O objetivo deste trabalho foi avaliar os indicadores de prognóstico para o mastocitoma canino, na tentativa de melhorar o tratamento oferecido aos animais acometidos por esta neoplasia. Foram analisados os parâmetros clínicos dos animais e do mastocitoma, parâmetros de evolução da doença e parâmetros histológicos, imunoistoquímicos, estereológicos e de expressão gênica. Para isso, 81 cães portadores de mastocitoma cutâneo foram submetidos à intervenção cirúrgica para excisão dos tumores. Após a operação os cães foram acompanhados e avaliados por um período mínimo de 12 meses para a colheita de dados sobre a evolução. Os tumores foram graduados por histopatologia, tiveram seus bordos investigados quando a eficiência da cirurgia e foram submetidos à imunomarcação com ki-67, c-kit. Destes, 20 casos passaram por avaliação estereológica, nos parâmetros volume total do tumor, densidade numérica, número total de mastócitos, volume de célula e de núcleo e a relação núcleo/célula. Além disso, 22 casos tiveram seus tumores quantificados quanto a expressão gênica de c-kit e do seu ligante. Os parâmetros clínicos avaliados foram: idade, sexo, raça, localização, tempo de evolução, tamanho do tumor, velocidade de crescimento, presença de ulceração, sangramento, eritema, temperatura, hiperpigmentação, consistência tumoral, base de inserção, abrangência de tecido, aderência a planos profundos, formato, superfície, presença de alopecia, número de tumores, além de investigar a presença de alteração em linfonodos, sinais clínicos e metástases e calcular a taxa de crescimento tumoral. Foram associados a piora do prognóstico os pacientes que apresentaram os parâmetros: animais idosos, presença de eritema, ulceração, aumento de temperatura, presença de aderência, superfície irregular, tumores firmes, crescimento rápido, presença de sinais clínicos e metástases. O diâmetro foi associado a maior taxa de óbito. As demais variáveis clínicas não foram associadas ao prognóstico. O grau histopatológico, a presença de sinais clínicos e metástase foram considerados marcadores preditivos independentes de recidiva e óbito, pela análise multivariada. Na avaliação por estereologia, as variáveis: volume do tumor e número total de mastócitos neoplásicos também foram associados à redução de sobrevida livre de recidiva, assim como uma maior porcentagem de núcleos imunomarcados com ki-67. A diminuição da expressão do ligante de c-kit, também conhecido como fator de crescimento do c-kit também foi associado aos eventos combinados óbito e recidiva. Os resultados demonstraram que mesmo parâmetros clínicos, de fácil investigação fornecem dados para a interpretação dos mastocitomas de grau intermediário, podendo oferecer melhor tratamento aos animais acometidos. Os parâmetros laboratoriais como imunomarcação com ki-67, parâmetros estereológicos podem ajudar a estabelecer uma nova classificação destes tumores que ficam à margem de classificação. Porém, mais uma vez a graduação preconizada por Patnaik e al. (1984) se mostrou de grande eficiência na previsão da evolução dos mastocitomas. As expressões gênicas de c-kit e seu ligante e a expressão por imunomarcação de c-kit devem ser melhores avaliadas e comparadas a análises de mutação destes genes. / The aim of this study was to evaluate prognostic markers for canine mast cell tumors to improve treatment options offered to animals affected by this cancer. We evaluated animals and clinical parameters of the tumors, parameters from clinical outcomes, and histological, immunohistochemical, stereological, and gene expression parameters. To this end, 81 dogs with cutaneous mast cell tumors underwent surgical excision of tumors. After surgery, the dogs were monitored and evaluated for a minimum of 12 months for the collection of outcome data. The tumors were graded histologically, had their surgical margins investigated regarding the efficacy of surgery, and were immunostained with Ki-67 and c-kit. Of these specimens, 20 cases underwent stereological assessment: total tumor volume, numerical density, total number of mast cells, cell and nucleus