RESUMEN
Objective: This study was designed to examine the supplementation of a carotenoid-rich carrot powder, on retina function and carotenoid metabolism in non-diabetic control and type 1 diabetic animals. Methods: Male Wistar rats (n = 30) were randomly assigned to diets supplemented with (n = 15) or without (n = 15) carrot powder enriched diets (150â g/kg diet). After 3 weeks of diet adaptation, 8 rats in each group were treated with streptozotocin (iv) to induce type 1 diabetes and fed for a further 9â wk. Retinal function was assessed with the electroretinogram (ERG). Hepatic and plasma retinoids and carotenoids were measured by ultra-performance liquid chromatography. Results: Non-diabetic control rats fed the carrot diet had significantly (p < 0.02) higher rod- and cone- driven post-synaptic b-wave amplitudes, respectively, compared to those fed the control diet. These functional changes correlated with higher (p < 0.05) liver levels of carotenoids (α- and ß- carotene) and retinoids. In diabetic rats, carrot diet exacerbated retina dysfunction; the amplitudes for most of rod- and cone-driven ERG components were the lowest amplitudes among all groups (p < 0.02). Diabetic rats fed the carrot diet had lower hepatic retinol and retinyl palmitate, while having higher α- and ß-carotene levels, indicating diminished hepatic conversion of carotenoids into retinoids. Discussion: Dietary supplementation of high dose dietary carotenoids plays a beneficial role on healthy rat retina function, but exerts a detrimental effect in diabetes, which warrants undertaking detailed mechanistic studies.
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Carotenoides/administración & dosificación , Diabetes Mellitus Experimental/fisiopatología , Retina/fisiopatología , Animales , Carotenoides/sangre , Diabetes Mellitus Experimental/metabolismo , Electrorretinografía , Masculino , Ratas Wistar , Retinoides/sangreRESUMEN
Insulin-secreting pancreatic ß-cells adapt to obesity-related insulin resistance via increases in insulin secretion and ß-cell mass. Failed ß-cell compensation predicts the onset of type 2 diabetes (T2D). However, the mechanisms of ß-cell compensation are not fully understood. Our previous study reported changes in ß-cell mass during the progression of T2D in the Nile rat (NR; Arvicanthis niloticus) fed standard chow. In the present study, we measured other ß-cell adaptive responses, including glucose metabolism and ß-cell insulin secretion in NRs at different ages, thus characterizing NR at 2 months as a model of ß-cell compensation followed by decompensation at 6 months. We observed increased proinsulin secretion in the transition from compensation to decompensation, which is indicative of impaired insulin processing. Subsequently, we compared adaptive unfolded protein response in ß-cells and demonstrated a positive role of endoplasmic reticulum (ER) chaperones in insulin secretion. In addition, the incidence of insulin-positive neogenic but not proliferative cells increased during the compensation phase, suggesting nonproliferative ß-cell growth as a mechanism of ß-cell mass adaptation. In contrast, decreased neogenesis and ß-cell dedifferentiation were observed in ß-cell dysfunction. Furthermore, the progression of T2D and pathophysiological changes of ß-cells were prevented by increasing fibre content of the diet. Novelty Our study characterized a novel model for ß-cell compensation with adaptive responses in cell function and mass. The temporal association of adaptive ER chaperones with blood insulin and glucose suggests upregulated chaperone capacity as an adaptive mechanism. ß-Cell neogenesis but not proliferation contributes to ß-cell mass adaptation.
