Made available in DSpace on 2015-07-22T20:50:09Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-07-29 / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / INTRODUÇÃO: O papel da angiogênese como indicador prognóstico em câncer é estudado frequentemente e resultados demonstram correlação entre angiogênese e prognostico em neoplasias malignas. Em osteossarcoma poucos estudos foram realizados e permanecem controversos. OBJETIVO: avaliar a densidade de microvasos (MVD) e a expressão do VEGF (vascular endothelial growth factor) como marcadores de angiogenese e a sua correlação com fatores prognósticos. MATERIAL E METODO: blocos de parafina de 50 pacientes pré-quimioterapia ( 22 deles metastáticos ao diagnostico) com osteossarcoma central de alto grau tratados na nossa instituição foram estudados pela técnica de imuno-histoquimica para VEGF e densidade de microvasos determinada por CD 34 e CD 31. A expressão de ambos foi correlacionada com aspectos clinicopatologicos e fatores relacionados ao prognostico (metástases e óbito). As neoplasias foram consideradas positivas para VEGF quando mais que 64,7% (media) das células demonstravam positividade. MVD foi obtida em cada caso pela média de contagem de microvasos em 10 campos de grande aumento (400x) em cada biopsia. Foi considerada alta e baixa usando a media como ponto de corte. RESULTADOS: apesar da alta vascularização não houve significância estatística entre MVD e fatores prognósticos para CD 31 e CD 34. Em relação ao VEGF houve correlação entre VEGF melhor sobrevida global e livre de doença para o total de pacientes. Quando separamos os pacientes metastaticos ao diagnostico os que foram VEGF negativos tiveram melhor sobrevida global e livre de doença. CONCLUSÃO: osteossarcomas são neoplasias bem vascularizadas e a imunoexpressão de VEGF dá informações prognosticas importantes principalmente em pacientes metastaticos ao diagnostico. / INTRODUCTION: The role of angiogenesis as a prognostic indicator in cancer has been extensively studied in recent times with several studies demonstrating a positive correlation for various malignant tumours. However, the role of angiogenesis in osteosarcoma remains a topic of debate. AIM: evaluate the significance of intratumoral microvessel density (MVD) and the degree of vascular endothelial growth factor (VEGF) expression as markers of angiogenesis and correlate with disease outcome. MATERIAL AND METHODS: Pretherapeutic paraffin- embedded biopsies from 50 patients diagnosed high grade central osteosarcoma at our institution with clinicopathological data were studied by immunohistochemical tecnique for VEGF and microvessel density (MVD) determined by CD 34and CD 31expression. VEGF expression and MVD were correlated with clinicopathological factors and patient outcome in terms of metastasis and death. The degree of angiogenesis was assessed, as determined by the microvessel density (MVD). Tumors were considered VEGF positive if immunoexpression was positive in greater than 64,7% ( mean) of the tumor cells. MVD was obtained by microvessels counting 10 high power fields (X400 field) in each biopsy sample. The mean MVD of the entire group was predetermined to classify patients into two groups with high and low MVD.RESULTS: Despite the moderate to high vascularity, there was no significant difference between the MVD and disease outcome factors for both CD 31and CD 34. In terms of VEGF expression, patients with metastases at diagnosis VEGF negative had better overall survival and disease-free survival .CONCLUSION: We were able to demonstrate that osteosarcoma is a relatively vascular tumor and VEGF expression provides prognostic information in osteosarcoma; however, the degree of MVD does not provide prognostic information. Angiogenesis has a potential target for anti-angiogenic therapies. / TEDE / BV UNIFESP: Teses e dissertações

