RESUMEN
Leber hereditary optic neuropathy (LHON) is caused by mitochondrial DNA mutations and is the most common inherited mitochondrial disease. It is responsible for central vision loss in young adulthood. However, the precise mechanisms of onset are unknown. This study aimed to elucidate the mechanisms underlying LHON pathology and to discover new therapeutic agents. First, we assessed whether rotenone, a mitochondrial complex â inhibitor, induced retinal degeneration such as that in LHON in a mouse model. Rotenone decreased the thickness of the inner retina and increased the expression levels of 8-hydroxy-2'-deoxyguanosine (8-OHdG) and immunoglobulin heavy-chain binding protein (BiP). Second, we assessed whether rotenone reproduces LHON pathologies on RGC-5, a neural progenitor cell derived from the retina. Rotenone increased the cell death rate, ROS production and the expression levels of ER stress markers. During chemical compounds screening, we used anti-oxidative compounds, ER stress inhibitors and anti-inflammatory compounds in a rotenone-induced in vitro model. We found that SUN N8075, an ER stress inhibitor, reduced mitochondrial ROS production and improved the mitochondrial membrane potential. Consequently, the ER stress response is strongly related to the pathologies of LHON, and ER stress inhibitors may have a protective effect against LHON.
Asunto(s)
Compuestos de Anilina/farmacología , Descubrimiento de Drogas , Estrés del Retículo Endoplásmico/efectos de los fármacos , Estrés del Retículo Endoplásmico/fisiología , Atrofia Óptica Hereditaria de Leber/tratamiento farmacológico , Atrofia Óptica Hereditaria de Leber/genética , Piperazinas/farmacología , Rotenona/efectos adversos , Animales , Células Cultivadas , ADN Mitocondrial/genética , Modelos Animales de Enfermedad , Evaluación Preclínica de Medicamentos , Estrés del Retículo Endoplásmico/genética , Masculino , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Potencial de la Membrana Mitocondrial/genética , Ratones Endogámicos C57BL , Terapia Molecular Dirigida , Mutación , Atrofia Óptica Hereditaria de Leber/inducido químicamente , Atrofia Óptica Hereditaria de Leber/patología , Especies Reactivas de Oxígeno/metabolismo , Retina/efectos de los fármacos , Retina/metabolismo , Retina/patología , Degeneración Retiniana/inducido químicamente , Degeneración Retiniana/genética , Degeneración Retiniana/patologíaRESUMEN
Fertilization begins with interaction between the sperm and the egg. The surface of the vertebrate oocyte is covered with the egg envelope, which is composed of ZP (zona pellucida) glycoproteins. We have identified two glycoproteins, ZP1/gp97 and ZPC/gp42, as the major components of the chicken egg envelope. In the present study, another 42 kDa protein, designated ZPD, has been found as a new major component of the chicken egg envelope. ZPD was specifically released from the egg envelope by ultrasonication treatment without urea. ZPD cDNA was cloned using a chicken granulosa cell cDNA pool. The deduced amino acid sequence showed that preproprotein of ZPD is composed of 418 amino acid residues with four potential N-glycosylation sites and includes a ZP domain, common in vertebrate ZP glycoproteins, and a transmembrane domain. ZPD belongs phylogenetically to a distinct group from known ZP glycoprotein subfamilies, ZPA, ZPB, and ZPC. In two-dimensional gel electrophoresis ZPD proteins were identified to be several isoforms with different pI values between 5 and 7. ZP1, ZPC and the newly identified ZPD were confirmed to be the major components of chicken egg envelope by MS of proteolytic digests of whole egg envelope. The in vitro incubation of chicken sperm with calcium ionophore A23187 induced sperm activation, resulting in the fragmentation and release of a 41 kDa PNA (peanut agglutinin)-positive glycoprotein and the decrease or loss of sperm PNA-stainability. The incubation with ZPD and dimeric ZP1, but not ZPC and monomeric ZP1, also induced the decrease or loss of sperm PNA-stainability, suggesting the in vitro sperm activation by these ZP components. Collectively, ZPD might bind loosely to egg envelope matrix and play a key role in the sperm activation on avian sperm-egg interaction.