<p>Medulloblastoma is the most common malignant brain tumor in children. The understanding of the genetic alterations in this tumor is emergent, and many such genetic driver events have yet to be functionally-characterized. Our studies have sought to understand the causes and consequences of OTX2 dysregulation in established medulloblastomas and in its putative cellular origins. Using a tumor genetic approach, we have uncovered frequent OTX2 copy number gains driving expression of this oncogene in a subset of medulloblastomas. However, OTX2 is frequently expressed in medulloblastomas independent of genomic copy number gain, and we thus sought to understand the transcriptional regulation of this gene in these tumors. We have found that chromatin accessibility, promoter DNA methylation, and activity of a distal downstream enhancer is distinct between OTX2-expressing and -nonexpressing medulloblastomas. Notably, autoregulation serves to maintain OTX2 expression in some medulloblastomas, whereas DNA methylation actively suppresses OTX2 in tumors not expressing this gene. Finally, we describe the effect of expressing Otx2 (the mouse homolog of OTX2) aberrantly in the developing mouse hindbrain, revealing that Otx2 disrupts spatiotemporal restrictions of neuronal progenitor cell proliferation. The effect of Otx2 in vivo is transient, with ectopically-proliferating cells give way to differentiated neurons. We found that OTX2 expression was not able to give rise to high penetrance medulloblastoma when combined with P53 deletion or double heterozygosity for P53 and PTEN. Thus, although Otx2 alters migration and proliferation dynamics of hindbrain neuronal progenitor cells, further studies are needed to identify the genetic alterations that cooperate with this oncogene to give rise to medulloblastoma.</p> / Dissertation

Diss. (sammanfattning) Stockholm : Karol inst., 2003. / Härtill 4 uppsatser.

Diss. (sammanfattning) Stockholm : Karol. inst. / Härtill 4 uppsatser.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2002. / Härtill 5 uppsatser.

Thesis (Ph. D.)--University of Washington, 2000. / Vita. Includes bibliographical references (leaves 155-179).

Keloid disease (KD) is a cutaneous fibroproliferative tumour characterised by heterogeneity, locally aggressive invasion and therapeutic resistance. Clinical, histological and molecular differences between the keloid scar centre and margin as well as recent evidence of the importance of epithelial-mesenchymal interactions (EMI) in KD pathobiology contribute to the complexity and diversity of KD, which coupled with the lack of a validated animal model have hindered research and effective management. Despite significant progress in the field of KD research, reliance on conventional monolayer cell culture and whole tissue analysis methods have failed to fully reflect the natural architecture, pathology and complexity of KD in vivo. In order to address these challenges, a site-specific in situ approach was therefore employed here for the first time in KD research. The first aim of this work was to compare the value of this contemporary approach with traditional methods of tissue dissection. The second aim was to compare the genomic expression between well-defined, distinct keloid sites and normal skin (NS). The third aim was to develop and explore hypotheses arising from this site-specific gene expression profiling approach, so as to enhance understanding of KD pathobiology as a basis for improved diagnostic and therapeutic strategies in future KD management. The fourth aim was to probe these hypotheses with relevant functional in vitro studies. The current site-specific in situ approach was achieved through a combination of laser capture microdissection and whole genome microarray, allowing separation of epidermis from dermis for keloid centre, margin and extralesional sites compared with NS. This in situ approach yielded selective, accurate and sensitive data, exposing genes that were overlooked with alternative methods of dissection. Identification of significant upregulation of the aldo-keto reductase enzyme AKR1B10 in all three sites of the keloid epidermis (KE) in situ, implicated dysregulation of the retinoic acid (RA) pathway in KD pathogenesis. This hypothesis was supported by showing that induced AKR1B10 overexpression in NS keratinocytes reproduced the keloid RA pathway expression pattern. Moreover, co-transfection with a luciferase reporter plasmid revealed reduced RA response element activity. Paracrine signals released by AKR1B10-overexpressing keratinocytes into conditioned medium resulted in TGFβ1 and collagen upregulation in keloid fibroblasts, suggesting the disturbed RA metabolism exerts a pro-fibrotic effect through pathological EMI, thus further supporting the hypothesis of RA deficiency in KE. Gene expression profiling further revealed an upregulation of NRG1 and ErbB2 in keloid margin dermis. Exogenous NRG1 led to enhanced keloid fibroblast migration with increased Src and PTK2 expression, which were attenuated with ErbB2 siRNA studies. Together with the observed failure to recover this expression with NRG1 treatment, suggested the novel KD pathobiology hypothesis that NRG1/ErbB2/Src/PTK2 signaling plays a role in migration at the keloid margin. In addition to these hypotheses, LCM methodology with comprehensive analysis of the data permitted the development of additional novel working hypotheses that will inform future KD research, including inflammatory gene dysregulation and cancer-like stem cells that may contribute to the therapeutic resistance characteristic of KD.