Objetivo: Este estudo foi realizado com objetivo de traduzir para o português do Brasil e validar a ferramenta Musculoskeletal Tumor Society Rating Scale (MSTS) para avaliação da função em pacientes com sarcomas ósseos dos membros inferiores. Método: A ferramenta MSTS foi traduzida da língua inglesa para o português do Brasil. As versões traduzidas foram sintetizadas em uma versão de consenso que foi retrotraduzida para o inglês e analisada por um comitê multidisciplinar para posterior aplicação da ferramenta. O questionário foi aplicado em 67 pacientes operados por tumores musculoesqueléticos ósseos malignos de membros inferiores com cirurgia preservadora de membro ou amputação. Este grupo também preencheu uma versão validada em Português do Toronto Extremity Salvage Score (TESS). Foram realizadas as análises de confiabilidade do teste e reteste, confiabilidade interobservador, consistência interna, validação do instrumento e correlação com o TESS. Resultados: A confiabilidade do teste e reteste e interobservadores foi feita através da análise dos coeficientes de correlação intraclasse (ICC) com valores de 0,92 e 0,98 respectivamente (intervalo de confiança de 95%). Na análise da consistência interna, o valor do coeficiente alfa de Cronbach foi de 0,84. A correlação entre as ferramentas MSTS e TESS foi moderada. Conclusão: A ferramenta MSTS foi traduzida e validada para o Português do Brasil e mostrou ser confiável para aplicação em pacientes com sarcomas ósseos de membros inferiores. Isto possibilitará a avaliação funcional dos pacientes brasileiros e comparação com resultados de estudos multicêntricos / Objective: This study was designed to translate and validate the Brazilian version of the Musculoskeletal Tumor Society Rating Scale (MSTS) for functional evaluation in patients with lower extremity bone sarcomas. Method: The MSTS was translated from English into Brazilian Portuguese. Translations were synthesized, translated back into English and reviewed by a multidisciplinary committee for further implementation. The questionnaire was administered to 67 patients treated for malignant lower extremity bone tumors who were submitted to limb salvage surgery or amputation. They also completed a Brazilian version of the Toronto Extremity Salvage Score (TESS). Scale reliability and validity were evaluated. Results: Testretest reliability and interobserver analysis were performed using the intraclass correlation coefficients (ICC) with values of 0,92 and 0,98 respectively (confidence interval 95%). Cronbach\'s alpha was used to estimate internal consistency (0,84). There was mild correlation between MSTS and TESS. Conclusion: The Brazilian version of the MSTS was translated and validated. It is a reliable tool to assess functional outcome in patients with lower extremity bone sarcomas. It can be used for functional evaluation of Brazilian patients and cross-cultural comparisons

A partir de células de osteossarcoma humano (MG-63) que tem características de osteoblastos imaturos produzimos anticorpos monoclonais (Mabs) nomeados PSP 4-5, PSP 42-22 e PSP 85-9. Esses anticorpos reconhecem antígenos de 100, 26 e 20 kDa respectivamente. Avaliamos a especificidade dos mesmos, testando suas expressões em cortes congelados de tecidos oriundos da mesma célula mesenquimal, ou seja, osso, cartilagem, músculo cardíaco e tecido adiposo. Os anticorpos marcaram o periósteo, com distintos padrões de expressão, porém o PSP 42-22 foi o mais específico marcando a camada osteogênica do periósteo. Os anticorpos marcaram também células ósseas. Diante desses resultados nosso objetivo, no