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Diabetes Mellitus Tipo 2/fisiopatología , Dieta/efectos adversos , Estrés del Retículo Endoplásmico , Células Secretoras de Insulina/fisiología , Animales , Diabetes Mellitus Tipo 2/inducido químicamente , Fibras de la Dieta , Glucosa/metabolismo , Insulina/metabolismo , Masculino , Murinae , Respuesta de Proteína DesplegadaRESUMEN
BACKGROUND: The Nile rat (Arvicanthis niloticus) is an emerging laboratory model of type 2 diabetes. When fed standard rodent chow, the majority of males progress from hyperinsulinemia by 2 months to hyperglycemia by 6 months, while most females remain at the hyperinsulinemia-only stage (prediabetic) from 2 months onward. Since diabetic cardiomyopathy is the major cause of type-2 diabetes mellitus (T2DM)-related mortality, we examined whether sexual dimorphism might entail cardiac functional changes. Our ultimate goal was to isolate the effect of diet as a modifiable lifestyle factor. MATERIALS AND METHODS: Nile rats were fed either standard rodent chow (Chow group) or a high-fiber diet previously established to prevent type 2 diabetes (Fiber group). Cardiac function was determined with echocardiography at 12 months of age. To isolate the effect of diet alone, only the small subset of animals resistant to both hyperinsulinemia and hyperglycemia were included in this study. RESULTS: In males, Chow (compared to Fiber) was associated with elevated heart rate and mitral E/A velocity ratio, and with lower e'-wave velocity, isovolumetric relaxation time, and ejection time. Of note, these clinically atypical types of diastolic dysfunction occurred independently of body weight. In contrast, females did not exhibit changes in cardiovascular function between diets. CONCLUSIONS: The higher prevalence of T2DM in males correlates with their susceptibility to develop subtle diastolic cardiac dysfunction when fed a Western style diet (throughout most of their lifespan) despite no systemic evidence of metabolic syndrome, let alone T2DM.
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Diabetes Mellitus Tipo 2/patología , Dieta , Animales , Glucemia/análisis , Peso Corporal , Fibras de la Dieta , Modelos Animales de Enfermedad , Femenino , Frecuencia Cardíaca , Ventrículos Cardíacos/química , Ventrículos Cardíacos/patología , Masculino , Ratas , Caracteres Sexuales , Función Ventricular Izquierda/fisiologíaRESUMEN
For many neurodegenerative disorders, expression of a pathological protein by one cell type impedes function of other cell types, which in turn contributes to the death of the first cell type. In transgenic mice modelling Stargardt-like (STGD3) maculopathy, human mutant ELOVL4 expression by photoreceptors is associated with defects in the underlying retinal pigment epithelium (RPE). To examine how photoreceptors exert cytotoxic effects on RPE cells, transgenic ELOVL4 (TG1-2 line; TG) and wild-type (WT) littermates were studied one month prior (preclinical stage) to onset of photoreceptor loss (two months). TG photoreceptor outer segments presented to human RPE cells are recognized and internalized into phagosomes, but their digestion is delayed. Live RPE cell imaging pinpoints decreased numbers of acidified phagolysomes. In vivo, master regulator of lysosomal genes, transcription factor EB (TFEB), and key lysosomal enzyme Cathepsin D are both unaffected. Oxidative stress, as ruled out with high-resolution respirometry, does not play a role at such an early stage. Upregulation of CRYBA1/A3 and phagocytic cells (microglia/macrophages) interposed between RPE and photoreceptors support adaptive responses to processing delays. Impaired phagolysosomal maturation is observed in RPE of mice expressing human mutant ELOVL4 in their photoreceptors prior to photoreceptor death and associated vision loss.
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Lisosomas/patología , Degeneración Macular/congénito , Fagosomas/patología , Células Fotorreceptoras/patología , Epitelio Pigmentado de la Retina/patología , Animales , Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice/metabolismo , Catepsina D/metabolismo , Línea Celular , Modelos Animales de Enfermedad , Proteínas del Ojo/metabolismo , Humanos , Lisosomas/metabolismo , Degeneración Macular/metabolismo , Degeneración Macular/patología , Proteínas de la Membrana/metabolismo , Ratones , Ratones Transgénicos , Mutación/fisiología , Fagosomas/metabolismo , Células Fotorreceptoras/metabolismo , Enfermedades de la Retina/metabolismo , Enfermedades de la Retina/patología , Epitelio Pigmentado de la Retina/metabolismo , Cadena A de beta-Cristalina/metabolismoRESUMEN
Purpose: To characterize retinal mitochondrial respiration associated with type 2 diabetes (T2D) progression in a cone-rich diurnal rodent, the Nile rat (genus Arvicanthis, species niloticus). Methods: Nile rats were fed a standard rodent diet that resulted in rising glucose levels from 6 months. Age-matched control animals were fed a high-fiber diet that prevented diabetes up to 18 months. The functional status of specific retinal mitochondrial components and mitochondrial outer membrane integrity were studied by using high-resolution respirometry. Ocular complications were documented with funduscopy, electroretinography (ERG), and trypsin digestion of retinal vasculature. Results: Mitochondrial functional changes were detected during hyperinsulinemia with maintained normoglycemia (2 months), corresponding to stage 1 of human T2D. Our data showed increased contribution of mitochondrial respiration through the NADH pathway relative to maximal oxidative phosphorylation capacity, with simultaneous electron entry into NADH (Complex I and related dehydrogenases) and succinate (Complex II) pathways. These compensatory events coincided with compromised mitochondrial outer membrane integrity. The first clinical sign of retinopathy (pericyte loss) was only detected at 12 months (after 6 months of sustained hyperglycemia) alongside a common ocular complication of diabetes, cataractogenesis. Further prolongation of hyperglycemia (from 12 to 18 months) led to capillary degeneration and delayed photopic ERG oscillatory potentials. Conclusions: Oxidative phosphorylation compensatory changes in the retina can be detected as early as 2 months, before development of hyperglycemia, and are associated with reduced mitochondrial outer membrane integrity.
