RESUMO
Malignant melanomas are characterized by the ability of early metastatic dissemination to regional lymph nodes and the detection of sentinel lymph node metastases serves as an important prognostic parameter. There is clear evidence that melanoma cells and stromal cells of tumor environment can induce lymphangiogenesis, e.g. growth of lymphatic vessels, and this phenomenon is correlated with lymph node metastases. Vascular endothelial growth factor (VEGF) C represents the most potent and well-recognized lymphangiogenic growth factor secreted in tumor milieu by melanoma cells and tumor-associated macrophages, however the mechanism underlying VEGF-C secretion is not completely understood. We demonstrate that an acidic extracellular pH promotes the expression of VEGF-C in A375P melanoma cells and in melanoma cells isolated from a human spontaneous metastatic lesion, through the NF-κB transcription factor. We also demonstrate that esomeprazole, a proton pump inhibitor which requires acidosis to be activated, is able to prevent VEGF-C expression in acidic melanoma cells by interfering with NF-κB activation. Furthermore, we show that esomeprazole abrogates the enhanced VEGF-C expression in tumor cells grown in a acidic medium and stimulated by IL-1ß. On the whole, the present study reveals that acidity may be considered a strong promoter of VEGF-C expression in melanoma cells and provides a new pharmacological target to limit the development of tumor lymphangiogenesis.
Assuntos
Acidose , Neoplasias da Mama/patologia , Melanoma/patologia , NF-kappa B/metabolismo , Neoplasias da Próstata/patologia , Fator C de Crescimento do Endotélio Vascular/metabolismo , Western Blotting , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Esomeprazol/farmacologia , Feminino , Imunofluorescência , Humanos , Concentração de Íons de Hidrogênio , Técnicas Imunoenzimáticas , Masculino , Melanoma/genética , Melanoma/metabolismo , NF-kappa B/antagonistas & inibidores , NF-kappa B/genética , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Inibidores da Bomba de Prótons/farmacologia , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/efeitos dos fármacos , Células Tumorais Cultivadas , Fator C de Crescimento do Endotélio Vascular/genéticaRESUMO
Chronic myeloid leukaemia has a specific therapy: BCR/ABL inhibitor imatinib. Resistance due to BCR/ABL dependent and independent mechanisms is partially reversible by histone deacetylase inhibitors. We analysed by 2D-electrophoresis and anti-pan-acetylated and anti-phosphotyrosine immunoblots, followed by spot-matching and MALDI-TOF mass spectrometry, which proteome modifications would parallel restoration of sensitivity to imatinib by valproic acid (VPA). VPA plus imatinib significantly increased acetylation of HSP90 and hnRNP L and decreased phosphorylation of HSPs and hnRNPs in imatinib resistant cells. VPA was able to modify profoundly acetylome and phosphoproteome of CML cells, while reverting resistance to imatinib.
Assuntos
Resistencia a Medicamentos Antineoplásicos , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Fosfotirosina/metabolismo , Piperazinas/efeitos adversos , Proteoma/análise , Pirimidinas/efeitos adversos , Ácido Valproico/uso terapêutico , Acetilação , Apoptose , Benzamidas , Western Blotting , Proliferação de Células , Eletroforese em Gel Bidimensional , Proteínas de Fusão bcr-abl/genética , Proteínas de Fusão bcr-abl/metabolismo , Inibidores de Histona Desacetilases/uso terapêutico , Humanos , Mesilato de Imatinib , Imunoprecipitação , Inibidores de Proteínas Quinases/efeitos adversos , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Células Tumorais CultivadasRESUMO
Basal and H(2)O(2)-induced DNA breaks as well as DNA repair activity and efficacy of the antioxygenic system were determined in human dermal fibroblasts explanted from either (i) young donors and passaged serially to reach replicative senescence or (ii) young, old and centenarian donors and shortly propagated in culture. These fibroblasts have been employed as an in vitro and ex vivo model, respectively, to evaluate comparatively DNA integrity during senescence (increasing population doubling levels) and aging (increasing donor age). Constitutive levels of DNA total strand breaks, as determined by the alkaline extraction procedure, changed moderately among the different cell lines, which exhibited, however, significant differences in the amount of either single or double strand breaks. The former decreased along with both aging and senescence; the latter augmented during senescence while being virtually steady during aging. Moreover, fibroblasts from centenarians showed to be less sensitive to H(2)O(2)-induced DNA damage than other ex vivo fibroblasts. This feature could not account for either increased DNA repair activity or higher efficacy of the antioxygenic system and pointed, instead, to an intrinsic nuclear stability which might be typical of centenarian fibroblasts and potentially functional to longevity.