presente estudo, foi identificar os antígenos reconhecidos por esses anticorpos usando clonagem e sequenciamento dos genes em biblioteca de cDNA com capacidade de expressão proteica. Outro objetivo foi testar os anticorpos, pela técnica de imunohistoquímica (IHQ), em tumores ósseos primários visando estabelecer os padrões de marcação e se diferenciavam os diferentes tipos de tumores. Os antígenos reconhecidos pelos anticorpos PSP 42-22 e PSP 85-9 foram isolados, purificados e sequenciados. Devido ao elevado peso molecular do PSP 4-5 necessitaremos de outras técnicas para isolar o clone que codifica seu antígeno. O sequenciamento do clone isolado pelo PSP 42-22, mostrou uma sequência com 99% de homologia descrita no \"Homo sapiens\" como sendo \"serologically defined colon cancer antigen 3 (SDCCAG3), transcript variant 3\". A proteína SDCCAG3 foi descrita pela primeira vez em 1998 como um antígeno de câncer de cólon reconhecido por anticorpo autólogo. Vale lembrar que antígeno reconhecido pelo nosso anticorpo tem um peso molecular de aproximadamente 26 kDa, inferior ao da proteína SDCCAG3. Essa diferença poderia ocorrer devido a uma diferença no local da tradução do mRNA das células MG-63, produzindo uma proteína menor (isoforma), ou no clone isolado (3B2-3-1), poderia ocorrer uma mutação produzindo uma proteína mais curta que se expressaria na membrana celular ao contrário de uma proteína mais longa. Já o alinhamento das sequências obtidas dos clones isolados utilizando o PSP 85-9, mostrou uma sequência com 100% de concordância descrita com a porção não codante da proteína \"Homo sapiens microtubule associated protein, RP/EB family, member 1 (MAPRE1)\" portanto um falso positivo; já o clone 1A5-1-3, mostrou uma sequência de 99% de homologia com a proteína \"Homo sapiens Yip1 interacting factor homolog B (S. cerevisiae) (YIF1B), transcript variant 8\". Trata-se de uma proteína de membrana, com 6 isoformas diferentes que pode interagir com o receptor de serotonina. O resultado da IHQ nos tumores ósseos testados mostrou que o PSP 4-5 se expressou predominantemente no citoplasma de osteossarcoma, condrossarcoma e leiomiossarcoma e no núcleo de osteoblastoma. O anticorpo PSP 42-22 não reconheceu nenhum antígeno nos tumores avaliados. Quanto ao anticorpo PSP 85-9 nos condrossarcomas e leiomiossarcomas a expressão foi predominante citoplasmática e nos osteossarcomas e osteoblastoma foi nuclear. Em conclusão, identificamos os antígenos de dois dos anticorpos estudados que apresentam potencial diagnóstico diferencial em tumores ósseos primários / From human osteosarcoma cells (MG-63), which have characteristics of immature osteoblasts, we produced monoclonal antibodies (Mabs) named PSP 4-5, PSP 42-22 and PSP 85-9. These antibodies recognize antigens with 100, 26 and 20 kDa, respectively. We evaluated their specificity by testing their expression in frozen tissue sections from the same mesenchymal cells, that is, bone, cartilage, cardiac muscle and adipose tissue. The antibodies stained the periosteum, with distinct patterns of expression, but PSP 42-22 was the most specific, scoring the osteogenic layer of the periosteum. The antibodies also marked bone cells. With these results, our goal in this study was to identify the antigens recognized by these antibodies using cloning and sequencing of the genes in the cDNA library with capacity of protein expression. Another objective was to test the antibodies by immunohistochemistry (IHC) in primary bone tumors to establish standards for marking and whether they set apart different types of tumors. The antigens recognized by the PSP 42-22 and PSP 85-9 antibodies were isolated, purified and sequenced. Due to the high molecular weight of PSP 4-5 we