Dataset Information

A scoring system for the evaluation of the mutated Crb1/rd8-derived retinal lesions in C57BL/6N mice.

ABSTRACT: As part of the International Mouse Phenotyping Consortium (IMPC) programme, the MRC Harwell is conducting a large eye morphology phenotyping screen on genetically modified mice compared to the baseline phenotype observed in the background strain of C57BL/6NTac. The C57BL/6NTac strain is known to carry a spontaneous mutation in the Crb1 gene that causes retinal degeneration characterized by the presence of white spots (flecks) in the fundus. These flecks potentially represent a confounding factor, masking similar retinal phenotype abnormalities that may be detected in mutants. Therefore we investigated the frequency, position and extent of the flecks in a large population of C57BL/6NTac mice to provide the basis for evaluating the presence of flecks in mutant mice with the same genetic background. We found that in our facility males were more severely affected than females and that in both males and females the most common localisation of the flecks was in the inferior hemicycle of the fundus.

SUBMITTER: Concas D

PROVIDER: S-EPMC5580417 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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A scoring system for the evaluation of the mutated <i>Crb1/rd8</i>-derived retinal lesions in C57BL/6N mice.

Concas Danilo D Cater Heather H Wells Sara S

F1000Research 20170331

As part of the International Mouse Phenotyping Consortium (IMPC) programme, the MRC Harwell is conducting a large eye morphology phenotyping screen on genetically modified mice compared to the baseline phenotype observed in the background strain of C57BL/6NTac. The C57BL/6NTac strain is known to carry a spontaneous mutation in the <i>Crb1</i> gene that causes retinal degeneration characterized by the presence of white spots (flecks) in the fundus. These flecks potentially represent a confounding ...[more]

PMID: 28928942

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Project description:Mutations in crumb homologue 1 (CRB1) in humans are associated with Leber's congenital amaurosis (LCA) and retinitis pigmentosa (RP). There is no clear genotype-phenotype correlation for human CRB1 mutations in RP and LCA. The high variability in clinical features observed in CRB1 mutations suggests that environmental factors or genetic modifiers influence severity of CRB1 related retinopathies. Retinal degeneration 8 (rd8) is a spontaneous mutation in the Crb1 gene (Crb1(rdr/rd8)). Crb1(rdr/rd8) mice present with focal disruption in the outer retina manifesting as white spots on fundus examination. Mild retinal dysfunction with decreased b-wave amplitude has been reported in Crb1(rdr/rd8) mice at 18 months. Methylene tetrahydrofolate reductase (MTHFR) is a crucial enzyme of homocysteine metabolism. MTHFR mutations are prevalent in humans and are linked to a broad spectrum of disorders including cardiovascular and neurodegenerative diseases. We recently reported the retinal phenotype in Mthfr-deficient (Mthfr(+/-)) heterozygous mice. At 24 weeks the mice showed decreased RGC function, thinner nerve fiber layer, focal areas of vascular leakage and 20% fewer cells in the ganglion cell layer (GCL). Considering the variability in CRB1-related retinopathies and the high occurrence of human MTHFR mutations we evaluated whether Mthfr deficiency influences rd8 retinal phenotype. Mthfr heterozygous mice with rd8 mutations (Mthfr(+/-)(rd8/rd8)) and Crb(rd8/rd8) mice (Mthfr(+/+rd8/rd8)) mice were subjected to comprehensive retinal evaluation using ERG, fundoscopy, fluorescein angiography (FA), morphometric and retinal flat mount immunostaining analyses of isolectin-B4 at 8-54 wks. Assessment of retinal function revealed a significant decrease in the a-, b- and c-wave amplitudes in Mthfr(+/-)(rd8/rd8) mice at 52 wks. Fundoscopic evaluation demonstrated the presence of signature rd8 spots in Mthfr(+/+rd8/rd8) mice and an increase in the extent of these rd8 spots in Mthfr(+/-)(rd8/rd8) mice at 24 weeks and beyond. FA revealed marked vascular leakage, ischemia and vascular tortuosity in Mthfr(+/-)(rd8/rd8) mice at 24 and 52 weeks. Retinal dysplasia was observed in ?14-33% Mthfr(+/-)(rd8/rd8) mice by morphometric analysis. This was accompanied by a ?20% reduction in cells of the GCL of Mthfr(+/-)(rd8/rd8) mice at 24 and 52 weeks. Retinal flat mount immunostaining with isolectin-B4 showed neovascularization and loss of blood vessel integrity in Mthfr(+/-)(rd8/rd8) mice in contrast to mild vasculopathy in Mthfr(+/+rd8/rd8) mice. Taken together, our data support an earlier onset and worsened retinal phenotype when Mthfr and rd8 mutations coexist. Our study sets the stage for future studies to investigate the role of MTHFR deficiency in human CRB1 retinopathies.

Project description:PURPOSE:The purpose of this study is to determine the effect of Cytoglobin (Cygb) deficiency on Crb1-related retinopathy. The Crb1 cell polarity complex is required for photoreceptor function and survival. Crb1-related retinopathies encompass a broad range of phenotypes which are not completely explained by the variability of Crb1 mutations. Genes thought to modify Crb1 function are therefore important targets of research. The biological function of Cygb involves oxygen delivery, scavenging of reactive oxygen species, and nitric oxide metabolism. However, the relationship of Cygb to diseases involving the Crb1 cell polarity complex is unknown. METHODS:Cygb knockout mice homozygous for the rd8 mutation (Cygb-/-rd8/rd8) were screened for ocular abnormalities and imaged using optical coherence tomography and fundus photography. Electroretinography was performed, as was histology and immunohistochemistry. Quantitative PCR was used to determine the effect of Cygb deficiency on transcription of Crb1 related cell polarity genes. RESULTS:Cygb-/-rd8/rd8 mice develop an abnormal retina with severe lamination abnormalities. The retina undergoes progressive degeneration with the ventral retina more severely affected than the dorsal retina. Cygb expression is in neurons of the retinal ganglion cell layer and inner nuclear layer. Immunohistochemical studies suggest that cell death predominates in the photoreceptors. Electroretinography amplitudes show reduced a- and b-waves, consistent with photoreceptor disease. Cygb deficient retinas had only modest transcriptional perturbations of Crb1-related cell polarity genes. Cygb-/- mice without the rd8 mutation did not exhibit obvious retinal abnormalities. CONCLUSIONS:Cygb is necessary for retinal lamination, maintenance of cell polarity, and photoreceptor survival in rd8 mice. These results are consistent with Cygb as a disease modifying gene in Crb1-related retinopathy. Further studies are necessary to investigate the role of Cygb in the human retina.

Dataset Information

A scoring system for the evaluation of the mutated Crb1/rd8-derived retinal lesions in C57BL/6N mice.

Publications

A scoring system for the evaluation of the mutated <i>Crb1/rd8</i>-derived retinal lesions in C57BL/6N mice.

Similar Datasets

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