RESUMO
Photoreceptor degeneration is the most critical cause of visual impairment in age-related macular degeneration (AMD). In neovascular form of AMD, severe photoreceptor loss develops with subretinal hemorrhage due to choroidal neovascularization (CNV), growth of abnormal blood vessels from choroidal circulation. However, the detailed mechanisms of this process remain elusive. Here we demonstrate that neovascular AMD with subretinal hemorrhage accompanies a significant increase in extracellular ATP, and that extracellular ATP initiates neurodegenerative processes through specific ligation of Purinergic receptor P2X, ligand-gated ion channel, 7 (P2RX7; P2X7 receptor). Increased extracellular ATP levels were found in the vitreous samples of AMD patients with subretinal hemorrhage compared to control vitreous samples. Extravascular blood induced a massive release of ATP and photoreceptor cell apoptosis in co-culture with primary retinal cells. Photoreceptor cell apoptosis accompanied mitochondrial apoptotic pathways, namely activation of caspase-9 and translocation of apoptosis-inducing factor (AIF) from mitochondria to nuclei, as well as TUNEL-detectable DNA fragmentation. These hallmarks of photoreceptor cell apoptosis were prevented by brilliant blue G (BBG), a selective P2RX7 antagonist, which is an approved adjuvant in ocular surgery. Finally, in a mouse model of subretinal hemorrhage, photoreceptor cells degenerated through BBG-inhibitable apoptosis, suggesting that ligation of P2RX7 by extracellular ATP may accelerate photoreceptor cell apoptosis in AMD with subretinal hemorrhage. Our results indicate a novel mechanism that could involve neuronal cell death not only in AMD but also in hemorrhagic disorders in the CNS and encourage the potential application of BBG as a neuroprotective therapy.
Assuntos
Trifosfato de Adenosina/farmacologia , Degeneração Macular/metabolismo , Células Fotorreceptoras/efeitos dos fármacos , Receptores Purinérgicos P2X7/metabolismo , Hemorragia Retiniana/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Apoptose/efeitos dos fármacos , Fator de Indução de Apoptose/genética , Fator de Indução de Apoptose/metabolismo , Caspase 9/genética , Caspase 9/metabolismo , Corioide/irrigação sanguínea , Corioide/metabolismo , Corioide/patologia , Neovascularização de Coroide , Técnicas de Cocultura , Fragmentação do DNA/efeitos dos fármacos , Humanos , Degeneração Macular/patologia , Masculino , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Células Fotorreceptoras/metabolismo , Células Fotorreceptoras/patologia , Cultura Primária de Células , Antagonistas do Receptor Purinérgico P2X/farmacologia , Receptores Purinérgicos P2X7/genética , Hemorragia Retiniana/patologia , Epitélio Pigmentado da Retina , Corantes de Rosanilina/farmacologia , Corpo Vítreo/irrigação sanguínea , Corpo Vítreo/metabolismo , Corpo Vítreo/patologiaRESUMO
In young puppies, the retina, which is incompletely vascularized at birth becomes fully vascularized at approximately four weeks of age. During this period of vessel growth the total content of vitreous-soluble protein was found closely associated with the rate of retinal vessel growth. As vascularization progressed toward completion, the protein originally present at birth decreased to a negligible or undetected amount. Intravitreal neovascularization was produced in young puppies by exposure to 85 per cent oxygen for four days, then removal to room air. This form of neovascularization resembles closely that observed in human proliferative diabetic retinopathy. The oxygen treatment, which initially produced retinal capillary closure, then neovascularization, was associated with a retention of vitreous-soluble protein at a high level for several days. The results raise the possibility that the vitreous protein(s) may be fundamentally involved in the process of normal vascularization of the retina and in retinal neovascularization.