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Respiración de la Célula/fisiología , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Retinopatía Diabética/metabolismo , Mitocondrias/metabolismo , Retina/metabolismo , Animales , Glucemia/metabolismo , Retinopatía Diabética/fisiopatología , Electrorretinografía , Hemoglobina Glucada/metabolismo , Insulina/sangre , Masculino , RatasRESUMEN
We compared the evolution of insulin resistance, hyperglycemia, and pancreatic ß-cell dysfunction in the Nile rat (Arvicanthis niloticus), a diurnal rodent model of spontaneous type 2 diabetes (T2D), when maintained on regular laboratory chow versus a high-fiber diet. Chow-fed Nile rats already displayed symptoms characteristic of insulin resistance at 2 months (increased fat/lean mass ratio and hyperinsulinemia). Hyperglycemia was first detected at 6 months, with increased incidence at 12 months. By this age, pancreatic islet structure was disrupted (increased α-cell area), insulin secretion was impaired (reduced insulin secretion and content) in isolated islets, insulin processing was compromised (accumulation of proinsulin and C-peptide inside islets), and endoplasmic reticulum (ER) chaperone protein ERp44 was upregulated in insulin-producing ß-cells. By contrast, high-fiber-fed Nile rats had normoglycemia with compensatory increase in ß-cell mass resulting in maintained pancreatic function. Fasting glucose levels were predicted by the α/ß-cell ratios. Our results show that Nile rats fed chow recapitulate the five stages of progression of T2D as occurs in human disease, including insulin-resistant hyperglycemia and pancreatic islet ß-cell dysfunction associated with ER stress. Modification of diet alone permits long-term ß-cell compensation and prevents T2D.
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Diabetes Mellitus Tipo 2/fisiopatología , Células Secretoras de Insulina/fisiología , Animales , Glucemia/metabolismo , Diabetes Mellitus Tipo 2/dietoterapia , Diabetes Mellitus Tipo 2/etiología , Fibras de la Dieta/administración & dosificación , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Estrés del Retículo Endoplásmico , Humanos , Hiperglucemia/etiología , Técnicas In Vitro , Insulina/sangre , Insulina/metabolismo , Resistencia a la Insulina , Secreción de Insulina , Células Secretoras de Insulina/patología , Masculino , MurinaeRESUMEN
PURPOSE: We investigated the interplay between photoreceptors expressing mutant ELOVL4 (responsible for Stargardt-like disease, STGD3) and RPE in the initial stages of retinal degeneration. METHODS: Using electron microscopy and electroretinogram (ERG), we assessed RPE and photoreceptor ultrastructure and function in transgenic ELOVL4 (TG1-2 line; TG) and wild-type (WT) littermates. Experiments were done at P30, 1 month before photoreceptor loss in TG and at P90, a time point with approximately 30% rod loss. To further elucidate the mechanism underlying our ultrastructural and functional results, we undertook Western blotting and immunohistochemistry of key proteins involved in phagocytosis of outer segments by RPE cells. RESULTS: Firstly, we showed that in TG mouse photoreceptors, endogenous ELOVL4 protein is not mislocalized in the presence of the mutated ELOVL4 protein. Secondly, we found evidence of RPE toxicity at P30, preceding any photoreceptor loss. Pathology in RPE cells was exacerbated at P90. Furthermore, higher proportions of phagosomes remained at the apical side of RPE cells. Subretinal lysosomal deposits were immunopositive for phagocytic proteins. Ultrastructural analysis of photoreceptor (rod) outer segments showed disrupted surface morphology consisting of disc spacing irregularities. Finally, rods and RPE exhibited signs of dysfunction as measured by the ERG a-wave leading edge (P30) and c-wave (P90), respectively. CONCLUSIONS: The presence of human mutant ELOVL4 in transgenic mouse photoreceptors leads to early outer segment disc pathology and RPE cytotoxicity. Defective processing of these abnormal discs by RPE cells ultimately may be responsible for outer segment truncation, photoreceptor death, and vision loss.