will need other techniques to recognize its antigen. Sequencing of the clone isolated using the PSP 42-22 showed a sequence with 99% homology described in \"Homo sapiens\" as being \"serologically defined colon cancer antigen 3 (SDCCAG3), transcript variant 3\". The SDCCAG3 protein was first described in 1998 as a colon cancer antigen recognized by autologous antibody. It is worth remembering that the antigen recognized by our antibody has a molecular weight of approximately 26 kDa, lower than the SDCCAG3 protein. This difference could be due to a difference in mRNA translation site of MG-63 cells producing a lower protein; or on the isolated clone (3B2-3-1) a mutation could occur producing a shorter protein that expresses in the cell membrane instead of a longer protein. The alignment of the sequences obtained from isolated clones using the PSP 85-9 showed a sequence with 100% of homology described with the non-coding protein portion \"Homo sapiens microtubule associated protein, RP/EB family, member 1 (MAPRE1), mRNA\" therefore, a false positive; 1A5-1-3 clone showed a 99% homology sequence with the protein \"Homo sapiens Yip1 interacting factor homolog B (S. cerevisiae) (YIF1B), transcript variant 8, mRNA\". It is a membrane protein with 6 different isoforms which can interact with the serotonin receptor. The result of IHC in bone tumors tested showed that PSP 4-5 expressed predominantly in the cytoplasm of osteosarcoma, chondrosarcoma, leiomyosarcoma and osteoblastoma in was in nucleus. The PSP 42-22 antibody did not recognize any antigen in the tumors examined. As for the PSP 85-9 antibody in chondrosarcoma and leiomyosarcoma, the expression was predominantly cytoplasmic and the osteoblastoma was nuclear in osteosarcomas. In conclusion, we have identified the antigens of two of the antibodies studied which have potential differential diagnosis in primary bone tumors

A ressecção ampla do trajeto de biópsia junto ao tumor faz parte da técnica cirúrgica oncológica para evitar os riscos de implante de células tumorais e recidiva local e/ou sistêmica da doença. O objetivo deste estudo foi verificar se havia risco de contaminação por células neoplásicas no trajeto da biópsia. Realizou-se um estudo prospectivo dos trajetos de biópsias de pacientes operados por tumores musculoesqueléticos malignos no Instituto de Ortopedia e Traumatologia da Faculdade de Medicina da Universidade de São Paulo no período de abril de 2006 a abril de 2007. Foram estudados por histopatologia 25 casos. Houve implante de células neoplásicas em 32% dos trajetos. Quando foram correlacionados os resultados dos estudos dos trajetos e o uso de quimioterapia neo-adjuvante (p=0,19) não houve significância, mostrando que mesmo com esse tratamento o risco de implante existe. As alterações histológicas mais comuns foram: fibrose acentuada, componente inflamatório leve e neovascularização acentuada. Sugerimos a ressecção tradicional oncológica do trajeto junto com a peça / Wide resection of biopsy tracts to tumors forms part of oncological surgical techniques because of the risk of tumor cell implantation and local and/or systemic disease recurrence. The aim of this study was to investigate the risk of seeding by neoplastic cells along biopsy tracts. This was a prospective histopathological study on biopsy tracts in 25 patients who underwent operations due to malignant musculoskeletal tumors, at the Institute of Orthopedics and Traumatology, School of Medicine of the University of São Paulo, between April 2006 and April 2007. Neoplastic cells were found implanted in 32% of the tracts. Correlation of the results from studying the tracts with the use of neoadjuvant chemotherapy did not present significant findings. This shows that, even with chemotherapy, the risk of implantation exists. The most common histological abnormalities in positive cases were classified as severe fibrosis, mild inflammatory component and severe neovascularization. We suggest that oncological resection of the tract should be carried out together with excision of the specimen

