RESUMO
Many viral infections are associated with the development of immunopathologies and autoimmune diseases, which are of difficult treatment and for which no vaccines are yet available. Obtaining compounds that conjugate both antiviral and immunomodulatory activities in the same molecule would be very useful for the prevention and/or treatment of these immunopathologies. The compound (22S,23S)-22,23-dihydroxystigmast-4-en-3-one (compound 1) displays anti-Herpes simplex virus type 1 activity in vitro and reduces the incidence of herpetic stromal keratitis (HSK) in mice, a chronic inflammatory syndrome induced by ocular HSV-1 infection. In the present study, compound 1 showed opposite immunomodulatory properties in vitro. It induced the release of pro-inflammatory cytokines in HSV-1-infected epithelial cells of ocular origin, and significantly reduced the production of these cytokines in LPS-activated macrophages. RNA microarrays revealed various overexpressed and repressed genes in compound 1 treated infected epithelial cells and activated macrophages, many of which are associated with innate immune responses and inflammatory processes. These immunomodulatory properties of compound 1, together with its previously reported antiviral activity, make it a potential drug for the treatment of HSK and many other immunopathologies of viral and non-viral origin.
Assuntos
Antivirais/farmacologia , Colestenonas/química , Herpesvirus Humano 1/efeitos dos fármacos , Fatores Imunológicos/química , Estigmasterol/análogos & derivados , Animais , Antivirais/química , Antivirais/uso terapêutico , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Colestenonas/farmacologia , Colestenonas/uso terapêutico , Substância Própria/citologia , Substância Própria/virologia , Citocinas/genética , Citocinas/metabolismo , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Células Epiteliais/virologia , Perfilação da Expressão Gênica , Humanos , Fatores Imunológicos/farmacologia , Fatores Imunológicos/uso terapêutico , Ceratite Herpética/tratamento farmacológico , Ceratite Herpética/imunologia , Ceratite Herpética/veterinária , Camundongos , NF-kappa B/genética , NF-kappa B/metabolismo , Estigmasterol/química , Estigmasterol/farmacologia , Estigmasterol/uso terapêutico , Ativação Transcricional/efeitos dos fármacosRESUMO
The white coat colour of sheep is an important economic trait. For unknown reasons, some animals are born with, and others develop with time, black skin spots that can also produce pigmented fibres. The presence of pigmented fibres in the white wool significantly decreases the fibre quality. The aim of this work was to study gene expression in black spots (with and without pigmented fibres) and white skin by microarray techniques, in order to identify the possible genes involved in the development of this trait. Five unrelated Corriedale sheep were used and, for each animal, the three possible comparisons (three different hybridisations) between the three samples of interest were performed. Differential gene expression patterns were analysed using different t-test approaches. Most of the major genes with well-known roles in skin pigmentation, e.g. ASIP, MC1R and C-KIT, showed no significant difference in the gene expression between white skin and black spots. On the other hand, many of the differentially expressed genes (raw P-value < 0.005) detected in this study, e.g. C-FOS, KLF4 and UFC1, fulfil biological functions that are plausible to be involved in the formation of black spots. The gene expression of C-FOS and KLF4, transcription factors involved in the cellular response to external factors such as ultraviolet light, was validated by quantitative polymerase chain reaction (PCR). This exploratory study provides a list of candidate genes that could be associated with the development of black skin spots that should be studied in more detail. Characterisation of these genes will enable us to discern the molecular mechanisms involved in the development of this feature and, hence, increase our understanding of melanocyte biology and skin pigmentation. In sheep, understanding this phenomenon is a first step towards developing molecular tools to assist in the selection against the presence of pigmented fibres in white wool.
Assuntos
Perfilação da Expressão Gênica , Carneiro Doméstico/genética , Pigmentação da Pele/genética , Proteína Agouti Sinalizadora/genética , Animais , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Regulação da Expressão Gênica , Genes fos , Fator 4 Semelhante a Kruppel , Fatores de Transcrição Kruppel-Like/genética , Melanócitos/citologia , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Fenótipo , Proteínas Proto-Oncogênicas c-kit/genética , Receptor Tipo 1 de Melanocortina/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Carneiro Doméstico/fisiologia , Pele/citologia , Pigmentação da Pele/fisiologia , Fatores de Transcrição/genética , Enzimas de Conjugação de Ubiquitina/genética , Lã/fisiologiaRESUMO
Las nuevas técnicas de biología molecular aplicadas al diagnóstico genético y el uso de marcadores moleculares posibilitan el estudio de los mecanismos que subyacen en la predisposición individual y familiar a padecer determinadas enfermedades. Para llevar a cabo estos estudios, es necesario en primera instancia establecer cuál es la prevalencia de dichos marcadores en la población general. El estudio se realizó empleando una muestra de 108 individuos seleccionados por muestreo simple del banco de ADN de 500 individuos representativos de nuestra población, que pertenece al Departamento de Citogenética del Instituto de Investigaciones Biológicas Clemente Estable (IIBCE). Para establecer el genotipo del gen de la enzima convertidora de angiotensina (ECA) en cada una de las muestras, se amplificó un fragmento de ADN perteneciente al intrón 16 de este gen mediante la técnica de reacción en cadena de la polimerasa (PCR). El genotipo predominante en esta población control es el heterocigota I/D (50,9 por ciento), encontrándose el genotipo homocigota para la delección (D/D) (30,6 por ciento) en mayoría con respecto al genotipo homocigota para la inserción (I/I) (18,5 por ciento). Los resultados sugieren que existe, por tanto, un predominio del alelo D con respecto al alelo I en la población montevideana, habiéndose hallado diferencias significativas con respecto a poblaciones de origen asiático y americano, pero no con poblaciones europeas.