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Proteínas del Ojo/genética , Degeneración Macular/congénito , Mutación , ARN/genética , Epitelio Pigmentado de la Retina/ultraestructura , Segmento Externo de la Célula en Bastón/ultraestructura , Animales , Western Blotting , Células Cultivadas , Análisis Mutacional de ADN , Modelos Animales de Enfermedad , Electrorretinografía , Proteínas del Ojo/metabolismo , Humanos , Inmunohistoquímica , Degeneración Macular/diagnóstico , Degeneración Macular/metabolismo , Degeneración Macular/fisiopatología , Ratones , Ratones Transgénicos , Microscopía Electrónica de Transmisión , Epitelio Pigmentado de la Retina/metabolismo , Epitelio Pigmentado de la Retina/fisiopatología , Segmento Externo de la Célula en Bastón/metabolismoRESUMEN
We sought to study the expression pattern of Disabled-1 (Dab1; an adaptor protein in the reelin pathway) in the cone-rich retina of a diurnal murine rodent. Expression was examined by western blotting and immunohistochemistry using well-established antibodies against Dab1 and various markers of retinal neurons. Western blots revealed the presence of Dab1 (80 kDa) in brain and retina of the Nile grass rat. Retinal immunoreactivity was predominant in soma and dendrites of horizontal cells as well as in amacrine cell bodies aligned at the INL/IPL border. Dab1(+) neurons in the inner retina do not stain for parvalbumin, calbindin, protein kinase C-alpha, choline acetyltransferase, glutamic acid decarboxylase, or tyrosine hydroxylase. They express, however, the glycine transporter GlyT1. They have small ovoid cell bodies (7.1 ± 1.06 µm in diameter) and bistratified terminal plexii in laminas a and b of the IPL. Dab1(+) amacrine cells are evenly distributed across the retina (2600 cells/mm(2)) in a fairly regular mosaic (regularity indexes ≈3.3-5.5). We conclude that retinal Dab1 in the adult Nile grass rat exhibits a dual cell patterning similar to that found in human. It is expressed in horizontal cells as well as in a subpopulation of glycinergic amacrine cells undetectable with antibodies against calcium-binding proteins. These amacrine cells are likely of the AII type.
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Proteínas Adaptadoras Transductoras de Señales/metabolismo , Murinae/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Retina/metabolismo , Células Amacrinas/citología , Células Amacrinas/metabolismo , Animales , Western Blotting , Recuento de Células , Inmunohistoquímica , Ratones Endogámicos C57BL , Modelos Animales , Proteína Reelina , Retina/citologíaRESUMEN
PURPOSE: To examine the pattern of cone degeneration in the retina of a transgenic mouse model of Stargartd-like dystrophy (STGD3). METHODS: Investigations were performed on ELOVL4/TG1-2 transgenic (TG) mice and wild-type (WT) littermates from 1 to 24 months of age. Phenotypes were assessed by fundus imaging, fatty acid analysis, and electroretinogram (ERG) recording. Cone degeneration pattern was determined on retina whole mounts using immunohistochemistry and Voronoi domain analyses. RESULTS: Consistent with low transgene expression, photoreceptors degenerate very slowly. At 1 month, anatomical structure and fatty acid composition of the TG retina is comparable with WT. Rod loss appears at 2 months, exhibiting a central to peripheral gradient, and fundus defects are observed at 3 months. In contrast, cone morphology, distribution and function are still normal at 12 months. Cone loss becomes apparent at 15 months when the outer nuclear layer is reduced to 3 to 4 photoreceptor rows. This process starts at the center of the retina and affects cone subtypes similarly. Very few cones remain at 24 months, after all rods have disappeared (18 months). Quantitative studies focusing on cones expressing M-opsin show a net increase in Voronoi domains and a significant decrease in regularity indexes only beyond 15 months. CONCLUSIONS: Photoreceptor degeneration in this STGD3 mouse model follows the time course of a slow rod-cone dystrophy. The cone mosaic is preserved for almost 1 year after the onset of rod loss. This long delay provides an opportunity to examine rod-cone interactions during retinal degeneration and to test therapeutic effectiveness at protracting cone dysfunction.