Protein kinase A (PKA) is an important enzyme inside the body; it is responsible for phosphorylation of gene regulatory elements and thus regulation of gene expression inside the nucleus. Malfunction of PKA affects transcriptional and translational levels of cell signaling ligands, leading to abnormal activity of various signaling pathways. PKA holoenzyme is composed of two regulatory and two catalytic subunits; four main regulatory subunit isoforms (R1α, R1β, R2α and R2β) and four main catalytic subunit isoforms (Cα, Cβ, Cγ and Prkx) of PKA have been identified. Mutations in these subunits lead to altered total PKA activities and PKAT-I to PKAT-II ratios, leading to diseases both in human and mice. These diseases include Carney Complex (CNC), fibrous dysplasia (FD) and Cushing syndrome. We studied the effect of PKA subunit mutations on intracellular PKA activities, PKAT-I to PKAT-II ratios, and bone and adrenal gland phenotypes in transgenic mouse models. Firstly, we generated whole-body transgenic mice single or double heterozygous for PKA regulatory subunits. Tail vertebral bone lesions including osteosarcomas, osteochondromas and osteochondrosarcomas were found in these mice and we found that mutations in different PKA subunits affect bone lesion formation, new bone generation, and bone organization and mineralization in mouse tail vertebrae. Elevated Cβ subunit expression in Parkar1a+/-Prkar2a+/- and Prkar1a+/-Prkar2b+/-double heterozygous mice leads to a less severe vertebral bone lesion phenotype, an increased osteogenic activity and a better bone regeneration activity. We then studied mice with tissue specific knock out of Prkar1a, the gene coding for type I regulatory subunit, specifically in adrenal cortex (AdKO). AdKO mice developed pituitary-independent Cushing syndrome with increased PKA activity. They also demonstrated increased plasma corticosterone levels resistant to dexamethasone suppression. Dietary treatment of both mice with bone lesions and mice with adrenal lesions with COX2 inhibitor Celecoxib led to partial rescue of phenotypes; this is due to inhibition of the positive feedback loop between PKA signaling and inflamasome pathway at COX2 induction level by Celecoxib. / 蛋白激酶A（PKA）是人體中重要的蛋白酶， 它通過燐酸化基因調控元件來實現對細胞核內基因表達的調節。PKA異常影響細胞內信號傳遞因子的基因轉錄和蛋白翻譯水平，從而導致各細胞信號通路的異常活動。PKA全酶由兩個調節亞基和兩個催化亞基組成，目前已經發現的有四個調節亞基 (R1α, R1β, R2α 和R2β) 以及四個催化亞基(Cα, Cβ, Cγ和Prkx)。發生在這些亞基中的基因突變會改變總的PKA活動水平，PKA-I 和PKA-II的比例，在人類和實驗鼠中引起疾病。這些疾病包括卡尼綜合症 （CNC），骨纖維性發育不良（FD）和庫欣綜合症。我們在轉基因鼠模型中研究PKA亞基突變對細胞中PKA總活性， PKA-I和PKA-II比例的影響，以及由此帶來的骨和腎上腺表型的改變和病變。我們首先製造了有一個或兩個PKA亞基雜合性缺失的全身轉基因鼠。在這些轉基因鼠中，我們發現了包括骨肉瘤，骨軟骨瘤和骨軟骨肉瘤在內的尾椎骨病變。研究發現在不同PKA亞基中的基因變異對實驗鼠尾椎骨病變的發生，新骨的形成和骨的結構和纖維化均有影響。在Prkar1a+/-Prkar2a+/-和Prkar1a+/-Prkar2b+/-實驗鼠中我們發現了較高的Cβ催化亞基表達，這兩個基因型因此具有更輕度的骨病變和更強的骨再生能力。我們繼續研究了在腎上腺中敲除了標記PKA 第一調節亞基的Prkar1a基因的實驗鼠 （AdKO）。AdKO實驗鼠中產生了與垂體無關的庫欣綜合症，並伴隨PKA活性的增加。它們還表現出耐地塞米松抑制的血漿皮質酮水平增加。對骨病變或腎上腺病變的實驗鼠通過飲食進行COX2抑制劑塞來昔布的治療可以部分緩解病變表型。這是由對PKA和炎性體的正反饋機制在COX2誘導步驟的抑制造成的。 / Liu, Sisi. / Thesis Ph.D. Chinese University of Hong Kong 2015. / Includes bibliographical references (leaves 115-130). / Abstracts also in Chinese. / Title from PDF title page (viewed on 09, September, 2016). / Detailed summary in vernacular field only.

Dissertation (Ph.D.)--University of South Florida, 2007. / Title from PDF of title page. Document formatted into pages; contains 192 pages. Includes vita. Includes bibliographical references.