volume, and the nucleus/cell relationship were investigated. In addition, gene expression of c-kit and its ligand was measured in tumors from 22 cases. Examined clinical parameters included age, sex, breed, location, duration of evolution, tumor size, growth rate, presence of ulceration, bleeding, erythema, temperature, hyperpigmentation, tumor consistency, insertion base, range of tissues, adherence to deep levels, shape, surface, presence of alopecia, and number of tumors. We also investigated the presence of changes in lymph nodes, metastases, clinical signs and metastases. We observed that tumors with erythema, ulceration, increased temperature, adherence, irregular surface, solidity, rapid growth, presence of metastases and clinical signs were associated with worse prognostic. A larger tumor diameter was associated with a higher death rate. The other clinical variables were not associated with the prognosis. A multivariate analysis revealed that the histopathological grade and the presence of clinical signs and metastasis were independent predictive markers of the progression-free survival time. The stereological analysis indicated that a larger tumor volume, a larger total number of neoplastic mast cells, and a higher percentage of nuclei immunostained for Ki-67 were also associated with a reduced survival time and a higher recurrence rate. The reduction of c-kit ligand expression, also called the growth factor c-kit, was also associated with the death and recurrence rates. Our results demonstrate that clinical parameters that are easy to determine can provide useful data for assessing intermediate-grade mast-cell tumors and thereby be useful in determining better treatments for affected animals. Stereological parameters and laboratory parameters such as immunostaining for Ki-67 could help to establish a new classification of those tumors that are at the margin of existing classifications. However, the histopathological classification established by Patnaik et al. (1984) proved very efficient in predicting the evolution of mast-cell tumors. The gene expressions of c-kit and its ligand, and the KIT immunostaining should be further evaluated and compared in order to analyze mutations in these genes.

Mastzellen sind ubiquitär im Bindegewebe vorkommende Zellen, welche eine Vielzahl von Mediatoren physiologischer und pathologischer Prozesse exprimieren. Ihre Zahl ist in gesunden Geweben gering, am höchsten jedoch in der Haut und Schleimhäuten von Nase, Auge und Gastrointestinaltrakt sowie in der Lunge. Die Ergebnisse unserer Arbeitsgruppe zeigen, dass unter pathologischen Bedingungen, insbesondere bei entzündlichen Reaktionen und Erkrankungen des atopischen Formenkreises, die Mastzellzahlen um ein Vielfaches steigen. Dabei werden die Vorläuferzellen des Blutes, welche aus einer CD34+/c-Kit+ hämatopoetischen Stammzelle des Knochenmarkes rekrutiert werden, aktiviert und durch chemotaktisch wirkende Faktoren zur Einwanderung in die Gewebe stimuliert. Unter Verwendung von in vitro Kulturmodellen konnte gezeigt werden, dass diese Vorläuferzellen im Blut die Expression von c-Kit und CD34 herunterregulieren und als Zellpool im peripheren Blut zirkulieren. Nach Aktivierung der Zellen wurde c-Kit wieder nachgewiesen. Die Zellen wandern ins Gewebe ein und differenzieren dort unter Einfluss von Zytokinen, welche durch andere Gewebszellen freigesetzt werden, aus. Es wurden Wechselwirkungen in der Haut zwischen Mastzellen und Fibroblasten, Melanozyten, Keratinozyten und Nervenzellen gezeigt. Als Mittler dieser Wechselwirkungen konnten, neben den in der Literatur beschriebenen Faktoren, von uns SCF, GM-CSF, NGF und andere Neurotrophine zum Beispiel BDNF, NT-3 und NT-4 gezeigt werden. Die Regulation und Freisetzung dieser Faktoren ist bei pathologisch veränderter Haut, wie bei Atopischer Dermatitis, Psoriasis, Haarwachstumsstörungen, Tumoren der Haut und in der Wundheilung verändert. Die Modulation der Expression dieser Faktoren und ihrer Rezeptoren durch verschiedene Therapeutika, wie Glukokortikoide, Antihistaminika, Retinoide und UV-Bestrahlung konnte in