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Degeneración Macular/congénito , Células Fotorreceptoras Retinianas Conos/patología , Animales , Western Blotting , Supervivencia Celular , Modelos Animales de Enfermedad , Electrorretinografía , Proteínas del Ojo/genética , Proteínas del Ojo/metabolismo , Estudios de Seguimiento , Inmunohistoquímica , Degeneración Macular/diagnóstico , Degeneración Macular/genética , Degeneración Macular/fisiopatología , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Microscopía Fluorescente , Retina/patología , Retina/fisiopatología , Células Fotorreceptoras Retinianas Conos/metabolismo , Factores de TiempoRESUMEN
BACKGROUND: Intra-uterine growth restriction (IUGR) has been associated with increased predisposition to age-related complications. We tested the hypothesis that rat offspring models of IUGR would exhibit exacerbated, age-related retinal dysfunction. METHODS: Female Sprague-Dawley rats (maintained at 11.5% O2 from gestational day 15 to 21 to induce IUGR) and control offspring (maintained at 21% O2 throughout pregnancy) had retinal function assessed at 2 months (young) and 14 months of age (aged) with electroretinogram (ERG) recordings. Retinal anatomy was assessed by immunofluorescence. RESULTS: Deficits in rod-driven retina function were observed in aged IUGR offspring, as evidenced by reduced amplitudes of dark-adapted mixed a-wave V(max) (by 49.3%, P < 0.01), b-wave V(max) (by 42.1%, P < 0.001) and dark-adapted peak oscillatory potentials (by 42.3%, P < 0.01). In contrast to the rod-driven defects specific to aged IUGR offspring, light adapted ERG recordings revealed cone defects in young animals, that were stationary until old age. At 2 months, IUGR offspring had amplitude reductions for both b-wave (V(max) by 46%, P < 0.01) and peak oscillatory potential (V(max) by 38%, P < 0.05). Finally, defects in cone-driven responses were further confirmed by reduced maximal photopic flicker amplitudes at 2 (by 42%, P < 0.001) and 14 months (by 34%, P â=â 0.06) and critical flicker fusion frequencies at 14 months ( CONTROL: 42 ± 1 Hz, IUGR: 35 ± 2 Hz, P < 0.05). These functional changes were not paralleled by anatomical losses in IUGR offspring retinas. CONCLUSIONS: These data support that the developing retina is sensitive to stressors, and that pathways governing cone- and rod-driven function differ in their susceptibilities. In the case of prenatal hypoxia, cone- and rod-driven dysfunction manifest at young and old ages, respectively. We must, therefore, take into account the specific impact that fetal programming might exert on age-related retinal dystrophies when considering related diagnoses and therapeutic applications.
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Hipoxia/embriología , Efectos Tardíos de la Exposición Prenatal/fisiopatología , Retina/fisiopatología , Adaptación Fisiológica/efectos de la radiación , Envejecimiento/patología , Envejecimiento/fisiología , Animales , Oscuridad , Femenino , Embarazo , Efectos Tardíos de la Exposición Prenatal/patología , Ratas , Ratas Sprague-Dawley , Retina/embriología , Retina/patología , Retina/efectos de la radiación , Células Fotorreceptoras Retinianas Conos/patología , Células Fotorreceptoras Retinianas Conos/fisiología , Células Fotorreceptoras Retinianas Conos/efectos de la radiación , Células Fotorreceptoras Retinianas Bastones/patología , Células Fotorreceptoras Retinianas Bastones/fisiología , Células Fotorreceptoras Retinianas Bastones/efectos de la radiaciónRESUMEN
PURPOSE: With age, retina function progressively declines and A2E, a constituent of the toxin lipofuscin, accumulates in retinal pigment epithelial (RPE) cells. Both events are typically exacerbated in age-related retina diseases. We studied the effect of dietary docosahexaenoic acid (DHA, C22:6n-3) supplementation on these events, using a transgenic mouse model (mutant human ELOVL4; E4) displaying extensive age-related retina dysfunction and massive A2E accumulation. METHODS: Retina function was assessed with the electroretinogram (ERG) and A2E levels were measured in E4 and wildtype (WT) mice. Dietary DHA was manipulated from 1 to 3, 1 to 6, 6 to 12, and 12 to 18 months: 1% DHA over total fatty acids (E4+, WT+) or similar diet without DHA (E4-, WT-). RESULTS: Increased omega-3/6 ratios (DHA/arachidonic acid) in E4+ and WT+ retinas were confirmed for the 1- to 3-month and 1- to 6-month trials. Although 1- to 3-month intervention had no effects, when prolonged to 1 to 6 months, RPE function (ERG c-wave) was preserved in E4+ and WT+. Intervention from 6 to 12 months led to maintained outer and inner retina function (ERG a- and b-wave, respectively) in E4+. At 12 to 18 months, a similar beneficial effect on retina function occurred in WT+; A2E levels were reduced in E4+ and WT+. CONCLUSIONS: DHA supplementation was associated with: preserved retina function at mid-degenerative stages in E4 mice; prevention of age-related functional losses in WT mice; and reduced A2E levels in E4 and WT mice at the oldest age examined. These findings imply that dietary DHA could have broad preventative therapeutic applications (acting on pathologic and normal age-related ocular processes).
