RESUMEN
PURPOSE: Abnormal retinal angiogenesis leads to visual impairment and blindness. Understanding how retinal vessels develop normally has dramatically improved treatments for people with retinal vasculopathies, but additional information about development is required. Abnormal neuron patterning in the outer retina has been shown to result in abnormal vessel development and blindness, for example, in people and mouse models with Crumbs homologue 1 (CRB1) mutations. In this study, we report and characterize a mouse model of inner retinal lamination disruption and bleeding, the Down syndrome cell adhesion molecule (Dscam) mutant, and test how neuron-neurite placement within the inner retina guides development of intraretinal vessels. METHODS: Bax mutant mice (increased neuron cell number), Dscam mutant mice (increased neuron cell number, disorganized lamination), Fat3 mutant mice (disorganized neuron lamination), and Dscam gain-of-function mice (Dscam(GOF)) (decreased neuron cell number) were used to manipulate neuron placement and number. Immunohistochemistry was used to assay organization of blood vessels, glia, and neurons. In situ hybridization was used to map the expression of angiogenic factors. RESULTS: Significant changes in the organization of vessels within mutant retinas were found. Displaced neurons and microglia were associated with the attraction of vessels. Using Fat3 mutant and Dscam(GOF) retinas, we provide experimental evidence that vessel branching is induced at the neuron-neurite interface, but that other factors are required for full plexus layer formation. We further demonstrate that the displacement of neurons results in the mislocalization of angiogenic factors. CONCLUSIONS: Inner retina neuron lamination is required for development of intraretinal vessels.
Asunto(s)
Modelos Animales de Enfermedad , Retina/anomalías , Hemorragia Retiniana/etiología , Neovascularización Retiniana/etiología , Neuronas Retinianas/patología , Animales , Western Blotting , Cadherinas/genética , Moléculas de Adhesión Celular/genética , Recuento de Células , Proteínas del Citoesqueleto , Glicoproteínas/metabolismo , Hibridación in Situ , Proteínas de la Membrana/metabolismo , Ratones , Ratones Mutantes , Microscopía Confocal , Hemorragia Retiniana/metabolismo , Hemorragia Retiniana/patología , Neovascularización Retiniana/metabolismo , Neovascularización Retiniana/patología , Vasos Retinianos/patología , Semaforinas/metabolismo , Factor A de Crecimiento Endotelial Vascular/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Proteína X Asociada a bcl-2/genéticaRESUMEN
Humans are exposed to a number of "heavy metals" such as cadmium, mercury and its organic form methylmercury, uranium, lead, and other metals as wel as metalloids, such as arsenic, in the environment, workplace, food, and water supply. Exposure to these metals may result in adverse health effects, and national and international health agencies have methodologies to set health-based guidance values with the aim to protect the human population. This chapter introduces the general principles of chemical risk assessment, the common four steps of chemical risk assessment: hazard identification, hazard characterization, exposure assessment, risk characterization, and toxicokinetic and toxicity aspects. Finally, the risk assessments performed by international health agencies such as the World Health Organisation, the Environmental Protection Agency of the United States, and the European Food Safety Authority are reviewed for cadmium, lead, mercury, uranium, and arsenic.
Asunto(s)
Exposición a Riesgos Ambientales/efectos adversos , Metales Pesados/análisis , Metales Pesados/toxicidad , Arsénico/análisis , Arsénico/farmacocinética , Arsénico/toxicidad , Cadmio/análisis , Cadmio/farmacocinética , Cadmio/toxicidad , Carcinógenos/análisis , Carcinógenos/farmacocinética , Carcinógenos/toxicidad , Contaminantes Ambientales/análisis , Contaminantes Ambientales/farmacocinética , Contaminantes Ambientales/toxicidad , Inocuidad de los Alimentos , Humanos , Mercurio/análisis , Mercurio/farmacocinética , Mercurio/toxicidad , Metaloides/análisis , Metaloides/farmacocinética , Metaloides/toxicidad , Metales Pesados/farmacocinética , Farmacocinética , Medición de Riesgo , Uranio/análisis , Uranio/farmacocinética , Uranio/toxicidad , Organización Mundial de la SaludRESUMEN
Posttranscriptional regulation of gene expression is an elaborate and intricate process, constituting an important mechanism for the control of protein expression. During its existence, mRNA is escorted by proteins and other RNAs, which control the maturation, transportation, localization, translational efficiency, and ultimately its degradation. Without changes at the transcription level, mRNA steady-state levels can vary dramatically by just small changes in mRNA stability. By influencing the metabolism of specific mRNAs, the abundance of specific mRNAs can be controlled in organisms from bacteria to mammals. In eukaryotic cells, the control of mRNA stability is exerted through specific cis-acting elements (sequence-specific control elements) and trans-acting factors (mRNA binding proteins and some miRNAs). mRNA stability appears to be a key regulator in controlling the expression of many proteins. Dysregulation of mRNA stability has been associated with human diseases, including cancer, inflammatory disease, and Alzheimer's. These observations suggest that modulating the stability of specific mRNAs may represent a viable strategy for pharmaceutical intervention. The literature already describes several compounds that influence mRNA stability. Measuring mRNA stability by conventional methods is labor intensive and time-consuming. However, several systems have been described that can be used to screen for modulators of mRNA levels in a high-throughput format. Thus, these assay systems offer a novel approach for screening targets that at present appear to be poorly "drugable." This review describes the utility of mRNA stability as a novel approach to drug discovery, focusing on assay methods and tool compounds available to monitor mRNA stability. The authors describe mRNA stability assays and issues related to this approach.
Asunto(s)
Bioensayo/métodos , Descubrimiento de Drogas/métodos , Estabilidad del ARN/genética , Evaluación Preclínica de Medicamentos , HumanosRESUMEN
Proteasome inhibition is a therapeutic concept of current interest in anticancer research. We report here the design, synthesis, and biological characterization of prototypes of a new class of noncovalent proteasome inhibitors showing high activity in biochemical and cellular assays.
Asunto(s)
Antineoplásicos/química , Antineoplásicos/farmacología , Inhibidores de Cisteína Proteinasa/química , Inhibidores de Cisteína Proteinasa/farmacología , Complejos Multienzimáticos/antagonistas & inhibidores , Sitios de Unión , División Celular/efectos de los fármacos , Cisteína Endopeptidasas/química , Cisteína Endopeptidasas/metabolismo , Inhibidores de Cisteína Proteinasa/metabolismo , Ensayos de Selección de Medicamentos Antitumorales/métodos , Humanos , Enlace de Hidrógeno , Concentración 50 Inhibidora , Modelos Moleculares , Estructura Molecular , Complejos Multienzimáticos/química , Complejos Multienzimáticos/metabolismo , Oligopéptidos/química , Oligopéptidos/metabolismo , Oligopéptidos/farmacología , Complejo de la Endopetidasa Proteasomal , Relación Estructura-Actividad , Células Tumorales CultivadasRESUMEN
We have identified 2-aminobenzylstatine derivatives that inhibit non-covalently the chymotrypsin-like activity of the human 20S proteasome. A structure-based optimisation approach has allowed us to improve the potency of this structural class of proteasome inhibitors from micromolar to nanomolar level. The new derivatives showed good selectivity against the trypsin-like and post-glutamyl-peptide hydrolytic activities of this enzyme.