verschiedenen Kulturmodellen gezeigt werden. Diese Erkenntnisse können in Zukunft bei der Entwicklung neuer Therapeutika zur Behandlung von verschiedenen Erkrankungen der Haut, Lunge sowie Darm und, da in zunehmendem Maße auch von Mastzellfunktionen in anderen Organen, wie Hirn, Herz, Leber und Niere berichtet wird, auch hier weiterführend beitragen. / Mast cells are ubiquitary connective tissue cells derived from bone marrow CD34+/c-Kit+ stem cells. They are able to produce various regulators of physiological and pathological processes. Normally, they are present in low numbers with highest density in skin and nasal, ophthalmic, gastrointestinal and pulmonary mucosa. The number is increased up to 100-fold in various pathological processes as inflammatory reactions and atopic diseases. During this processes mast cell precursors from peripheral blood are activated, migrate in the tissues by the effects of chemotactically acting factors. In order to elucidate the mechanisms involved in this processes, we established different in vitro cell culture models. Our results suggest that the precursor cells circulating in the peripheral blood do not express c-Kit. When the cells are activated, c-Kit expression is upregulated and the cells are able to migrate in the tissue, where they differentiate influenced by cytokines released from tissue cells. The interaction between mast cells and fibroblasts, melanocytes, keratinocytes and nerve cells were studied. Stem cell factor, GM-CSF, nerve growth factor and other neurotrophins as BDNF, NT-3 and NT-4 could be demonstrated as mediators of this interactions. The regulation and release of these factors are modified in pathological skin diseases as atopic dermatitis, psoriasis, changes in the hair cycle and skin tumors and in wound healing. Modulation of expression of these factors and its receptors by therapeutics as glucocorticoids, antihistamines, retinoids and UV-radiation was shown in different culture models. Our results may contribute to develop new therapeutics for skin, pulmonary and intestinal diseases and give new insights in mast cell functions in brain, liver and kidney.

Die Mechanismen der Mastzell(MZ)-Homöostase in peripheren Geweben sind weitgehend unbekannt. Die Regulation der MZ-Zahlen schließt wahrscheinlich die lokale Proliferation der Gewebe-MZ sowie die Rekrutierung und Differenzierung von MZ-Vorläuferzellen ein, wobei das Gewebe den MZ-Phänotyp beeinflusst. Fibroblasten (Fb) induzieren die Proliferation und Differenzierung unreifer Knochenmark-generierter MZ (BMCMC) zu Bindegewebs-MZ (CTMC). Da BMCMC an Fb adhärieren, wurde untersucht ob dieser direkte Zellkontakt für die Fb-induzierte Proliferation und die Differenzierung zu CTMC entscheidend ist. Cokultur-Experimente mit BMCMC und Fb zeigten, dass die verstärkte Proliferation, der erhöhte Histamingehalt und die Induktion der Expression der MZ Protease 4 (MCPT-4) mRNA (beides Marker der CTMC) vom direkten Zellkontakt abhängen. Adhäsionsversuche mit blockierenden Antikörpern demonstrierten, dass die Interaktion von alpha4beta7 Integrin auf den BMCMC mit Vascular Cell Adhesion Molecule 1 (VCAM-1), auf den Fb, an der Adhäsion beteiligt ist, die Proliferation und Differenzierung der MZ selbst jedoch nicht auslöst. Interessanterweise zeigten BMCMC die Kit, den Rezeptor für den wichtigen MZ-Wachstumsfaktor Stammzellfaktor (SCF), nicht exprimieren, ebenfalls die kontaktabhängigen Fb-induzierte Veränderungen, wenngleich im geringeren Ausmaß. Eine genomweite Expressionsanalyse wies schließlich die kontaktabhängige Hochregulation der Expression weiterer Gene, die mit dem Phänotyp der CTMC assoziiert sind, nach. Des Weiteren konnte die Verringerung der Expression bestimmter Gene gezeigt werden, die von unreifen MZ oder MZ-Vorläufern exprimiert werden. Insgesamt zeigen die Daten der hier vorliegenden Arbeit, dass für die durch Fb ausgelöste umfassenden Differenzierung und Ausreifung von unreifen MZ zu CTMC, eine teilweise durch VCAM-1/alpha4beta7 vermittelte direkte Zell-Zell-Interaktion notwendig ist, bei der sowohl Kit-abhängige, als auch Kit-unabhängige