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Envejecimiento/fisiología , Trastornos de los Cromosomas/prevención & control , Grasas Insaturadas en la Dieta/administración & dosificación , Modelos Animales de Enfermedad , Ácidos Docosahexaenoicos/administración & dosificación , Degeneración Macular/prevención & control , Compuestos de Piridinio/metabolismo , Retina/metabolismo , Retinoides/metabolismo , Animales , Trastornos de los Cromosomas/metabolismo , Trastornos de los Cromosomas/fisiopatología , Cromosomas Humanos Par 6/metabolismo , Adaptación a la Oscuridad , Electrorretinografía , Ácidos Grasos Omega-3/metabolismo , Ácidos Grasos Omega-6/metabolismo , Femenino , Técnica del Anticuerpo Fluorescente Indirecta , Degeneración Macular/congénito , Degeneración Macular/metabolismo , Degeneración Macular/fisiopatología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Factores de TiempoRESUMEN
PURPOSE: To investigate differentially expressed genes in eyecup and retina of the ELOVL4 transgenic mouse, a model of Stargardt-like macular dystrophy (STGD3). METHODS: We examined gene and protein expression in known pathways relevant to retinal degeneration using PCR arrays, Western blotting, and immunohistochemistry. Investigations were performed on ELOVL4 transgenic mice at 9 months, when 50% of rod (but no cone) photoreceptors had degenerated. Age-matched wild-type littermates served as controls. RESULTS: Significant expression level changes were found in only 17 of the 252 genes examined. Nine were upregulated (Fgf2, Fgfr1, Ntf5, Cbln1, Ngfr, Ntrk1, Trp53, Tlr6, and Herpud1), and eight were downregulated (Ccl22, Ccr3, Il18rap, Nf1, Ccl11, Atf6ß, Rpn1, and Serp1). Overexpression of FGF2 was detected at 1 month, before rod loss onset, and was maintained at high levels until cone loss (18 months). By 9 months, FGF2 overexpression was seen in photoreceptor cell bodies. Increased glial fibrillary acidic protein (GFAP) expression due to glial cell reactivity followed the same time course. Levels of NGFR/p75NTR remained invariant. Although present in rod outer segments at 1 month, the macrophage chemoattracting chemokine CCL22 became undetectable by 9 months, a likely consequence of progressive rod outer segment truncation. CONCLUSIONS: At a mid-degeneration stage, major changes in gene expression in the ELOVL4 transgenic mouse retina included upregulation of Fgf2 and Fgfr1 and downregulation of Ccl22. Modulation of FGF2 occurred very early, concomitant with an increase in GFAP expression. Future studies will address which factors upstream of Fgf2 could provide potential therapeutic targets to slow photoreceptor degeneration in STGD3.
