RESUMEN
Dysregulation of the complement system has been implicated in the pathogenesis of age-related macular degeneration. To investigate consequences of altered complement regulation in the eye with age, we examined Cd59a complement regulator deficient (Cd59a(-/-)) mice between 4 and 15 months. In vivo imaging revealed an increased age-related accumulation of autofluorescent spots in Cd59a(-/-) mice, a feature that reflects accumulation of subretinal macrophages and/or microglia. Despite this activation of myeloid cells in the eye, Cd59a(-/-) mice showed normal retinal histology and function as well as normal choroidal microvasculature. With age, they revealed increased expression of activators of the alternative complement pathway (C3, Cfb, Cfd), in particular in the retinal pigment epithelium (RPE)-choroid but less in the retina. This molecular response was not altered by moderately-enhanced light exposure. Cd59a deficiency therefore leads to a preferential age-related dysregulation of the complement system in the RPE-choroid, that alone or in combination with light as a trigger, is not sufficient to cause choroidal vascular changes or retinal degeneration and dysfunction. This data emphasizes the particular vulnerability of the RPE-choroidal complex to dysregulation of the alternative complement pathway during aging.
Asunto(s)
Envejecimiento/genética , Antígenos CD59/metabolismo , Coroides/metabolismo , Factores Inmunológicos/metabolismo , Degeneración Macular , Epitelio Pigmentado de la Retina/metabolismo , Análisis de Varianza , Animales , Antígenos CD59/genética , Coroides/patología , Activación de Complemento , Modelos Animales de Enfermedad , Electrorretinografía , Regulación de la Expresión Génica/genética , Macrófagos/metabolismo , Macrófagos/patología , Degeneración Macular/genética , Degeneración Macular/metabolismo , Degeneración Macular/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microglía/metabolismo , Microglía/patología , Proteínas del Tejido Nervioso/genética , ARN Mensajero/metabolismo , Retina/patología , Epitelio Pigmentado de la Retina/patologíaRESUMEN
Monocytes, macrophages, dendritic cells and microglia play critical roles in the local immune response to acute and chronic tissue injury and have been implicated in the pathogenesis of age-related macular degeneration. Defects in Ccl2-Ccr2 and Cx3cl1-Cx3cr1 chemokine signalling cause enhanced accumulation of bloated subretinal microglia/macrophages in senescent mice and this phenomenon is reported to result in the acceleration of age-related retinal degeneration. The purpose of this study was to determine whether defects in CCL2-CCR2 and CX3CL1-CX3CR1 signalling pathways, alone or in combination, cause age-dependent retinal degeneration. We tested whether three chemokine knockout mouse lines, Ccl2(-/-), Cx3cr1(-/-) and Ccl2(-/-)/Cx3cr1(-/-), in comparison to age-matched C57Bl/6 control mice show differences in subretinal macrophage accumulation and loss of adjacent photoreceptor cells at 12-14 months of age. All mouse lines are derived from common parental strains and do not carry the homozygous rd8 mutation in the Crb1 gene that has been a major confounding factor in previous reports. We quantified subretinal macrophages by counting autofluorescent lesions in fundus images obtained by scanning laser ophthalmoscopy (AF-SLO) and by immunohistochemistry for Iba1 positive cells. The accumulation of subretinal macrophages was enhanced in Ccl2(-/-), but not in Cx3cr1(-/-) or Ccl2(-/-)/Cx3cr1(-/-) mice. We identified no evidence of retinal degeneration in any of these mouse lines by TUNEL staining or semithin histology. In conclusion, CCL2-CCR2 and/or CX3CL1-CX3CR1 signalling defects may differentially affect the trafficking of microglia and macrophages in the retina during ageing, but do not appear to cause age-related retinal degeneration in mice.
Asunto(s)
Quimiocina CCL2/fisiología , Degeneración Macular/metabolismo , Receptores de Quimiocina/fisiología , Animales , Receptor 1 de Quimiocinas CX3C , Proteínas de Unión al Calcio/metabolismo , Recuento de Células , Genotipo , Etiquetado Corte-Fin in Situ , Macrófagos/metabolismo , Degeneración Macular/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas de Microfilamentos/metabolismo , Oftalmoscopía , Células Fotorreceptoras de Vertebrados/metabolismo , Células Fotorreceptoras de Vertebrados/patología , Reacción en Cadena de la PolimerasaRESUMEN
Uniquely for non-primate mammals, three classes of cone photoreceptors have been previously identified by microspectrophotometry in two marsupial species: the polyprotodont fat-tailed dunnart (Sminthopsis crassicaudata) and the diprotodont honey possum (Tarsipes rostratus). This report focuses on the genetic basis for these three pigments. Two cone pigments were amplified from retinal cDNA of both species and identified by phylogenetics as members of the short wavelength-sensitive 1 (SWS1) and long wavelength-sensitive (LWS) opsin classes. In vitro expression of the two sequences from the fat-tailed dunnart confirmed the peak absorbances at 363 nm in the UV for the SWS1 pigment and 533 nm for the LWS pigment. No additional expressed cone opsin sequences that could account for the middle wavelength cones could be amplified. However, amplification from the fat-tailed dunnart genomic DNA with RH1 (rod) opsin primer pairs identified two genes with identical coding regions but sequence differences in introns 2 and 3. Uniquely therefore for a mammal, the fat-tailed dunnart has two copies of an RH1 opsin gene. This raises the possibility that the middle wavelength cones express a rod rather than a cone pigment.