Signalwege involviert sind. / The precise mechanisms of mast cell (MC) homeostasis in peripheral tissues are largely unknown. Regulation of MC numbers is assumed to involve the proliferation of local MCs and the recruitment and subsequent differentiation of MC precursors, whereas the tissues determine the MC phenotype. Fibroblasts (Fb) induce proliferation and differentiation of bone marrow-derived cultured MCs (BMCMCs) towards connective tissue type MCs (CTMCs). Since BMCMCs exhibit adhesion to Fbs, it had been analyzed, whether this direct cell-to-cell crosstalk is mandatory for the differentiation towards CTMCs. It was found that Fb-cocultured immature MCs exhibit increased proliferation and histamine content and the induced expression of mast cell protease 4 (mcpt4) mRNA, both markers for mature CTMC, and that these changes required a directed cell-to-cell interaction. Adhesion Assays using blocking mAbs revealed that the interaction of alpha4beta7 integrin on BMCMCs with Fb-expressed Vascular Cell Adhesion Molecule 1 (VCAM-1) is largely responsible for the adhesion, which itself didn’t induce proliferation and differentiation of MCs. Most notably, MCs deficient for Kit, the receptor for the MC growth factor stem cell factor (SCF), also showed a significant Fb-induced increase in histamine content, mcpt4 expression, and proliferation, albeit to a lesser extent than wildtype BMCMCs. Whole genome expression analysis showed contact-dependently upregulated expression of several genes associated with CTMC phenotype and functions. Furthermore, downregulation of genes associated with MC progenitors had been shown. Collectively, the data show that the Fb-induced substantial differentiation of immature MCs towards CTMCs requires a partly VCAM-1/alpha4beta7-mediated cell-to-cell interaction and involves both Kit-dependent and -independent signaling pathways.

O objetivo deste trabalho foi avaliar os indicadores de prognóstico para o mastocitoma canino, na tentativa de melhorar o tratamento oferecido aos animais acometidos por esta neoplasia. Foram analisados os parâmetros clínicos dos animais e do mastocitoma, parâmetros de evolução da doença e parâmetros histológicos, imunoistoquímicos, estereológicos e de expressão gênica. Para isso, 81 cães portadores de mastocitoma cutâneo foram submetidos à intervenção cirúrgica para excisão dos tumores. Após a operação os cães foram acompanhados e avaliados por um período mínimo de 12 meses para a colheita de dados sobre a evolução. Os tumores foram graduados por histopatologia, tiveram seus bordos investigados quando a eficiência da cirurgia e foram submetidos à imunomarcação com ki-67, c-kit. Destes, 20 casos passaram por avaliação estereológica, nos parâmetros volume total do tumor, densidade numérica, número total de mastócitos, volume de célula e de núcleo e a relação núcleo/célula. Além disso, 22 casos tiveram seus tumores quantificados quanto a expressão gênica de c-kit e do seu ligante. Os parâmetros clínicos avaliados foram: idade, sexo, raça, localização, tempo de evolução, tamanho do tumor, velocidade de crescimento, presença de ulceração, sangramento, eritema, temperatura, hiperpigmentação, consistência tumoral, base de inserção, abrangência de tecido, aderência a planos profundos, formato, superfície, presença de alopecia, número de tumores, além de investigar a presença de alteração em linfonodos, sinais clínicos e metástases e calcular a taxa de crescimento tumoral. Foram associados a piora do prognóstico os pacientes que apresentaram os parâmetros: animais idosos, presença de eritema, ulceração, aumento de temperatura, presença de aderência, superfície irregular, tumores firmes, crescimento rápido, presença de sinais clínicos e metástases. O diâmetro foi associado a maior taxa de óbito. As demais variáveis clínicas não foram associadas ao prognóstico. O grau histopatológico, a presença de sinais clínicos e metástase foram considerados marcadores preditivos independentes de recidiva e óbito, pela análise multivariada. Na avaliação por estereologia, as variáveis: volume do tumor e número total de