Made available in DSpace on 2014-06-11T19:25:37Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-02-10Bitstream added on 2014-06-13T18:29:05Z : No. of bitstreams: 1 oliveira_lc_me_botfm.pdf: 220276 bytes, checksum: 96205056c501bc976044abaee0c89592 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O objetivo deste estudo foi avaliar o papel do ácido retinóico na remodelação cardíaca induzida pela exposição à fumaça do cigarro. Os ratos foram divididos em três grupos: controle (C, n = 8); expostos animais expostos à fumaça do cigarro (EFC, n = 9); animais expostos à fumaça de cigarro e suplementados com ácido retinóico (EFC-AR, n = 9). Após dois meses, foram submetidos ao ecocardiograma e análise morfométrica. Os dados estão expressos em média ± desvio padrão ou medianas e intervalo interquartílico (Q1-Q3). Não houve diferença na pressão sistólica caudal entre os grupos (C = 116 ± 27 mmHg; EFC = 129 ± 17 mmHg; EFC-AR = 139 ± 19 mmHg; p = 0,102). O grupo EFC mostrou maior diâmetro diastólico do ventrículo esquerdo ajustado ao peso corporal (C = 18,42 ± 3,57 mm/kg; EFC = 23 ± 1,85 mm/kg; EFC-AR = 19,51 ± 0,99 mm/kg; p = 0,001) e sistólico ajustado ao peso corporal (C = 8,25 ± 2,16 mm/kg; EFC = 11,53 ± 1,31 mm/kg; EFC-AR = 8,25 ± 0,71 mm/kg; p = <0,001) quando comparado com C grupo e EFC-AR. Por outro lado, não houve diferença entre C e EFC-AR. A espessura da parede do ventrículo esquerdo ajustado ao diâmetro diastólico do ventrículo esquerdo foi maior no grupo C do que nos EFC (C = 0,20 (0,18-0,23); EFC = 0,15 (0,14-0,18); EFC-AR = 0,19 (0,17-0,22; p = 0,003). A fração de ejeção (C = 0,91 ± 0,02; EFC = 0,87 ± 0,03; EFC-AR = 0,92 ± 0,03; p = 0,003) e a fração de encurtamento (C = 55,77 ± 4,41%; EFC = 49,73 ± 4,43%; EFC-AR = 57,60 ± 5,15%; p = 0,005) foram mais elevados nos animais do grupo C e EFC-AR em relação aos animais do grupo EFC. Por outro lado, não houve diferenças entre C e EFC-AR. Além disso, o tabagismo está associado com um aumento significativo da área seccional do miócito (C = 294 ± 21 μm2; EFC = 347 ± 44 μm2; EFC-AR = 310 ± 37 μm2; p = 0,016). Por outro lado... / The objective of this study was to evaluate the role of retinoic acid in the myocardial remodeling induced by tobacco smoke exposure. Rats were allocated into three groups: control (C, n=8); exposed to tobacco smoke (ETS, n=9); exposed to tobacco smoke and all-transretinoic acid (ETS-AR, n = 9). After two months, were submitted to echocardiogram, and morphometric analyses. Data are expressed as mean ± SD or medians (including the lower quartile and upper quartile). There were no differences in the tail systolic pressure among the groups (C = 116 ± 27 mmHg, ETS = 129 ± 17 mmHg, ETS-AR = 139 ± 19 mmHg; p = 0,102). ETS showed higher normalized left ventricular diastolic (C = 18,42 ± 3,57 mm/kg, ETS = 23 ± 1,85 mm/kg, ETS-AR = 19,51 ± 0,99 mm/kg; p = 0,001) and systolic (C = 8,25 ± 2,16 mm/kg, ETS = 11,53 ± 1,31 mm/kg, ETS-AR = 8,25 ± 0,71mm/kg; p = <0,001) diameters than C and ETS-AR. On the other hand, there were no differences between C and ETS-RA. LVWT/LVEDD was higher in C than ETS (C = 0,20 (0,18-0,23), ETS = 0,15 (0,14-0,18), ETS-RA = 0,19 (0,17-0,22; p = 0,030). The ejection fraction (C = 0,91 ± 0,02, ETS = 0,87 ± 0,03, ETS-AR = 0,92 ± 0,03; p = 0,003) and fractional shortening (C = 55,77 ± 4,41 %, ETS = 49,73 ± 4,43 %, ETS-AR = 57,60 ± 5,15 %; p = 0,005) were higher in C and ETS-RA animals than ETS animals. On the other hand, there were no differences between C and ETS-RA. Also, smoking was associated with significantly increased myocyte cross-sectional area (C = 294 ± 21 μm2, ETS = 347 ± 44 μm2, ETS-RA = 310 ± 37 μm2; p = 0,016). On the other hand, there were no differences between C and ETS-RA. Retinoic acid did not affect other functional or morphological variables. In conclusion, ...(Complete abstract click electronic access below)

