It is the purpose of this investigation to determine the variation of glyoxalase activity in liver of normal mice and in the liver of mice bearing a lymphosarcoma and to compare the glyoxalase activity of the lymphosarcoma with values previously reported in the literature for other tumor types. Further, if there is indeed a variation in liver glyoxalase activity between normal and tumor-bearing mice, it will be compared to the variation in the activity of two other enzymes present in liver tissue in relatively high concentration.

The problem with which this investigation is concerned is that of determining the nature of events leading to the change of normal cells into malignant cells. The design of the study is multi-phasic: (A) to establish the presence or absence of an oncongenic virion, (B) to demonstrate by use of the electron microscopy any ultracellular alteration in malignant or transformed tissues, (C) to investigate the nature of the transforming agent in the murine lymphosarcoma, and (D) to employ various methods to demonstrate cellular transformations in vivo and in vitro.

Calcium/phospholipid-dependent protein kinase (PKC) was purified from P1798 lymphosarcoma. It was demonstrated that uncomplexed calcium and uncomplexed phosphatidylserine are the activators, that the activation of PKC requires that calcium bind first, that the activation of PKC requires that calcium bind first, that high calcium concentrations inhibit PKC activation, and that calcium inhibition can be overcome by phosphatidylserine.

This investigation was concerned with characterizing effects of this tumor line on lipid metabolism in DBA/lJ mice and serum protein levels and cellular changes in DBA/lJ and Swiss white mice. Total lipids, lipid phosphorus, neutral lipids, and changes in fatty acids were determined in liver, spleen, skin, and tumor of DBA/lJ mice bearing the lymphosarcoma at various days after injection of tumor cells.

The investigation was concerned with characterizing effects of this tumor line on lipid metabolism in DBA/1J mice and serum protein levels and cellular changes in DBA/1J and Swiss white mice. Total lipids, lipid phosphorus, neutral lipids, and changes in fatty acids were determined in liver, spleen, skin and tumor of DBA/1J mice bearing the lymphosarcoma at various days after injection of tumor cells.

The purpose of this investigation was (1) to isolate and purify glyoxalase I from the livers of normal DBA/lJ mice and the livers from mice bearing a lymphosarcoma tumor; and (2) to determine, at least with respect to glyoxalase I, if the tumor has an effect on the chemical properties or structure of macromolecules in an organ removed from tumor locale and not histologically affected by its presence.

The problem with which this investigation is concerned is that of determining the nature of events leading to the change. of normal cells into malignant cells. The design of the study is multi-phasic: (A) to establish the presence or absence of an oncogenic virion, (B) to demonstrate by use of the electron microscopy any ultracellular alteration in malignant or transformed tissues, (C) to investigate the nature of the transforming agent in the murine lymphosarcoma, and (D) to employ various methods to demonstrate cellular transformations in vivo and in vitro. It is concluded that the transforming and tumorinducing agent in this' investigation was not a virion, but an infectious ribonucleic acid genome or a segment of a viral genome which had become integrated into the genome of the mouse cells. The vision has lost its ability to form a protein coat; therefore it is not demonstrable as a virion. But the ribonucleic acid is able to infect other cells and transform them from normal to neoplastic tissues.