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Trastornos de los Cromosomas/metabolismo , Degeneración Macular/metabolismo , Envejecimiento/metabolismo , Envejecimiento/fisiología , Animales , Western Blotting , Quimiocina CCL22/metabolismo , Trastornos de los Cromosomas/genética , Cromosomas Humanos Par 6/genética , Cromosomas Humanos Par 6/metabolismo , Modelos Animales de Enfermedad , Factor 2 de Crecimiento de Fibroblastos/metabolismo , Regulación de la Expresión Génica , Proteína Ácida Fibrilar de la Glía/metabolismo , Degeneración Macular/genética , Ratones , Ratones Transgénicos , Reacción en Cadena de la Polimerasa , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/metabolismo , Receptor de Factor de Crecimiento Nervioso/metabolismo , Receptores de Factor de Crecimiento Nervioso/metabolismoRESUMEN
Purpose. To investigate the impact of progressive age-related photoreceptor degeneration on retinal integrity in Stargardt-like macular dystrophy (STGD3). Methods. The structural design of the inner retina of the ELOVL4 transgenic mouse model of STGD3 was compared with that of age-matched littermate wild-type (WT) mice from 1 to 24 months of age by using immunohistofluorescence and confocal microscopy and by relying on antibodies against cell-type-specific markers, synapse-associated proteins, and neurotransmitters. Results. Müller cell reactivity occurred at the earliest age studied, before photoreceptor loss. This finding is perhaps not surprising, considering the cell's ubiquitous roles in retina homeostasis. Second-order neurons displayed salient morphologic changes as a function of photoreceptoral input loss. Age-related sprouting of dendritic fibers from rod bipolar and horizontal cells into the ONL did not occur. In contrast, with the loss of photoreceptor sensory input, these second-order neurons progressively bore fewer synapses. After rod loss, the few remaining cones showed abnormal opsin expression, revealing tortuous branched axons. After complete ONL loss (beyond 18 months of age), localized areas of extreme retinal disruptions were observed in the central retina. RPE cell invasion, dense networks of strongly reactive Müller cell processes, and invagination of axons and blood vessels were distinctive features of these regions. In addition, otherwise unaffected cholinergic amacrine cells displayed severe perturbation of their cell bodies and synaptic plexi in these areas. Conclusions. Remodeling in ELOVL4 transgenic mice follows a pattern similar to that reported after other types of hereditary retinopathies in animals and humans, pointing to a potentially common pathophysiologic mechanism.
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Modelos Animales de Enfermedad , Degeneración Macular/fisiopatología , Células Fotorreceptoras de Vertebrados/patología , Neuronas Retinianas/patología , Envejecimiento/fisiología , Animales , Biomarcadores/metabolismo , Proteínas Portadoras/genética , Electrorretinografía , Proteínas del Ojo/genética , Proteínas del Ojo/metabolismo , Femenino , Técnica del Anticuerpo Fluorescente Indirecta , Genotipo , Degeneración Macular/genética , Degeneración Macular/metabolismo , Masculino , Proteínas de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Microscopía Confocal , Compuestos de Piridinio/metabolismo , Retinoides/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , cis-trans-IsomerasasRESUMEN
PURPOSE: The retina of Arvicanthis niloticus, a diurnal murine rodent closely related to Rattus (rats) and Mus (mice), contains approximately 30% to 35% cones and has several cone-driven functional characteristics found in humans. In this study the organization of these cone photoreceptors was examined, with emphasis on those expressing the S-opsin photopigment (S-cones). METHODS: Cones were labeled with antibodies against M- and S-opsins. Their topographic arrangement was examined on images of retinal flatmounts using density measures, nearest-neighbor distance, and Voronoi domain analysis. Partial sequencing of the S-opsin DNA was also performed to determine whether this visual pigment was blue/violet or UV sensitive. RESULTS: Cone photoreceptors (estimated total population approximately 1.450 million) came in two distinct types that express either M/L- or S-opsin. Both types were present across the retinal surface. S-cones (approximately 7-8% of the total cone population) achieved a higher density in a discrete temporodorsal sector of the retina. The S-cone mosaic was irregular. Finally, S-cones were likely to be UV sensitive, according to genetic analysis. CONCLUSIONS: The topographic arrangement of cone photoreceptors in the retina of the diurnal Nile grass rat A. niloticus represents a highly pertinent model to improve understanding of the pathologic course of and related therapy for retinal disease involving cones.
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Ritmo Circadiano/fisiología , Células Fotorreceptoras Retinianas Conos/citología , Células Fotorreceptoras Retinianas Conos/metabolismo , Opsinas de Bastones/metabolismo , Animales , Western Blotting , Recuento de Células , Femenino , Técnica del Anticuerpo Fluorescente Indirecta , Masculino , Ratas , Transducina/metabolismoRESUMEN
The most widespread models to study blindness, rats and mice, have retinas containing less than 3% cones. The diurnal rodent Arvicanthis niloticus retina has around 35% cones. Using ERG recordings, we studied retina function in this species. Several features differed from that reported in rats and mice: (a) fivefold larger photopic a-wave amplitudes; (b) photopic hill effect in Nile grass rats only; and (c) flicker amplitude plateau between 5 to 35 Hz with fusion beyond 60 Hz in Nile grass rats only. We conclude that A. niloticus might complement rats and mice for studying retinal function and pathologies involving cones.