mastócitos neoplásicos também foram associados à redução de sobrevida livre de recidiva, assim como uma maior porcentagem de núcleos imunomarcados com ki-67. A diminuição da expressão do ligante de c-kit, também conhecido como fator de crescimento do c-kit também foi associado aos eventos combinados óbito e recidiva. Os resultados demonstraram que mesmo parâmetros clínicos, de fácil investigação fornecem dados para a interpretação dos mastocitomas de grau intermediário, podendo oferecer melhor tratamento aos animais acometidos. Os parâmetros laboratoriais como imunomarcação com ki-67, parâmetros estereológicos podem ajudar a estabelecer uma nova classificação destes tumores que ficam à margem de classificação. Porém, mais uma vez a graduação preconizada por Patnaik e al. (1984) se mostrou de grande eficiência na previsão da evolução dos mastocitomas. As expressões gênicas de c-kit e seu ligante e a expressão por imunomarcação de c-kit devem ser melhores avaliadas e comparadas a análises de mutação destes genes. / The aim of this study was to evaluate prognostic markers for canine mast cell tumors to improve treatment options offered to animals affected by this cancer. We evaluated animals and clinical parameters of the tumors, parameters from clinical outcomes, and histological, immunohistochemical, stereological, and gene expression parameters. To this end, 81 dogs with cutaneous mast cell tumors underwent surgical excision of tumors. After surgery, the dogs were monitored and evaluated for a minimum of 12 months for the collection of outcome data. The tumors were graded histologically, had their surgical margins investigated regarding the efficacy of surgery, and were immunostained with Ki-67 and c-kit. Of these specimens, 20 cases underwent stereological assessment: total tumor volume, numerical density, total number of mast cells, cell and nucleus volume, and the nucleus/cell relationship were investigated. In addition, gene expression of c-kit and its ligand was measured in tumors from 22 cases. Examined clinical parameters included age, sex, breed, location, duration of evolution, tumor size, growth rate, presence of ulceration, bleeding, erythema, temperature, hyperpigmentation, tumor consistency, insertion base, range of tissues, adherence to deep levels, shape, surface, presence of alopecia, and number of tumors. We also investigated the presence of changes in lymph nodes, metastases, clinical signs and metastases. We observed that tumors with erythema, ulceration, increased temperature, adherence, irregular surface, solidity, rapid growth, presence of metastases and clinical signs were associated with worse prognostic. A larger tumor diameter was associated with a higher death rate. The other clinical variables were not associated with the prognosis. A multivariate analysis revealed that the histopathological grade and the presence of clinical signs and metastasis were independent predictive markers of the progression-free survival time. The stereological analysis indicated that a larger tumor volume, a larger total number of neoplastic mast cells, and a higher percentage of nuclei immunostained for Ki-67 were also associated with a reduced survival time and a higher recurrence rate. The reduction of c-kit ligand expression, also called the growth factor c-kit, was also associated with the death and recurrence rates. Our results demonstrate that clinical parameters that are easy to determine can provide useful data for assessing intermediate-grade mast-cell tumors and thereby be useful in determining better treatments for affected animals. Stereological parameters and laboratory parameters such as immunostaining for Ki-67 could help to establish a new classification of those tumors that are at the margin of existing classifications. However, the histopathological classification established by Patnaik et al. (1984) proved very efficient in predicting the evolution of mast-cell tumors. The gene expressions of c-kit and its ligand, and the KIT immunostaining should be further evaluated and compared in order to analyze mutations in these genes.