Made available in DSpace on 2014-06-11T19:32:13Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-02-16Bitstream added on 2014-06-13T19:42:37Z : No. of bitstreams: 1 freire_cmm_dr_botfm.pdf: 388273 bytes, checksum: 9c831a245404da97a807b7720aa9ae7a (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Devido à escassez de trabalhos sobre a influência do todo-trans-ácido retinóico (ATRA) no coração de ratos normais, foi realizado experimento com ratos jovens adultos, que receberam diferentes doses de ATRA. O objetivo deste estudo foi verificar se as alterações causadas pelo ATRA no coração dos ratos, são fisiológicas ou patológicas e se apresentam relação dose-dependente. Para este estudo, foram utilizados 72 ratos Wistar, distribuídos em 4 grupos: 1) grupo com 18 animais que receberam a dieta sem ATRA (controle), 2) grupo com 18 animais que receberam 0,3 mg de ATRA / kg de dieta/dia (AR 0,3), 3) grupo com 18 animais que receberam 10 mg de ATRA /kg de dieta/dia (AR 10), 4) grupo com 18 animais que receberam 50 mg de ATRA /kg de dieta/dia (AR 50). Após 4 semanas de tratamento, os animais foram avaliados pelo ecocardiograma e após eutanásia, as amostras de sangue, do fígado e do coração, foram coletadas. As amostras dos ventrículos esquerdos (VE) foram separadas para a avaliação do teor de água, da hipertrofia, da fibrose, da conexina 43, das enzimas do metabolismo energético cardíaco e para a realização das dosagens de vitamina A total e ATRA. No fígado, foram feitas as dosagens da vitamina A total e do ATRA. No sangue, foram realizadas as quantificações de retinol, triglicerídeos e colesterol total. Neste trabalho, foi observado, por meio do ecocardiograma, que os animais que receberam 50 mg de ATRA apresentaram hipertrofia cardíaca caracterizada pelo aumento proporcional entre a espessura da parede do ventrículo esquerdo e o diâmetro da sua cavidade (EPVE/DDVE), sem o aparecimento de fibrose. A função sistólica melhorou e a função diastólica permaneceu normal. O teste de tendência linear mostrou aumento gradual nas seguintes variáveis: diâmetro do átrio esquerdo (DAE), diâmetro diastólico do ventrículo... / In the current medical bibliography there are only a few papers concerning the influence of all-trans-retinoic acid (ATRA) on the hearts of normal rats. To supply this lack was carried out a study with young adult rats, that received different doses of ATRA. The aim of this work was to evaluate if the cardiac alterations caused by ATRA are physiological or pathological and if these alterations are dose-dependent. In this research were used 72 Wistar rats allocated in 4 groups: 1) group with 18 animals that received a diet without ATRA (control), 2) group with animals that received 0,3mg of ATRA/ kg of the diet (RA 0,3), 3) group with animals that received 10mg of ATRA/ kg of the diet (RA 10) and 4) group with animals that received 50 mg of ATRA/ kg of the diet (RA 50). After 4 weeks, the animals were evaluated by echocardiography exam and in the next day, the rats were euthanasiated and the samples of blood, liver and heart were collected. The left ventricle (LV) samples were separated for evaluation of water content, hypertrophy, fibrosis, connexin 43, enzymes of energetic metabolism and for dosages of total vitamin A and ATRA. In the livers were determined the total vitamin A and ATRA and in the blood were carried out the dosages of the retinol, triglycerides and total cholesterol. In was observed ,by echocardiogram, that the animals which received 50mg of ATRA presented cardiac hypertrophy with proportional increase in the wall thickness of the left ventricle in relation to its diameter (LVWT/LVDD), but it was observed fibrosis. The systolic function increased and diastolic function was normal. In relation to the trend test was observed gradual increase in the left atrium diameter (LAD), left ventricle diastolic diameter (LVDD), left ventricle systolic diameter (LVDS)... (Complete abstract click electronic access below)