Asunto(s)
Ritmo Circadiano/fisiología , Muridae/fisiología , Células Fotorreceptoras Retinianas Conos/fisiología , Adulto , Animales , Adaptación a la Oscuridad/fisiología , Modelos Animales de Enfermedad , Electrorretinografía , Femenino , Humanos , Masculino , Persona de Mediana Edad , Muridae/anatomía & histología , Estimulación Luminosa/métodos , Células Fotorreceptoras Retinianas Conos/citología , Umbral Sensorial/fisiología , Especificidad de la Especie , Adulto JovenRESUMEN
Unlike laboratory rats and mice, muridae of the Arvicanthis family (A. ansorgei and A. niloticus) are adapted to functioning best in daylight. To date, they have been used as experimental models mainly in studies of circadian rhythms. However, recent work aimed at optimizing photoreceptor-directed gene delivery vectors (Khani et al. [2007] Invest Ophthalmol Vis Sci 48:3954-3961) suggests their potential usefulness for studying retinal pathologies and therapies. In the present study we analyzed the retinal anatomy and visual performance of the Nile grass rat (A. niloticus) using immunohistofluorescence and the optokinetic response (OKR). We found that approximately 35-40% of photoreceptors are cones; that many neural features of the inner retina are similar to those in other diurnal mammals; and that spatial acuity, measured by the OKR, is more than two times that of the usual laboratory rodents. These observations are consistent with the known diurnal habits of this animal, and further support its pertinence as a complementary model for studies of structure, function, and pathology in cone-rich mammalian retinae.
Asunto(s)
Muridae , Retina/anatomía & histología , Células Fotorreceptoras Retinianas Conos/fisiología , Agudeza Visual/fisiología , Animales , Conducta Animal/fisiología , Biomarcadores/metabolismo , Femenino , Masculino , Muridae/anatomía & histología , Muridae/fisiología , Retina/fisiología , Células Fotorreceptoras Retinianas Conos/citologíaRESUMEN
Necdin and Magel2 are related proteins inactivated in Prader-Willi syndrome (PWS), a sporadic chromosomal deletion disorder. We demonstrate that necdin and Magel2 bind to and prevent proteasomal degradation of Fez1, a fasciculation and elongation protein implicated in axonal outgrowth and kinesin-mediated transport, and also bind to the Bardet-Biedl syndrome (BBS) protein BBS4 in co-transfected cells. The interactions among necdin, Magel2, Fez1 and BBS4 occur at or near centrosomes. Centrosomal or pericentriolar dysfunction has previously been implicated in BBS and may also be important in the features of PWS that overlap with BBS, such as learning disabilities, hypogonadism and obesity. Morphological abnormalities in axonal outgrowth and fasciculation manifest in several regions of the nervous system in necdin null mouse embryos, including axons of sympathetic, retinal ganglion cell, serotonergic and catecholaminergic neurons. These data demonstrate that necdin mediates intracellular processes essential for neurite outgrowth and that loss of necdin impinges on axonal outgrowth. We further suggest that loss of necdin contributes to the neurological phenotype of PWS, and raise the possibility that co-deletion of necdin and the related protein Magel2 may explain the lack of single gene mutations in PWS.
Asunto(s)
Axones/metabolismo , Proteínas del Tejido Nervioso/genética , Proteínas Nucleares/genética , Síndrome de Prader-Willi/genética , Animales , Antígenos de Neoplasias , Centrosoma/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Humanos , Inmunohistoquímica , Ratones , Proteínas Asociadas a Microtúbulos , Proteínas del Tejido Nervioso/metabolismo , Proteínas Nucleares/metabolismo , Síndrome de Prader-Willi/metabolismo , Proteínas/genética , Proteínas/metabolismo , Técnicas del Sistema de Dos Híbridos , Tirosina 3-Monooxigenasa/metabolismoRESUMEN
Pulsed-field gel electrophoresis and gene typing were able to differentiate among 3,597 Bordetella pertussis isolates circulating in Alberta and Québec Provinces, Canada, from 1985 to 1994 and distinguish them from the strains used in vaccine production. This study provides a baseline for continued surveillance of prevalent and emerging strains of B. pertussis in Canada.