue samples from healthy individuals submitted to endoscopic routine examination, not related to cancer or esophageal disorders, at HUPE-UERJ were also included in this study. The expression of proteins in tissues was evaluated by immunohistochemistry, while mRNA expression of GLUT-1 was also evaluated in the control samples. It was observed that the control samples express HK2, PKM1, PKM2, HIF1&#945; layers of the esophageal epithelium. GLUT-1 and Ki-67 are seen only in the basal layer. Furthermore, expression of GLUT-1 mRNA did not correlate with disease etiological factors. In ESCC expression of HK2, PKM2 and GLUT-1 was seen in all tumors, and the expression of HIF1&#945; and PKM1 was variable. We found that increased expression of HIF-1&#945; correlates with lymph node invasion and differentiation, whereas the expression of HK2 is related to survival, and differentiation with PKM1. The clinical correlations found suggest that alterations in energy metabolism are an interesting subject of study for development of biomarkers that help clinical practice.