Pulmonary arterial hypertension (PAH) is a group of diseases characterized by avascular obstruction leading to a progressive increase of the resistances in the pulmonary blood flow. The recent progress in the understanding of mechanisms at the origin of this disease underlines the role of extrapulmonary cells, such as circulating stem cells and bone marrow-derived progenitor cells in vascular remodeling and in PAH development. In this thesis we have shown implication of the progenitor cells and chemotactic axis in the vascular remodeling in human and experimental PAH. This work could help to develop new therapies allowing more specific and more effective treatments leading to improved survival of PAH patients. / L’hypertension artérielle pulmonaire (HTAP) est un groupe de maladies qui se caractérise par une obstruction vasculaire conduisant à une augmentation progressive des résistances à l’écoulement sanguin. Le remodelage vasculaire qui implique toutes les couches de la paroi du vaisseau est considéré comme un élément clé dans la pathogenèse de l'HTAP. Les progrès récents dans la compréhension des mécanismes à l'origine de cette maladie soulignent le rôle de cellules extrapulmonaires, telles que des cellules souches circulantes et des progéniteurs dérivés de la moelle osseuse, dans le remodelage vasculaire et dans le développement de l’HTAP. Les thérapeutiques ciblées sur la dysfonction endothéliale ne permettent pas à l'heure actuelle de guérir cette maladie, il est donc nécessaire d’identifier d'autres mécanismes physiopathologiques permettant de développer de nouvelles stratégies thérapeutiques. C’est pourquoi nous avons exploré l’implication des cellules progénitrices et des signaux chimiotactiques dans le remodelage vasculaire au cours de l’HTAP humaine et expérimentale. Nous avons mis en évidence un recrutement de cellules c-kit+ (incluant des progéniteurs et des mastocytes) ainsi que l’expansion de vasa vasorum dans les poumons des patients atteints d’HTAP. Grâce à l’utilisation du modèle expérimentale d’HTAP induit par l’hypoxie chez la souris, nous avons montré le rôle central de la chimiokine CXCL12 et de ses deux récepteurs CXCR4 et CXCR7 dans le recrutement des progéniteurs c-kit+. Le traitement combiné par les antagonistes AMD3100 et CCX771, grâce à leurs actions synergiques, inhibe le remodelage vasculaire pulmonaire, l’hypertrophie cardiaque droite, ainsi que le recrutement des progéniteurs c-kit+, induits par l’hypoxie. Par ailleurs, le blocage de c-kit par l’imatinib améliore également les paramètres d’hémodynamique et diminue le recrutement périvasculaire des cellules c-kit+, probablement en inhibant leurs expansion dans la moelle osseuse. Nous avons également mis en évidence une altération d’une population de progéniteurs mésenchymateux d’origine hématopoïétique, appelés fibrocytes, dans le sang des patients souffrant d’HTAP.Ce travail pourrait contribuer à développer des actions thérapeutiques ciblées permettant la mise en place de traitements à la fois plus spécifiques et plus efficaces susceptibles d’améliorer la survie des patients HTAP.

O objetivo deste trabalho foi avaliar a eficácia do mesilato de imatinibe, em comparação com a quimioterapia usual com vimblastina e prednisona, no tratamento do mastocitoma canino e descrever os efeitos colaterais apresentados pelas medicações. Bem como analisar a expressão do VEGF (fator de crescimento endotelial), a relação da expressão do gene c-kit por RT-PCR e marcação imunoistoquímica do KIT com a presença de mutações na justamembrana e a relação desta mutação com a resposta à terapia. Para tanto foram incluídos 29 animais com diagnóstico citológico de mastocitoma, estes animais foram submetidos a tomografia computadorizada para determinação das medidas das formações cutâneas e em seguida divididos em 2 grupos. O grupo 1 foi tratado com o protocolo quimioterápico de vimblastina e prednisona por 12 semanas e o grupo 2 com o mesilato de imatinibe na dose de 10 mg/Kg a cada 24 horas por 8 semanas. A avaliação da resposta ao tratamento foi realizada com mensurações periódicas das formações com paquímetro e nova tomografia ao final do tratamento para mensuração do maior diâmetro e volume tumoral. Um fragmento das formações cutâneas foi coletado antes do início do tratamento para graduação histológica da neoplasia, determinação do índice mitótico e imunomarcação para KIT, VEGF e Ki- 67. Parte do material coletado teve o DNA e RNA extraídos e posterior sequenciamento dos exons 11 do gene c-kit e determinação da expressão deste e do seu ligante por