Resumo: Sabendo-se da influência das mutações no gene TP53 no desenvolvimento das neoplasias e da discrepância entre os resultados obtidos pelas técnicas de sequenciamento e imunoistoquímica, esta pesquisa teve como objetivo avaliar a sequência do TP53 e a imunorreatividade da p53, relacionando-as com fatores clínicopatológicos em cães com linfoma. Foram avaliados 12 cães com linfoma e cinco cães sadios. Amostras de neoplasia/linfonodo e de bulbo piloso foram obtidas. O diagnóstico histopatológico foi determinado pela classificação de Kiel. O imunofenótipo e a imunomarcação da p53 foram determinados por imunoistoquímica. Para a imunomarcação com a p53, utilizou-se anticorpo policlonal anti-p53 (CM1) na diluição de 1:500. A região do gene TP53 compreendida entre os exons quatro e nove (incluindo íntrons) foi amplificada por PCR e submetida ao sequenciamento. A média da imunorreatividade da p53 nos cães com linfoma foi 27,33%±12,14%, diferente da média do grupo controle (0,75±0,5) (P<0,05). A ausência de marcação da p53 foi observada em apenas um caso de linfoma de baixo grau de malignidade, cujo cão apresentou um ano de sobrevida. O outro caso de linfoma de baixo grau de malignidade apresentou a menor marcação da p53 (14,12%). O linfoma com maior marcação da p53 (50,56%) consistiu em um linfoma alimentar cujo cão evoluiu para o óbito no dia da consulta. As maiores médias de marcação foram observadas nos linfomas cutâneos, estádio clínico V, subestádio "b", imunofenótipo B e tempo de sobrevida de até trinta dias, sem diferença estatística (P>0,05). Apesar dos resultados obtidos pela imunoistoquímica, nenhuma mutação foi encontrada nas sequências analisadas ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: TP53 mutations are usually involved in cancer, but sequencing and immunohistochemistry results are often controversial. Thus, the aim of this study was to analyse both TP53 sequence and p53 immunostaining and to associate their findings to clinical-pathological features in dogs with lymphoma. Twelve dogs with lymphoma and five healthy dogs were included in this study. Tumor and hair sample was collected. Histopathological diagnosis was performed according to Kiel classification. Immunohistochemistry was performed to identify immunophenotype as well as p53 expression. Polyclonal antibody anti-p53 (CM1) was used at a 1:500 dilution. The region that encompasses exons 4-9 was amplified by means of PCR reactions and sequencing was then performed. p53 immunolabeling mean was 27,33%±12,14 in canine lymphoma group. It was statistically different from control group mean (0,75±0,5) (P<0,05). Only one p53-negative case was observed in a dog with low grade lymphoma, which presented one-year survival. Other low grade lymphoma presented lowest p53 immunoreactivity (14,12%). Highest reactivity (50,56%) was observed in dog with alimentary lymphoma, which died in the day of diagnosis. The highest immunolabeling means were verified in cutaneous presentation, clinical stage V, substage "b", immunophenotype B and survival time up to 30 days, but there was no significant difference (P>0,05). Nevertheless, gene mutations were not observed in any sequence. In conclusion, p53 immunostaining is not essentially related to mutation in the gene TP53, particularly to the region between exons 4-9 / Orientador: Carlos Roberto Daleck / Coorientador: Janete Apparecida Desidério Sena / Banca: Andrigo Barboza De Nardi / Banca: Julio Lopes Sequeira / Banca: Renée Laufer Amorim / Banca: Manoel Victor Franco Lemos / Doutor