Conselho Nacional de Desenvolvimento Científico e Tecnológico / This doctoral thesis involved the study of three groups of neoplasms that affect bones of dogs (primary bone neoplasms, bone metastases, and multicentric neoplasms with bone involvement) and a bone lesion, often paraneoplastic, known as hypertrophic osteopathy. The study of primary bone neoplasms covered important pathological and epidemiological aspects for the diagnosis of this group of tumors, with emphasis on osteosarcomas. It was retrospectively performed, covering a period of 22 years. Reports of biopsy and necropsy cases of dogs received at the Laboratório de Patologia Veterinária, Universidade Federal de Santa Maria (LPVUFSM) were analyzed. Out of the 90 primary bone neoplasms diagnosed in this period, 89 were malignant. Osteosarcoma was the most prevalent (86.7%) neoplasm. Regarding osteosarcomas, most cases occurred in large and giant breed dogs, between six and 10 years of age. The neoplasms predominantly involved the appendicular skeleton and were 3.5 times more prevalent in the forelimbs than in the hindlimbs. The predominant histologic subtype was the osteoblastic. For the study of neoplasms that comprise the second and third groups, i.e., neoplasms with bone metastases or with bone involvement by multicentric neoplasms, a prospective study was conducted over a period of three years. The skeleton of 110 dogs, with 118 malignant neoplasms of different origins received in the necropsy service of the LPV-UFSM were examined for bone lesions. Twenty-one cases of bone metastases or bone involvement by multicentric neoplasms (19.1%) were detected. In general, the bone lesions affected more female dogs. However, when mammary gland neoplasms were not considered, the distribution of cases according to the sex was very similar. The mean age was 9-years-old and dogs of different breeds were affected. The mammary gland was the primary site of most bone metastases, followed by neoplasms of the musculoskeletal and respiratory systems. Most metastases were observed grossly and occurred in multiple bones. However, in 23% of the cases metastases could only be observed microscopically. Vertebrae and humerus were the mosdt frequently affected bones. Simultaneously, seven cases of hypertrophic osteopathy, diagnosed in a period of 11 years at the LPV-UFSM, were retrospectively and prospectively studied. Affected dogs had clinical signs of bone involvement and lesions mainly in the long bones of the limbs. The lesions consisted of periosteal bone neoformation, detected on radiographs, bone inspection during necropsy, and with great level of detail, in macerated bone specimens. The bone proliferation was partially circumferential and occurred mainly in the diaphysis of long bones. It consisted of bone trabeculae of irregular size and thickness, which were arranged perpendicularly to the original cortical bone. In all cases, the lesions of hypertrophic osteopathy were associated with lung neoplasms (primary or metastatic). In two of the seven cases, the lung metastases were of primary bone sarcomas and, in one case, there was a primary lung osteosarcoma (extra-skeletic). / Esta tese envolveu o estudo de três grupos de neoplasmas que afetam os ossos de cães (neoplasmas ósseos primários, metástases ósseas e neoplasmas multicêntricos com envolvimento ósseo) e uma alteração óssea, muitas vezes paraneoplásica, conhecida como osteopatia hipertrófica. O estudo dos neoplasmas ósseos primários envolveu aspectos epidemiológicos e patológicos importantes para o diagnóstico deste grupo de tumores, com ênfase nos osteossarcomas. Foi realizado de forma retrospectiva, compreendo um período de 22 anos. Foram analisados laudos de casos de biópsias e necropsias de cães recebidos no Laboratório de Patologia Veterinária da Universidade Federal de Santa Maria (LPV-UFSM). Dos 90 neoplasmas ósseos primários diagnosticados neste período, 89 eram malignos, sendo os osteossarcomas os mais prevalentes (86,7%). Em relação aos osteossarcomas, a maioria dos casos ocorreu em cães de raças grandes e gigantes e entre seis e 10 anos de idade. Os neoplasmas envolvendo o esqueleto apendicular predominaram e foram 3,5 vezes mais prevalentes nos membros anteriores que nos