RT-PCR. A toxicidade a medicação foi avaliada segundo as normas do VCOG 1.1.A taxa de resposta do grupo VP foi de 7,7 % e no grupo MI de 28,6%, embora os pacientes tratados com mesilato de imatinibe tenham apresentado maior chance de resposta a terapia, não foi observado diferença entre os dois grupos. Os dois protocolos foram bem tolerados, os pacientes do grupo MI d menor número de efeitos colaterais. O grau histológico, Indice mitótico, padrão imunohistoquimico do KIT, além da quantificação do ki-67 foram homogêneos nos dois grupos e não influenciaram na resposta ao tratamento. A quantificação do VEGF foi mais intensa nos pacientes com remissão parcial e total. Não foi observado relação entre a quantificação do KIT e a expressão do gene c-kit, que foi maior nos pacientes que responderam ao tratamento, porém a associação desta com a resposta a terapia não pode ser determinada. Mutações ativantes no exon11 do gene c-kit não foram identificadas. O tratamento com o mesilato de imatinibe é bem tolerado pelos animais, no entanto este não se mostrou superior ao protocolo padrão de quimioterapia para o tratamento do mastocitoma; este resultado pode ter sido influenciado pelo número de animais incluídos no estudo. Mutações em outros domínios do receptor KIT e a ação do ITK em receptores como do PDGF e o VEGF podem estar relacionados a resposta a esta classe de fármacos observada neste estudo, a despeito da ausência de mutações ativantes no exon 11 do gene c-kit. / The objective of this study was to evaluate the efficacy of imatinib mesylate, compared with the usual chemotherapy with vinblastine and prednisone in the treatment of canine mast cell tumor and describe the side effects submitted by medications. Well as analyzing the expression of VEGF (vascular endothelial growth factor), the relationship between the expression of c-kit gene by RT-PCR and immunohistochemical staining of KIT with the presence of mutations in the juxtamembrane and the relationship of this mutation with response to therapy. For both 29 animals with cytological diagnosis of mast cell tumor were included, these animals underwent computed tomography to determine the measured skin formations and then divided into 2 groups. Group 1 was treated with the chemotherapeutic protocol vinblastine and prednisone for 12 weeks and the second group with the imatinib mesylate in a dose of 10 mg / kg every 24 hours for 8 weeks. The assessment of response to treatment was performed with periodic measurements of the formations Caliper and a new computed tomography in the end of treatment to measure the largest tumor diameter and volume. A fragment of skin formations was collected before the initiation of treatment for histological grading, determination of mitotic index, KIT and VEGF staining patterns and the proliferation marker Ki67. Part of the collected material was extracted RNA and DNA and subsequent sequencing of 11 exons of the c-kit gene and determination and expression of its ligand by RT-PCR. The medication toxicity was evaluated according to the standards of VCOG 1.1.A response rate of the VP group was 7.7% and 28.6% MI group, although patients treated with imatinib had a higher chance of response therapy, no difference in response between the two groups was observed. The two protocols were well tolerated, patients in the MI group had a smaller number of side effects. The histological grade, mitotic index, staining patterns KIT, beyond the quantification of Ki-67 were homogeneous in both groups and did not influence the response to treatment. Quantification of VEGF was intensely in patients with partial and total remission. It was no relationship between KIT and quantification of the expression of c-kit gene, which was higher in patients who responded to treatment, but its association with response to therapy cant be determined. Exon11 activating mutations in the c-kit gene were not identified. Treatment with imatinib mesylate is well tolerated by the animals, however this was not superior to standard chemotherapy protocol for the treatment of mast cell tumors; this result may have been influenced by the number of animals included in the study. Mutations in other domains of the KIT receptor and action in ITK receptors as PDGF and VEGF may be related to response to this class of drugs in this study, despite the absence of activating mutations in exon 11 of c-kit gene.

No description available.

No description available.

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Diss. (sammanfattning) Stockholm : Karol. inst., 2002. / Härtill 4 uppsatser.