Resumo: Devido à escassez de trabalhos sobre a influência do todo-trans-ácido retinóico (ATRA) no coração de ratos normais, foi realizado experimento com ratos jovens adultos, que receberam diferentes doses de ATRA. O objetivo deste estudo foi verificar se as alterações causadas pelo ATRA no coração dos ratos, são fisiológicas ou patológicas e se apresentam relação dose-dependente. Para este estudo, foram utilizados 72 ratos Wistar, distribuídos em 4 grupos: 1) grupo com 18 animais que receberam a dieta sem ATRA (controle), 2) grupo com 18 animais que receberam 0,3 mg de ATRA / kg de dieta/dia (AR 0,3), 3) grupo com 18 animais que receberam 10 mg de ATRA /kg de dieta/dia (AR 10), 4) grupo com 18 animais que receberam 50 mg de ATRA /kg de dieta/dia (AR 50). Após 4 semanas de tratamento, os animais foram avaliados pelo ecocardiograma e após eutanásia, as amostras de sangue, do fígado e do coração, foram coletadas. As amostras dos ventrículos esquerdos (VE) foram separadas para a avaliação do teor de água, da hipertrofia, da fibrose, da conexina 43, das enzimas do metabolismo energético cardíaco e para a realização das dosagens de vitamina A total e ATRA. No fígado, foram feitas as dosagens da vitamina A total e do ATRA. No sangue, foram realizadas as quantificações de retinol, triglicerídeos e colesterol total. Neste trabalho, foi observado, por meio do ecocardiograma, que os animais que receberam 50 mg de ATRA apresentaram hipertrofia cardíaca caracterizada pelo aumento proporcional entre a espessura da parede do ventrículo esquerdo e o diâmetro da sua cavidade (EPVE/DDVE), sem o aparecimento de fibrose. A função sistólica melhorou e a função diastólica permaneceu normal. O teste de tendência linear mostrou aumento gradual nas seguintes variáveis: diâmetro do átrio esquerdo (DAE), diâmetro diastólico do ventrículo ...(Resumo completo, clicar acesso eletrônico abaixo) / Abstract: In the current medical bibliography there are only a few papers concerning the influence of all-trans-retinoic acid (ATRA) on the hearts of normal rats. To supply this lack was carried out a study with young adult rats, that received different doses of ATRA. The aim of this work was to evaluate if the cardiac alterations caused by ATRA are physiological or pathological and if these alterations are dose-dependent. In this research were used 72 Wistar rats allocated in 4 groups: 1) group with 18 animals that received a diet without ATRA (control), 2) group with animals that received 0,3mg of ATRA/ kg of the diet (RA 0,3), 3) group with animals that received 10mg of ATRA/ kg of the diet (RA 10) and 4) group with animals that received 50 mg of ATRA/ kg of the diet (RA 50). After 4 weeks, the animals were evaluated by echocardiography exam and in the next day, the rats were euthanasiated and the samples of blood, liver and heart were collected. The left ventricle (LV) samples were separated for evaluation of water content, hypertrophy, fibrosis, connexin 43, enzymes of energetic metabolism and for dosages of total vitamin A and ATRA. In the livers were determined the total vitamin A and ATRA and in the blood were carried out the dosages of the retinol, triglycerides and total cholesterol. In was observed ,by echocardiogram, that the animals which received 50mg of ATRA presented cardiac hypertrophy with proportional increase in the wall thickness of the left ventricle in relation to its diameter (LVWT/LVDD), but it was observed fibrosis. The systolic function increased and diastolic function was normal. In relation to the trend test was observed gradual increase in the left atrium diameter (LAD), left ventricle diastolic diameter (LVDD), left ventricle systolic diameter (LVDS)... (Complete abstract click electronic access below) / Orientador: Sergio Alberto Rupp de Paiva / Coorientador: Leonardo Antônio Mamed Zornoff / Banca: Luís Shiguero Matsubara / Banca: Paula Schmidt Azevedo Gaiolla / Banca: Felix José Alvarez Ramires / Banca: Fernando Bacal / Doutor

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