posteriores. O subtipo histológico predominante foi o osteoblástico. Para o estudo dos neoplasmas que compreenderam o segundo e o terceiro grupos, ou seja, neoplasmas com metástases ósseas ou o envolvimento ósseo em neoplasmas multicêntricos, foi realizado um estudo prospectivo durante um período de três anos. Neste período, cães provenientes do serviço de necropsias do LPV-UFSM foram avaliados. O esqueleto de 110 cães portadores de 118 neoplasmas malignos de diferentes origens foi examinado em busca de lesões ósseas. Foram encontrados vinte e um casos de metástases ou neoplasmas multicêntricos com envolvimento ósseo (19,1%). Em geral, as lesões ósseas afetaram mais as fêmeas. No entanto, quando os neoplasmas originados na glândula mamária foram desconsiderados, a distribuição dos casos de acordo com o sexo foi muito semelhante. Foram afetados cães com idade média de 9 anos e de diferentes raças. A glândula mamária foi a origem da maioria dos tumores que metastatizaram para os ossos, seguida do sistema músculo-esquelético e respiratório. A maioria das metástases foi detectada macroscopicamente e ocorreu em múltiplos ossos. Entretanto, em 23% dos casos as metástases só puderam ser observadas microscopicamente. Dentre os ossos afetados, vértebras e úmero foram os mais frequentemente acometidos. Paralelamente foram estudados, de forma retrospectiva, sete casos de osteopatia hipertrófica diagnosticados em um período de 11 anos no LPV-UFSM. Os cães afetados apresentavam sinais clínicos indicativos de envolvimento ósseo e lesões macroscópicas principalmente nos ossos longos dos membros. As lesões consistiram de neo-formação óssea periosteal, detectada em exame radiográfico, na inspeção óssea durante a necropsia e, com grande nível de detalhamento, em espécimes ósseos macerados. A proliferação óssea observada era parcialmente circunferencial e ocorreu principalmente na diáfise dos ossos longos. Era constituída por trabéculas ósseas de tamanho e espessura irregulares que estavam dispostas de forma perpendicular ao córtex ósseo original. Em todos os casos, as lesões de osteopatia hipertrófica foram associadas a neoplasmas pulmonares (primários ou metastáticos). Em dois dos sete casos, as metástases pulmonares eram de sarcomas ósseos e, em um caso, havia um osteossarcoma primário pulmonar (extraesquelético).

BACKGROUND: Scythe/BAT3 is a member of the BAG protein family whose role in apoptosis has been extensively studied. However, since the developmental defects observed in Bat3-null mouse embryos cannot be explained solely by defects in apoptosis, we investigated whether BAT3 is also involved in cell-cycle progression. METHODS/PRINCIPAL FINDINGS: Using a stable-inducible Bat3-knockdown cellular system, we demonstrated that reduced BAT3 protein level causes a delay in both G1/S transition and G2/M progression. Concurrent with these changes in cell-cycle progression, we observed a reduction in the turnover and phosphorylation of the CDK inhibitor p21, which is best known as an inhibitor of DNA replication; however, phosphorylated p21 has also been shown to promote G2/M progression. Our findings indicate that in Bat3-knockdown cells, p21 continues to be synthesized during cell-cycle phases that do not normally require p21, resulting in p21 protein accumulation and a subsequent delay in cell-cycle progression. Finally, we showed that BAT3 co-localizes with p21 during the cell cycle and is required for the translocation of p21 from the cytoplasm to the nucleus during the G1/S transition and G2/M progression. CONCLUSION: Our study reveals a novel, non-apoptotic role for BAT3 in cell-cycle regulation. By maintaining a low p21 protein level during the G1/S transition, BAT3 counteracts the inhibitory effect of p21 on DNA replication and thus enables the cells to progress from G1 to S phase. Conversely, during G2/M progression, BAT3 facilitates p21 phosphorylation by cyclin A/Cdk2, an event required for G2/M progression. BAT3 modulates these pro- and anti-proliferative roles of p21 at least in part by regulating cyclin A abundance, as well as p21 translocation between the cytoplasm and the nucleus to ensure that it functions in the appropriate intracellular compartment during each phase of the cell cycle. / Dissertation