Made available in DSpace on 2014-06-11T19:31:09Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-09-18Bitstream added on 2014-06-13T19:20:13Z : No. of bitstreams: 1 calazans_sg_dr_jabo.pdf: 11128452 bytes, checksum: 6057c220293ff16d0d56a83a1af6d1e5 (MD5) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Sabendo-se da influência das mutações no gene TP53 no desenvolvimento das neoplasias e da discrepância entre os resultados obtidos pelas técnicas de sequenciamento e imunoistoquímica, esta pesquisa teve como objetivo avaliar a sequência do TP53 e a imunorreatividade da p53, relacionando-as com fatores clínicopatológicos em cães com linfoma. Foram avaliados 12 cães com linfoma e cinco cães sadios. Amostras de neoplasia/linfonodo e de bulbo piloso foram obtidas. O diagnóstico histopatológico foi determinado pela classificação de Kiel. O imunofenótipo e a imunomarcação da p53 foram determinados por imunoistoquímica. Para a imunomarcação com a p53, utilizou-se anticorpo policlonal anti-p53 (CM1) na diluição de 1:500. A região do gene TP53 compreendida entre os exons quatro e nove (incluindo íntrons) foi amplificada por PCR e submetida ao sequenciamento. A média da imunorreatividade da p53 nos cães com linfoma foi 27,33%±12,14%, diferente da média do grupo controle (0,75±0,5) (P<0,05). A ausência de marcação da p53 foi observada em apenas um caso de linfoma de baixo grau de malignidade, cujo cão apresentou um ano de sobrevida. O outro caso de linfoma de baixo grau de malignidade apresentou a menor marcação da p53 (14,12%). O linfoma com maior marcação da p53 (50,56%) consistiu em um linfoma alimentar cujo cão evoluiu para o óbito no dia da consulta. As maiores médias de marcação foram observadas nos linfomas cutâneos, estádio clínico V, subestádio “b”, imunofenótipo B e tempo de sobrevida de até trinta dias, sem diferença estatística (P>0,05). Apesar dos resultados obtidos pela imunoistoquímica, nenhuma mutação foi encontrada nas sequências analisadas... / TP53 mutations are usually involved in cancer, but sequencing and immunohistochemistry results are often controversial. Thus, the aim of this study was to analyse both TP53 sequence and p53 immunostaining and to associate their findings to clinical-pathological features in dogs with lymphoma. Twelve dogs with lymphoma and five healthy dogs were included in this study. Tumor and hair sample was collected. Histopathological diagnosis was performed according to Kiel classification. Immunohistochemistry was performed to identify immunophenotype as well as p53 expression. Polyclonal antibody anti-p53 (CM1) was used at a 1:500 dilution. The region that encompasses exons 4-9 was amplified by means of PCR reactions and sequencing was then performed. p53 immunolabeling mean was 27,33%±12,14 in canine lymphoma group. It was statistically different from control group mean (0,75±0,5) (P<0,05). Only one p53-negative case was observed in a dog with low grade lymphoma, which presented one-year survival. Other low grade lymphoma presented lowest p53 immunoreactivity (14,12%). Highest reactivity (50,56%) was observed in dog with alimentary lymphoma, which died in the day of diagnosis. The highest immunolabeling means were verified in cutaneous presentation, clinical stage V, substage “b”, immunophenotype B and survival time up to 30 days, but there was no significant difference (P>0,05). Nevertheless, gene mutations were not observed in any sequence. In conclusion, p53 immunostaining is not essentially related to mutation in the gene TP53, particularly to the region between exons 4-9

Um mecanismo de tolerância periférica estudado é mediado por células T reguladoras (Tregs) que expressa o marcador CD25 e o fator de transcrição FoxP3. Estudamos o papel das Tregs nos fenômenos de imunossupressão na resistência e susceptibilidade genética dos hospedeiros contra o fungo. Animais A/J e B10.A foram depletados de células CD25 e outro grupo recebeu Ig de rato. Na 2ª semana de infecção, a depleção de células CD25 resultou em doença menos grave em ambas as linhagens com intenso afluxo de linfócitos T e macrófagos somente nos animais A/J. Na 10ª semana, a depleção de células CD25 também resultou em doença menos grave com aumento dos níveis de citocinas em ambas as linhagens. Neste período, entre os grupos controle, somente os animais A/J apresentaram aumento de linfócitos T e macrófagos. Em conclusão, os animais A/J apresentaram aumento da ativação da resposta imune enquanto que animais B10.A não desenvolveram eficiente ativação da resposta adaptativa mas que podem ter apresentado uma imunidade inata mais eficiente no controle do crescimento fúngico. / A mechanism for studying peripheral tolerance is mediated by regulatory T cells (Tregs) expressing the marker CD25 and the transcription factor FoxP3. We studied the role of Tregs in the mechanism of immunosuppression in genetic susceptibility and resistance of host against the fungus. A/J and B10.A mice were depleted of CD25 and control group received rat Ig. In the second week post infection, the depletion of CD25 cells resulted in low severity of disease in both mouse strains, with an intense influx of T lymphocytes and macrophages only in A/J. At 10 weeks, the depletion of CD25 also resulted in low severity disease with increased levels of cytokines in both mouse strains. Moreover, between the control group, only A/J mice showed an increase of T lymphocytes and macrophages. In conclusion, A/J mice showed increased activation of the immune response, whereas B10.A mice that had not developed efficient activation of the adaptive immune response, but that probably had an innate immunity that was more effective in controlling fungal growth.